Diabetic-like conditions modulate the expression of thiamine transporters in human retinal cells

Design of Study: Thiamine, a co-factor for transketolase and other glycolytic enzymes, counteracts high glucose-induced damage in microvascular cells in vitro and prevents progression of retinopathy and nephropathy in diabetic animals. Two single nucleotide polymorphisms (SNPs) located in the gene encoding for the thiamine transporter THTR2 are associated with resistance to development of proliferative diabetic retinopathy and end-stage renal disease in type 1 diabetic subjects. The protective effects of these SNPs may work either through loss of function, decreasing the discharge of thiamine, or gain of function, by increasing its uptake in target tissues. Purpose: The objective of this work was to investigate if, in human retinal cells, a diabetic-like microenvironment is able to modulate the expression of the two thiamine transporters THTR1 and THTR2 and their transcription factor Sp1. Methods: Human retinal pericytes (HRP), human microvascular endothelial cells (HMEC) and human Műller cells (MIO-M1) were cultivated for 8 days in physiological (NG), stable high glucose (HG) or intermittent physiological/high glucose conditions (intHG) (n=10). To investigate substrate influence on the expression of transporters, cells were also cultured in thiamine-deficient medium (noT) or in high thiamine conditions (50µmol/l, HT) (n=6). THTR1, THTR2 and Sp1 mRNA expression was checked by relative quantitative RT-PCR and protein expression by Western blotting. Results: We found that both transporters and Sp1 are expressed in HRP, HMEC and MIO-M1, THTR1 being more expressed than THTR2 in all cases. THTR2 and Sp1 expression decreased in HRP cultured in HG and intHG (THTR2: -20.8 and -36.1% respectively, p<0.05 vs NG; Sp1: -17.1 and -19.9%, p<0.05), while THTR1 expression was unchanged. Instead, THTR2 expression increased in MIO-M1 cells in intHG (+36.4%, p<0.05), and was unchanged in HMEC. Different thiamine concentrations did not influence THTR1, THTR2 or Sp1 expression. Conclusions: Diabetic-like conditions are able to modulate the expression of thiamine transporters in retinal cells. This may lead to decreased intracellular availability of thiamine, with consequent metabolic damage due to the accumulation of toxic metabolites.