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Manish Mishra, Renu Kowluru; Mismatch Mitochondrial DNA Sequence in Diabetic Retinopathy. Invest. Ophthalmol. Vis. Sci. 2014;55(13):5832.
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In diabetic retinopathy, reactive oxygen species (ROS) are elevated in the retina, mitochondrial DNA (mtDNA) is oxidatively modified, and the base excision repair (BER) machinery is compromised, raising a possibility of mismatch base pairs in the DNA. To repair the base mismatches incorporated during faulty BER, the cell is also equipped with mismatch repair (MMR) system consisting of the key regulatory proteins Mlh1 and Msh2. While Msh2 is largely associated with nuclear DNA polymerase B, Mlh1 is mainly associated with mitochondrial polymerase gamma (POLG), and we have shown that its expression in the retina is downregulated in diabetes. The aim of present study is to investigate the role of MMR system, and subsequently mtDNA sequence mismatch, in diabetic retinopathy.
Gene expressions of Mlh1 and Msh2 were quantified in the retina from streptozotocin-induced diabetic rats by q-PCR, and their protein expressions by western blot analysis. Sequence mismatch of mtDNA was examined using Surveyor nuclease assay kit for mismatch specific nuclease digestion assay. To confirm the role of Mlh1 and Msh2 in diabetic retinopathy, effect of high glucose on mtDNA mismatch and cell apoptosis was investigated in retinal endothelial cells overexpressing Mlh1 or Msh2.
Diabetes decreased the expression (gene and protein) of Mlh1 and Msh2 in the retina, and increased mtDNA sequence mismatch and fragmentation. Overexpression of Mlh1 in retinal endothelial cells significantly ameliorated glucose-induced increase in mtDNA sequence mismatch and fragmentation, and also prevented increase in cell apoptosis. However, overexpression of Msh2 had partial beneficial effect on mtDNA fragmentation and cell apoptosis.
Better protection of mtDNA fragmentation and cell apoptosis by Mlh1 than by Msh2 suggests that impaired repair of the mismatch base pairs in the mtDNA plays significant role in the development of diabetic retinopathy. Thus, regulation of mitochondrial mismatch repair system could ameliorate mtDNA damage, and inhibit the development of this blinding disease.
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