Insulin and Cancer – biblio growing and growing

Study protocol: Insulin and its role in cancer.

Harish K, Dharmalingam M, Himanshu M.

ABSTRACT: BACKGROUND: Studies have shown that metabolic syndrome and its consequent biochemical derangements in the various phases of diabetes may contribute to carcinogenesis. A part of this carcinogenic effect could be attributed to hyperinsulinism. High levels of insulin decrease the production of IGF-1 binding proteins and hence increase levels of free IGF-1. It is well established that bioactivity of free insulin growth factor 1 (IGF-1) increases tumor turnover rate. The objective is to investigate the role of insulin resistance/sensitivity in carcinogenesis by studying the relation between insulin resistance/sensitivity and IGF-1 levels in cancer patients. We postulate that hyperinsulinaemia which prevails during initial phases of insulin resistance (condition prior to overt diabetes) increases bioactivity of free IGF-1, which may contribute to process of carcinogenesis. Methods / Design: Based on our pilot study results and power analysis of the same, we have designed a two group case-control study. 800 proven untreated cancer patients (solid epithelial cell tumors) under age of 50 shall be recruited with 200 healthy subjects serving as controls. Insulin resistance/sensitivity and free IGF-1 levels shall be determined in all subjects. Association between the two parameters shall be tested using suitable statistical methods.

DISCUSSION: Well controlled studies in humans are essential to study the link between insulin resistance, hyperinsulinaemia, IGF-1 and carcinogenesis. This study could provide insights to the role of insulin, insulin resistance, IGF-1 in carcinogenesis although a precise role and the extent of influence cannot be determined. In future, cancer prevention and treatment strategies could revolve around insulin and insulin resistance.

Ann Surg Oncol. 1998 Mar;5(2):194-201.Links

Altered serum levels of insulin-like growth-factor binding proteins in breast cancer patients.

Ng EH, Ji CY, Tan PH, Lin V, Soo KC, Lee KO.

Department of Surgery, Singapore General Hospital, Republic of Singapore.

BACKGROUND: Insulin-like growth factor 1 (IGF-1) has mitogenic properties for breast cancer cell lines and has been proposed to be an important factor in breast carcinogenesis. We hypothesized that differences in IGF-1 or its binding proteins might increase susceptibility to breast cancer. This case-control study was designed to investigate whether patients with breast cancer have altered levels of either IGF-1 or its intermediary modulatory proteins, the IGF binding proteins (BP). METHODS: Serum was collected from 90 patients (63 with breast cancer and 27 with benign breast disease) after an overnight fast and before surgery. IGF-1, BP1, and BP3 levels were determined by immunoradiometric assays. In a subset of 66 patients, Western ligand blots were also performed for a semiquantitative measurement of functioning BP levels. A forward stepwise logistic regression model to adjust for other confounding variables (age, menopausal status, parity, age at menarche, use of oral contraceptives, history of breast biopsy, family history of breast cancer, hormone replacement therapy, and body-mass index) was used in the multivariate analysis.

RESULTS: Serum IGF-1 levels were similar in cases and controls. However, levels of BP3 (p < 0.001), BP4 (p < 0.01), and BP1 (p < 0.05) were significantly associated with risk of breast cancer. The level of BP3 was the most significant factor predictive of breast cancer. The odds ratio for breast cancer in women with BP3 levels >2066 ng/ml was 0.18 (95% CI, 0.05-0.55). Correspondingly, women with BP1 levels higher than 39 ng/ml had an odds ratio of 0.21 (95% CI, 0.07-0.68) for breast cancer. When considering only cancer patients (n = 63), decreasing levels of BP4 (p < 0.01) and increasing levels of BP1 (p < 0.02) were significantly associated with progesterone receptor positivity (PR+) in the tumor. The odds ratio of PR+ in patients with BP1 levels higher than 34 ng/ml was 7.49 (95% CI, 1.5-37.4). Better grade of tumor (well and moderately differentiated) was observed in patients with higher levels of BP3 (p < 0.03).

CONCLUSIONS: Distinct differences in BP profiles exist among patients with breast cancer and also among those with high-grade, hormonal receptor-negative tumors. These findings suggest that the bioavailability of IGF-1 as mediated by its binding proteins may participate in both breast carcinogenesis and selection of more aggressive breast carcinomas.

 

added 12-07

INSULIN RECEPTORS AND CANCER

1: J Immunol  2002 Jun 15;168(12):6215-23 

Over-expression of insulin receptor substrate-1, but not insulin receptor substrate-2, protects a T cell hybridoma from activation-induced cell death.

Li L, Qi X, Williams M, Shi Y, Keegan AD.

Department of Immunology, Jerome Holland Laboratories, American Red Cross, Rockville, MD 20852, USA.

The insulin receptor substrate (IRS) family of signaling molecules is expressed in lymphocytes, although their functions in these cells is largely unknown. To investigate the role of IRS in the protection of T cells from activation-induced cell death (AICD), we transfected the T cell hybridoma A1.1, which is IL-4 responsive but lacks expression of IRS family members with cDNA encoding IRS1 or IRS2. Stimulation of these clones with immobilized anti-CD3-induced expression of CD69 to the same level as the parental A1.1 cells. However, the A1.1 IRS1-expressing cells were markedly resistant to AICD, while the A1.1

IRS2-expressing cells were not. Inhibition of phosphatidylinositol 3'-kinase in

the A1.1 IRS1-expressing cells did not abrogate their resistance to AICD. Fas

mRNA was induced similarly by anti-CD3 in A1.1, A1.1 IRS1-expressing, and A1.1

IRS2-expressing cells. However, induction of Fas ligand (FasL) mRNA and

functional FasL protein was delayed and decreased in IRS1-expressing cells, but

not in IRS2-expressing cells. The induction of transcription from a 500-bp FasL

promoter and a minimal 16-mer early growth response element linked to luciferase

was also impaired in the IRS1-expressing cells. These results suggest that

overexpression of IRS1, but not IRS2, protects A1.1 cells from AICD by

diminishing FasL transcription through a pathway that is independent of the

tyrosine phosphorylation of IRS1 and phosphatidylinositol 3'-kinase activity.

2: Int J Obes Relat Metab Disord  2002 Jun;26(6):747-53 

Upper abdominal obesity, insulin resistance and breast cancer risk.

Stoll BA.

Oncology Department, St Thomas' Hospital, London, UK.

PURPOSE: A majority of prospective studies show breast cancer risk to be higher

in obese postmenopausal women with upper abdominal adiposity than in those with

overall adiposity. The evidence is more limited and inconsistent in the case of

premenopausal women. The review examines evidence that aberrant insulin

signalling may be involved in the promotion of mammary carcinogenesis. The

aetiology and concomitants of abdominal visceral obesity are examined.

MECHANISMS: Clinical and experimental evidence suggests that the higher breast

cancer risk associated with greater abdominal visceral obesity may be related to

aberrant insulin signalling through the insulin receptor substrate 1 pathway,

leading to insulin resistance, hyperinsulinaemia and increased concentrations of

endogenous oestrogen and androgen. The putative role of aberrant insulin

signalling in the promotion of mammary carcinogenesis may help to explain

clinical relationships between breast cancer risk and age at menarche,

pregnancies and onset of obesity. CONCLUSION: Overall adiposity in women

adversely affects breast cancer risk mainly by greater exposure of mammary

epithelial tissue to endogenous oestrogen. Upper abdominal adiposity appears to

involve an additional effect related to the presence of insulin resistance.

Aetiological factors in the development of hyperinsulinaemic insulin resistance

are still uncertain but may involve aberrant susceptibility genes in adipocyte

insulin receptors or in the insulin receptor substrate 1 pathway. Epigenetic

factors are also likely to contribute, including high free fatty acid levels and

obesity. Dietary fatty acids, particularly polyunsaturated fatty acids, are

known to regulate adipocyte differentiation through the nuclear peroxisome

proliferator-activated receptor gamma, and may also have a role in insulin

resistance. These aetiological factors are likely to be relevant to the high

risk of postmenopausal breast cancer in industrialised Western populations.

3: Am J Med Sci  2002 Mar;323(3):140-5 

Insulin: a novel factor in carcinogenesis.

Gupta K, Krishnaswamy G, Karnad A, Peiris AN.

Department of Internal Medicine, Mountain Home VA Medical Center and East

Tennessee State University, Johnson City, Tennessee, USA.

Cancer is a leading cause of mortality in the United States. Despite much

research on specific carcinogens, the cause of many cancers remains unclear. The

identification of novel causative agents offers the potential for cancer

prevention. Diseases such as obesity and diabetes mellitus, characterized by

hyperinsulinemia, are associated with increased risk of endometrial, colorectal,

and breast carcinomas. There is increasing evidence that insulin is a growth

factor for tumor formation. The mechanisms underlying insulin-mediated neoplasia

may include enhanced DNA synthesis with resultant tumor cell growth, inhibition

of apoptosis, and altered sex hormone milieu. The reduced insulin levels seen

with physical activity, weight loss, and a high fiber diet may account for

decreased cancer risk. The role of newer drugs that restore sensitivity to

insulin, thereby reducing hyperinsulinemia, is an exciting potential area of

cancer prevention. In this review, we discuss the potential role of insulin as a

tumor growth factor.

4: J Clin Endocrinol Metab  2002 Jan;87(1):245-54 

A novel autocrine loop involving IGF-II and the insulin receptor isoform-A

stimulates growth of thyroid cancer.

Vella V, Pandini G, Sciacca L, Mineo R, Vigneri R, Pezzino V, Belfiore A.

Istituto di Medicina Interna e di Malattie Endocrine e del Metabolismo, Cattedra

di Endocrinologia, University of Catania, Ospedale Garibaldi, 95123 Catania,

Italy.

The insulin receptor (IR) occurs in two isoforms (IR-A and IR-B) resulting from

alternative splicing of exon 11 of the gene. The IR-A isoform is predominantly

expressed in fetal tissues and malignant cells and binds IGF-II with high

affinity. We previously observed that IRs are overexpressed in thyroid cancer

cells; now we evaluated whether these cells preferentially express IR-A and

produce IGF-II, which would activate a growth-promoting autocrine loop. The IR

content ranged 6.0-52.6 ng/100 microg cell membrane protein in thyroid cancer

primary cultures (n = 8) and permanent cell lines (n = 6) vs. 1.2-1.7 in normal

thyroid cells (n = 11 primary cultures; P < 0.0001). IR-A isoform relative

abundance ranged from 36-79% in cancer cells (with the highest values in

undifferentiated cancers) vs. 27-39% in normal cells. Similar results were

obtained in normal vs. cancer thyroid tissue specimens. IGF-II caused IR

autophosphorylation with an ED(50) of 1.5-40.0 nM in cancer cells vs. more than

100 nM in normal cells; IGF-II affinity correlated with the relative abundance

of IR-A (r = 0.628; P < 0.0001). IGF-II was expressed in all cancer cells,

highly expressed in anaplastic cells, and less expressed in normal cells. In

conclusion, malignant thyrocytes, especially when poorly differentiated, produce

IGF-II and overexpress IR, predominantly as IGF-II-sensitive isoform A. A

growth-promoting autocrine loop is activated, therefore, and may affect thyroid

cancer biology.

5: Diabetes Metab  2001 Sep;27(4 Pt 2):S7-12 

[Resistance to insulin and polycystic ovary syndrome]

[Article in French]

Ducluzeau PH, Laville M, Vidal H, Pugeat M.

Federation d'Endocrinologie, Hopital de l'Antiquaille, Hospices Civils de Lyon.

michel.pugeat@chu-lyon.fr

6: Brain Res Mol Brain Res  2001 Dec 30;97(2):161-70 

Altered insulin receptor processing and function in scrapie-infected

neuroblastoma cell lines.

Ostlund P, Lindegren H, Pettersson C, Bedecs K.

Department of Neurochemistry and Neurotoxicology, University of Stockholm,

Svante Arrhenius v. 21A, S-10691 Stockholm, Sweden.

The underlying neurochemical changes contributing to prion-induced

neurodegeneration remain largely unknown. This study shows that scrapie

infection induced a 2-fold increase of insulin receptor (IR) protein and

aberrantly processed IR beta-chain in scrapie-infected N2a neuroblastoma cells

(ScN2a) as measured by Western blot of immunoprecipitated IR, in the absence of

increased IR mRNA. Elevated IR protein level was further confirmed in an

independently scrapie-infected neuroblastoma cell line N1E-115 (ScN1E-115).

Proliferation studies showed that the increased IR level in ScN2a did not result

in an increased insulin-mediated cell growth compared to normal N2a cells.

Binding studies indicated that this apparent paradox was due to a 65% decrease

in specific [(125)I]insulin binding sites in ScN2a when compared to the amount

of immunoreactive IR, although the IR binding affinity was unchanged. Analysis

of insulin stimulated IR tyrosine phosphorylation showed a slight but not

significant reduction in ScN2a, when related to the increased level of

immunoreactive IR. However, comparing the IR tyrosine phosphorylation to the

loss of binding sites in ScN2a, we demonstrated an increased IR tyrosine

phosphorylation of the remaining functional IR. In addition to these differences

in IR properties, the basal extracellular signal regulated kinase-2 (ERK2)

phosphorylation detected by Western blot, was significantly elevated and the

insulin stimulated ERK2 phosphorylation was subsequently decreased in ScN2a.

Together, these data show that scrapie infection affects the level and

processing of the IR and signal transduction mediated by the IR in neuroblastoma

cells, as well as induces an elevated basal ERK2 phosphorylation. Aberrant

regulation of neuroprotective receptors may contribute to neurodegeneration in

prion diseases.

7: J Endocrinol  2001 Nov;171(2):285-92 

Expression of insulin target genes in skeletal muscle and adipose tissue in

adult patients with growth hormone deficiency: effect of one year recombinant

human growth hormone therapy.

Khalfallah Y, Sassolas G, Borson-Chazot F, Vega N, Vidal H.

INSERM U.449, Rene Laennec Faculty of Medicine, Lyon, France.

Our aim was to investigate the effects of one year recombinant human growth

hormone (rhGH) therapy on the regulation by insulin of gene expression in muscle

and adipose tissue in adults with secondary GH deficiency (GHD). Six GHD

subjects without upper-body obesity were submitted to a 3-h euglycemic

hyperinsulinemic clamp before and after one year of rhGH therapy. Muscle and

abdominal subcutaneous adipose tissue biopsies were taken before and at the end

of each clamp. The mRNA levels of insulin receptor, p85

alpha-phosphatidylinositol-3 kinase (p85 alpha PI-3K), insulin dependent glucose

transporter (Glut4), hexokinase II, glycogen synthase, lipoprotein lipase (LPL)

in muscle and in adipose tissue, hormone sensitive lipase and peroxisome

proliferator-activated receptor gamma (PPAR gamma) in adipose tissue were

quantified by RT-competitive PCR. One year treatment with rhGH (1.25 IU/day)

increased plasma IGF-I concentrations (54+/-7 vs 154+/-11 ng/ml, P<0.01) but did

not affect insulin-stimulated glucose disposal rate measured during the

hyperinsulinemic clamp (74+/-9 vs 85+/-5 micromol/kg free fat mass/min). Insulin

significantly increased p85 alpha PI-3K, hexokinase II and Glut4 mRNA levels in

muscle both before and after rhGH treatment. One year of GH therapy increased

LPL mRNA levels in muscle (38+/-2 vs 70+/-7 amol/microg total RNA, P<0.05) and

in adipose tissue (2490+/-260 vs 4860+/-880 amol/microg total RNA, P<0.05), but

did not change the expression of the other mRNAs. We conclude from this study

that GH therapy did not alter whole body insulin sensitivity and the response of

gene expression to insulin in skeletal muscle of adult GHD patients, but it did

increase LPL expression in muscle and adipose tissue. This result could be

related to the documented beneficial effect of GH therapy on lipid metabolism.

8: J Biol Chem  2001 Oct 26;276(43):40362-7 

Regulation of insulin/insulin-like growth factor-1 signaling by

proteasome-mediated degradation of insulin receptor substrate-2.

Rui L, Fisher TL, Thomas J, White MF.

Howard Hughes Medical Institute, Joslin Diabetes Center, Harvard Medical School,

One Joslin Place, Boston, MA 02215, USA.

Insulin and insulin-like growth factor-1 (IGF-1) regulate metabolism and body

growth through homologous receptor tyrosine kinases that phosphorylate the

insulin receptor substrate (IRS) proteins. IRS-2 is an important IRS protein, as

it mediates peripheral insulin action and beta-cell survival. In this study, we

show that insulin, IGF-1, or osmotic stress promoted

ubiquitin/proteasome-mediated degradation of IRS-2 in 3T3-L1 cells, Fao

hepatoma, cells and mouse embryo fibroblasts; however, insulin/IGF-1 did not

promote degradation of IRS-1 in 3T3-L1 preadipocytes or mouse embryo

fibroblasts. MG132 or lactacystin, specific inhibitors of 26S proteasome,

blocked insulin/IGF-1-induced degradation of IRS-2 and enhanced the detection of

ubiquitinated IRS-2. Insulin/IGF1-induced ubiquitination and degradation of

IRS-2 was blocked by inhibitors of phosphatidylinositol 3-kinase (wortmannin or

LY294002) or mTOR (rapamycin). Chronic insulin or IGF-1 treatment of

IRS-1-deficient mouse embryo fibroblasts inhibited IRS-2-mediated activation of

Akt and ERK1/2, which was reversed by lactacystin pretreatment. By contrast,

IRS-1 activation of Akt and ERK1/2 was not inhibited by chronic insulin/IGF-1

stimulation in IRS-2-deficient mouse embryo fibroblasts. Thus, we identified a

novel negative feedback mechanism by which the ubiquitin/proteasome-mediated

degradation of IRS-2 limits the magnitude and duration of the response to

insulin or IGF-1.

9: Physiologist  1993 Feb;36(1 Suppl):S160-1 

Enhancement of insulin receptor mediated endocytosis in cultured osteoblastic

cells under hypergravity.

Malouvier A, Holy X, Zerath E, Marie PJ, Schmitt DA.

CERMA, Division Histophysiologie, Centre d'Essais en Vol, Bretigny Sur Orge,

France.

10: Clin Endocrinol (Oxf)  2001 Aug;55(2):191-9 

Tissue insulin sensitivity and body weight in polycystic ovary syndrome.

Marsden PJ, Murdoch AP, Taylor R.

Department of Obstetrics and Gynaecology, University Hospital of North Durham,

Durham, UK. Philippa_Marsden@hotmail.com

OBJECTIVE: Polycystic ovarian syndrome (PCOS) and obesity both affect insulin

sensitivity. This study was designed to investigate the biochemical indices of

PCOS and tissue insulin sensitivity in groups of lean and obese women with

clinically equivalent degrees of the syndrome, relative to control subjects.

DESIGN: A prospective study of in vivo parameters and in vitro study of

adipocytes to assess insulin sensitivity. PATIENTS: Six lean and 14 overweight

patients fulfilling formal diagnostic criteria for PCOS were studied. The degree

of hirsutism and amenorrhoea was similar in each group. Eight control subjects

were also studied. MEASUREMENTS: Endocrine and metabolic parameters were

measured in lean and overweight patients with PCOS and control subjects. In

vitro studies of adipocyte insulin receptor binding and adipocyte insulin action

were performed. RESULTS: The mean plasma LH level was elevated in both groups of

PCOS but was significantly higher in the lean group (LH levels were 25.1 +/- 3.1

and 14.5 +/- 1.6 iu/l in lean PCOS and obese PCOS, respectively (P = 0.01)).

There was a strong inverse correlation between BMI and LH levels (R = - 0.70, P

= 0.001). Fasting insulin levels were elevated in both lean and obese groups

(11.5 +/- 2.8 and 26.8 +/- 8.1 mU/l, respectively; P = 0.068). Mean serum

testosterone and serum androstenedione levels were also elevated in PCOS

compared to control subjects but there was no difference between the two groups

of PCOS subjects. Insulin receptor binding in amenorrhoeic subjects with PCOS

was low in both lean and obese patients with PCOS but was not significantly

different between the two groups (0.79 +/- 0.17% and 0.66 +/- 0.07% per 10 cm2

cell membrane, respectively). Maximally insulin-stimulated rates of

3-O-methylglucose transport were low in both groups compared to previously

studied normal subjects (0.96 +/- 0.21 and 0.64 +/- 0.07 pmol per 10 cm2

membrane in lean and obese PCOS subjects, respectively; P = NS). CONCLUSIONS:

Lean subjects with a given phenotypic expression of PCOS have an equivalent

degree of tissue insulin resistance compared to obese PCOS subjects. This

implies that the insulin resistance may be a primary feature of PCOS. If this is

so, a similar clinical degree of the syndrome may be brought about by

genetically determined insulin resistance in lean subjects or by insulin

resistance which is secondary to obesity.

PMID: 11531925 [PubMed - indexed for MEDLINE]

11: Med Hypotheses  2001 Aug;57(2):146-50 

Insulin secretion as a determinant of pancreatic cancer risk.

McCarty MF.

Pantox Laboratories, 4622 Santa Fe St, San Diego, CA 92109, USA.

New epidemiology confirms that glucose intolerance is a risk factor for

pancreatic cancer, and that this association cannot be accounted for by an

adverse impact of early pancreatic cancer on beta cell function. Previous

reports indicate that risk for pancreatic cancer is increased in adult-onset

diabetics. Since streptozotocin diabetes inhibits carcinogen-mediated induction

of pancreatic cancer in hamsters, the most reasonable interpretation of these

findings is that insulin (or some other beta cell product) acts as a promoter

for pancreatic carcinogenesis. This view is consistent with a report that human

pancreatic adenocarcinomas express insulin receptors that can stimulate mitosis;

an additional possibility is that high insulin levels indirectly promote

pancreatic carcinogenesis by boosting effective IGF-I activity via hepatic

actions. In international ecologic epidemiology, pancreatic cancer rates

correlate tightly with dietary intake of animal products; this may reflect the

fact that vegan diets are associated with low diurnal insulin secretion. There

is also suggestive evidence that macrobiotic vegan diets, which are low in

glycemic index, may increase mean survival time in pancreatic cancer. However,

other types of diets associated with decreased postprandial insulin response,

such as high-protein diets or 'Mediterranean' diets high in oleic acid, may also

have the potential for pancreatic cancer prevention. The huge increases of

age-adjusted pancreatic cancer mortality in Japan and among African-Americans

during the last century imply that pancreatic cancer is substantially

preventable; a low-insulin-response diet coupled with exercise training, weight

control, and smoking avoidance, commendable for a great many other reasons, may

slash pancreatic cancer mortality dramatically. Copyright 2001 Harcourt

Publishers Ltd.

PMID: 11461162 [PubMed - indexed for MEDLINE]

12: Eur J Endocrinol  2001 Aug;145(2):213-21 

Effects of IGF-I and -II, IGF binding protein-3 (IGFBP-3), and transforming

growth factor-beta (TGF-beta) on growth and apoptosis of human osteosarcoma

Saos-2/B-10 cells: lack of IGF-independent IGFBP-3 effects.

Schmid C, Ghirlanda-Keller C, Zapf J.

Division of Endocrinology and Diabetes, Department of Medicine, University

Hospital, 8091 Zurich, Switzerland. ndozaj@usz.unizh.ch

OBJECTIVE: Insulin-like growth factor binding protein-3 (IGFBP-3) inhibits cell

growth. Previous reports have suggested the existence of plasma membrane IGFBP-3

receptors that could mediate direct, IGF-independent effects. Thus far, however,

the only well-defined putative IGFBP-3 receptor is the type V transforming

growth factor-beta (TGF-beta) receptor, a membrane glycoprotein that mediates

TGF-beta-induced growth inhibition in selected cells. The aim of the study was

to test whether IGFBP-3 and TGF-beta exert short-term effects in an osteosarcoma

cell line that produces no IGF but contains type 1 IGF receptors. DESIGN: DNA

synthesis and apoptosis in Saos-2/B-10 cells were measured in response to IGF-I,

IGF-II, IGFBP-3 and TGF-beta2, and to type 1 IGF receptor ligands with poor

affinity for IGFBP-3 ([QAYL]-IGF-I and insulin). RESULTS: IGF-I and IGF-II

stimulated thymidine incorporation into DNA and suppressed apoptosis in a

dose-dependent manner with maximal effects at 1 and 3 nM respectively. TGF-beta2

slightly increased thymidine incorporation into DNA but had no effect on

apoptosis. IGFBP-3 had no effect by itself. Whereas it blocked the above effects

of 1 nmol/l IGF-I, it did not block those of 1 nmol/l [QAYL]-IGF-I or 100 nmol/l

insulin. CONCLUSIONS: IGFBP-3 does not affect DNA synthesis or apoptosis in an

IGF-independent manner in IGF-responsive osteosarcoma cells. It therefore

appears to act essentially by sequestration of IGF.

PMID: 11454519 [PubMed - indexed for MEDLINE]

13: Am J Physiol Endocrinol Metab  2001 Aug;281(2):E392-9 

Defects in insulin receptor signaling in vivo in the polycystic ovary syndrome

(PCOS).

Dunaif A, Wu X, Lee A, Diamanti-Kandarakis E.

Division of Women's Health, Department of Medicine, Brigham and Women's

Hospital, Boston, Massachusetts 02115, USA.

Women with polycystic ovary syndrome (PCOS) are insulin resistant secondary to a

postbinding defect in insulin signaling. Sequential euglycemic glucose clamp

studies at 40 and 400 mU. m(-2). min(-1) insulin doses with serial skeletal

muscle biopsies were performed in PCOS and age-, weight-, and ethnicity-matched

control women. Steady-state insulin levels did not differ, but insulin-mediated

glucose disposal was significantly decreased in PCOS women (P < 0.05). Insulin

receptor substrate (IRS)-1-associated phosphatidylinositol 3-kinase (PI 3K)

activity was significantly decreased in PCOS (n = 12) compared with control

skeletal muscle (n = 8; P < 0.05). There was no significant difference in the

abundance of IR, IRS-1, or the p85 regulatory subunit of PI 3K in PCOS (n = 14)

compared with control (n = 12) muscle. The abundance of IRS-2 was significantly

increased (P < 0.05) in PCOS skeletal muscle, suggesting a compensatory change.

We conclude that there is a physiologically relevant defect in insulin receptor

signaling in PCOS that is independent of obesity and type 2 diabetes mellitus.

Publication Types:

Clinical Trial

Controlled Clinical Trial

PMID: 11440917 [PubMed - indexed for MEDLINE]

14: Mol Cell Biol Res Commun  2000 Oct;4(4):234-8 

Complexes formation between insulin receptor and extracellular signal-regulated

kinases ERKs.

Lin YL, Mettling C, Chou CK.

Institut de Genetique Humaine, Centre National de la Recherche Scientifique, 141

rue de la Cardonille, Montpellier Cedex 5, 34396, France

A property of signal transduction pathways that might explain their efficiency

and specificity is the formation of signaling complexes. The recent

demonstration that adaptor proteins can interact with many components of the

extracellular signal-regulated kinases (ERKs) signaling cascade leads us to

investigate whether such complexes may include the transmembrane receptor. The

present work shows that in human hepatoma Hep3B cells, insulin receptor (IR) can

be coimmunoprecipitated with other components of the ERKs cascade: insulin

receptor substrate (IRS), Raf-1, and ERKs. Furthermore, these complexes formed

near the cytoplasmic membrane even prior to insulin stimulation. Copyright 2001

Academic Press.

PMID: 11409918 [PubMed - indexed for MEDLINE]

15: Mol Pathol  2001 Jun;54(3):138-44 

The IGF axis and hepatocarcinogenesis.

Scharf JG, Dombrowski F, Ramadori G.

Department of Medicine, Division of Gastroenterology and Endocrinology,

Georg-August-Universitat, D-37075 Gottingen, Germany.

jscharf@med.uni-goettingen.de

Deregulation of the insulin-like growth factor (IGF) axis, including the

autocrine production of IGFs, IGF binding proteins (IGFBPs), IGFBP proteases,

and the expression of the IGF receptors, has been identified in the development

of hepatocellular carcinoma (HCC). Characteristic alterations detected in HCC

and hepatoma cell lines comprise the increased expression of IGF-II and the

IGF-I receptor (IGF-IR), which have emerged as crucial events in malignant

transformation and the growth of tumours. Alterations of IGFBP production and

the proteolytic degradation of IGFBPs resulting in an excess of bioactive IGFs,

as well as the defective function of the IGF degrading IGF-II/mannose

6-phosphate receptor (IGF-II/M6PR), may further potentiate the mitogenic effects

of IGFs in the development of HCC.

Publication Types:

Review

Review, Tutorial

PMID: 11376124 [PubMed - indexed for MEDLINE]

16: Mol Pathol  2001 Jun;54(3):121-4 

The IGF system in thyroid cancer: new concepts.

Vella V, Sciacca L, Pandini G, Mineo R, Squatrito S, Vigneri R, Belfiore A.

Cattedra di Endocrinologia, Istituto di Medicina Interna, Malattie Endocrine e

del Metabolismo, University of Catania, Ospedale Garibaldi, Catania, Italy.

In recent years, the activation of the insulin-like growth factor (IGF) system

in cancer has emerged as a key factor for tumour progression and resistance to

apoptosis. Therefore, a variety of strategies have been developed to block the

type I IGF receptor (IGF-I-R), which is thought to mediate the biological

effects of both IGF-I and IGF-II. However, recent data suggest that the IGF

signalling system is complex and that other receptors are involved. To unravel

the complexity of the IGF system in thyroid cancer, IGF-I and IGF-II production,

and the expression and function of their cognate receptors were studied. Both

IGFs were found to be locally produced in thyroid cancer: IGF-I by stromal cells

and IGF-II by malignant thyrocytes. Values were significantly higher in

malignant tissue than in normal tissue. IGF-I-Rs were overexpressed in

differentiated papillary carcinomas but not in poorly differentiated or

undifferentiated tumours, whereas insulin receptors (IRs) were greatly

overexpressed in all tumour hystotypes, with a trend for higher values in

dedifferentiated tumours. As a consequence of IR overexpression, high amounts of

IR/IGF-I-R hybrids (which bind IGF-I with high affinity) were present in all

thyroid cancer histotypes. Because of recent evidence that isoform A of IR

(IR-A) is a physiological receptor for IGF-II in fetal life, the relative

abundance of IR-A in thyroid cancer was measured. Preliminary data indicate that

overexpressed IRs mainly occur as IR-A in thyroid cancer. These data indicate

that both IR/IGF-I-R hybrids and IR-A play an important role in the

overactivation of the IGF system in thyroid cancer and in IGF-I mitogenic

signalling in these tumours. J Clin PATHOL: Mol Pathol

Publication Types:

Review

Review, Tutorial

PMID: 11376121 [PubMed - indexed for MEDLINE]

17: J Biol Chem  2001 Apr 27;276(17):14459-65 

Insulin stimulates PKCzeta -mediated phosphorylation of insulin receptor

substrate-1 (IRS-1). A self-attenuated mechanism to negatively regulate the

function of IRS proteins.

Liu YF, Paz K, Herschkovitz A, Alt A, Tennenbaum T, Sampson SR, Ohba M, Kuroki

T, LeRoith D, Zick Y.

Department of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot

76100, Israel.

Incubation of rat hepatoma Fao cells with insulin leads to a transient rise in

Tyr phosphorylation of insulin receptor substrate (IRS) proteins. This is

followed by elevation in their P-Ser/Thr content, and their dissociation from

the insulin receptor (IR). Wortmannin, a phosphatidylinositol 3-kinase (PI3K)

inhibitor, abolished the increase in the P-Ser/Thr content of IRS-1, its

dissociation from the IR, and the decrease in its P-Tyr content following 60 min

of insulin treatment, indicating that the Ser kinases that negatively regulate

IRS-1 function are downstream effectors of PI3K. PKCzeta fulfills this

criterion, being an insulin-activated downstream effector of PI3K.

Overexpression of PKCzeta in Fao cells, by infection of the cells with

adenovirus-based PKCzeta construct, had no effect on its own, but it accelerated

the rate of insulin-stimulated dissociation of IR.IRS-1 complexes and the rate

of Tyr dephosphorylation of IRS-1. The insulin-stimulated negative regulatory

role of PKCzeta was specific and could not be mimic by infecting Fao cells with

adenoviral constructs encoding for PKC alpha, delta, or eta. Because the

reduction in P-Tyr content of IRS-1 was accompanied by a reduced association of

IRS-1 with p85, the regulatory subunit of PI3K, it suggests that this negative

regulatory process induced by PKCzeta, has a built-in attenuation signal. Hence,

insulin triggers a sequential cascade in which PI3K-mediated activation of

PKCzeta inhibits IRS-1 functions, reduces complex formation between IRS-1 and

PI3K, and inhibits further activation of PKCzeta itself. These findings

implicate PKCzeta as a key element in a multistep negative feedback control

mechanism of IRS-1 functions.

PMID: 11278339 [PubMed - indexed for MEDLINE]

18: Exp Cell Res  2001 Apr 1;264(2):388-96 

Loss of insulin-like growth factor II receptor expression promotes growth in

cancer by increasing intracellular signaling from both IGF-I and insulin

receptors.

Osipo C, Dorman S, Frankfater A.

Division of Molecular and Cellular Biochemistry, Loyola University Medical

Center, Maywood, Illinois 60153, USA.

The insulin-like growth factor-II receptor (IGF-IIR) is frequently mutated or

deleted in some malignant human tumors, suggesting that the IGF-IIR is a tumor

suppressor. However, the exact mechanism by which IGF-IIR suppresses growth in

tumors has not been definitively established. We demonstrate that

IGF-IIR-deficient murine L cells (D9) have higher growth rates than

IGF-IIR-positive L cells (Cc2) in response to IGF-II. IGF-II levels are higher

in growth-conditioned medium from D9 versus Cc2 cells. Receptor neutralization

studies and measurements of insulin receptor substrate 1 phosphorylation confirm

that the enhanced growth of D9 cells is due to increased stimulation of the

IGF-I and insulin receptors by IGF-II. In contrast, the levels of secreted

latent and active transforming growth factor beta (TGF-beta) are similar for

both D9 and Cc2 cells, indicating that the slower growth of Cc2 cells is not due

to activation of latent TGF-beta by IGF-IIR and growth inhibition. The results

directly demonstrate that down regulation of the IGF-IIR promotes the growth of

transformed D9 cells by sustaining IGF-II, which binds to and activates IGF-IR

and insulin receptor to increase intracellular growth signals. Copyright 2001

Academic Press.

PMID: 11262195 [PubMed - indexed for MEDLINE]

19: Srp Arh Celok Lek  2000 Sep-Oct;128(9-10):335-9 

[The role of insulin and hyperinsulinemia in the pathogenesis of the polycystic

ovary syndrome]

[Article in Serbo-Croatian (Cyrillic)]

Vasiljevic M.

Publication Types:

Review

Review, Academic

PMID: 11255689 [PubMed - indexed for MEDLINE]

20: Cancer Lett  2001 Jan 26;162(2):253-60 

Expression levels of insulin-like growth factor binding proteins and insulin

receptor isoforms in hepatoblastomas.

von Horn H, Tally M, Hall K, Eriksson T, Ekstrom TJ, Gray SG.

Laboratory for Molecular Development and Tumor Biology, Experimental Alcohol and

Drug Addiction Research Section, Department of Clinical Neuroscience, Karolinska

Institute, CMM, L8 01, S-171 76, Stockholm, Sweden.

Hepatoblastoma, a rare pediatric liver tumour, is a poorly understood disease.

While expression studies for some members of the Insulin-like growth factor axis

have been studied in hepatoblastoma, a systematic analysis of the IGF-axis has

not been carried out. We have examined a series of hepatoblastomas with matched

normal liver tissue for gene expression differences with emphasis on members of

the insulin-like growth factor binding proteins. The expression profiles

obtained reveal that the expression of these genes are altered in these tumors.

The results indicate that the IGF-axis is seriously disturbed in the tumors.

PMID: 11146233 [PubMed - indexed for MEDLINE]

21: Toxicol Sci  2001 Jan;59(1):178-84 

Effects of dichloroacetate (DCA) on serum insulin levels and insulin-controlled

signaling proteins in livers of male B6C3F1 mice.

Lingohr MK, Thrall BD, Bull RJ.

Washington State University, Pullman, Washington 99164-6510, USA.

DCA is hepatocarcinogenic in rodents. At carcinogenic doses, DCA causes a large

accumulation of liver glycogen. Thus, we studied the effects of DCA treatment on

insulin levels and expression of insulin-controlled signaling proteins in the

liver. DCA treatment (0.2-2.0 g/l in drinking water for 2 weeks) reduced serum

insulin levels. The decrease persisted for at least 8 weeks. In livers of mice

treated with DCA for 2-, 10-, and 52-week periods, insulin receptor (IR) protein

levels were significantly depressed. Additionally, protein kinase B (PKBalpha)

expression decreased significantly with DCA treatment. In normal liver, glycogen

levels were increased as early as at 1 week, and this effect preceded changes in

insulin and IR and PKBalpha. In contrast to normal liver, IR protein was

elevated in DCA-induced liver tumors relative to that in liver tissue of

untreated animals and to an even greater extent when compared to adjacent normal

liver in the treated animal. Mitogen-activated protein kinase (MAP kinase)

phosphorylation was also increased in tumor tissue relative to normal liver

tissue and tissue from untreated controls. These data suggest that normal

hepatocytes down-regulate insulin-signaling proteins in response to the

accumulation of liver glycogen caused by DCA. Furthermore, these results suggest

that the initiated cell population, which does not accumulate glycogen and is

promoted by DCA treatment, responds differently from normal hepatocytes to the

insulin-like effects of this chemical. The differential sensitivity of the 2

cell populations may contribute to the tumorigenic effects of DCA in the liver.

PMID: 11134557 [PubMed - indexed for MEDLINE]

22: Laryngorhinootologie  2000 Jul;79(7):438-41 

[Detection of overexpression of insulin receptor gene in laryngeal carcinoma

cells by using differential display method]

[Article in German]

Maune S, Gorogh T.

Klinik fur Hals-, Nasen-, Ohrenheilkunde, Kopf- und Halschirurgie,

Christian-Albrechts-Universitat zu Kiel. maune@hno.uni-kiel.de

BACKGROUND: The insulin receptor (IR) is an essential protein localized on the

surface of almost all cell types. IR belongs to the tyrosine-kinase growth

factor receptor family. Insulin mediates proliferative responses in a variety of

tumor cells, however, the role of the IR molecule in carcinogenesis has not yet

exactly been established. METHODS: Messenger RNA from mucosal keratinocytes and

laryngeal carcinoma cells were transcribed in cDNA using reverse transcriptase

and amplified by PCR by means of a number of oligonucleotides. PCR products were

analyzed electrophoretically. RESULTS: Comparing the electrophoretic pattern of

both cell types the overexpression of a 127 bp fragment could be detected in

laryngeal carcinoma cells in contrast to benign keratinocytes. Cloning and

sequencing this fragment exact homologous match was found with exon-2 of the IR

gene. The overexpression of the IR gene in laryngeal carcinoma cells was

confirmed by Northern hybridization. CONCLUSIONS: The results show up-regulation

of IR-mRNA in laryngeal carcinoma cells suggesting that the number of IR is

enhanced in these cells. Hence it follows, that the overexpression of IR plays a

possible role in laryngeal cancer initiation and/or progression.

PMID: 11005099 [PubMed - indexed for MEDLINE]

23: Eur J Nutr  2000 Apr;39(2):67-70 

Paleolithic vs. modern diets--selected pathophysiological implications.

Eaton SB, Eaton SB 3rd.

Dept Anthropology, Emory University, Atlanta, GA 30327, USA. sboydeaton@aol.com

The nutritional patterns of Paleolithic humans influenced genetic evolution

during the time segment within which defining characteristics of contemporary

humans were selected. Our genome can have changed little since the beginnings of

agriculture, so, genetically, humans remain Stone Agers--adapted for a

Paleolithic dietary regimen. Such diets were based chiefly on wild game, fish

and uncultivated plant foods. They provided abundant protein; a fat profile much

different from that of affluent Western nations; high fibre; carbohydrate from

fruits and vegetables (and some honey) but not from cereals, refined sugars and

dairy products; high levels of micronutrients and probably of phytochemicals as

well. Differences between contemporary and ancestral diets have many

pathophysiological implications. This review addresses phytochemicals and

cancer; calcium, physical exertion, bone mineral density and bone structural

geometry; dietary protein, potassium, renal acid secretion and urinary calcium

loss; and finally sarcopenia, adiposity, insulin receptors and insulin

resistance. While not, yet, a basis for formal recommendations, awareness of

Paleolithic nutritional patterns should generate novel, testable hypotheses

grounded in evolutionary theory and it should dispel complacency regarding

currently accepted nutritional tenets.

Publication Types:

Historical Article

PMID: 10918987 [PubMed - indexed for MEDLINE]

24: J Biol Chem  2000 Jul 7;275(27):20880-6 

Selective attenuation of metabolic branch of insulin receptor down-signaling by

high glucose in a hepatoma cell line, HepG2 cells.

Nakajima K, Yamauchi K, Shigematsu S, Ikeo S, Komatsu M, Aizawa T, Hashizume K.

Department of Aging Medicine and Geriatrics, Shinshu University School of

Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan.

The effects of a high concentration of glucose on the insulin receptor-down

signaling were investigated in human hepatoma (HepG2) cells in vitro to

delineate the molecular mechanism of insulin resistance under glucose toxicity.

Treatment of the cells with high concentrations of glucose (15-33 mm) caused

phosphorylation of serine residues of the insulin receptor substrate 1 (IRS-1),

leading to reduced electrophoretic mobility of it. The phosphorylation of IRS-1

with high glucose treatment was blocked only by protein kinase C (PKC)

inhibitors. The high glucose treatment attenuated insulin-induced association of

IRS-1 and phosphatidylinositol 3-kinase and insulin-stimulated phosphorylation

of Akt. A metabolic effect of insulin, stimulation of glycogen synthesis, was

also inhibited by the treatment. In contrast, insulin-induced association of Shc

and Grb2 was not inhibited. Treatment of the cells with high glucose promoted

the translocation of PKCepsilon and PKCdelta from the cytosol to the plasma

membrane but not that of other PKC isoforms. Finally, PKCepsilon and PKCdelta

directly phosphorylated IRS-1 under cell-free conditions. We conclude that a

high concentration of glucose causes phosphorylation of IRS-1, leading to

selective attenuation of metabolic signaling of insulin. PKCepsilon and PKCdelta

are involved in the down-regulation of insulin signaling, and they may lie in a

pathway regulating the phosphorylation of IRS-1.

PMID: 10764799 [PubMed - indexed for MEDLINE]

25: J Clin Endocrinol Metab  2000 Mar;85(3):1232-8 

The intracellular mechanism of insulin resistance in pancreatic cancer patients.

Liu J, Knezetic JA, Strommer L, Permert J, Larsson J, Adrian TE.

Department of Biomedical Sciences, Creighton University School of Medicine,

Omaha, Nebraska 68178, USA.

The diabetes that frequently occurs in pancreatic cancer patients is

characterized by profound peripheral insulin resistance. The intracellular

mechanism of this insulin resistance was investigated in skeletal muscle

biopsies from pancreatic cancer patients with or without diabetes and control

subjects. Insulin receptor (IR) binding, tyrosine kinase activity, IR messenger

RNA (mRNA), IR substrate-1 content, GLUT-4, and GLUT-4 mRNA content were all

normal in pancreatic cancer patients. In contrast, multiple defects in glycogen

synthesis were found in pancreatic cancer patients, especially in those with

diabetes. Glycogen synthase I activity, total activity, and mRNA levels were

significantly decreased in pancreatic cancer patients compared with controls.

The fractional velocity of glycogen synthase was decreased only in the diabetic

pancreatic cancer group. Glycogen phosphorylase a and b activities were

increased in diabetic pancreatic cancer patients, but glycogen phosphorylase

mRNA levels were not significantly different. The insulin resistance associated

with pancreatic cancer is associated with a post-IR defect, which impairs

skeletal muscle glycogen synthesis and glycogen storage.

Publication Types:

Clinical Trial

PMID: 10720068 [PubMed - indexed for MEDLINE]

26: Diabetes Metab Res Rev  2000 Jan-Feb;16(1):26-32 

High insulin levels do not influence PC-1 gene expression and protein content in

human muscle tissue and hepatoma cells.

Frittitta L, Sbraccia P, Costanzo BV, Tassi V, D'Adamo M, Spampinato D, Ercolino

T, Purrello F, Tamburrano G, Vigneri R, Trischitta V.

Institute of Internal Medicine, Endocrine and Metabolic Diseases, University of

Catania, Garibaldi Hospital, Catania, Italy. segmeint@mbox.unict.it

BACKGROUND: To verify whether insulin levels influence PC-1 tissue content, we

studied PC-1 gene expression and protein content in skeletal muscle of patients

with insulinoma, a model of primary hyperinsulinemia. Data were compared with

those obtained in matched insulin sensitive or resistant healthy subjects. In

addition, the effect of high insulin concentration on PC-1 protein content was

studied in HepG2 cells. METHODS: The following measurements were performed:

insulin sensitivity by euglycemic clamp; PC-1 protein content and insulin

receptor autophosphorylation by specific ELISAs; PC-1 gene expression by

competitive polymerase chain reaction (PCR); phosphatidyl-inositol-3 kinase by

immunoprecipitation and thin layer chromatography; glycogen synthesis by

(14)C-glucose incorporation. RESULTS: Muscle PC-1 content was similar in the

insulinoma patients and in insulin sensitive controls but higher (p<0.01) in

insulin resistant controls (21.9+/-4.6 ng/mg protein, 23.8+/-3.9, 48.0+/-8.7,

respectively). PC-1 protein content was inversely correlated with insulin

sensitivity (r=-0.5, p<0.015) but with neither plasma insulin nor glucose

levels. PC-1 protein content was correlated with PC-1 gene expression (r=0.53,

p<0.05, n=14). Exposure to high insulin (100 nmol/l for 16 h) caused a

significant (p<0.05-0.01) impairment of insulin receptor autophosphorylation,

phosphatidyl-inositol-3 kinase activity and glycogen synthesis, but not of PC-1

protein content (114+/-3 vs 102+/-14 ng/mg protein) in HepG2 cells. CONCLUSION:

These findings suggest that chronic high insulin levels do not influence PC-1

expression. Copyright 2000 John Wiley & Sons, Ltd.

PMID: 10707036 [PubMed - indexed for MEDLINE]

27: J Cell Biochem  1999;Suppl 32-33:68-75 

Differentiation and malignant transformation: two roads diverged in a wood.

Baserga R, Morrione A.

Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania

19107, USA. r_baserga@lac.jci.tju.edu

Growth factors and their receptors are known to send at times contradictory

signals, such as proliferation or differentiation. Recent developments in our

knowledge of growth factor receptors and their signaling pathways have clarified

the basis for this puzzling behavior. Separate domains of a receptor and/or the

availability of specific substrates determine the fate of a cell stimulated by

the same growth factor. In some models, the difference between malignant

transformation and differentiation (leading to cell death) may depend on the

presence or absence of a single agonist or antagonist molecule. The type 1

insulin-like growth factor receptor will serve as the paradigm for this review.

J. Cell. Biochem. Suppls. 32/33:68-75, 1999. Copyright 1999 Wiley-Liss, Inc.

Publication Types:

Review

Review, Tutorial

PMID: 10629105 [PubMed - indexed for MEDLINE]

28: FEBS Lett  1999 Dec 31;464(3):159-63 

Insulin selectively activates STAT5b, but not STAT5a, via a JAK2-independent

signalling pathway in Kym-1 rhabdomyosarcoma cells.

Storz P, Doppler H, Pfizenmaier K, Muller G.

Institute of Cell Biology, University of Stuttgart, Allmandring 31, 70569,

Stuttgart, Germany. peter.storz@po.uni-stuttgart.de

The STAT multigene family of transcriptional regulators conveys signals from

several cytokines and growth factors upon phosphorylation by janus kinases

(JAK). Activation of STAT5 is typically mediated by JAK2, but more recent data

indicate a direct activation by the insulin receptor kinase. STAT5 exists in two

closely homologous isoforms, STAT5a and b. We here describe the selective

tyrosine phosphorylation of STAT5b in Kym-1 cells in response to insulin.

Blocking insulin signalling by HNMPA-(AM)(3), an insulin receptor kinase

inhibitor, resulted in the loss of insulin-induced STAT5b tyrosine

phosphorylation, whereas the inhibition of JAK2 by the JAK selective inhibitor

tyrphostin AG490 had no effect. By contrast, in the same cells, IFNgamma-induced

STAT5b activation was JAK2-dependent, indicating that this signal pathway is

functional in Kym-1 cells. We conclude from this rhabdomyosarcoma model that

STAT5b, but not STAT5a is a direct target of the insulin receptor kinase.

PMID: 10618497 [PubMed - indexed for MEDLINE]

29: Hum Reprod  1999 Sep;14(9):2216-22 

Adipocyte insulin action following ovulation in polycystic ovarian syndrome.

Marsden PJ, Murdoch AP, Taylor R.

Department of Obstetrics and Gynaecology, Sunderland Royal Hospital, Kayll Road,

Sunderland SR4 7TP, UK.

The role of anovulation and insulin resistance in the pathogenesis of polycystic

ovarian syndrome (PCOS) remains to be determined. The aim of this study was to

investigate whether the metabolic abnormality of insulin resistance in PCOS

reflects, rather than causes, the ovarian dysfunction. Eight subjects with

classical PCOS were studied on two occasions. Adipocyte insulin sensitivity

together with hormonal and metabolic changes were investigated in patients with

PCOS following prolonged amenorrhoea and then again in the early follicular

phase after ovulation. Insulin receptor binding in amenorrhoeic subjects with

PCOS was low at 0.78 +/- 0.08% and this increased to 1.18 +/- 0.19% after an

ovulatory cycle (P < 0.05). Maximal insulin stimulated 3-O-methylglucose uptake

was 0.70 +/- 0. 14 during amenorrhoea and increased to 1.08 +/- 0.25 pmol/10

cm(2) cell membrane (P < 0.05). Plasma testosterone fell (4.0 +/- 0.4 to 2. 3

+/- 0.2 nmol/l; P < 0.001), luteinizing hormone fell (17.6 +/- 2.3 to 6.7 +/-

0.8 IU/l; P < 0.001) but plasma insulin concentrations remained unchanged

following ovulation (14.6 +/- 1.9 and 15.7 +/- 3. 8 pmol/l during amenorrhoea

and after ovulation respectively). The results of this study suggest that

chronic anovulation per se appears to modify the factors contributing to

cellular insulin resistance seen in PCOS.

PMID: 10469683 [PubMed - indexed for MEDLINE]

30: Clin Endocrinol (Oxf)  1999 Aug;51(2):247-53 

Comment in:

 Clin Endocrinol (Oxf). 1999 Aug;51(2):147.

Hypoglycaemia associated with the production of insulin-like growth factor II

and insulin-like growth factor binding protein 6 by a haemangiopericytoma.

Hoekman K, van Doorn J, Gloudemans T, Maassen JA, Schuller AG, Pinedo HM.

Department of Medical Oncology, Free University Hospital, Amsterdam, The

Netherlands. hoekman@azvu.nl

Non-islet-cell tumour-induced hypoglycaemia (NICTH) is, in most cases,

attributable to tumour production of insulin-like growth factor II (IGF-II).

Tumour-derived IGF-II has a higher than normal molecular weight (big 'IGF-II')

and an impaired ability to form the normal ternary 150 kD complex with IGF

binding protein-3 (IGFBP-3) and the acid-labile subunit (ALS). Consequently,

tumoral IGF-II circulates mainly in smaller binary complexes which have a higher

bioavailability than the ternary complex. We had the opportunity to analyze IGFs

and IGF-related factors in both pre- and post-operative blood, tumour tissue and

tumour cyst fluid from a patient with a disseminated haemangiopericytoma and

severe hypoglycaemia. In addition, the effect of serum and tumour cyst fluid on

autophosphorylation of the insulin receptor was examined. Patient serum

contained low levels of IGF-I, IGFBP-3 and ALS, while the concentrations of

IGFBP-2 and IGFBP-6 were markedly elevated. The total level of circulating

IGF-II was within the normal range, but Biogel P-60 gel filtration of patient

serum revealed that 77% of the IGF-II was present in high molecular weight forms

(normal: 10-15%), which decreased to 53% after partial removal of the tumour.

Most of the IGF-II immunoreactivity in pre- and post-operative patient serum was

associated with 50-60 kD complexes with only a minimal contribution (<10%) from

the 150 kD complex. Tumour cyst fluid contained excessive amounts of both big

IGF-II and IGFBP-6. Northern blot analysis of total mRNA isolated from the

tumour demonstrated high expression of the IGF-II gene and abundant 1.1 kb

IGFBP-6 transcript, while the genes encoding IGFBP-3, -4 and -5 were only weakly

expressed and mRNA of IGFBP-1, -2 and IGF-I could not be detected. mRNAs for the

IGF type II receptor could be easily demonstrated, whereas those for the

insulin- and IGF type I receptor were hardly detectable. In contrast to patient

serum tumour cyst fluid strongly stimulated the insulin receptor in vitro. The

present study suggests an important role of the simultaneous production of

IGF-II and IGFBP-6 in the pathophysiology of tumour-induced hypoglycaemia.

PMID: 10468998 [PubMed - indexed for MEDLINE]

31: Endocr Rev  1999 Aug;20(4):535-82 

The insulin-related ovarian regulatory system in health and disease.

Poretsky L, Cataldo NA, Rosenwaks Z, Giudice LC.

Department of Medicine, New York Presbyterian Hospital and Weill Medical College

of Cornell University, New York, New York 10021, USA.

Publication Types:

Review

Review, Academic

PMID: 10453357 [PubMed - indexed for MEDLINE]

32: Exp Cell Res  1999 Aug 25;251(1):22-32 

Characterization of an antibody that can detect an activated IGF-I receptor in

human cancers.

Rubini M, D'Ambrosio C, Carturan S, Yumet G, Catalano E, Shan S, Huang Z,

Criscuolo M, Pifferi M, Baserga R.

University of Ferrara, Via L. Borsari 46, Ferrara, 44100, Italy.

The type 1 insulin-like growth factor receptor (IGF-IR) plays an important role

in malignant transformation and in apoptosis. Its role in human cancer has now

been firmly established. IGF-IR signaling occurs only when the receptor is

activated by its ligands, which induce autophosphorylation of the receptor at

several tyrosine residues. Although the IGF-IR (phosphorylated or not) can be

detected in human cancers with conventional antibodies, it would be desirable to

obtain antibodies that can detect the IGF-IR only when activated by its ligands.

We describe and characterize in this paper such an antibody and show that it can

be used in sections of human cancers to detect an autophosphorylated IGF-IR.

This antibody will be useful in detecting autocrine or paracrine influences on

normal and tumor cells and could eventually be also useful in diagnostic and

prognostic studies of human primary and metastatic cancer. Copyright 1999

Academic Press.

PMID: 10438568 [PubMed - indexed for MEDLINE]

33: Clin Cancer Res  1999 Jul;5(7):1935-44 

Insulin and insulin-like growth factor-I (IGF-I) receptor overexpression in

breast cancers leads to insulin/IGF-I hybrid receptor overexpression: evidence

for a second mechanism of IGF-I signaling.

Pandini G, Vigneri R, Costantino A, Frasca F, Ippolito A, Fujita-Yamaguchi Y,

Siddle K, Goldfine ID, Belfiore A.

Istituto di Medicina Interna, Malattie Endocrine e del Metabolismo, Universita

di Catania, Ospedale Garibaldi, Italy.

The insulin receptor (IR) form hybrids with the closely related insulin-like

growth factor-I (IGF-I) receptor (IGF-I-R). Because most human breast carcinomas

overexpress both the IR and the IGF-I-R, we evaluated whether the insulin/IGF-I

hybrid receptor (Hybrid-R) is also overexpressed in these tumors and what role

it plays in breast cancer biology. Using specific ELISAs and Western blots, we

measured Hybrid-R content and function in 8 human cultured breast cancer cell

lines and 39 human breast cancer specimens. Hybrid-R content and function were

also compared to the content and function of the IR and the IGF-I-R. Hybrid-R

content exceeded the IGF-I-R content in >75% of breast cancer specimens and was

directly related to the molar ratio of both the IR and IGF-I-R content,

suggesting that Hybrid-R formation occurred by random assembly of IR and IGF-I-R

half-receptors. Hybrid-Rs became tyrosine autophosphorylated when breast cancer

cells were exposed to IGF-I but not when they were exposed to insulin. In cells

with an elevated Hybrid-R content, Hybrid-R autophosphorylation in response to

IGF-I exceeded IGF-I-R autophosphorylation, suggesting that most of the IGF-I

effect occurred via the Hybrid-R. Furthermore, Hybrid-Rs mediated growth in

response to IGF-I, as indicated by experiments with blocking antibodies to the

IGF-I-R. These data indicated therefore that: (a) Hybrid-Rs are present and play

a major role in mediating the IGF-I signal in breast cancer; (b) their

expression is directly related to IR overexpression; and (c) potential therapies

designed to block IGF-I actions in breast cancer must take into account the role

of these Hybrid-Rs.

PMID: 10430101 [PubMed - indexed for MEDLINE]

34: Cancer Res  1999 Jul 15;59(14):3527-34 

Differential reconstitution of mitochondrial respiratory chain activity and

plasma redox state by cysteine and ornithine in a model of cancer cachexia.

Ushmorov A, Hack V, Droge W.

Deutsches Krebsforschungszentrum, Division of Immunochemistry, Heidelberg,

Germany.

The mechanism of wasting, as it occurs in malignant diseases and various

etiologically unrelated conditions, is still poorly understood. We have,

therefore, studied putative cause/effect relationships in a murine model of

cancer cachexia, C57BL/6 mice bearing the fibrosarcoma MCA-105. The plasma of

these mice showed decreased albumin and increased glutamate levels, which are

typically found in practically all catabolic conditions. Skeletal muscles from

tumor-bearing mice were found to have an abnormally low mitochondrial

respiratory chain activity (mito.RCA) and significantly decreased glutathione

(GSH) levels. The decrease in mito.RCA was correlated with an increase in the

i.m. GSH disulfide/GSH ratio, the plasma cystine/thiol ratio, and the GSH

disulfide/GSH ratio in the bile. This is indicative of a generalized shift in

the redox state extending through different body fluids. Treatment of

tumor-bearing mice with ornithine, a precursor of the radical scavenger

spermine, reversed both the decrease in mito.RCA and the change in the redox

state, whereas treatment with cysteine, a GSH precursor, normalized only the

redox state. Treatment of normal mice with difluoromethyl-ornithine, a specific

inhibitor of ornithine decarboxylase and spermine biosynthesis, inhibited the

mito.RCA in the skeletal muscle tissue, thus illustrating the importance of the

putrescine/spermine pathway in the maintenance of mito.RCA. Ornithine, cysteine,

and N-acetyl-cysteine (NAC) also reconstituted the abnormally low concentrations

of the GSH precursor glutamate in the skeletal muscle tissue of tumor-bearing

mice. Higher doses, however, enhanced tumor growth and increased the plasma

glucose level in normal mice. In the latter, cysteine and NAC also decreased

i.m. catalase and GSH peroxidase activities. Taken together, our studies on the

effects of ornithine, cysteine, and NAC illuminate some of the mechanistic

pathways involved in cachexia and suggest targets for therapeutic intervention.

PMID: 10416620 [PubMed - indexed for MEDLINE]

35: Biochimie  1999 Apr;81(4):403-7 

Insulin/IGF-I hybrid receptors play a major role in IGF-I signaling in thyroid

cancer.

Belfiore A, Pandini G, Vella V, Squatrito S, Vigneri R.

Istituto di Medicina Interna, Malattie Endocrine e del Metabolismo, Universita

di Catania, Ospedale Garibaldi, Italy.

The insulin-like growth factor-I (IGF-I) plays an important role in determining

the biological behavior of a variety of malignancies. We measured IGF-I, its

receptor and related receptors in thyroid cancer. IGF-I was present both in

normal thyroid tissue and in thyroid cancer tissue and it was produced by

stromal cells but not by thyrocytes. Values were significantly higher in

malignant than in normal tissue. IGF-I receptors (IGF-I-Rs) and the homologous

insulin receptors (IRs) were found overexpressed in both thyroid cancer cell

lines (n = 4) and specimens (n = 17) as compared to normal values. In addition,

high levels of hybrid IGF-I/insulin receptors (IR/IGF-I-Rs) were present in both

thyroid cancer specimens and cell lines. IR/IGF-I-R hybrids were the most

represented type of receptor in 14/17 specimens and exceeded the IGF-I-R content

in all cases. Hybrid content correlated with the IR and IGF-I-R content,

suggesting that in thyroid tissue hybrid formation occurs by random assembly of

IR and IGF-I-R half receptors. Hybrid receptor autophosphorylation was

stimulated by IGF-I with high affinity. In cells with a high IR/IGF-I-Rs

content, blocking antibodies specific to these receptors substantially inhibited

IGF-I induced cell growth. These data indicate that the IGF-I system is

overactivated in thyroid cancer and that IR/IGF-I-R hybrid receptors play an

important role in IGF-I mitogenic signaling in these tumors.

PMID: 10401676 [PubMed - indexed for MEDLINE]

36: J Intern Med  1999 Jun;245(6):621-5 

The role of TNFalpha and TNF receptors in obesity and insulin resistance.

Hotamisligil GS.

Department of Nutrition, Harvard University, School of Public Health, Boston,

Massachusetts 02115, USA. ghota-mis@hsph.harvard.edu

Insulin resistance, a smaller than expected response to a given dose of insulin,

is associated with many common diseases including, ageing, polycystic ovarian

disease, syndrome X, cancer, infections, trauma and, most significantly, obesity

and type 2 diabetes mellitus. The biochemical basis of insulin resistance in

type 2 diabetes has been the subject of many studies. Earlier studies have

indicated that quantitative regulation of the insulin sensitive glucose

transporters (Glut-4) and insulin receptors themselves may contribute to this

disorder, however, these two factors are probably inadequate to explain the

extent of insulin resistance. This point also became apparent by the development

of only mild hyperinsulinaemia in mice with a targeted mutation in the Glut-4

gene. Studies on postreceptor defects in type 2 diabetes has recently focused on

the intrinsic catalytic activity of the insulin receptor and downstream

signalling events. A reduction in tyrosine phosphorylation of both the insulin

receptor (IR) and the insulin receptor substrate-1 (IRS-1) has been noted in

both animal and human type 2 diabetes. Importantly, this appears to occur in all

of the major insulin-sensitive tissues, namely the muscle, fat and liver. It is

now clear that decreased signalling capacity of the insulin receptor is an

important component of this disease. I will review some of the potential

mechanisms underlying this deficiency.

Publication Types:

Review

Review, Tutorial

PMID: 10395191 [PubMed - indexed for MEDLINE]

37: Endocrinol Metab Clin North Am  1999 Jun;28(2):361-78 

Insulin action in the normal and polycystic ovary.

Franks S, Gilling-Smith C, Watson H, Willis D.

Department of Reproductive Science and Medicine, Imperial College School of

Medicine, St. Mary's Hospital, London, United Kingdom. s.franks@ic.ac.uk

Insulin has a stimulatory effect on steroidogenesis by granulosa cells of normal

and polycystic ovaries and interacts with gonadotropins in an additive or, as in

the case of LH, a synergistic manner. These actions seem to be mediated

specifically by the insulin receptor rather than by cross-reaction with the type

I IGF receptor, even in tissue obtained from women with PCOS with biochemical

evidence of insulin resistance. The authors suggest that hyperinsulinemia makes

a significant contribution to premature arrest of follicle growth, which is

characteristic of anovulation in women with PCOS, and that the interaction of

insulin with LH is a key element in this process. Insulin may also have a role

in amplifying LH-induced androgen production by theca cells, which may help

explain the prominence of symptoms of hyperandrogenism in obese subjects with

PCOS. The results of recent clinical studies of insulin-sensitizing agents such

as metformin and the thiazoladinedione troglitazone in PCOS have provided

encouragement that improvement of insulin sensitivity and consequent lowering of

circulating insulin levels by these agents may be of therapeutic value in the

management of both anovulation and hirsutism.

Publication Types:

Review

Review, Tutorial

PMID: 10352923 [PubMed - indexed for MEDLINE]

38: Biochemistry  1999 May 4;38(18):5896-904 

Reversible change in thiol redox status of the insulin receptor alpha-subunit in

intact cells.

Garant MJ, Kole S, Maksimova EM, Bernier M.

Diabetes Section, Laboratory of Clinical Investigation, National Institute on

Aging, National Institutes of Health, Baltimore, Maryland 21224-6825, USA.

In this study, we used maleimidobutyrylbiocytin to examine possible alteration

that may occur in the redox state of the insulin receptor (IR) sulfhydryl groups

in response to reduced glutathione (GSH) or N-acetyl-L-cysteine (NAC).

Short-term treatment of intact cells expressing large numbers of IR with GSH or

NAC led to a rapid and reversible reduction of IR alpha-subunit disulfides,

without affecting the receptor beta-subunit thiol reactivity. The overall

integrity of the oligomeric structure of IR was maintained, indicating that

neither class I nor class II disulfides were targeted by these agents. Similar

findings were obtained in cells transfected with IR mutants lacking cysteine524,

one of the class I disulfides that link the two IR alpha-subunits.

Membrane-associated thiols did not participate in GSH- or NAC-mediated reduction

of IR alpha-subunit disulfides. No difference in insulin binding was observed in

GSH-treated cells; however, ligand-mediated increases in IR autophosphorylation,

tyrosine phosphorylation of cellular substrates, and dual phosphorylation of the

downstream target mitogen-activated protein kinase were inhibited at

concentrations of GSH (10 mM or greater) that yielded a significant increase in

IR alpha-subunit thiol reactivity. GSH did not affect IR signaling in the

absence of insulin. Our results provide the first evidence that the IR

alpha-subunit contains a select group of disulfides whose redox status can be

rapidly altered by the reducing agents GSH and NAC.

PMID: 10231542 [PubMed - indexed for MEDLINE]

39: Oncogene  1999 Apr 15;18(15):2471-9 

Insulin receptor activation by IGF-II in breast cancers: evidence for a new

autocrine/paracrine mechanism.

Sciacca L, Costantino A, Pandini G, Mineo R, Frasca F, Scalia P, Sbraccia P,

Goldfine ID, Vigneri R, Belfiore A.

Istituto di Medicina Interna, Malattie Endocrine e del Metabolismo, Universita

di Catania, Ospedale Garibaldi, Italy.

IGF-II, produced by breast cancer epithelial and stromal cells, enhances tumor

growth by activating the IGF-I receptor (IGF-I-R) via autocrine and paracrine

mechanisms. Previously we found that the insulin receptor (IR), which is related

to the IGF-I-R, is overexpressed in breast cancer cells. Herein, we find that,

in breast cancer the IR is activated by IGF-II. In eight human breast cancer

cell lines studied there was high affinity IGF-II binding to the IR, with

subsequent IR activation. In these lines, IGF-II had a potency up to 63% that of

insulin. In contrast, in non malignant human breast cells, IGF-II was less than

1% potent as insulin. Via activation of the IR tyrosine kinase IGF-II stimulated

breast cancer cell growth. Moreover, IGF-II also activated the IR in breast

cancer tissue specimens; IGF-II was 10-100% as potent as insulin. The IR occurs

in two isoforms generated by alternative splicing of exon 11; these isoforms are

IR-A (Ex11-) and IR-B (Ex11+). IR-A was predominantly expressed in breast cancer

cells and specimens and the potency of IGF-II was correlated to the expression

of this isoform (P<0.0001). These data indicate, therefore, that the IR-A, which

binds IGF-II with high affinity, is predominantly expressed in breast cancer

cells and represents a new autocrine/paracrine loop involved in tumor biology.

PMID: 10229198 [PubMed - indexed for MEDLINE]

40: J Surg Res  1999 May 1;83(1):32-5 

Human insulin receptor and insulin signaling proteins in hepatic disease.

Spector SA, Olson ET, Gumbs AA, Friess H, Buchler MW, Seymour NE.

Surgical Service, VA Connecticut Healthcare System, West Haven, Connecticut

06516, USA.

Insulin regulates hepatocellular metabolism and growth following insulin

receptor (IR) autophosphorylation and activation of the intracellular adapter

protein, insulin receptor substrate 1 (IRS-1). IRS-1 activates SH2 domain

proteins such as Grb2, which may be vital to hepatocyte growth. To determine if

these substances are abnormally expressed under pathophysiologic conditions, IR,

IRS-1, Grb2 protein, and IR mRNA were studied in normal human liver (n = 10),

cirrhotic liver (n = 10), and hepatocellular carcinoma (HCC) (n = 10) that had

been procured during operative procedures. IR mRNA was quantified by S1-nuclease

assay using a 195-bp digoxigenin-labeled IR DNA probe and normalized to the

level of expression of the glyceraldehyde 3-phosphate dehydrogenase (GAPDH)

gene. Protein concentrations were determined by immunoblot analysis following

SDS-PAGE of liver homogenate samples. Labeled DNA and antibody-complexed protein

were detected by chemiluminescent means and quantified by densitometric analysis

(mean densitometric units +/- standard error). Similar levels of IR mRNA were

observed in normal tissue, cirrhosis, and HCC. IR protein concentration was

significantly greater in HCC than in normal liver (1.82 +/- 0.2 vs 1. 25 +/-

0.17; P < 0.05). IRS-1 was significantly increased in cirrhosis compared to

normal liver (1.61 +/- 0.31 vs 0.86 +/- 0.21; P < 0.05). No differences were

observed in Grb2 in the three tissue types. Insulin receptor overexpression,

previously seen in other tumor types, may confer an insulin-mediated growth

advantage in HCC if added receptors reflect functional high affinity binding

sites. Although an altered mass of IRS-1 protein was not observed in HCC, an

IRS-1 increase in cirrhosis may favor hepatic regeneration. Copyright 1999

Academic Press.

PMID: 10210639 [PubMed - indexed for MEDLINE]

41: Diabetologia  1999 Mar;42(3):317-25 

Modulation of insulin receptor signalling by pancreastatin in HTC hepatoma

cells.

Sanchez-Margalet V.

Department of Medical Biochemistry and Molecular Biology, School of Medicine,

Virgen Macarena Hospital, University of Seville, Spain.

Pancreastatin, a neuropeptide derived from chromogranin A, has a glycogenolytic

and counterregulatory effect to insulin in the rat liver. This effect is

mediated by calcium and protein kinase C activity. Our aim was to study the

possible cross-talk between pancreastatin and the insulin signalling system, by

using the well-studied insulin sensitive rat hepatoma HTC cells. First, we

checked the counterregulatory effect of pancreastatin on insulin action.

Pancreastatin dose-dependently inhibited insulin stimulated glycogen synthesis.

This effect was not due to competition for insulin receptors. Moreover, when

protein kinase C activation was blocked with staurosporine, this effect of

pancreastatin was not observed. Next, we found a dose-dependent inhibition of

insulin receptor autophosphorylation by pancreastatin. In addition,

phosphorylation of the major substrates of insulin receptor in HTC, i. e.

insulin-receptor substrate (IRS)-1/IRS-2 and p62 was also blunted and so was its

association with p85 regulatory subunit of phosphatidylinositol-3-kinase.

Moreover, the insulin activation of S6 kinase was also blocked by pancreastatin.

Again, all these inhibitory effects of pancreastatin were prevented by

staurosporine. Furthermore, pancreastatin produced Ser/Thr phosphorylation of

insulin receptor by a staurosporine-sensitive mechanism. Finally, we checked the

pancreastatin activation of protein kinase C in HTC cells and found that a

"classical" isoform of this protein is translocated to the plasma membrane.

These findings suggest that pancreastatin could exert its anti-insulin effect in

the hepatocyte by interrupting the stimulation of early insulin receptor

signalling as a result of phosphorylation. This interaction might have a role in

the mechanisms of insulin resistance.

PMID: 10096784 [PubMed - indexed for MEDLINE]

42: Cell Mol Life Sci  1999 Jan;55(1):142-7 

Insulin activates G alpha il,2 protein in rat hepatoma (HTC) cell membranes.

Sanchez-Margalet V, Gonzalez-Yanes C, Santos-Alvarez J, Najib S.

Departamento de Bioquimica Medica y Biologia Molecular, Facultad de Medicina,

Hospital Universitario Virgen Macarena, Sevilla, Spain.

Insulin action is initiated by binding to its cognate receptor, which then

triggers multiple cellular responses by activating different signaling pathways.

There is evidence that insulin receptor signaling may involve G protein

activation in different target cells. We have studied the activation of G

proteins in rat hepatoma (HTC) cells. We found that insulin stimulated binding

of guanosine 5'-O-(3-thiotriphosphate) (GTP-gamma-35S) to plasma membrane

proteins of HTC cells, in a dose-dependent manner. This effect was completely

blocked by pertussis toxin treatment of the membranes, suggesting the

involvement of G proteins of the G alpha i/G alpha o family. The expression of

these G alpha proteins was checked by Western blotting. Next, we used blocking

antibodies to sort out the specific G alpha protein activated by insulin

stimulation. Anti-G alpha il,2 antibodies completely prevented

insulin-stimulated GTP binding, whereas anti-G alpha o,i3 did not modify this

effect of insulin on GTP binding. Moreover, we found physical association of the

insulin receptor with G alpha il,2 by copurification studies. These results

further support the involvement of a pertussis toxin-sensitive G protein in

insulin receptor signaling and provides some evidence of specific association

and activation of G alpha il,2 protein by insulin. These findings suggest that G

alpha il,2 proteins might be involved in insulin action.

PMID: 10065161 [PubMed - indexed for MEDLINE]

43: Cancer  1999 Jan 15;85(2):492-8 

Functional insulin receptors are overexpressed in thyroid tumors: is this an

early event in thyroid tumorigenesis?

Frittitta L, Sciacca L, Catalfamo R, Ippolito A, Gangemi P, Pezzino V, Filetti

S, Vigneri R.

Istituto di Medicina Interna, Malattie Endocrine e del Metabolismo, Universita

di Catania, Ospedale Garibaldi, Italy.

BACKGROUND: Insulin receptor (IR), a member of the receptor tyrosine kinase

family, is expressed in normal thyroid cells and affects thyroid cell

proliferation and differentiation. METHODS: The authors measured IR content in

benign and malignant thyroid tumors by three independent methods: a specific

radioimmunoassay, 125I-insulin binding studies, and immunohistochemistry. The

results obtained were compared with the IR content in paired, adjacent, normal

thyroid tissue. To assess IR function in thyroid carcinoma cells, glucose uptake

responsiveness to insulin was also studied in a human transformed thyroid cell

line (B-CPAP) and in follicular carcinoma cells in primary culture. RESULTS: In

9 toxic adenomas, the average IR content was similar to that observed in the 9

paired normal thyroid tissue specimens from the same patients (2.2+/-0.3 vs.

2.1+/-0.3). In 13 benign nonfunctioning, or "cold," adenomas, the average IR

content was significantly higher (P < 0.001) than in paired normal tissue

specimens (4.3+/-0.5 vs. 1.8+/-0.1). In 12 papillary and 10 follicular

carcinomas, IR content was significantly higher (P < 0.001) than in the adjacent

normal thyroid tissue (4.0+/-0.4 vs. 1.6+/-0.2 and 5.6+/-1.0 vs. 1.8+/-0.2,

respectively). The finding of a higher IR content in benign "cold" adenomas and

in thyroid carcinomas was confirmed by both binding and immunostaining studies.

CONCLUSIONS: The current studies indicate that 1) IR content is elevated in most

follicular and papillary differentiated thyroid carcinomas, and 2) IR content is

also elevated in most benign follicular adenomas ("cold" nodules) but not in

highly differentiated, hyperfunctioning follicular adenomas ("hot" nodules),

which very rarely become malignant. This observation suggests that increased IR

expression is not restricted to the thyroid malignant phenotype but is already

present in the premalignant "cold" adenomas. It may contribute, therefore, to

thyroid tumorigenesis and/or represent an early event that gives a selective

growth advantage to transformed thyroid cells.

PMID: 10023720 [PubMed - indexed for MEDLINE]

44: Biochem Biophys Res Commun  1998 Dec 18;253(2):315-9 

Role of IRS-1 signaling in insulin-induced modulation of estrogen receptors in

breast cancer cells.

Ando' S, Panno ML, Salerno M, Sisci D, Mauro L, Lanzino M, Surmacz E.

Dipartimento di Biologia Cellulare, Universita' degli Studi della Calabria,

Cosenza, Italy. sando@diemme.it

Cross-talk between steroid hormones and polypeptide growth factors regulates the

growth of hormone-responsive breast cancer cells. For example, in the MCF-7

human breast cancer cell line, insulin up-regulates estrogen receptor (ER)

content and binding capacity. Since the insulin receptor (IR) substrate 1

(IRS-1) is one of the core signaling elements transmitting mitogenic and

metabolic effects of insulin, we investigated whether IRS-1 is also required for

the insulin-induced function of the ER. The effects of insulin on the ER were

compared in MCF-7 cells and MCF-7-derived cell lines with decreased levels (by

approximately 80%) of IRS-1 due to the expression of IRS-1 antisense RNA. The

severe IRS-1 deficiency in MCF-7 cells was associated with (1) reduced mitogenic

response to 20 ng/ml insulin and 10% calf serum (CS), but not to 1 nM estradiol

(E2); (2) loss of insulin-E2 synergism; (3) up-regulation of ER protein

expression and binding capacity; and (4) loss of insulin-induced regulation of

ER tyrosine phosphorylation. In conclusion, the data confirm the existence of

the IR-ER cross-talk and suggest that IRS-1-dependent signaling may contribute

to the negative regulation of the ER expression and function in MCF-7 cells.

Copyright 1998 Academic Press.

PMID: 9878535 [PubMed - indexed for MEDLINE]

45: Pancreas  1998 Nov;17(4):359-66 

The intracellular mechanism of insulin resistance in the hamster pancreatic

ductal adenocarcinoma model.

Liu J, Kazakoff K, Pour PM, Adrian TE.

Department of Biomedical Sciences, Creighton University School of Medicine,

Omaha, Nebraska 68178, USA.

Diabetes associated with pancreatic cancer is characterized by profound

peripheral insulin resistance. The intracellular mechanism of insulin resistance

was investigated in skeletal muscles from N-nitrosobis(2-oxopropyl)amine

(BOP)-treated hamsters. Effects of high-fat diet and exercise also were studied.

BOP (20 mg/kg body weight) was administrated weekly for 2 weeks.

Hyperinsulinemia was found in BOP-treated hamsters at 20 weeks after BOP

treatment, suggesting the peripheral insulin resistance is an early feature in

pancreatic cancer. Hamsters were killed at 42 weeks, and soleus muscles were

taken for the analysis. Skeletal muscle insulin-receptor binding and insulin

receptor tyrosine kinase activities were similar between the control and

BOP-treated hamsters. However, maximal muscle glycogen synthase activity was

significantly reduced in BOP-treated hamsters compared with the control group.

Muscle glycogen phosphorylase activity was increased in the BOP-treated group

fed with high-fat diet as well as in BOP-treated groups with exercise. These

findings indicate that insulin resistance in the hamster pancreatic cancer model

is caused by a postreceptor defect, which led to significant decrease of muscle

glycogen synthase activity. Whereas a high-fat diet causes more severe insulin

resistance in BOP-treated hamsters, high-fat diet and exercising had no

significant effects on skeletal muscle insulin-receptor function and glycogen

synthase activity. Furthermore, both high-fat diet and exercise enhanced

glycogen phosphorylase activity in BOP-treated hamsters.

PMID: 9821177 [PubMed - indexed for MEDLINE]

46: Clin Cancer Res  1995 Nov;1(11):1429-36 

Overexpression of insulin receptor substrate 1 (IRS-1) in the human breast

cancer cell line MCF-7 induces loss of estrogen requirements for growth and

transformation.

Surmacz E, Burgaud JL.

Jefferson Cancer Institute, Thomas Jefferson University, Philadelphia,

Pennsylvania 19107, USA.

The synergistic action of estrogens and insulin-like growth factors (IGFs)

promotes the growth of many human breast cancer cell lines. This synergistic

effect involves estrogen-dependent induction of the IGF system, i.e., estrogens

augment the number of IGF-I receptors, stimulate the secretion of IGF-II, and

promote the synthesis of certain IGF-binding proteins. On the other hand, the

sustained activation of the IGF-I receptor (IGF-IR) by the overexpression of

IGF-II has been found to contribute to the development of the

estrogen-independent phenotype in breast cancer cells. In this study, we have

investigated whether the amplification of the IGF-IR intracellular signaling in

MCF-7 cells can abolish or reduce estrogen requirements for growth and

transformation. To this end we developed several MCF-7 clones that overexpressed

insulin receptor substrate 1 (IRS-1), one of the principal substrates of the

IGF-IR. We report here that in MCF-7 cells overexpressing IRS-1, estrogen

requirements for growth in monolayer culture as well as in soft agar were

reduced. The decreased estrogen requirements depended on the level of the

overexpressed IRS-1 protein, and in cells which contained several-fold more

functional IRS-1 than the parental cells, we observed total loss of estrogen

dependence for growth. In addition, the importance of IRS-1 in proliferation of

MCF-7 cells has been confirmed through the use of antisense strategies.

PMID: 9815941 [PubMed - indexed for MEDLINE]

47: Am J Physiol  1998 Nov;275(5 Pt 1):E748-56 

Adenovirus-mediated transfer of a modified human proinsulin gene reverses

hyperglycemia in diabetic mice.

Short DK, Okada S, Yamauchi K, Pessin JE.

Department of Physiology and Biophysics, The University of Iowa, Iowa City, Iowa

52242-1109, USA.

The human proinsulin cDNA was introduced into a replication-defective adenovirus

and was found to confer proinsulin expression to a hepatocyte (H4-II-E) cell

line upon infection. A second virus was constructed in which the dibasic

prohormone convertase recognition sequence was mutated to a tetrabasic furin

cleavage site. Cells infected with this virus synthesized both proinsulin and

mature insulin. Gel filtration chromatography, competition of insulin binding,

and activation of the insulin receptor kinase activity demonstrated that this

mature insulin was functionally identical to that of authentic processed

insulin. Injection of these viral constructs into the external jugular vein of

mice resulted in insulin gene expression in the liver. Expression from the

mutated proinsulin virus dramatically improved the glycemic state of diabetic

mice. However, the effects of the viral infection were transient, being maximal

at approximately 5-7 days and returning to steady-state levels by 14-21 days.

These data demonstrate that somatic cell insulin gene delivery by the use of

recombinant adenovirus can be used to transiently reverse the diabetic state in

mice.

PMID: 9814992 [PubMed - indexed for MEDLINE]

48: Am J Med  1998 Oct;105(4):331-45 

Insulin action and insulin resistance: diseases involving defects in insulin

receptors, signal transduction, and the glucose transport effector system.

Hunter SJ, Garvey WT.

Department of Medicine, Medical University of South Carolina, Ralph H. Johnson

Veterans Affairs Medical Center, Charleston 29425, USA.

Publication Types:

Review

Review, Tutorial

PMID: 9809695 [PubMed - indexed for MEDLINE]

49: Gynecol Endocrinol  1998 Aug;12(4):277-92 

Insulin and polycystic ovary syndrome: a new look at an old subject.

Ciampelli M, Lanzone A.

Department of Obstetrics and Gynecology, Catholic University of Sacred Heart,

Rome, Italy.

In recent years the metabolic implications of polycystic ovary syndrome (PCOS)

have received a great deal of attention; in fact 50% of women with PCOS are

obese and a similar percentage of subjects was found to show exaggerated insulin

secretion and reduced insulin-stimulated glucose uptake. The presence of these

features in women with PCOS has profound clinical implications in terms of

morbidity due to diabetes mellitus, dyslipidemia, hypertension and

cardiovascular disease. Moreover, hyperinsulinemia has recently been proposed as

a possible independent risk factor for endometrial and breast cancer. In the

light of these considerations, the importance of metabolic screening in patients

with PCOS in order to improve their quality of life cannot be underestimated. In

this review we analyze all the clinical pathologies in which hyperinsulinemia of

PCOS could be involved. Furthermore, in order to clarify the possible mechanisms

leading to the insulin disorders of the syndrome, we review the available data

about the insulin receptor abnormalities, as well as those concerning the

insulin resistance and the exaggerated insulin secretion. Finally, we examine

the main therapeutic strategies to ameliorate the insulinemic status of PCOS

patients in order to potentially be able to prevent the long-term consequences

of this syndrome.

Publication Types:

Review

Review, Academic

PMID: 9798138 [PubMed - indexed for MEDLINE]

50: Head Neck  1998 Oct;20(7):634-9 

Focal adhesion kinase expression in oral cancers.

Kornberg LJ.

Department of Otolaryngology, University of Florida, Gainesville 32610, USA.

BACKGROUND: Evidence suggests that focal adhesion kinase (FAK) is involved in

the pathogenesis of certain cancers. Focal adhesion kinase is overexpressed in

invasive and metastatic cancers of the breast, colon, thyroid, and prostate. The

objective of this study was to determine the presence and cellular distribution

of FAK in oral cancer and determine whether there is a difference in FAK

expression in preinvasive and invasive oral cancers. METHODS:

Immunohistochemistry was used to detect FAK expression in 20 archival oral

cancer specimens. RESULTS: Focal adhesion kinase immunoreactivity was detected

in all specimens that were examined. In tumors, there was an increase in the

intensity of FAK staining and in the percentage of FAK-positive cells.

Preinvasive tumors had populations of cancer cells which stained more intensely

than neighboring cancer cells. CONCLUSIONS: Focal adhesion kinase expression

appears to be increased in invasive and preinvasive oral cancers. It is

speculated that enhanced expression of FAK may contribute to the aggressive

phenotype of oral cancers.

PMID: 9744465 [PubMed - indexed for MEDLINE]

51: Am J Obstet Gynecol  1998 Jul;179(1):6-12 

Specific binding and growth-promoting activity of insulin in endometrial cancer

cells in culture.

Nagamani M, Stuart CA.

Department of Obstetrics and Gynecology, University of Texas Medical Branch,

Galveston 77555, USA.

OBJECTIVE: Insulin is known to be mitogenic to a variety of cells in culture.

The purpose of this study was to investigate the possible role of insulin in the

growth and development of endometrial cancers. STUDY DESIGN: Specific binding

and growth effects of insulin were studied in 5 different human endometrial

cancer cell lines derived from cancers with different degrees of

differentiation: HEC-1-A and HEC-1-B (from a moderately well-differentiated

adenocarcinoma), RL95-2 (from a moderately well-differentiated adenosquamous

carcinoma), KLE (from poorly differentiated carcinoma), and AN3 CA (from a

metastatic undifferentiated endometrial carcinoma). The receptors were further

characterized by competitive binding and chemical cross-linking studies.

RESULTS: Binding studies with 125I-insulin revealed the presence of

high-affinity binding sites for insulin on all the 5 cell lines. Binding of

insulin was found to be highly specific. Competitive binding studies with

125I-insulin revealed that insulin was most effective in displacing the labeled

hormone, whereas insulin-like growth factor-I and insulin-like growth factor-II

competed for binding only at very high concentrations. Scatchard analysis of the

binding data revealed that the association constant for the high-affinity

binding sites ranged from 0.72 to 1.91 x 10(9) L/mol. Estrogen-receptor-negative

cell lines HEC-1-A and HEC-1-B had the highest number of insulin receptors,

whereas the estrogen-receptor-positive cell line RL95-2 had the least number of

receptors. The effect of insulin on cell proliferation was studied by monitoring

cell number and incorporating [3H]thymidine into deoxyribonucleic acid of the

cells. Insulin stimulated cell growth of all the cell lines. CONCLUSIONS: The

results of this study indicate the potential role of hyperinsulinemia in the

growth and development of endometrial cancer.

PMID: 9704758 [PubMed - indexed for MEDLINE]

52: J Clin Endocrinol Metab  1998 Jun;83(6):2001-5 

Insulin stimulates testosterone biosynthesis by human thecal cells from women

with polycystic ovary syndrome by activating its own receptor and using

inositolglycan mediators as the signal transduction system.

Nestler JE, Jakubowicz DJ, de Vargas AF, Brik C, Quintero N, Medina F.

Department of Internal Medicine, Medical College of Virginia/Virginia

Commonwealth University, Richmond 23298, USA. nestler@hsc.vcu.edu

To determine whether insulin stimulates human ovarian testosterone production in

the polycystic ovary syndrome by activating its own receptor and using

inositolglycan mediators as the signal transduction system, thecal cells from

polycystic ovary syndrome women were isolated and cultured. Insulin and

insulin-like growth factor I stimulated thecal testosterone biosynthesis.

Antibody blockade of the insulin receptor abolished insulin's stimulatory

action, whereas effective antibody blockade of the insulin-like growth factor I

receptor did not alter insulin's stimulation of thecal testosterone

biosynthesis. A chiro-inositol containing glycan (INS-2) increased thecal

testosterone biosynthesis. Preincubation of cells with an antiinositolglycan

antibody (A23939 or alpha IGP) abolished insulin's stimulatory effect, but not

that of hCG. These findings suggest that inositolglycans serve as the signal

transduction system for insulin's stimulation of human thecal testosterone

biosynthesis.

PMID: 9626131 [PubMed - indexed for MEDLINE]

53: J Biol Chem  1998 Apr 17;273(16):9994-10003 

Insulin receptor substrate-1 is the predominant signaling molecule activated by

insulin-like growth factor-I, insulin, and interleukin-4 in estrogen

receptor-positive human breast cancer cells.

Jackson JG, White MF, Yee D.

Division of Medical Oncology, Department of Medicine, University of Texas Health

Science Center, San Antonio, Texas 78284-7884, USA.

Because insulin-like growth factor-I (IGF-I), insulin, and interleukin-4 (IL-4)

have known biological effects in breast cancer cells and signal through

insulin-receptor substrate (IRS) adaptor proteins, we examined the expression

and function of IRS-1 and IRS-2 in breast tumors and cell lines. IRS-1 and IRS-2

were expressed by cell lines and primary breast tumor specimens. IGF-I, insulin,

and IL-4 treatment of MCF-7 and ZR-75, and IGF-I treatment of T47-D breast

cancer cells, resulted in much greater tyrosine phosphorylation of IRS-1

compared with IRS-2. Furthermore, IGF-I stimulated greater tyrosine

phosphorylation of IRS-1 than either insulin or IL-4. IGF-I treatment also

enhanced association of the p85 regulatory subunit of phosphatidylinositol

3-kinase with IRS-1 and stimulated increased enzymatic activity compared with

IL-4 and insulin in all three cell lines. Similarly, mitogen-activated protein

kinase activity was greater in IGF-I-stimulated cells. To determine the

functional significance of the activation of these pathways, we inhibited

activation of phosphatidylinositol 3-kinase with wortmannin and

mitogen-activated protein kinase with PD098059. Both compounds inhibited

IGF-stimulated growth, suggesting that both pathways contributed to the

mitogenic response to IGF-I. We conclude that IRS-1, and not IRS-2, is the

predominant signaling molecule activated by IGF-I, insulin, and IL-4.

Furthermore, enhanced tyrosine phosphorylation of IRS-1 by IGF-I, compared with

either insulin or IL-4, is associated with greater activation of mitogenic

downstream signaling pathways resulting in enhanced cell growth.

PMID: 9545345 [PubMed - indexed for MEDLINE]

54: Surgery  1998 Mar;123(3):315-20 

Variable effect of streptozotocin-diabetes on the growth of hamster pancreatic

cancer (H2T) in the Syrian hamster and nude mouse.

Fisher WE, Muscarella P, Boros LG, Schirmer WJ.

Department of Surgery, Ohio State University, Columbus 43210, USA.

BACKGROUND: Streptozotocin-diabetes prevents induction of pancreatic tumors in

several animal models and inhibits the growth of established human pancreatic

cancer implants in nude mice. However, it also promotes growth of the hamster

pancreatic cancer cell line, H2T, in the Syrian hamster. To test the hypothesis

that these contradictory effects are due to tumor host differences, the growth

of the H2T cell line was examined in the streptozotocin-diabetic nude mouse.

METHODS: H2T cells were implanted subcutaneously into streptozotocin-diabetic

nude mice (n = 10) and untreated control mice (n = 10). After 21 days, tumors

were excised and weighed. Plasma insulin and somatostatin were determined by

radioimmunoassay. RESULTS: After 3 weeks, tumors in the control group weighed

118 mg and tumors in the diabetic group weighed 28 mg (p < 0.001). Plasma

insulin was significantly decreased in the streptozotocin-treated animals

compared with control animals (insulin, 23 microU/ml vs 31 microU/ml; p <

0.001). In contrast, somatostatin was significantly elevated in the

streptozotocin-diabetic group compared with the control group (somatostatin, 179

pg/ml versus 54 pg/ml, p < 0.001). Competitive binding studies revealed specific

cell surface receptors for insulin (Kd, 15.5 nmol/L), and somatostatin (Kd, 2.5

nmol/L) on the H2T cells. In an in vitro cell proliferation assay, cell division

was promoted by insulin (p < 0.01, maximum +11%) and inhibited by somatostatin

(p < 0.01, maximum -18%). CONCLUSIONS: The variable effect of

streptozotocin-diabetes on pancreatic cancer growth is due to differences in the

tumor host. The growth of pancreatic cancer, particularly in

streptozotocin-diabetic nude mice, may be influenced by gut peptides in a

receptor-dependent fashion.

PMID: 9526524 [PubMed - indexed for MEDLINE]

55: Cancer Res  1998 Mar 15;58(6):1159-64 

Elevated insulin-like growth factor I receptor autophosphorylation and kinase

activity in human breast cancer.

Resnik JL, Reichart DB, Huey K, Webster NJ, Seely BL.

Department of Medicine, University of California, San Diego, La Jolla 92093,

USA.

Insulin-like growth factor I action has been implicated in the pathogenesis of

many different malignancies, including breast cancer. Insulin-like growth factor

I receptors (IGF-IRs) are overexpressed in virtually all breast cancer cell

lines, in which they are believed to enhance growth and inhibit apoptosis. In

this study, the functional activity of IGF-IRs from normal and malignant human

breast tissue was assessed. IGF-IR expression was 14-fold higher in malignant

breast tissue than in normal breast tissue. IGF-IR autophosphorylation and

kinase activity were 2-4-fold higher in purified receptor preparations from

malignant breast tissue as compared to normal breast tissue when normalized for

receptor number. This increase in receptor function, coupled with the enhanced

receptor expression, amounts to a 40-fold elevation in IGF-IR tyrosine kinase

activity in malignant breast tissue. The enhanced receptor autophosphorylation

and kinase activity were observed in the absence of hormonal stimulation and

seem to result from an alteration in the intrinsic activity of the receptor

itself. Protein tyrosine phosphatase activity is also increased in malignant

breast tissue. These data suggest that the IGF-IR is an important target for

breast cancer therapy.

PMID: 9515800 [PubMed - indexed for MEDLINE]

56: Diabetes  1998 Jan;47(1):87-92 

Expression of insulin/IGF-I hybrid receptors is increased in skeletal muscle of

patients with chronic primary hyperinsulinemia.

Federici M, Lauro D, D'Adamo M, Giovannone B, Porzio O, Mellozzi M, Tamburrano

G, Sbraccia P, Sesti G.

Laboratory of Molecular Medicine, University of Rome, Italy.

The insulin receptor (IR) shares structural and functional homology with the

IGF-I receptor (IGF-IR). Hybrid receptors composed of an IR

alphabeta-heterodimer and an IGF-IR alphabeta-heterodimer are formed in tissues

expressing both molecules. Hybrids behave as IGF-IR rather than IR with respect

to ligand binding affinity, receptor autophosphorylation, and hormone

internalization and degradation. Factors regulating hybrid formation in vivo are

unknown. We recently reported that in skeletal muscle of NIDDM patients,

expression of hybrids is increased and correlated with a decrease in IR number

and an increase in fasting insulin levels. However, it is not clear whether

increased expression of hybrid receptors is a primary defect specifically

associated with NIDDM or a secondary event caused by hyperinsulinemia. To

address this issue, we used a quantitative microwell-based immunoassay to

measure hybrid receptor abundance in skeletal muscle of 11 normal subjects and

12 patients with insulinoma, a state of primary nongenetically determined

hyperinsulinemia. Total insulin binding was lower in insulinoma patients than in

normal subjects (0.70 +/- 0.18 vs. 4.59 +/- 0.77; P < 0.0001). Total IGF-I

binding did not differ between the two groups (0.81 +/- 0.27 and 0.85 +/- 0.10,

respectively). The amount of hybrids, expressed as bound/total (B/T), was higher

in patients with insulinoma than in normal subjects (0.57 +/- 0.19 vs. 0.36 +/-

0.03; P < 0.0006) and was inversely correlated with total insulin binding (r =

-0.64, P < 0.0004). Increased abundance of hybrid receptors was positively

correlated with insulin levels (r = -0.82, P < 0.0009) and inversely correlated

with insulin-mediated glucose uptake (r = -0.80, P < 0.01). No correlations were

observed between insulin-mediated glucose uptake and maximal specific insulin

binding (r = 0.19, P = 0.64). These results indicate that insulin-induced IR

downregulation may lead to the formation of a higher proportion of hybrid

receptors, whose abundance is negatively correlated with in vivo insulin

sensitivity. These results, therefore, support a role for insulin in the

regulation of hybrid receptors formation and suggest that increased expression

of hybrids in NIDDM may be a secondary event caused by hyperinsulinemia rather

than a primary defect.

PMID: 9421379 [PubMed - indexed for MEDLINE]

57: J Soc Gynecol Investig  1994 Jan-Mar;1(1):74-8 

Enhanced post-receptor insulin effects in women following dehydroepiandrosterone

infusion.

Schriock ED, Buffington CK, Givens JR, Buster JE.

Department of Obstetrics and Gynecology, University of California, San

Francisco, USA.

OBJECTIVE: We hypothesized that intravenous dehydroepiandrosterone (DHEA) would

decrease insulin resistance in normal and insulin-resistant women. METHODS: Five

insulin-resistant women diagnosed as having polycystic ovaries (PCO) with

elevated testosterone and normal dehydroepiandrosterone sulfate (DHEAS) with

amenorrhea were recruited. Obese controls (OC) with normal menses and normal

testosterone and DHEAS were recruited and matched to each PCO woman for age and

weight. The PCO women had a mean testosterone of 3.2 +/- 0.4 nmol/L, fasting

serum insulin level of 330 +/- 55 pmol/L, and DHEAS level of 3.4 +/- 1.3

mumol/L. An oral glucose tolerance test (OGTT) was performed at 8 AM after an

overnight fast. A DHEA infusion (1 mg/hour for 17 hours) was begun at 6 PM and

continued until the completion of the second OGTT performed the following

morning at 8 AM. T-lymphocytes were drawn at 8 AM each morning. RESULTS: The

DHEA infusion had no significant effect on any of the in vivo indices of insulin

sensitivity, ie, basal and OGTT insulin, C-peptide, and ratios of

insulin/glucose. In vitro, DHEA significantly increased insulin binding to

T-lymphocytes of PCO women but caused no significant change in OC women. There

was, however, marked enhancement of T-lymphocyte pyruvate dehydrogenase (PDH)

activities in both groups of study subjects following DHEA. CONCLUSION: We

conclude that a 17-hour infusion of DHEA enhanced T-lymphocyte insulin binding

and PDH activity while producing no detectable improvements in in vivo indices

of insulin sensitivity.

Publication Types:

Clinical Trial

Controlled Clinical Trial

PMID: 9419751 [PubMed - indexed for MEDLINE]

58: J Endocrinol Invest  1997 Oct;20(9):531-6 

Sporadic amplification of the insulin receptor gene in human breast cancer.

Papa V, Milazzo G, Goldfine ID, Waldman FM, Vigneri R.

Cattedra di Endocrinologia, Universita di Catania, Ospedale Garibaldi, Italy.

Insulin receptor (IR) content is increased in most human breast carcinomas when

compared to normal breast tissue. In the present study we investigated IR gene

copy number by using both conventional DNA analysis (slot blot) and fluorescence

in situ hybridization (FISH). Cultured human breast cell lines and primary

breast carcinoma specimens were analyzed. In 6 breast cell lines in culture both

techniques gave similar results: the relative IR copy number determined by FISH

strongly correlated with slot blot results (r = 0.831), even if probes for

different reference loci were used in the 2 methods. We find that in human

breast cancer IR gene amplification is a sporadic event. It occurred in 1/5

cultured breast cancer cell lines (MDA-MB 231) and in 8/93 (8.6%) breast cancer

specimens. In contrast an increased copy number of the entire chromosome 19

(which contains IR gene) was frequently observed in both breast cancer cell

lines (100%) and breast cancer specimens (45%). When present, IR gene

amplification always occurred at low level. These data indicate that IR gene

amplification is an uncommon event in human breast carcinomas and that

mechanisms other than gene amplification are responsible for IR protein

overexpression in most human breast cancers.

PMID: 9413807 [PubMed - indexed for MEDLINE]

59: Proc Assoc Am Physicians  1997 Nov;109(6):565-71 

Insulin receptor expression and clinical outcome in node-negative breast cancer.

Mathieu MC, Clark GM, Allred DC, Goldfine ID, Vigneri R.

Department of Medicine, Mount Zion Medical Center, University of California, San

Francisco 94143-1616, USA.

The insulin receptor (IR), a ligand-activated tyrosine kinase, is present in

breast cancers, but its relationship to patient survival is unknown. The IR was

measured in 584 tumor specimens from patients with node-negative breast

carcinoma by frozen-section immunohistochemistry and light microscopy. The

immunostaining signal was quantitated in relation to both the staining intensity

and the proportion of positive malignant epithelial cells. Analyses indicated

that patients with tumors with undetectable IR content in malignant epithelial

cells (260 cases) had a relatively lower predicted 5-year disease-free survival

(DFS) (69% +/- 3%) than did patients with tumors with detectable IR content (324

cases; DFS 76% +/- 3%, p = .032). The significance of IR content in these breast

malignant epithelial cells was then analyzed along with patient age, tumor size,

progesterone and estrogen receptor status, p53 accumulation, and S-phase.

Multivariate analysis of these data revealed that after adjustment for these

other variables, IR content was the strongest independent predictive factor for

DFS (relative risk = 1.73, p = .005). Interestingly, in a small subset of

patients with very high IR content (n = 62), DFS was decreased. These data

indicate that IR content in node-negative breast cancers is a significant major

predictor of reduced DFS. Moreover, they raise the possibility that the

measurement of IR content might provide important information concerning breast

cancer biology.

PMID: 9394418 [PubMed - indexed for MEDLINE]

60: Life Sci  1997;61(16):1583-92 

Bovine fetuin is an inhibitor of insulin receptor tyrosine kinase.

Mathews ST, Srinivas PR, Leon MA, Grunberger G.

Department of Internal Medicine, Wayne State University, Detroit, MI 48201, USA.

Fetuin has been identified earlier as the bovine homolog of the human plasma

protein, alpha2-Heremans Schmid glycoprotein (alpha2-HSG). Although bovine

fetuin shares over 70% amino acid sequence similarity with alpha2-HSG and rat

fetuin, no common function(s) have been identified. We report that

immunoaffinity purified bovine fetuin acts as an inhibitor of insulin receptor

tyrosine kinase activity (IR-TKA) with half-maximal inhibition at 1.5 microM. In

vitro, bovine fetuin (1.5 microM) blocked insulin-induced autophosphorylation of

the human IR completely and the half-maximal inhibitory effect was observed at

0.5 microM. Incubation of HIRcB cells (rat1 fibroblasts transfected with

wild-type human insulin receptor cDNA) with bovine fetuin (1.5 microM) inhibited

insulin-induced tyrosine phosphorylation of the IR beta-subunit by 40%. In

addition, bovine fetuin (2 microM) completely blocked insulin-stimulated DNA

synthesis in H-35 rat hepatoma cells. Our results, together with earlier reports

on rat fetuin and human alpha2-HSG, indicate a common IR-TK inhibitory function

for fetuin homologs.

PMID: 9353167 [PubMed - indexed for MEDLINE]

61: Endocrinology  1997 Nov;138(11):4821-9 

Vanadate, but not insulin, inhibits insulin receptor gene expression in rat

hepatoma cells.

Bortoli S, Amessou M, Collinet M, Desbuquois B, Lopez S.

Unite 30 INSERM, Hopital Necker-Enfants Malades, Paris, France.

Insulin and vanadate treatments have recently been shown to reverse the

overexpression of the hepatic insulin receptor (IR) gene in

streptozotocin-induced diabetic rats. To better understand the mechanisms

underlying these effects, the abilities of insulin and vanadate to affect IR

gene expression have been comparatively examined in Fao hepatoma cells, an

insulin-responsive cell line. Exposure of Fao cells to insulin (1 microM) or

vanadate (500 microM) for 24 h led to a 2-fold decrease in IR number in total

cellular membranes. Insulin treatment did not affect IR messenger RNA (mRNA)

level regardless of time of exposure and concentration. In contrast, vanadate

treatment caused a time- and dose-dependent decrease in IR mRNA level, which was

maximal (4-fold change) after a 24-h exposure to 500 microM vanadate and was

fully reversible. Insulin treatment increased from 28 to 39% the relative

expression of isotype A IR mRNA, but vanadate treatment did not significantly

affect this parameter. Vanadate treatment did not modify mRNA half-life (3.5 h)

in 5, 6 dichlorobenzimidazole riboside-treated cells but decreased by 4-fold the

transcriptional activity of the IR gene. These data show for the first time

that, although both insulin and vanadate decrease total cellular IR number in

Fao cells, only vanadate decreases IR mRNA level. It does so by inhibiting

transcription of the IR gene, suggesting an action on the gene promoter which

could be mediated by changes in the level of expression and/or of

phosphorylation of trans-acting factors.

PMID: 9348211 [PubMed - indexed for MEDLINE]

62: Breast Cancer Res Treat  1997 Sep;45(2):141-7 

The insulin receptor content is increased in breast cancers initiated by three

different oncogenes in transgenic mice.

Frittitta L, Cerrato A, Sacco MG, Weidner N, Goldfine ID, Vigneri R.

Istituto di Medicina Interna, Endocrinologia e Malattie del Metabolismo,

Universita di Catania, Ospedale Garibaldi, Italy.

The Insulin Receptor (IR) is a potential oncogene for mammary epithelial cells

since its content is increased in most human breast cancer specimens, and both

ligand-dependent malignant transformation and ligand-dependent enhanced growth

occurs in cultured breast cells overexpressing the IR. To better understand

whether the IR plays a role in mammary carcinogenesis which is independent of

other initiation factors, we measured IR content in transgenic mouse models of

breast cancer induced by 3 known oncogenes (Wnt-1, Neu, and Ret). Insulin

receptor content was measured by a specific radioimmunoassay. In normal mammary

gland tissues IR content was 14.6 +/- 1.4 ng/mg of protein (mean +/- SEM, n =

6). In the 3 cancers IR content was elevated (Neu = 36.1 +/- 4.6, n = 8, p <

0.002; Wnt-1 = 38.3 +/- 2.6, n = 13, p < 0.001; and Ret = 53.6 +/- 7.1, n = 7, p

< 0.001). These data indicate that IR overexpression, in addition to being a

potential oncogene, is increased in mouse tumors initiated by other oncogenes,

and therefore may also play a supportive role in the growth of breast cancers.

PMID: 9342439 [PubMed - indexed for MEDLINE]

1: Mol Cell Endocrinol  1998 Feb;137(2):177-86 

Insulin receptor substrate 1 antisense expression in an hepatoma cell line

reduces cell proliferation and induces overexpression of the Src homology 2

domain and collagen protein (SHC).

Taouis M, Dupont J, Gillet A, Derouet M, Simon J.

Endocrinologie de la Croissance et du Metabolisme, Station de Recherches

Avicoles, INRA, Nouzilly, France. taouis@tours.inra.fr

In mammalian cells, the insulin receptor substrate 1 protein (IRS-1) is a

specific substrate for insulin and IGF-1 receptor tyrosine kinases which is

involved in mediating metabolic and mitogenic actions of insulin and IGFs. In

order to determine if IRS-1 is also essential in a chicken derived hepatoma cell

line (LMH cells), IRS-1 gene has been invalidated in these cells. For this, we

subcloned chicken IRS-1 gene in an antisense orientation into a mammalian

expression vector driven by the cytomegalovirus early promoter. LMH cells were

stably transfected with this construct or with the empty vector carrying only

the neomycin resistance gene and selected for cIRS-1 expression. One subclone,

C2, showed a complete repression of cIRS-1 expression at both protein and mRNA

levels. Proliferation of C2 cells was dramatically reduced (54%) compared with

Neo(r) cells. Furthermore this reduction was accompanied by a decrease in

insulin-dependent [3H]thymidine incorporation, indicating a reduction in DNA

synthesis. Insulin-dependent [U-14C]glucose incorporation into cellular lipids

was also significantly reduced in C2 cell line suggesting an alteration in

lipogenesis. In wild type LMH cells, SHC which is involved in Ras pathway, also

served as a substrate for insulin receptor tyrosine kinase. In C2 cells, SHC

expression, its association with the insulin receptor and its tyrosine

phosphorylation were largely increased. Two forms of the regulatory subunit of

PI 3-kinase were present: p85 and p55 forms. Furthermore, C2 cells displayed

increased basal phosphatidylinositol (PI) 3'-kinase activity. This report

demonstrates a role for cIRS-1 in the metabolic and mitogenic actions of insulin

in LMH cells. However, the overexpression of cIRS-1 antisense did not completely

abolish cell proliferation. This may be explained by the exacerbation of an

alternative pathway that only partly compensate for the knocking out of cIRS-1

gene: the overexpression of SHC.

PMID: 9605520 [PubMed - indexed for MEDLINE]

2: J Biol Chem  1998 Apr 17;273(16):9994-10003 

Insulin receptor substrate-1 is the predominant signaling molecule activated by

insulin-like growth factor-I, insulin, and interleukin-4 in estrogen

receptor-positive human breast cancer cells.

Jackson JG, White MF, Yee D.

Division of Medical Oncology, Department of Medicine, University of Texas Health

Science Center, San Antonio, Texas 78284-7884, USA.

Because insulin-like growth factor-I (IGF-I), insulin, and interleukin-4 (IL-4)

have known biological effects in breast cancer cells and signal through

insulin-receptor substrate (IRS) adaptor proteins, we examined the expression

and function of IRS-1 and IRS-2 in breast tumors and cell lines. IRS-1 and IRS-2

were expressed by cell lines and primary breast tumor specimens. IGF-I, insulin,

and IL-4 treatment of MCF-7 and ZR-75, and IGF-I treatment of T47-D breast

cancer cells, resulted in much greater tyrosine phosphorylation of IRS-1

compared with IRS-2. Furthermore, IGF-I stimulated greater tyrosine

phosphorylation of IRS-1 than either insulin or IL-4. IGF-I treatment also

enhanced association of the p85 regulatory subunit of phosphatidylinositol

3-kinase with IRS-1 and stimulated increased enzymatic activity compared with

IL-4 and insulin in all three cell lines. Similarly, mitogen-activated protein

kinase activity was greater in IGF-I-stimulated cells. To determine the

functional significance of the activation of these pathways, we inhibited

activation of phosphatidylinositol 3-kinase with wortmannin and

mitogen-activated protein kinase with PD098059. Both compounds inhibited

IGF-stimulated growth, suggesting that both pathways contributed to the

mitogenic response to IGF-I. We conclude that IRS-1, and not IRS-2, is the

predominant signaling molecule activated by IGF-I, insulin, and IL-4.

Furthermore, enhanced tyrosine phosphorylation of IRS-1 by IGF-I, compared with

either insulin or IL-4, is associated with greater activation of mitogenic

downstream signaling pathways resulting in enhanced cell growth.

PMID: 9545345 [PubMed - indexed for MEDLINE]

3: Surgery  1998 Mar;123(3):315-20 

Variable effect of streptozotocin-diabetes on the growth of hamster pancreatic

cancer (H2T) in the Syrian hamster and nude mouse.

Fisher WE, Muscarella P, Boros LG, Schirmer WJ.

Department of Surgery, Ohio State University, Columbus 43210, USA.

BACKGROUND: Streptozotocin-diabetes prevents induction of pancreatic tumors in

several animal models and inhibits the growth of established human pancreatic

cancer implants in nude mice. However, it also promotes growth of the hamster

pancreatic cancer cell line, H2T, in the Syrian hamster. To test the hypothesis

that these contradictory effects are due to tumor host differences, the growth

of the H2T cell line was examined in the streptozotocin-diabetic nude mouse.

METHODS: H2T cells were implanted subcutaneously into streptozotocin-diabetic

nude mice (n = 10) and untreated control mice (n = 10). After 21 days, tumors

were excised and weighed. Plasma insulin and somatostatin were determined by

radioimmunoassay. RESULTS: After 3 weeks, tumors in the control group weighed

118 mg and tumors in the diabetic group weighed 28 mg (p < 0.001). Plasma

insulin was significantly decreased in the streptozotocin-treated animals

compared with control animals (insulin, 23 microU/ml vs 31 microU/ml; p <

0.001). In contrast, somatostatin was significantly elevated in the

streptozotocin-diabetic group compared with the control group (somatostatin, 179

pg/ml versus 54 pg/ml, p < 0.001). Competitive binding studies revealed specific

cell surface receptors for insulin (Kd, 15.5 nmol/L), and somatostatin (Kd, 2.5

nmol/L) on the H2T cells. In an in vitro cell proliferation assay, cell division

was promoted by insulin (p < 0.01, maximum +11%) and inhibited by somatostatin

(p < 0.01, maximum -18%). CONCLUSIONS: The variable effect of

streptozotocin-diabetes on pancreatic cancer growth is due to differences in the

tumor host. The growth of pancreatic cancer, particularly in

streptozotocin-diabetic nude mice, may be influenced by gut peptides in a

receptor-dependent fashion.

PMID: 9526524 [PubMed - indexed for MEDLINE]

4: Cancer Res  1998 Mar 15;58(6):1159-64 

Elevated insulin-like growth factor I receptor autophosphorylation and kinase

activity in human breast cancer.

Resnik JL, Reichart DB, Huey K, Webster NJ, Seely BL.

Department of Medicine, University of California, San Diego, La Jolla 92093,

USA.

Insulin-like growth factor I action has been implicated in the pathogenesis of

many different malignancies, including breast cancer. Insulin-like growth factor

I receptors (IGF-IRs) are overexpressed in virtually all breast cancer cell

lines, in which they are believed to enhance growth and inhibit apoptosis. In

this study, the functional activity of IGF-IRs from normal and malignant human

breast tissue was assessed. IGF-IR expression was 14-fold higher in malignant

breast tissue than in normal breast tissue. IGF-IR autophosphorylation and

kinase activity were 2-4-fold higher in purified receptor preparations from

malignant breast tissue as compared to normal breast tissue when normalized for

receptor number. This increase in receptor function, coupled with the enhanced

receptor expression, amounts to a 40-fold elevation in IGF-IR tyrosine kinase

activity in malignant breast tissue. The enhanced receptor autophosphorylation

and kinase activity were observed in the absence of hormonal stimulation and

seem to result from an alteration in the intrinsic activity of the receptor

itself. Protein tyrosine phosphatase activity is also increased in malignant

breast tissue. These data suggest that the IGF-IR is an important target for

breast cancer therapy.

PMID: 9515800 [PubMed - indexed for MEDLINE]

5: Diabetes  1998 Jan;47(1):87-92 

Expression of insulin/IGF-I hybrid receptors is increased in skeletal muscle of

patients with chronic primary hyperinsulinemia.

Federici M, Lauro D, D'Adamo M, Giovannone B, Porzio O, Mellozzi M, Tamburrano

G, Sbraccia P, Sesti G.

Laboratory of Molecular Medicine, University of Rome, Italy.

The insulin receptor (IR) shares structural and functional homology with the

IGF-I receptor (IGF-IR). Hybrid receptors composed of an IR

alphabeta-heterodimer and an IGF-IR alphabeta-heterodimer are formed in tissues

expressing both molecules. Hybrids behave as IGF-IR rather than IR with respect

to ligand binding affinity, receptor autophosphorylation, and hormone

internalization and degradation. Factors regulating hybrid formation in vivo are

unknown. We recently reported that in skeletal muscle of NIDDM patients,

expression of hybrids is increased and correlated with a decrease in IR number

and an increase in fasting insulin levels. However, it is not clear whether

increased expression of hybrid receptors is a primary defect specifically

associated with NIDDM or a secondary event caused by hyperinsulinemia. To

address this issue, we used a quantitative microwell-based immunoassay to

measure hybrid receptor abundance in skeletal muscle of 11 normal subjects and

12 patients with insulinoma, a state of primary nongenetically determined

hyperinsulinemia. Total insulin binding was lower in insulinoma patients than in

normal subjects (0.70 +/- 0.18 vs. 4.59 +/- 0.77; P < 0.0001). Total IGF-I

binding did not differ between the two groups (0.81 +/- 0.27 and 0.85 +/- 0.10,

respectively). The amount of hybrids, expressed as bound/total (B/T), was higher

in patients with insulinoma than in normal subjects (0.57 +/- 0.19 vs. 0.36 +/-

0.03; P < 0.0006) and was inversely correlated with total insulin binding (r =

-0.64, P < 0.0004). Increased abundance of hybrid receptors was positively

correlated with insulin levels (r = -0.82, P < 0.0009) and inversely correlated

with insulin-mediated glucose uptake (r = -0.80, P < 0.01). No correlations were

observed between insulin-mediated glucose uptake and maximal specific insulin

binding (r = 0.19, P = 0.64). These results indicate that insulin-induced IR

downregulation may lead to the formation of a higher proportion of hybrid

receptors, whose abundance is negatively correlated with in vivo insulin

sensitivity. These results, therefore, support a role for insulin in the

regulation of hybrid receptors formation and suggest that increased expression

of hybrids in NIDDM may be a secondary event caused by hyperinsulinemia rather

than a primary defect.

PMID: 9421379 [PubMed - indexed for MEDLINE]

6: J Soc Gynecol Investig  1994 Jan-Mar;1(1):74-8 

Enhanced post-receptor insulin effects in women following dehydroepiandrosterone

infusion.

Schriock ED, Buffington CK, Givens JR, Buster JE.

Department of Obstetrics and Gynecology, University of California, San

Francisco, USA.

OBJECTIVE: We hypothesized that intravenous dehydroepiandrosterone (DHEA) would

decrease insulin resistance in normal and insulin-resistant women. METHODS: Five

insulin-resistant women diagnosed as having polycystic ovaries (PCO) with

elevated testosterone and normal dehydroepiandrosterone sulfate (DHEAS) with

amenorrhea were recruited. Obese controls (OC) with normal menses and normal

testosterone and DHEAS were recruited and matched to each PCO woman for age and

weight. The PCO women had a mean testosterone of 3.2 +/- 0.4 nmol/L, fasting

serum insulin level of 330 +/- 55 pmol/L, and DHEAS level of 3.4 +/- 1.3

mumol/L. An oral glucose tolerance test (OGTT) was performed at 8 AM after an

overnight fast. A DHEA infusion (1 mg/hour for 17 hours) was begun at 6 PM and

continued until the completion of the second OGTT performed the following

morning at 8 AM. T-lymphocytes were drawn at 8 AM each morning. RESULTS: The

DHEA infusion had no significant effect on any of the in vivo indices of insulin

sensitivity, ie, basal and OGTT insulin, C-peptide, and ratios of

insulin/glucose. In vitro, DHEA significantly increased insulin binding to

T-lymphocytes of PCO women but caused no significant change in OC women. There

was, however, marked enhancement of T-lymphocyte pyruvate dehydrogenase (PDH)

activities in both groups of study subjects following DHEA. CONCLUSION: We

conclude that a 17-hour infusion of DHEA enhanced T-lymphocyte insulin binding

and PDH activity while producing no detectable improvements in in vivo indices

of insulin sensitivity.

Publication Types:

Clinical Trial

Controlled Clinical Trial

PMID: 9419751 [PubMed - indexed for MEDLINE]

7: J Endocrinol Invest  1997 Oct;20(9):531-6 

Sporadic amplification of the insulin receptor gene in human breast cancer.

Papa V, Milazzo G, Goldfine ID, Waldman FM, Vigneri R.

Cattedra di Endocrinologia, Universita di Catania, Ospedale Garibaldi, Italy.

Insulin receptor (IR) content is increased in most human breast carcinomas when

compared to normal breast tissue. In the present study we investigated IR gene

copy number by using both conventional DNA analysis (slot blot) and fluorescence

in situ hybridization (FISH). Cultured human breast cell lines and primary

breast carcinoma specimens were analyzed. In 6 breast cell lines in culture both

techniques gave similar results: the relative IR copy number determined by FISH

strongly correlated with slot blot results (r = 0.831), even if probes for

different reference loci were used in the 2 methods. We find that in human

breast cancer IR gene amplification is a sporadic event. It occurred in 1/5

cultured breast cancer cell lines (MDA-MB 231) and in 8/93 (8.6%) breast cancer

specimens. In contrast an increased copy number of the entire chromosome 19

(which contains IR gene) was frequently observed in both breast cancer cell

lines (100%) and breast cancer specimens (45%). When present, IR gene

amplification always occurred at low level. These data indicate that IR gene

amplification is an uncommon event in human breast carcinomas and that

mechanisms other than gene amplification are responsible for IR protein

overexpression in most human breast cancers.

PMID: 9413807 [PubMed - indexed for MEDLINE]

8: Proc Assoc Am Physicians  1997 Nov;109(6):565-71 

Insulin receptor expression and clinical outcome in node-negative breast cancer.

Mathieu MC, Clark GM, Allred DC, Goldfine ID, Vigneri R.

Department of Medicine, Mount Zion Medical Center, University of California, San

Francisco 94143-1616, USA.

The insulin receptor (IR), a ligand-activated tyrosine kinase, is present in

breast cancers, but its relationship to patient survival is unknown. The IR was

measured in 584 tumor specimens from patients with node-negative breast

carcinoma by frozen-section immunohistochemistry and light microscopy. The

immunostaining signal was quantitated in relation to both the staining intensity

and the proportion of positive malignant epithelial cells. Analyses indicated

that patients with tumors with undetectable IR content in malignant epithelial

cells (260 cases) had a relatively lower predicted 5-year disease-free survival

(DFS) (69% +/- 3%) than did patients with tumors with detectable IR content (324

cases; DFS 76% +/- 3%, p = .032). The significance of IR content in these breast

malignant epithelial cells was then analyzed along with patient age, tumor size,

progesterone and estrogen receptor status, p53 accumulation, and S-phase.

Multivariate analysis of these data revealed that after adjustment for these

other variables, IR content was the strongest independent predictive factor for

DFS (relative risk = 1.73, p = .005). Interestingly, in a small subset of

patients with very high IR content (n = 62), DFS was decreased. These data

indicate that IR content in node-negative breast cancers is a significant major

predictor of reduced DFS. Moreover, they raise the possibility that the

measurement of IR content might provide important information concerning breast

cancer biology.

PMID: 9394418 [PubMed - indexed for MEDLINE]

9: Life Sci  1997;61(16):1583-92 

Bovine fetuin is an inhibitor of insulin receptor tyrosine kinase.

Mathews ST, Srinivas PR, Leon MA, Grunberger G.

Department of Internal Medicine, Wayne State University, Detroit, MI 48201, USA.

Fetuin has been identified earlier as the bovine homolog of the human plasma

protein, alpha2-Heremans Schmid glycoprotein (alpha2-HSG). Although bovine

fetuin shares over 70% amino acid sequence similarity with alpha2-HSG and rat

fetuin, no common function(s) have been identified. We report that

immunoaffinity purified bovine fetuin acts as an inhibitor of insulin receptor

tyrosine kinase activity (IR-TKA) with half-maximal inhibition at 1.5 microM. In

vitro, bovine fetuin (1.5 microM) blocked insulin-induced autophosphorylation of

the human IR completely and the half-maximal inhibitory effect was observed at

0.5 microM. Incubation of HIRcB cells (rat1 fibroblasts transfected with

wild-type human insulin receptor cDNA) with bovine fetuin (1.5 microM) inhibited

insulin-induced tyrosine phosphorylation of the IR beta-subunit by 40%. In

addition, bovine fetuin (2 microM) completely blocked insulin-stimulated DNA

synthesis in H-35 rat hepatoma cells. Our results, together with earlier reports

on rat fetuin and human alpha2-HSG, indicate a common IR-TK inhibitory function

for fetuin homologs.

PMID: 9353167 [PubMed - indexed for MEDLINE]

10: Endocrinology  1997 Nov;138(11):4821-9 

Vanadate, but not insulin, inhibits insulin receptor gene expression in rat

hepatoma cells.

Bortoli S, Amessou M, Collinet M, Desbuquois B, Lopez S.

Unite 30 INSERM, Hopital Necker-Enfants Malades, Paris, France.

Insulin and vanadate treatments have recently been shown to reverse the

overexpression of the hepatic insulin receptor (IR) gene in

streptozotocin-induced diabetic rats. To better understand the mechanisms

underlying these effects, the abilities of insulin and vanadate to affect IR

gene expression have been comparatively examined in Fao hepatoma cells, an

insulin-responsive cell line. Exposure of Fao cells to insulin (1 microM) or

vanadate (500 microM) for 24 h led to a 2-fold decrease in IR number in total

cellular membranes. Insulin treatment did not affect IR messenger RNA (mRNA)

level regardless of time of exposure and concentration. In contrast, vanadate

treatment caused a time- and dose-dependent decrease in IR mRNA level, which was

maximal (4-fold change) after a 24-h exposure to 500 microM vanadate and was

fully reversible. Insulin treatment increased from 28 to 39% the relative

expression of isotype A IR mRNA, but vanadate treatment did not significantly

affect this parameter. Vanadate treatment did not modify mRNA half-life (3.5 h)

in 5, 6 dichlorobenzimidazole riboside-treated cells but decreased by 4-fold the

transcriptional activity of the IR gene. These data show for the first time

that, although both insulin and vanadate decrease total cellular IR number in

Fao cells, only vanadate decreases IR mRNA level. It does so by inhibiting

transcription of the IR gene, suggesting an action on the gene promoter which

could be mediated by changes in the level of expression and/or of

phosphorylation of trans-acting factors.

PMID: 9348211 [PubMed - indexed for MEDLINE]

11: Breast Cancer Res Treat  1997 Sep;45(2):141-7 

The insulin receptor content is increased in breast cancers initiated by three

different oncogenes in transgenic mice.

Frittitta L, Cerrato A, Sacco MG, Weidner N, Goldfine ID, Vigneri R.

Istituto di Medicina Interna, Endocrinologia e Malattie del Metabolismo,

Universita di Catania, Ospedale Garibaldi, Italy.

The Insulin Receptor (IR) is a potential oncogene for mammary epithelial cells

since its content is increased in most human breast cancer specimens, and both

ligand-dependent malignant transformation and ligand-dependent enhanced growth

occurs in cultured breast cells overexpressing the IR. To better understand

whether the IR plays a role in mammary carcinogenesis which is independent of

other initiation factors, we measured IR content in transgenic mouse models of

breast cancer induced by 3 known oncogenes (Wnt-1, Neu, and Ret). Insulin

receptor content was measured by a specific radioimmunoassay. In normal mammary

gland tissues IR content was 14.6 +/- 1.4 ng/mg of protein (mean +/- SEM, n =

6). In the 3 cancers IR content was elevated (Neu = 36.1 +/- 4.6, n = 8, p <

0.002; Wnt-1 = 38.3 +/- 2.6, n = 13, p < 0.001; and Ret = 53.6 +/- 7.1, n = 7, p

< 0.001). These data indicate that IR overexpression, in addition to being a

potential oncogene, is increased in mouse tumors initiated by other oncogenes,

and therefore may also play a supportive role in the growth of breast cancers.

PMID: 9342439 [PubMed - indexed for MEDLINE]