Synthesis, induced-fit docking investigations, and in vitro aldose reductase inhibitory activity of non-carboxylic acid containing 2,4-thiazolidinedione derivatives.

In continuation of our studies, we here report a series of non-carboxylic acid containing 2,4-thiazolidinedione derivatives, analogues of previously synthesized carboxylic acids which we had found to be very active in vitro aldose reductase (ALR2) inhibitors. Although the replacement of the carboxylic group with the carboxamide or N-hydroxycarboxamide one decreased the in vitro ALR2 inhibitory effect, this led to the identification of mainly non-ionized derivatives with micromolar ALR2 affinity. The 5-arylidene moiety deeply influenced the activity of these 2,4-thiazolidinediones. Our induced-fit docking studies suggested that 5-(4-hydroxybenzylidene)-substituted derivatives may bind the polar recognition region of the ALR2 active site by means of the deprotonated phenol group, while their acetic chain and carbonyl group at position 2 of the thiazolidinedione ring form a tight net of hydrogen bonds with amino acid residues of the lipophilic specificity pocket of the enzyme.

The stimulus-secretion coupling of glucose-induced insulin release. Metabolic and functional effects of NH4+ in rat islets.

NH4+ caused a dose-related, rapid, and reversible inhibition of glucose-stimulated insulin release by isolated rat islets. It also inhibited glyceraldehyde-, Ba2+-, and sulfonylurea-stimulated insulun secretion. NH4+ failed to affect glucose utilization and oxidation, glucose-stimulated proinsulin biosynthesis, the concentration of ATP, AD, and AMP, and the intracellular pH. NH4+ also failed to affect the ability of theophylline and cytochalasin B to augment glucose-induced insulin release. However, in the presence and absence of glucose, accumulation of NH4+ in islet cells was associated with a fall in the concentration of NADH and HADPH and a concomitant alteration of 86Rb+ and 45Ca2+ (or 133Ba2+) handling. These findings suggest that reduced pyridine nucleotides, generated by the metabolism of endogenous of exogenous nutrients, may modulate ionophoretic processes in the islet cells and by doing so, affect the net uptake of Ca2+ and subsequent release of insulin.

Comparison of alpha- and beta-cell secretory responses in islets isolated with collagenase and in the isolated perfused pancreas of rats.

The inhibitory actions of somatostatin (100 ng./ml.) on insulin release, stimulated by high glucose (20 mM), and on glucagon release, stimulated by arginine (15 mM), were studied with two in vitro systems: the isolated perifused rat islets prepared by the collagenase procedure and the isolated perfused rat pancreas. Suppression of arginine-induced glucagon release by glucose (20 mM) and glyceraldehyde (5 mM) was also assessed in both systems. With the perfused pancreas, somatostatin caused 32 per cent inhibition of glucose-mediated insulin release and inhibited arginine-induced glucagon release by 72 per cent. In the same system, glucose and glyceraldehyde were similarly potent inhibitors of arginine-induced glucagon secretion. In contrast to the isolated perfused pancreas, there was no significant somatostation suppression of glucose-induced insulin release or arginine-induced glucagon release whether the inhibitor was present prior to or was added during stimulation by glucose or arginine. Furthermore, glucose was only minimally active and glyceraldehyde ineffective in inhibiting glucagon secretion due to arginine in the perifusion system. The most plausible explanation for the difference in the endocrine response of islet cells in the two types of widely used in vitro systems is that the alpha and beta cells have lost inhibitory receptors in the plasma membrane as a result of the collagenase isolation technic.

Isoflurane inhibits insulin secretion from isolated rat pancreatic islets of Langerhans.

We have investigated the effects of isoflurane on insulin secretion in vitro from rat isolated islets of Langerhans and found a significant, dose-related and reversible inhibition of insulin secretion. Isoflurane 2% decreased insulin secretion stimulated by glucose 20 mmol litre-1 to basal, nonstimulated values. In other studies to identify the stage in the stimulus secretion pathway for insulin at which the anaesthetic may exert an inhibitory action, we have stimulated insulin release using glyceraldehyde and a phorbol ester. Insulin secretion induced by these secretagogues was also blocked by isoflurane. This suggests that the inhibitory effect of the anaesthetic agent may be at a site distal to stimulation of insulin secretion by glyceraldehyde and phorbol esters.

Inhibitory effect of glycation on catalytic activity of alanine aminotransferase.

Non-enzymatic glycation is a common post-translational modification of tissue and plasma proteins which can impair their functions in living organisms. In this study, the authors have demonstrated for the first time an inhibitory effect of in vitro glycation on the catalytic activity of alanine aminotransferase (ALT, EC 2.6.1.2), a pyridoxal phosphate enzyme with several lysine residues in the molecule. The porcine heart enzyme was incubated with 50 mmol/l D-fructose, D-glucose, D,L-glyceraldehyde, or D-ribose in 0.1 mol/l phosphate buffer (pH 7.4) at 25 degrees C for up to 20 days. The strongest glycation effect was shown by D,L-glyceraldehyde, which caused complete enzyme inhibition within 6 days. After 20 days of incubation, the ALT activity in samples with D-fructose and D-ribose was less than 7% of the initial enzyme activity. A statistically significant effect of D-glucose on the enzymatic activity of ALT was not found. Incubation of ALT with D-fructose, D,L-glyceraldehyde and D-ribose minimized its catalytic activity both in the glycated and non-glycated fractions of the samples. Markedly higher activity was found in the glycated fraction with glucose. The inhibitory effect of glycation of ALT with D-fructose and D-ribose was found to be more intensive in the presence of L-alanine and weaker in the presence of 2-oxoglutarate. The findings suggest that glycation of the epsilon-amino group of Lys313 as a crucial part of the catalytic site of ALT may contribute to ALT inactivation in the presence of glycating sugars. Nevertheless, glycation of lysine residues outside the active center of ALT seems to be primary.