Electronic Thesis and Dissertation Repository

Degree

Doctor of Philosophy

Program

Pathology

Supervisor

Chakrabarti, Subrata

2nd Supervisor

N.A

3rd Supervisor

N.A

Abstract

LncRNAs, recently found to be pervasively transcribed in the genome, can play key roles in epigenetic modifications during gene transcription. Their role in diabetic complications are however not clear.

Using array analysis, we identified alterations of two lncRNAs in high glucose (25mM/L,HG) treated retinal endothelial cells (HRECs). ANRIL expressions were further measured with or without siRNA knockdown in ECs. ANRIL knockout (KO) mice with or without streptozotocin (STZ)-induced diabetes were also investigated for mRNA and protein expression of VEGF and ECM proteins FN and Col1α4. ANRIL knockdown prevented glucose-induced increased VEGF, FN and Col1α4 levels. Comparable results were observed in the retina, heart and kidneys of diabetic ANRILKO mice compared to wildtype controls. We further showed that these alterations under ANRIL’s regulation are mediated by p300 and enhancer of zeste 2 (EZH2) of the PRC2 complex.

We also investigated the role of H19 in diabetes by silencing or overexpressing H19 in ECs exposed to various glucose levels. We extended our study to H19 knockout (KO) mice and vitreous samples from patients with proliferative DR. In both instances, diabetes caused downregulation of H19 expression. H19 overexpression prevented glucose-induced endothelial-mesenchymal transition (EndMT) through TGF-β in a Smad-independent pathway. Additional experiments showed a regulatory relationship between ANRIL and H19.

These data suggest that glucose and diabetes cause alteration of specific lncRNAs in ECs and mouse tissues respectively. Identification of such mechanisms help in a better understanding of the pathologies in diabetes and consequent development of RNA based therapies.

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