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Background and Aim: In type 2 diabetes, the interplay between cells and inflammatory mediators up-regulates CD36 expression in macrophages. The aim of this work was to investigate advanced glycation end products (AGE)-induced CD36 expression and its regulation by insulin in aortic vascular smooth muscle cells (VSMCs) from Goto-Kakisaki (GK) rats, a non-obese insulin model of both insulin resistance and type 2 diabetes. The context of overexpression of CD36 in aortas was also evaluated., Methods and Results: VSMCs were isolated and cultured from the aortas of GK rats and non-diabetic rats. The expression of proteins was evaluated by Western blot. The aortic production of superoxide anion (O(2)(.-)) was measured by luminescence on isolated tissue. AGEs and advanced oxidation protein products (AOPPs) were determined in plasma by fluorescence spectroscopy and spectrophotometry, respectively. AGE receptor (RAGE), NF-kappaB, and CD36 protein expression as well as O(2)(.-) production were higher in GK aortas than in control aortas, and AGEs and AOPPs were higher in GK plasma. In VSMCs from non-diabetic rats, insulin was able to reduce (10 nM) or suppress (100 nM) the protein overexpression of CD36 induced by AGEs-BSA. In contrast, in VSMCs from GK rats, insulin was unable to reduce AGEs-BSA-induced CD36 overexpression., Conclusions: The results suggest an overexpression of CD36 in VSMCs from GK rats and impaired control by insulin. In the context of increased plasma AGEs, aortic RAGE overexpression and increased oxidative stress markers, the data are compatible with an AGEs induced CD36 overexpression in diabetes.

In both type 2 diabetes and insulin-resistance syndromes, hyperglycemia and advanced glycation end products (AGEs) activate the transcription factor nuclear factor-kappaB (NF-kappaB) through a mechanism that partly involves the generation of reactive oxygen species (ROS). The contribution of hyperinsulinemia in this sequence has not been completely elucidated. In this work we investigated the actions of insulin and PPAR-gamma on the stimulation by AGEs of NF-kappaB protein expression in cultured aortic vascular smooth-muscle cells (VSMCs) from non-insulin-dependent diabetic rats and nondiabetic rats. The expression of proteins was evaluated with the use of Western immunoblotting. Incubations (24 hours) of VSMCs with 10 to 100 microg/mL glycated bovine serum albumin (AGE- BSA) increased NF-kappaB protein expression in both models. PPAR-gamma protein expression was only enhanced at concentrations of 500 to 1000 microg/mL (AGE-BSA). In the presence of insulin (10-100 nmol/L), the stimulation of NF-kappaB protein expression by AGE-BSA (100 microg/mL) was decreased, whereas the expression of PPAR-gamma, protein was enhanced. 15-Deoxyprostaglandin J2, a direct activator of PPAR-gamma, decreased AGE-BSA-stimulated NF-kappaB expression. These findings suggest that insulin decreases the incidence of alterations in VSMCs induced by AGEs through the reduction of NF-kappaB and an increase in PPAR-gamma protein expression (as far as the model could be extrapolated to in vivo situations). These data may help justify current therapeutic approaches involving the use of insulin and PPAR-gamma agonists.

Nephrocalcin is a urinary gamma-carboxyglutamic acid (Gla) containing protein that may be a physiological inhibitor of calcium oxalate nephrolithiasis. Nephrocalcin isolated from urine of stone formers seems to be abnormal in lacking Gla that is required for inhibitory activity. In order to study this hypothesis, we compared the protein-bound urinary Gla contents in 32 calcium oxalate stone formers and in 24 controls. Protein-bound Gla was resolved by reversed-phase high-performance liquid chromatography after elimination of free Gla, alkaline hydrolysis and precolumn derivatization with o-phthalaldehyde and mercaptoethanol. Protein-bound urinary Gla concentrations were similar in stone formers (0.83 +/- 0.38 mumol/l, mean +/- SD) and controls (0.81 +/- 0.27) and were less than 5% of free urinary Gla. However, excretion rates of free and protein-bound Gla (nmol/min) were higher in stone formers (P = 0.006 and P = 0.002). Positive correlations (P = 0.000) were observed between free and protein-bound Gla both in controls and in stone formers. These results do not support the hypothesis of a lacking Gla nephrocalcin in stone formers.

The best route of insulin administration by infusion pumps remains a subject of controversy. For that reason plasma lipids and apolipoproteins were compared in three groups of nine patients who had been treated for several months or years with conventional treatment (group I), continuous subcutaneous insulin infusion (CSII, group II) or continuous intraperitoneal insulin infusion (CPII, group III). Plasma cholesterol and apolipoprotein B remained increased on CPII compared with CSII even when similar satisfactory or even tight diabetic control was achieved with both techniques. This study suggests that cholesterol and perhaps apolipoprotein B biosynthesis by the liver is increased in patients treated with CPII compared to those treated with CSII.

We compared the fractional absorption of calcium (FACa, 6 h, % TD) and the radiocalcium transit (% TD per min) in seven glucocorticoid-treated patients (10-25 mg prednisolone per day) and in seven normal subjects, in the basal state and 12 h after an oral dose of synthetic 1,25-(OH)2D (3 micrograms). In the basal state, the radiocalcium transit was significantly decreased (P less than 0.02) at 15 min in patients treated with prednisolone, but FACa at 6 h was not significantly decreased (51 +/- 5 vs. 60 +/- 5% TD). 12 h after an oral dose of 1,25-(OH)2D which resulted in supraphysiologic plasma levels, FACa increased significantly (P less than 0.02) in both groups but the peak absorption rate of Ca remained lower in the corticoid-treated patients than in controls (P less than 0.02). The results suggest that glucocorticoids decrease the 1,25-(OH)2D-dependent transport of calcium across the proximal small intestine.

Under conditions in which vasoactive intestinal peptide (VIP) induces somatostatin release from cortical and diencephalic neuronal cultures, VIP causes large increases in intracellular cyclic AMP. Both the release of somatostatin and the increase in cyclic AMP elicited by VIP require exogenous calcium, can be blocked by cobalt ion, and can be qualitatively mimicked by depolarizating concentrations of exogenous potassium ion. Direct activation of adenylate cyclase by forskolin causes large increases in cyclic AMP content but does not induce somatostatin release. In the absence of VIP, the calcium ionophore, ionomycin, and the phorbol ester, phorbol 12-myristate-13-acetate, also stimulate somatostatin release. These results indicate that VIP-stimulation of cyclic AMP formation and VIP-stimulation of somatostatin release are calcium-dependent and that the two phenomena are dissociatable. Cyclic AMP formation is not a necessary condition for VIP-induced somatostatin release. Nucleotide formation may be a sufficient condition for release or, possibly in association with calcium influx, it may be an event unrelated to the release process.

The effect of somatostatin (SRIF) on VIP induction of SRIF secretion, cyclic AMP accumulation and 45Ca2+ influx was investigated in cultured diencephalic cells. [D-Trp8]SRIF suppressed VIP-stimulated SRIF release and decreased VIP-stimulated cyclic AMP accumulation in a dose-dependent manner. SRIF-14 blocked basal and VIP-stimulated 45Ca2+ entry into cells. The data suggest that the inhibitory effect of SRIF on VIP-induced SRIF release is partly due to a decrease in Ca2+ entry into cells.

In order to study the involvement of the hypothalamic corticotropin-releasing factor (CRF) in catecholamine-induced adrenocorticotropin (ACTH) secretion, we have measured CRF levels in rat hypophysial portal blood (HPB) after the pharmacological destruction of the ventral noradrenergic bundle (VNAB), using 6-hydroxydopamine (6-OHDA) stereotaxically injected into the VNAB. CRF levels in HPB were measured by radioimmunoassay, and the effects of 6-OHDA injection were controlled by the determination of catecholamine concentrations in the total hypothalamus. VNAB lesions induced a dramatic decrease in norepinephrine and epinephrine hypothalamic concentration. The CRF levels in HPB were also significantly reduced. These results suggest that central catecholamines exert a direct stimulatory control on the CRF release and play a major role in stress-induced ACTH secretion.