Thiamine transporter 2 is involved in high glucose-induced damage and altered thiamine availability in cell models of diabetic retinopathy

Abstract

Thiamine prevents high glucose-induced damage in microvasculature, and progression of retinopathy and nephropathy in diabetic animals. Impaired thiamine availability causes renal damage in diabetic patients. Two single-nucleotide polymorphisms in SLC19A3 locus encoding for thiamine transporter 2 are associated with absent/minimal diabetic retinopathy and nephropathy despite long-term type 1 diabetes. We investigated the involvement of thiamine transporter 1 and thiamine transporter 2, and their transcription factor specificity protein 1, in high glucose-induced damage and altered thiamine availability in cells of the inner blood–retinal barrier. Human endothelial cells, pericytes and Müller cells were exposed to hyperglycaemic-like conditions and/or thiamine deficiency/over-supplementation in single/co-cultures. Expression and localization of thiamine transporter 1, thiamine transporter 2 and transcription factor specificity protein 1 were evaluated together with intracellular thiamine concentration, transketolase activity and permeability to thiamine. The effects of thiamine depletion on cell function (viability, apoptosis and migration) were also addressed. Thiamine transporter 2 and transcription factor specificity protein 1 expression were modulated by hyperglycaemic-like conditions. Transketolase activity, intracellular thiamine and permeability to thiamine were decreased in cells cultured in thiamine deficiency, and in pericytes in hyperglycaemic-like conditions. Thiamine depletion reduced cell viability and proliferation, while thiamine over-supplementation compensated for thiamine transporter 2 reduction by restoring thiamine uptake and transketolase activity. High glucose and reduced thiamine determine impairment in thiamine transport inside retinal cells and through the inner blood–retinal barrier. Thiamine transporter 2 modulation in our cell models suggests its major role in thiamine transport in retinal cells and its involvement in high glucose-induced damage and impaired thiamine availability.

Reduced Thiamine Availability and Hyperglycemia Impair Thiamine Transport in Renal Glomerular Cells through Modulation of Thiamine Transporter 2

“…Starting from these assumptions, our aims were to investigate the modulation of thiamine transporter expression in renal cells involved in DN and exposed to conditions mimicking the diabetic microenvironment, such as glucose fluctuations and thiamine deficiency, and if these variations could influence thiamine uptake and transketolase activity, ultimately worsening DN. Our results confirmed that THTR2 is the thiamine transporter more involved in DN, as we already demonstrated in DR [3], and that lack of thiamine concurs with hyperglycemia in impairing thiamine transport.…”

“…Relative gene expression was determined using the 2 −∆∆CT method and normalized against β-actin. Primers used were: THTR1 forward: 5 -AGCCAGACCGTCTCCTTGTA-3 , reverse: 5 -TAGAGAGGGCCCACCACAC-3 ; THTR2 forward: 5 -CTGGCTCTGGTGGTCTTCTC-3 , reverse: 5 -AGGCATAGCGTTCCACATTC-3 ; Sp1 forward: 5 -TGCAGCAGAATTGAGTCACC-3 , reverse: 5 -CACAACATACTGCCC-ACCAG-3 [3]; TK forward: 5 -CCCAGCTACAAAGTTGGGGACAAG-3 ; reverse, 5 -GGTCATCCTTGCTCTTCAGGACC-3 .…”

“…The role of thiamine in the prevention of diabetic complications is well acknowledged in the literature [3]. Thiamine is an essential cofactor in intracellular glucose metabolism, especially for transketolase (TK), an enzyme that shifts toxic metabolites from glycolysis to the pentose phosphate shunt [4,5].…”

“…High glucose conditions and reduced thiamine availability synergically impair thiamine transport inside retinal cells and through the inner blood-retinal barrier. In particular, THTR2 expression is decreased in retinal pericytes, and increased in endothelial and Müller cells, suggesting a major role for THTR2 in thiamine transport in retinal cells, and its involvement in high-glucoseinduced damage and impaired thiamine availability [3]. It was also demonstrated that two single-gene polymorphisms (SNP) located in the SLC19A3 gene encoding for THTR2 are associated with resistance to the development of DR and DN, in subjects with long-term type 1 diabetes [16].…”

Analysis of the transcriptome and DNA methylome in response to acute and recurrent low glucose in human primary astrocytes

“…One of the most significant DMPs (cg07417745) is located in intron 1 of the SLC19A3 gene, which encodes a thiamine transporter [20], and showed a linear relationship between increased DNA methylation and the number of LG exposures. Interestingly, expression of this gene has previously been found to be modulated by hyperglycaemic-like conditions [21]. Conversely, methylation of cg11022541 and cg11692715 located within the genes NIPA1 and SLC8B1 respectively, decreased following RLG.…”