Poster #022, Childrens Hospital
Disabling SIRT3 Promotes Metabolic Dysfunction in the Proximal Tubular Cells
Mentor: Dr. Hasmik Soloyan
Introduction: Alport Syndrome is a hereditary kidney disease caused predominantly by COL4A5 mutations in the glomerulus that lead to progressive renal failure. Even if Alport Syndrome is considered a disease of the glomerulus, our studies indicate that damage to the tubules is also important in this disease’s progression. Indeed, our preliminary data shows downregulation of SIRT3 and PGC-1a expression in the proximal tubular cells. SIRT3 is a major protein deacetylase found primarily in the mitochondria that regulates ATP production. This alteration is associated with metabolic abnormalities and lipid accumulation. These findings suggest that disruption of the SIRT3-PGC-1a signaling may lead to mitochondrial dysfunction and contribute to proximal tubule injury.
Methods: To test our hypothesis, we transiently inhibited SIRT3 in cultured cells using siRNA. After silencing, we performed immunocytochemistry and immunoblotting on our experimental groups to evaluate the expression of SIRT3 and PGC-1a in the silenced cells compared to normal cells. In addition, we are currently assessing markers for mitochondrial function: Tomm20, ATP5A, and ACSL1.
Results: We efficiently silenced SIRT3. By light microscopy, we determined that silencing impaired tubular cell morphology and induced tubular injury. In addition, we collected cell protein, evaluated protein concentration, and performed immunoblotting. At the moment of submitting this abstract, the blots are developing. We are also performing immunocytochemistry for PGC-1a, megalin, and cubilin.
Conclusion: These studies will define a model mechanism linking SIRT3 and PGC-1a to proximal tubular injury in Alport. In addition, we are also evaluating the role of SIRT3 in tubular metabolic function since its expression regulates mitochondrial activity in these cells.