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Background: Maternal obesity represents a risk factor for pregnancy-related complications. Glucocorticoids are known to promote obesity in adults., Methods: We evaluated maternal and fetal metabolic changes during and after 3 weekly courses of betamethasone administered to pregnant baboons (Papio subspecies) at doses equivalent to those given to pregnant women., Results: Betamethasone administration during the second half of pregnancy increased maternal weight but neither maternal food intake nor fetal weight, as assessed at the end of gestation. Betamethasone increased maternal serum glucose concentration, the ratio of insulin-like growth factor-I and insulin-like growth factor binding protein-3, and serum leptin during treatment (normalized by 17, 35, and 45 days posttreatment, respectively, for each parameter). Maternal and fetal serum leptin concentrations did not differ between groups at the end of gestation., Conclusion: Prolonged maternal hyperleptinemia caused by betamethasone administration in the second half of gestation did not change fetal metabolic parameters measured and placental leptin distribution at the end of gestation.

Understanding mechanisms underlying apoptotic destruction of insulin-secreting cells is critical to validate therapeutic targets for type 1 diabetes mellitus. We recently reported insulin-like growth factor binding protein-3 (IGFBP-3) as a novel mediator of apoptosis in insulin-secreting cells. In light of emerging IGF-independent roles for IGFBP-3, we investigated the mechanisms underlying actions of the novel, recombinant human mutant G(56)G(80)G(81)-IGFBP-3, which lacks intrinsic IGF binding affinity. Using the rat insulinoma RINm5F cell line, we report the first studies in insulin-secreting cells that IGFBP-3 selectively suppresses multiple, key intracellular phosphorelays. By immunoblot, we demonstrate that G(56)G(80)G(81)-IGFBP-3 suppresses phosphorylation of c-raf-MEK-ERK pathway and p38 kinase in time-dependent and dose-dependent manners. SAPK/JNK signaling was unaffected. These data delineate several novel intracellular sites of action for IGFBP-3 in insulin-secreting cells.

Mesangial cells are critical for glomerular filtration. Mesangial cell dysfunction, the hallmark of diabetic nephropathy, results from disordered mesangial growth induced by cytokines, abnormal hemodynamic influence, and metabolic factors associated with chronic hyperglycemia. Insulin-like growth factors (IGFs) and their high affinity binding proteins (IGFBPs) exert major actions on mesangial cell survival, but their underlying mechanisms remain unclear. In light of emerging IGF-independent roles for IGFBP-3, we investigated IGFBP-3 actions during mesangial cell apoptosis induced by cytokine or high glucose concentration. Quantified by DNA fragmentation ELISA and Annexin V flow cytometry, apoptosis occurred in rat mesangial cells (RMC) exposed to 2 microg/mL IGFBP-3 for 24 h under high ambient or standard glucose. Anti-sense IGFBP-3 oligo at 10 microg/mL significantly inhibited apoptosis induced by 100 ng/mL TNF-alpha, serum-free conditions, or high (25 mM) glucose. Increased IGFBP-3 release associated with high ambient glucose or TNF-alpha was inhibited by pre-treatment with anti-sense oligo. Under serum-free conditions, recombinant human IGFBP-3 blocked Akt phosphorylation at threonine 308 (pThr308), whereas anti-sense oligo treatment was associated with enhanced pThr308 activity. In summary, these data support a novel mechanism for TNF-alpha-induced mesangial cell apoptosis mediated by IGFBP-3 and present regulation of pThr308 activity as a novel mechanism underlying IGFBP-3 action.

Insulin-like growth factor binding protein-3 (IGFBP-3) is emerging as a critical regulator of cell survival. There has been no study which directly examined the potential role for this major growth factor in the programmed cell death (apoptosis) of insulin-secreting cells. To determine whether IGFBP-3 mediates apoptosis in insulin-secreting cells, we performed a rigorous series of experiments with the rat insulinoma (RIN) cell line m5F and the hamster insulin-secreting tumor (HIT) T-15. Within 24 h exogenous IGFBP-3 induced significant DNA fragmentation in RIN and HIT cells, at doses ranging from 4.4 to 2000 ng/ml (P<0.05) without a classic dose-response relationship. DNA fragmentation induced by rhIGFBP-3 occurred in the presence of immunoglobulin to block the type 1 IGF receptor. As detected by flow cytometry for Annexin V exposure to the cell surface, rhIGFBP-3 treatment doubled the proportion of apoptotic HIT cells from 1.7 +/- 0.4% (serum-free control) to 3.4 +/- 0.2% (P<0.02), an effect completely reversed by co-treatment with 1000 ng/ml rhIGF-I. Immunofluorescent microscopy disclosed that pro-inflammatory Th1 cytokines increased intranuclear aggregation of endogenous IGFBP-3. Cytokine-induced DNA fragmentation was completely blocked by relatively brief pre-treatment with antisense IGFBP-3 phosphorothioate oligodeoxynucleotides. In conclusion, we have presented the first evidence that IGFBP-3 contributes to cytokine-mediated apoptosis in insulin-secreting cells.

Angiogenic factors, such as vascular endothelial-derived growth factor (VEGF) and IGF-I, play pivotal roles in endothelial proliferation and migration. IGF binding protein-3 (IGFBP-3) is emerging as a key regulator of cell growth and apoptosis, both as an IGF antagonist and as an independent molecule. We investigated the role of IGFBP-3 in VEGFmediated survival of human macrovascular umbilical vein endothelial cells (HUVEC). Specific commercial ELISAs quantified cell proliferation and apoptosis, and Akt phosphorylation was assessed by immunoblots and confocal microscopy. IGF-I and VEGF significantly stimulated HUVEC proliferation and survival. Addition of IGFBP-3 reversed both IGF- and VEGF-induced proliferation and prevented the survival induced by these factors. The antiproliferative and proapoptotic effects of exogenous IGFBP-3 upon VEGF-induced HUVEC survival were not inhibited by blockade of the type 1 IGF receptor with alpha IR-3 immunoglobulin, which fully prevented IGF actions. An IGFBP-3 mutant, which binds IGFs normally but has a substituted mid-region domain, lost the ability to inhibit VEGF actions. VEGF-induced phosphorylation of Akt, as evident by both specific immunoblots and confocal microscopy, was significantly and rapidly reduced in the presence of IGFBP-3, as well as wortmannin.