Mishra Manish, Kowluru Renu A
Kresge Eye Institute, Wayne State University, Detroit, Michigan, United States.
Invest Ophthalmol Vis Sci. 2014 Sep 23;55(10):6960-7. doi: 10.1167/iovs.14-15020.
Mitochondrial DNA (mtDNA) is damaged in the retina in diabetes, and mitochondria copy numbers are decreased. The displacement-loop (D-loop) of the mtDNA, the region with transcription/replication elements, experiences more damage than other regions of mtDNA. Our aim was to examine the role of DNA mismatch repair (MMR) in mitochondria homeostasis in diabetic retinopathy, and in its continued progression after cessation of hyperglycemia.
Effect of hyperglycemia on sequence variants in the D-loop region was investigated in retinal endothelial cells and in the retina from streptozotocin-induced diabetic rats using mismatch-specific surveyor nuclease. The role of MMR machinery in mtDNA damage and mitochondrial respiration was investigated in retinal endothelial cells overexpressing Mlh1, an MMR enzyme mainly associated with mtDNA polymerase gamma, or Msh2 (associated with nuclear polymerase beta).
Hyperglycemia increased sequence variants in the D-loop region. While overexpression of Mlh1 in endothelial cells ameliorated glucose-induced increase in D-loop sequence variants, decrease in respiration rate and increase in apoptosis, overexpression of Msh2 did not protect the mitochondria damage. Termination of hyperglycemia failed to reverse decrease in MMR enzymes and increase in D-loop sequence variants.
Due to a compromised MMR system, the sequence variants in the D-loop region were not repaired, and that resulted in impaired mtDNA transcription. Mitochondria become dysfunctional, and they continued to be dysfunctional even after hyperglycemia was terminated, contributing to the development, and progression of diabetic retinopathy. Thus, strategies targeting mitochondrial MMR machinery could help maintain mitochondria homeostasis, and inhibit the development of diabetic retinopathy and its continued progression.
糖尿病患者视网膜中的线粒体DNA（mtDNA）会受到损伤，线粒体拷贝数减少。mtDNA的位移环（D环）区域含有转录/复制元件，比mtDNA的其他区域更容易受到损伤。我们的目的是研究DNA错配修复（MMR）在糖尿病视网膜病变线粒体稳态中的作用，以及高血糖停止后其持续进展中的作用。
使用错配特异性Surveyor核酸酶，研究高血糖对视网膜内皮细胞和链脲佐菌素诱导的糖尿病大鼠视网膜中D环区域序列变异的影响。在过表达主要与mtDNA聚合酶γ相关的MMR酶Mlh1或与核聚合酶β相关的Msh2的视网膜内皮细胞中，研究MMR机制在mtDNA损伤和线粒体呼吸中的作用。
高血糖增加了D环区域的序列变异。虽然在内皮细胞中过表达Mlh1可改善葡萄糖诱导的D环序列变异增加、呼吸速率降低和细胞凋亡增加，但过表达Msh2并不能保护线粒体免受损伤。高血糖终止未能逆转MMR酶的减少和D环序列变异的增加。
由于MMR系统受损，D环区域的序列变异未得到修复，导致mtDNA转录受损。线粒体功能失调，即使高血糖终止后仍持续功能失调，这促进了糖尿病视网膜病变的发生和发展。因此，针对线粒体MMR机制的策略可能有助于维持线粒体稳态，并抑制糖尿病视网膜病变的发生及其持续进展。
