High glucose-induced retinal pericyte apoptosis depends on association of GAPDH and Siah1 [Cell Biology]

October 5th, 2015 by Suarez, S., McCollum, G. W., Jayagopal, A., Penn, J. S.

Diabetic Retinopathy (DR) is a leading cause of blindness worldwide, and its prevalence is growing. Current therapies for DR address only the later stages of the disease, are invasive and are of limited effectiveness. Retinal pericyte death is an early pathologic
feature of DR. Though it has been observed in diabetic patients and in animal models of DR, the cause of pericyte death remains unknown. A novel pro-apoptotic pathway initiated by the interaction between glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the E3 ubiquitin ligase, seven in absentia homolog 1 (Siah1), was recently identified in ocular tissues. In this manuscript we examined the involvement of the GAPDH/Siah1 interaction in human retinal pericyte (hRP) apoptosis. HRP were cultured in 5mM normal glucose, 25mM L- or D-glucose for 48hrs (osmotic control and high glucose treatments, respectively). Siah1 siRNA was used to downregulate Siah1 expression. TAT-FLAG GAPDH and/or Siah1 directed peptides were used to block GAPDH and Siah1 interaction. Co-immunoprecipitation assays were conducted to analyze the effect of high glucose on the association of GAPDH and Siah1. Apoptosis was measured by Annexin V staining and caspase-3 enzymatic activity assay. High glucose increased Siah1 total protein levels, induced the association between GAPDH and Siah1, and led to GAPDH nuclear translocation. Our findings demonstrate that dissociation of the GAPDH/Siah1 pro-apoptotic complex can block high glucose-induced pericyte apoptosis, widely considered a hallmark feature of DR. Thus, the work presented in this manuscript can provide a foundation to identify novel targets for early treatment of DR.