Anna Pleskačová

Vitamin D Status in Women with Gestational Diabetes Mellitus during Pregnancy and Postpartum.

Of many vitamin D extraskeletal functions, its modulatory role in insulin secretion and action is especially relevant for gestational diabetes mellitus (GDM). The aims of the present study were to determine midgestational and early postpartum vitamin D status in pregnant women with and without GDM and to describe the relationship between midgestational and postpartum vitamin D status and parallel changes of glucose tolerance. A total of 76 pregnant women (47 GDM and 29 healthy controls) were included in the study. Plasma levels of 25(OH)D were measured using an enzyme immunoassay. Vitamin D was not significantly decreased in GDM compared to controls during pregnancy; however, both groups of pregnant women exhibited high prevalence of vitamin D deficiency. Prevalence of postpartum 25(OH)D deficiency in post-GDM women remained significantly higher and their postpartum 25(OH)D levels were significantly lower compared to non-GDM counterparts. Finally, based on the oGTT repeated early postpartum persistent glucose abnormality was ascertained in 15% of post-GDM women; however, neither midgestational nor postpartum 25(OH)D levels significantly differed between subjects with GDM history and persistent postpartum glucose intolerance and those with normal glucose tolerance after delivery.

Simultaneous Determination of Uric Acid, Xanthine and Hypoxanthine in Human Plasma and Serum by HPLC-UV: Uric Acid Metabolism Tracking

A simple reversed phase HPLC method with UV detection in isocratic conditions was developed and validated for the simultaneous determination of hypoxanthine, xanthine and uric acid levels in human plasma and serum. One analysis run takes 6.5xa0min including a short organic mobile phase gradient for column regeneration. Concentrations of uric acid, xanthine and hypoxanthine in plasma and serum samples were highly comparable. However, hypoxanthine levels were increased in serum compared to plasma samples due to a prolonged time between serum and blood elements separation. The method was validated for linearity, precision, accuracy, sensitivity and robustness in a similar manner to that for pharmacokinetic data and it is appropriate for physiological and pathophysiological levels of all analytes. The stability of stock standard solutions was verified using spectrophotometric analysis in different conditions. The method is simple and robust with a good precision for the measurement of hypoxanthine, xanthine and uric acid in human plasma and serum.

Dysfunctional protection against advanced glycation due to thiamine metabolism abnormalities in gestational diabetes

While the pathogenic role of dicarbonyl stress and accelerated formation of advanced glycation end products (AGEs) to glucose intolerance and to the development of diabetic complications is well established, little is known about these processes in gestational diabetes mellitus (GDM), a condition pathogenically quite similar to type 2 diabetes. The aims of the present study were (i) to determine plasma thiamine and erythrocyte thiamine diphosphate (TDP) and transketolase (TKT) activity in pregnant women with and without GDM, (ii) to assess relationships between thiamine metabolism parameters and selected clinical, biochemical and anthropometric characteristics and, finally, (iii) to analyse relationship between variability in the genes involved in the regulation of transmembrane thiamine transport (i.e. SLC19A2 and SLC19A3) and relevant parameters of thiamine metabolism. We found significantly lower plasma BMI adjusted thiamine in women with GDM (Pxa0=xa00.002, Mann-Whitney) while levels of erythrocyte TDP (an active TKT cofactor) in mid-trimester were significantly higher in GDM compared to controls (Pxa0=xa00.04, Mann-Whitney). However, mid-gestational TKT activity - reflecting pentose phosphate pathway activity - did not differ between the two groups (Pxa0>xa00.05, Mann-Whitney). Furthermore, we ascertained significant associations of postpartum TKT activity with SNPs SLC19A2 rs6656822 and SLC19A3 rs7567984 (Pxa0=xa00.03 and Pxa0=xa00.007, resp., Kruskal-Wallis). Our findings of increased thiamine delivery to the cells without concomitant increase of TKT activity in women with GDM therefore indicate possible pathogenic role of thiamine mishandling in GDM. Further studies are needed to determine its contribution to maternal and/or neonatal morbidity.

Transketolase Activity but not Thiamine Membrane Transport Change in Response to Hyperglycaemia and Kidney Dysfunction

AIMnPentose phosphate pathway (PPP) with key enzyme transketolase (TKT), represents a potentially protective mechanism in hyperglycaemia. Diabetic kidney disease (DKD), a common complication of both type 1 and type 2 diabetes associated with significant morbidity and mortality, represents the most common cause of chronic kidney disease (CKD). We hypothesized that protective PPP action in diabetes and eventually even more severely in concomitant DKD might be compromised by limited intracellular availability of an active TKT cofactor thiamine diphosphate (TDP).nnnMETHODSnEffect of hyperglycaemia on gene expression and protein levels of key PPP loci was studied in vitro using human cell lines relevant to diabetes (HUVEC and HRGEC) and (together with measurement of TKT activity, plasma thiamine and erythrocyte TDP concentration) in vivo in diabetic vs. non-diabetic subjects with comparable renal function (n=83 in total).nnnRESULTSnHyperglycaemia significantly decreased protein levels of RFC-1, THTR1, THTR2 and TKT (P<0.05) in vitro. Analysis of blood samples from CKD patients with and without diabetes and from controls did not reveal any difference in gene expression and protein levels of thiamine transporters while TKT activity and TDP in erythrocytes gradually increased with decreasing kidney function being highest in patients with CKD3-4 of both diabetic and non-diabetic aetiology. Hyperglycaemia and uremic serum mimicking CKD in diabetes did not affect TKT activity in vitro (P<0.05).nnnCONCLUSIONnBoth in vitro and human experiments showed decrease or unchanged expression, respectively, of thiamine transporters induced by hyperglycaemia while TKT activity in parallel with intracellular TDP was increased in CKD patients with or without diabetes. Therefore, lack of adaptive increase of thiamine transmembrane transport allowing further increase of TKT activity might contribute to compromised PPP function in diabetes and CKD and to the development of glycotoxic injury.

Deleterious Effect of Advanced CKD on Glyoxalase System Activity not Limited to Diabetes Aetiology

Methylglyoxal production is increased in diabetes. Methylglyoxal is efficiently detoxified by enzyme glyoxalase 1 (GLO1). The aim was to study the effect of diabetic and CKD milieu on (a) GLO1 gene expression in peripheral blood mononuclear cells; (b) GLO1 protein levels in whole blood; and (c) GLO1 activity in RBCs in vivo in diabetic vs. non-diabetic subjects with normal or slightly reduced vs. considerably reduced renal function (CKD1-2 vs. CKD3-4). A total of 83 subjects were included in the study. Gene expression was measured using real-time PCR, and protein levels were quantified using Western blotting. Erythrocyte GLO1 activity was measured spectrophotometrically. GLO1 gene expression was significantly higher in subjects with CKD1-2 compared to CKD3-4. GLO1 protein level was lower in diabetics than in non-diabetics. GLO1 activity in RBCs differed between the four groups being significantly higher in diabetics with CKD1-2 vs. healthy subjects and vs. nondiabeticsfig with CKD3-4. GLO1 activity was significantly higher in diabetics compared to nondiabetics. In conclusion, both diabetes and CKD affects the glyoxalase system. It appears that CKD in advanced stages has prevailing and suppressive effects compared to hyperglycaemia. CKD decreases GLO1 gene expression and protein levels (together with diabetes) without concomitant changes of GLO1 activity.