title: High Glucose-induced Retinal Pericyte Apoptosis Depends on Association of GAPDH and Siah1.
abstract: 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 have limited effectiveness. Retinal pericyte death is an early pathologic feature of DR. Although 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 article we examined the involvement of the GAPDH/Siah1 interaction in human retinal pericyte (hRP) apoptosis. HRP were cultured in 5 mm normal glucose, 25 mm l- or d-glucose for 48 h (osmotic control and high glucose treatments, respectively). Siah1 siRNA was used to down-regulate 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 article can provide a foundation to identify novel targets for early treatment of DR.
authors.affiliation: Departments of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-8808. Electronic address: Sandra.suarez@vanderbilt.edu.
authors.forename: Sandra
authors.initials: S
authors.lastname: Suarez
authors.affiliation: Departments of Ophthalmology and Visual Sciences, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-8808.
authors.forename: Gary W
authors.initials: GW
authors.lastname: McCollum
authors.affiliation: Pharma Research and Early Development (pRED), F. Hoffman-La Roche Ltd., Grenzacherstrasse 124, CH-4070 Basel, Switzerland.
authors.forename: Ashwath
authors.initials: A
authors.lastname: Jayagopal
authors.affiliation: Departments of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-8808; Departments of Ophthalmology and Visual Sciences, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-8808.
authors.forename: John S
authors.initials: JS
authors.lastname: Penn
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D002467:Cell Nucleus / Q000201:enzymology; D002478:Cells, Cultured; D055785:Gene Knockdown Techniques; D005947:Glucose / Q000008:administration & dosage* / Q000494:pharmacology; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism*; D006801:Humans; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D020286:Pericytes / Q000187:drug effects*; D021381:Protein Transport; D012160:Retina / Q000166:cytology / Q000187:drug effects*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*; D000098563:Seven in Absentia Proteins
chemical_list: D009687:Nuclear Proteins; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D044767:Ubiquitin-Protein Ligases; D000098563:Seven in Absentia Proteins; D005947:Glucose
keywords: GAPDH; Siah1; apoptosis; cell signaling; diabetes; diabetic retinopathy; glucose; retina