Amyloid Beta monomers regulate cyclic adenosine monophosphate response element binding protein functions by activating type-1 insulin-like growth factor receptors in neuronal cells.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder associated with synaptic dysfunction, pathological accumulation of ß-amyloid (Aß), and neuronal loss. The self-association of Aß monomers into soluble oligomers seems to be crucial for the development of neurotoxicity (J. Neurochem., 00, 2007 and 1172). Aß oligomers have been suggested to compromise neuronal functions in AD by reducing the expression levels of the CREB target gene and brain-derived neurotrophic factor (BDNF) (J. Neurosci., 27, 2007 and 2628; Neurobiol. Aging, 36, 2015 and 20406 Mol. Neurodegener., 6, 2011 and 60). We previously reported a broad neuroprotective activity of physiological Aß monomers, involving the activation of type-1 insulin-like growth factor receptors (IGF-IRs) (J. Neurosci., 29, 2009 and 10582, Front Cell Neurosci., 9, 2015 and 297). We now provide evidence that Aß monomers, by activating the IGF-IR-stimulated PI3-K/AKT pathway, induce the activation of CREB in neurons and sustain BDNF transcription and release.

MicroRNA-7 suppresses the homing and migration potential of human endothelial cells to highly metastatic human breast cancer cells.

BACKGROUND: MicroRNA-7 (miR-7) has been observed as a potent tumour suppressor in multiple cancer types including breast cancer. The aim of this study was to investigate the response sensitivities of metastatic breast cancer cells to miR-7 and the roles of miR-7 in the interaction of endothelial cells and metastatic cancer cells. METHODS: Expression profile of miRNAs in a breast cancer specimen cohort and breast cancer cells were determined using real-time quantitative miRNA assays. Effect of the altering expression of miR-7 on migration, invasion, proliferation, interaction and underlying molecular mechanism of breast cancer cells and endothelial cells was investigated after treatment with the synthesised mimic of miR-7. Luciferase activity analysis was performed to validate Wave-3 as a novel target of miR-7. RESULTS: miR-7 expression was negatively correlated with the stage, grade and survival of the breast cancer patients. There was also differential expression of miRNAs including miR-7 in the breast cancer cells. The synthesised mimic of miR-7 inhibits the motility and wound healing potential of breast cancer cells. The highly metastatic MDA-MB-231 cells are more sensitive to the miR-7 treatment than the poorly invasive MCF-7 cells. Treatment with miR-7 downregulated the expression of EGFR, IGF1R and Wave3 in MDA-MB-231 cells but not in MCF-7 cells. In addition, we further demonstrated that miR-7 inhibited the proliferation, migration and invasion of endothelial cells. And more importantly, miR-7 suppressed the homing and migration of endothelial cells to more aggressive tumour cell conditions. CONCLUSIONS: Given the dual inhibitory effect of miR-7 on metastatic breast cancer cells alone and the interaction of endothelial cells with the tumour-conditioned microenvironment, we suggest miR-7 may be a new therapeutic candidate for its capacity not only to prevent breast cancer cell spreading but also to inhibit tumour-associated angiogenesis in the metastatic breast cancer.

Adverse Events and Efficacy of Cixutumumab in Phase II Clinical Trials: A systematic Review and Meta-Analysis.

BACKGROUND AND OBJECTIVES: Cixutumumab is a monoclonal antibody targeting insulin-like growth factor 1 receptor (IGF1R). We sought to evaluate the efficacy of cixutumumab in the treatment of cancer, and to comprehensively assess the associated adverse events in phase II clinical trials. METHODS: Data were collected from PubMed, Embase, and Clinicaltrials.gov. The improvement on progression-free survival (PFS) was evaluated by hazard ratio (HR) and 95% confidence intervals (95% CIs). We also carried a meta-analysis to comprehensively evaluate the incidence of adverse events. RESULTS: The adverse events that were mentioned most frequently were hyperglycemia, anemia, nausea, fatigue, and thrombocytopenia. The most frequent adverse events were hyponatremia (40.28%), fatigue (35.18%), and skin rash (35.11%). Results showed that cixutumumab treatments did not benefit PFS (HR 1.03, 95% CI 0.83-1.26, p = 0.979). The complete response (CR) was rarely seen in phase II trials. CONCLUSIONS: Cixutumumab was well tolerated when used alone and in combination therapies, but its antitumor activity was low in the existing phase II clinical trials. An acceptable incidence of adverse effects supports further investigation of this drug, provided that it shows antitumor activity in combination with other drugs.

Assessment of insulin-like growth factor 1 receptor as an oncogene in esophageal squamous cell carcinoma and its potential implication in chemotherapy.

Insulin-like growth factor-1 receptor (IGF-1R) is a tyrosine kinase receptor implicated in the pathogenesis of multiple cancers. After ligand binding, IGF-1R can initiate the activation of the PI3K/AKT/mTOR and Ras/Raf/MEK/MAPK pathways to modulate cell proliferation, survival, differentiation, motility, invasion and angiogenesis. IGF-1R is a prerequisite for tumor progression and is one of the most attractive targets for therapeutic interventions in several types of cancer. In the present study, we determined the expression of IGF-1R in an esophageal squamous cell carcinoma (ESCC) cohort, investigated the detailed function of IGF-1R and screened the potential application of IGF-1R in the clinic. We verified the higher expression of IGF-1R in ESCC tumor tissues as compared to adjacent normal tissues. We also found that high expression of IGF-1R was associated with advanced tumor progression. We used ESCC cell lines and a mouse xenograft model to detect the function of IGF-1R in vitro and in vivo. Our results suggest the oncogenic function of IGF-1R in regulating cell proliferation, clonogenesis, the cell cycle and apoptosis. In addition, we found that IGF-1R was associated with the response to standard chemotherapy drugs 5-FU and cisplatin in an ESCC cell line. More importantly, we confirmed that the serum concentration of IGF-1/IGFBP3 can be used for predicting response to chemotherapy, and increased serum levels of IGF-1 and IGFBP-3 are associated with significantly higher rates of tumor response. In the present study, we demonstrated that IGF-1R is an important oncogene in ESCC and can be used to detect the chemotherapeutic response.

Targeting the insulin-like growth factor receptor: developing biomarkers from gene expression profiling.

Overwhelming evidence implicates insulin-like growth factor (IGF) signaling in the growth and survival of many types of human cancer cells. Numerous inhibitors of the IGF1 receptor (IGF1R) have been developed, and they displayed remarkable antineoplastic activity in preclinical models and promising success in early phase clinical trials. However, while responses have been observed in numerous cancer types, they have occurred in a minority of patients, and serious toxicities have been observed. Identifying patients likely to benefit from anti-IGF1R therapy requires further characterizing the role of IGF1 signaling in various stages of tumorigenesis in order to identify critical downstream factors that may be used as predictors of response, or to serve as novel therapeutic targets. Recent microarray analyses have begun to unravel expression "signatures" specific for IGF1 that correlate with poor breast cancer prognosis and with response to anti-IGFIR inhibitors. In this review we briefly discuss the history of the IGF1 family in neoplasia, how it is targeted, results from clinical trials, and the quest for biomarkers that will predict response to IGF1R-targeted therapy.

Receptor tyrosine kinase and phosphoinositide-3 kinase signaling in malignant mesothelioma.

OBJECTIVE: The phosphoinositide-3 kinase signaling pathway is implicated in the development of malignancy and promotes cell-cycle progression and resistance to apoptosis. Malignant mesothelioma tumor specimens demonstrate high levels of the phosphoinositide-3 kinase downstream mediator phosphorylated Akt. Exposure of mesothelioma cell lines to LY294002, a phosphoinositide-3 kinase inhibitor, results in apoptotic cell death and decreased phosphorylated Akt in vitro and tumor burden reduction in vivo. Phosphoinositide-3 kinase is activated by cell-surface receptor tyrosine kinases. We sought to determine which receptors are present in mesothelioma and their role in cellular survival and phosphoinositide-3 kinase signaling. METHODS: Western blot analysis was performed to determine the relative expression of epidermal growth factor receptor, insulin-like growth factor receptor, and platelet-derived growth factor receptor in the mesothelioma cell lines I-45 and REN and the mesothelial line Met5a. After exposure of mesothelioma lines to kinase inhibitors, a cell viability assay was performed, cell-cycle analysis was performed to determine the percentage of apoptosis, and Western blot analysis was performed for phosphorylated Akt. RESULTS: Inhibition of epidermal growth factor receptor resulted in apoptotic cell death and Akt hypophosphorylation in mesothelioma cell lines. Insulin-like growth factor receptor inhibition led to apoptotic cell death without affecting Akt phosphorylation. Platelet-derived growth factor receptor inhibition did not affect cellular survival or phosphoinositide-3 kinase signaling. CONCLUSION: In malignant mesothelioma constitutive activation of phosphoinositide-3 kinase/Akt results in cellular survival and contributes to the malignant phenotype. We have demonstrated that epidermal growth factor receptor inhibition leads to apoptotic cell death through downregulation of phosphoinositide-3 kinase signaling in mesothelioma cell lines, whereas insulin-like growth factor receptor inhibition leads to apoptosis independent of phosphoinositide-3 kinase. Epidermal growth factor receptor, insulin-like growth factor receptor, and phosphoinositide-3 kinase inhibition might be clinically relevant in malignant mesothelioma.

L-carnitine and isovaleryl L-carnitine fumarate positively affect human osteoblast proliferation and differentiation in vitro.

Age-related bone loss is characterized by decreased osteoblast activity, possibly related to the reduction of energy production. Carnitine promotes energy availability and its concentration declines with age; Therefore, two Carnitine derivatives, L-carnitine fumarate (LC) and isovaleryl L-carnitine fumarate (Iso-V-LC), have been tested on several parameters of human osteoblasts in vitro. Both compounds significantly increased osteoblast activity, but the new compound Iso-V-LC was more efficient than LC at lower concentrations. They both significantly enhanced cell proliferation, [3H]-proline incorporation and the expression of collagen type I (COLLI), and the bone sialoproteins (BSPs) and osteopontin (OPN). The percentage of alkaline phosphatase (ALP)-positive cells and the secretion of osteocalcin were not modified by LC and Iso-V-LC. Both molecules increased the formation of mineralized nodules, but Iso-V-LC reached the maximum effect at a concentration 10-fold lower than that of LC. Furthermore, we showed that insulin-like growth factor (IGF)-I and IGF-II mRNA levels were not modified by the treatment. However, the two compounds induced an increase of insulin-like growth factor binding protein (IGFBP)-3 and a decrease of IGFBP-5 in both osteoblast lysates and the extracellular matrix (ECM). In conclusion these data suggest that carnitine and, in particular, its new derivative, Iso-V-LC supplementation in the elderly may stimulate osteoblast activity and decrease age-related bone loss.

Ectodomain shedding-dependent transactivation of epidermal growth factor receptors in response to insulin-like growth factor type I.

Diverse extracellular stimuli activate the ERK1/2 MAPK cascade by transactivating epidermal growth factor (EGF) receptors. Here, we have examined the role of EGF receptors in IGF-I-stimulated ERK1/2 activation in several cultured cell lines. In human embryonic kidney 293 cells, IGF-I triggered proteolysis of heparin binding (HB)-EGF, increased tyrosine autophosphorylation of EGF receptors, stimulated EGF receptor inhibitor (AG1478)-sensitive ERK1/2 phosphorylation, and promoted EGF receptor endocytosis. In a mixed culture system that employed IGF-I receptor null murine embryo fibroblasts (MEFs) (R(-) cells) to detect paracrine signals produced by MEFs expressing the human IGF-I receptor (R(+) cells), stimulation of R(+) cells provoked rapid activation of green fluorescent protein-tagged ERK2 in cocultured R(-) cells. The R(-) cell response was abolished by either the broad-spectrum matrix metalloprotease inhibitor batimastat or by AG1478, indicating that it resulted from the proteolytic generation of an EGF receptor ligand from adjacent R(+) cells. These data suggest that the paracrine production of EGF receptor ligands leading to EGF receptor transactivation is a general property of IGF-I receptor signaling. In contrast, the contribution of transactivated EGF receptors to IGF-I-stimulated downstream events, such as ERK1/2 activation, varies in a cell type-dependent manner.

The complex biology of the receptor for the insulin-like growth factor-1.

The receptor for the insulin-like growth factor-1 (IGF-1R) controls a wide variety of cellular functions. IGF-1R is a tyrosine kinase receptor that is activated by the binding of a ligand to the extracellular domain. There are three ligands for IGF-1R: IGF-I, IGF-II and insulin at supraphysiological concentrations. The binding of the ligand activates the catalytic activity of the tyrosine kinase domain, which in turn causes the autophosphorylation of IGF-1R. As a consequence of autophosphorylation, the activated IGF-1R sends a potent mitogenic signal to the cell nucleus. It has been clearly shown that an overexpressed and activated IGF-1R is quasi-obligatory for the establishment of a malignant cell phenotype. Interestingly, the targeting of IGF-1R can reverse the malignant phenotype in cancer cells, without affecting the biology of normal cells. For these reasons, IGF-1R seems to be a very promising candidate target for cancer therapy. In addition, IGF-1R protects cells from apoptosis and promotes cell growth and proliferation. However, more recent studies have also shown that IGF-1R regulates cell adhesion and motility, and, in certain cellular contexts, can induce differentiation. Indeed, the biology of the IGF-1R appears more complex than previously thought. The goal of this review is to describe the multiple functions displayed by IGF-1R in cell biology.

Oral glutamine (AES-14) supplementation inhibits PI-3k/Akt signaling in experimental breast cancer.

BACKGROUND: 7,12-dimethylbenz [a] anthracene (DMBA) administration to pubertal rats causes breast tumors and inhibits glutathione (GSH) production. Our previous results have established that oral glutamine (GLN) supplementation significantly reduced tumor development, restored the depressed GSH production, and caused a significant decrease in the circulating levels of insulinlike growth factor-1 (IGF-1). The present study was designed to investigate the involvement of the IGF-1-activated phosphatidylinositol 3 kinase (PI-3K)/Akt apoptotic signaling pathway. MATERIALS AND METHODS: Forty female Sprague-Dawley rats were randomly divided into 4 groups: DMBA+GLN (n = 16), DMBA+water (n = 8), Oil+GLN (n = 8) and Oil+water (n = 8). At the age of 50 days, rats received a single dose of 100 mg/kg DMBA (n = 24) or sesame oil (n = 16) and were gavaged with a GLN suspension formulation (AES-14) or water for the duration of the entire experiment. The animals were killed 11 weeks after the DMBA application, and the levels of IGF-1, IGF-1 receptor (IGF-IR), Akt, Bcl-2 and Bad in tumorous and nontumorous breast tissue samples were measured by Western blot analysis. RESULTS: GLN supplementation resulted in a significant decrease in the levels of IGF-1, IGF-IR, Akt, and Bcl-2 in nontumorous samples. At the same time, the levels of pro-apoptotic protein Bad were significantly elevated. The samples collected from tumor tissues showed lower levels of IGF-1, Akt, Bcl-2, Bad, and IGF-IR in comparison with nontumorous tissues. CONCLUSIONS: GLN supplementation inhibited the PI-3K/Akt pathway that is thought to be important in increasing cell survival during tumorigenesis. These results are in agreement with our hypothesis that GLN counteracts the effects of DMBA and blocks carcinogenesis in vivo.

Suppression of insulin-like growth factor signalling pathway and collagen expression in keloid-derived fibroblasts by quercetin: its therapeutic potential use in the treatment and/or prevention of keloids.

BACKGROUND: Keloids are characterized by abnormal proliferation of fibroblasts and overproduction of collagen. Insulin-like growth factor (IGF)-I is mitogenic for fibroblasts and a stimulatory factor for collagen synthesis. OBJECTIVES: We have assessed the in vitro effects of quercetin on proliferation, collagen synthesis and the expression of the IGF system in keloid-derived fibroblasts. METHODS: Fibroblasts were isolated from earlobe keloids and exposed to quercetin at different concentrations. The inhibitory effects of quercetin on fibroblast proliferation were assayed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, Western and Northern blot analyses. RESULTS: Quercetin inhibited keloid fibroblast (KF) proliferation in a dose-dependent manner. Significant growth inhibition was observed on day 2 of culture. The dose required for 50% growth inhibition was approximately 25 microg mL-1. Collagen 1 expression was significantly decreased while collagen 3 was almost undetectable following quercetin treatment. Basal levels of IGF-I receptor (IGF-IR) beta subunits, p85 subunit of phosphatidylinositol 3-kinase, c-Raf, phospho-Raf-1, phospho-MEK 1/2, phospho-mitogen-activated protein kinase, phospho-Elk-1 and phospho-Akt-1 were significantly reduced when KF cells were exposed to quercetin for 24 h. Blocking IGF-IR activity with IGF-IR antibody or neutralizing endogenous IGF-I activity with IGF-I antibody led to significant growth inhibition suggesting the role of IGF-I in regulation of KF proliferation. CONCLUSIONS: Because the IGF system plays an important part in fibroblast cell proliferation and collagen production, the described activities of quercetin on the IGF system and collagen expression may provide a novel approach for the use of quercetin in treatment and/or prevention of hypertrophic scar and keloid.

Expression of insulin-like growth factor system constituents in differentiating rat osteoblastic cell populations.

The synthetic glucocorticoid dexamethasone (Dex) and insulin-like growth factor-I and -II (IGF-I and -II) stimulate osteoprogenitor proliferation and differentiation in bone cell populations isolated from adult rat vertebrae. Since glucocorticoids have been shown to regulate gene expression of IGFs and IGF binding proteins (IGFBPs) in several experimental models, we investigated whether Dex-stimulated osteoprogenitor proliferation and differentiation was associated with changes in mRNA levels of the IGF system components (i.e., IGF-I and -II, the type 1 and 2 IGF receptor, the insulin receptor and six IGFBPs). Osteoprogenitor-containing bone cell populations were isolated from the outgrowth of vertebral explant cultures of 3-month-old female rats and cultured for 20 days. Total RNA was extracted at day 8, 14, and 20, and mRNA levels of the IGF system constituents were compared between differentiating (Dex-treated) and non-differentiating (control) cultures. Northern hybridization data from 8- and 20-day cultures showed that mRNA levels of IGF-I were markedly lower in Dex-treated cultures than in control cultures at day 8 and 20. At day 20, mRNA levels of IGFBP-3 were also lower in Dex-treated cultures. Signals of IGFBP-5 mRNA were undetectable. To increase the sensitivity of our detection methods and therefore evaluate mRNA levels of all the components of the IGF system, we performed reverse transcription-polymerase chain reaction (RT-PCR) analysis of RNA extracted at day 8, 14, and 20 of culture. In agreement with the Northern data, IGF-I mRNA levels in Dex-treated cultures were lower than in control cultures at all three time points, and IGFBP-3 levels were lower in Dex-treated cultures at day 20 of culture. However, at day 8 and 14, IGFBP-3 mRNA levels were higher in Dex-treated cultures than in controls. Levels of the 2 IGF receptor mRNA and the insulin receptor mRNA were lower in Dex-treated cultures. Dex-treated cultures also had decreased levels of IGFBP-1 mRNA but increased levels of IGFBP-2 mRNA at all three time points. IGFBP-4 levels were lower at day 14 in Dex-treated cultures than in controls but higher at day 20. IGF-II and IGFBP-5 mRNA levels in control and Dex-treated cultures were similar. Signals for IGFBP-6 were undetectable. Our findings show that glucocorticoid-induced osteoprogenitor proliferation and differentiation in adult rat bone cell populations are associated with significant changes in the mRNA levels of virtually all components of the IGF system. Some of these changes are dependent on the stages of development (e.g., regulation of IGFBP-3 and -4) and some remain similar trends at all stages (e.g., regulation of IGF-I and the three receptors).

Enhanced proliferation of human fibroblasts, in the presence of dexamethasone, is accompanied by changes in p21Waf1/Cip1/Sdi1 and the insulin-like growth factor type 1 receptor.

The addition of dexamethasone (dex) to human fibroblast cultures has been found to elicit enhanced proliferation. This enhancement is manifested by an increase in the initial growth rate, saturation density, and proliferative life span of WI-38 fibroblast cultures grown in the presence of dex. We examined the acute effects of dex on a number of growth-related genes in WI-38 cells. Our results show a decrease in the level of the cyclin-dependent kinase inhibitor p21Waf1/Cip1/sdi1 in response to dex. In addition, the level of the insulin-like growth factor type 1 receptor (IGF-1R) is increased in dex-treated cells. These changes are correlated with changes in the activity of the p21waf1/Cip1/Sdi1 and IGF-1R promoters. The results presented in this report suggest that dex may delay growth arrest in response to contact inhibition, as well as during cellular senescence. Thus, dex may act at multiple levels to enhance cellular proliferation in WI-38 cells: first, to decrease the level of an inhibitor of cell-cycle progression, and second, to increase the sensitivity of WI-38 cells to the proliferative effects of IGF-1. These acute effects may cooperate with other, as yet uncharacterized effects, to result in the enhanced proliferation seen in the presence of dex.

Interleukin-1 blocks insulin and insulin-like growth factor-stimulated growth in MCF-7 human breast cancer cells by inhibiting receptor tyrosine kinase activity.

Interleukins-1 (IL-1s) are known to inhibit the growth of cultured breast cancer cells. We examined the effects of IL-1 alpha and IL-1 beta on insulin and insulin-like growth factor I (IGF-I) stimulation of cell growth and found that both IL-1s inhibited anchorage-dependent and independent growth of MCF-7 breast cancer cells. In cells incubated with IL-1 beta (100 U/ml), insulin receptor (IR) protein and messenger RNA were increased by 100%, while IGF-I receptor protein and transcript were not significantly changed. These data were confirmed by binding studies. Incubation of MCF-7 cells with IL-1s led, however, to a significant inhibition of IR and IGF-I receptor autophosphorylation (-55%) and phosphotransferase activity (-65%). Also, in 3T3/ HIR rat fibroblasts, transfected with and overexpressing IR, IL-1s decreased insulin-stimulated cell growth in soft agar and IR tyrosine kinase activity. The present findings suggest that IL-1s antagonize the insulin and IGF-I mitogenic effects in MCF-7 cells by blocking the receptor tyrosine kinase activity that is crucial for the mitogenic effect of these factors.

The early intracellular signaling pathway for the insulin/insulin-like growth factor receptor family in the mammalian central nervous system.

Several studies support the idea that the polypeptides belonging to the family of insulin and insulin-like growth factors (IGFs) play an important role in brain development and continue to be produced in discrete areas of the adult brain. In numerous neuronal populations within the olfactory bulb, the cerebral and cerebellar cortex, the hippocampus, some diencephalic and brainstem nuclei, the spinal cord and the retina, specific insulin and IGF receptors, as well as crucial components of the intracellular receptor signaling pathway have been demonstrated. Thus, mature neurons are endowed with the cellular machinery to respond to insulin and IGF stimulation. Studies in vitro and in vivo, using normal and transgenic animals, have led to the hypothesis that, in the adult brain, IGF-I not only acts as a trophic factor, but also as a neuromodulator of some higher brain functions, such as long-term potentiation and depression. Furthermore, a trophic effect on certain neuronal populations becomes clearly evident in the ischemic brain or neurodegenerative disorders. Thus, the analysis of the early intracellular signaling pathway for the insulin/IGF receptor family in the brain is providing us with new intriguing findings on the way the mammalian brain is sculpted and operates.

Involvement of Raf-1 kinase and protein kinase C zeta in insulin-like growth factor I-induced brown adipocyte mitogenic signaling cascades: inhibition by cyclic adenosine 3',5'-monophosphate.

In the present study we have examined the signaling cascades involved in insulin-like growth factor I (IGF-I)-induced mitogenesis in fetal rat brown adipocyte primary cultures, a model that constitutively expresses a high number of IGF-I receptors, where IGF-I is a complete mitogen at physiological concentrations. IGF-I rapidly stimulated beta-chain IGF-I receptor autophosphorylation, which peaked at a physiological/mitogenic concentration (1.4 nM) and also stimulated tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1). Tyrosine-phosphorylated IRS-1 bound and subsequently activated phosphatidylinositol 3-kinase by 3.5-fold, whereas the tyrosine-phosphorylated IGF-I receptor was not directly associated with the p85 subunit of the phosphatidylinositol 3-kinase. Moreover, mitogenic concentrations of IGF-I enhanced glucose transport by 2.5-fold. In addition, tyrosine phosphorylation of the 46- and 52-kDa SHC proteins was high in the basal state and doubled after IGF-I treatment, whereas IGF-I enhanced by 4-fold tyrosine phosphorylation of the 66-kDa SHC band. Furthermore, a 2-fold increase in the Ras. GTP active form was induced upon IGF-I stimulation. Downstream from Ras, IGF-I increased both Raf kinase and protein kinase C (PKC) zeta activities by 3.5-fold. (Bu)2cAMP, an inhibitor of IGF-I-induced mitogenesis in fetal brown adipocyte primary cultures, did not block the very early steps of the IGF-I-induced mitogenic cascade, such as IGF-I receptor autophosphorylation, IRS-1 or SHC tyrosine phosphorylation, and Ras activation to its GTP active form. However, (Bu)2cAMP disrupted IGF-I-Raf and IGF-I-PKC zeta signaling pathways by preventing IGF-I-induced Raf-1 kinase and PKC zeta enzymatic activities, respectively. Our results show the first characterization in situ of an IGF-I mitogenic signaling cascade that downstream Ras diverges to the nucleus through two different serine/threonine kinases (Raf-1 kinase and PKC zeta) in mammalian fetal primary cells under physiological conditions. Both kinases represent a point of regulation primarily described for IGF-I-induced, cAMP-inhibited mitogenic pathways.

Insulin-like growth factors in breast cancer.

Insulin-like growth factor (IGF)-I is one of the most potent mitogens to many breast cancer cell lines in vitro. Effective growth inhibition in vitro may be achieved by antibodies to the type I IGF receptor (IGF-IR) or by using antisense strategies. Most human breast cancers express IGF-IR in vivo. Thus, different therapeutic strategies aimed at inhibiting ligand stimulation of the IGF-IR may be an attractive treatment option against breast cancer. Several drugs commonly used in breast cancer influence the IGF system both in vitro and in vivo. While antioestrogens such as tamoxifen and droloxifene reduce the expression of IGF-IR in vitro and suppress plasma levels of IGF-I but elevate IGF-binding protein-1 in vivo, megestrol acetate may reduce the delivery of IGFs to the tissues by inhibition of IGFBP-3 protease activity.

Effect of insulin on area and Na+ channel density of apical membrane of cultured toad kidney cells.

1. The stimulation of transepithelial Na+ transport caused by insulin in A6 cultured toad kidney cells was investigated by determination of membrane capacitance (Cm), short circuit current (Isc) and current fluctuation analysis. Values of Cm are proportional to membrane area while blocker-induced current fluctuation analysis provides an estimate of the number of active amiloride-sensitive Na+ channels in the apical membrane. 2. Insulin simultaneously increased Cm, Isc and Gt (transepithelial conductance) in epithelia incubated with Na(+)-containing solutions on both sides. 3. Analysis of 6-chloro-3,5-diaminopyrazine-2-carboxamide (CDPC)-induced noise showed that insulin increased the number of active Na+ channels in the apical membrane, without altering the single channel current. 4. When nystatin was used to permeabilize the apical membrane the impedance data revealed the presence of a second time constant. Analysis of these data indicated that the basolateral membrane capacitance (Cb) is much larger than the apical membrane capacitance (Ca). Insulin administered to nystatin-treated epithelia increased the values for both capacitances. 5. We suggest that the stimulation of transepithelial Na+ transport caused by insulin may be associated with the exocytotic delivery of transporters to the apical membrane.

Effects of dietary insulin-like growth factor I on growth and insulin-like growth factor receptors in neonatal calf intestine.

Colostrum is rich in IGF-I and IGF-II, and the dietary effects of recombinant human (rh)IGF-I on the newborn are of interest. The objective of this study was to examine the effects of dietary rhIGF-I on intestinal tissue growth and populations of IGF receptors. Twenty-three male diary calves were fed one of three experimental diets: 1) milk replacer plus isolated colostrum-derived globulins (MR-), 2) same as 1 plus 750 ng of rhIGF-I/mL (MR+), or 3) pooled cow colostrum (COL). After the first four feedings, all calves received milk replacer without additional globulins; calves fed the MR+ diet continued to receive the addition of 750 ng of rhIGF-I/mL until the experiment ended at 7 d after birth. Calves were killed and intestinal tissue was collected for in vitro [3H]thymidine incorporation studies. Incorporation differed among intestinal regions (duodenum, jejunum, and ileum). The MR+ calves had greater (P < .01) [3H]thymidine incorporation per unit of DNA than either the COL or MR- calves (31.8 vs 18.6 and 11.5 x 10(3) dpm/microgram of DNA, respectively). Competitive binding analysis indicated the presence of specific type 1 and type 2 intestinal IGF receptors. The IGF-I was more potent than IGF-II and insulin at inhibiting [125I]rhIGF-I binding (ED50 was 1.84, 9.17, and 1.91 ng/mL, respectively). The IGF-II was the only ligand capable of inhibiting [125I]rhIGF-II binding (ED50 was .30 nmol/mL).(ABSTRACT TRUNCATED AT 250 WORDS)