Dr. Andrey Cybulsky
Complement C5b-9 signaling and mechanisms of podocyte injury
The glomerular visceral epithelial cell or podocyte plays an important role in maintaining glomerular permselectivity. The complement C5b-9 membrane attack complex is the key mediator of injury in a number of glomerulopathies, including membranous nephropathy, which is associated with podocyte injury. In experimental membranous nephropathy, C5b-9 injures the glomerular epithelial cell, but simultaneously activates pathways that are potentially cytoprotective. We have employed cultured cells and in vivo models, including knockout mice, to characterize a number of C5b-9-dependent mediators of podocyte injury, including protein kinases, phospholipases, and bioactive lipid mediators. Our studies have employed a structure-function approach to address the activation and regulation of these mediators. More recent work has focused on alleviation of protein misfolding, including induction of endoplasmic reticulum stress and activation of the ubiquitin-proteasome system as novel mechanisms that podocytes employ to recover from injury. Modulation of stress pathways offers therapeutic possibilities for complement-mediated glomerular diseases.
Extracellular matrix and cytoskeleton as a determinant of signaling responses in podocytes
Glomerular epithelial cell injury occurs in focal segmental glomerulosclerosis (FSGS), and may include apoptosis, detachment and proliferation. Injury may be acquired, or secondary to mutations in key podocyte structural proteins, e.g. a-actinin-4. Our studies have shown that following podocyte injury in vivo, signals from extracellular matrix, including focal adhesion kinase, Src and extracellular signal-regulated kinase, as well as growth factor receptors, are activated to maintain cell survival (or prevent apoptosis), limit injury, and promote healing. We have also characterized the functional effects of a FSGS-associated mutation in the cytoskeletal protein, a-actinin-4, and we have shown that this mutant protein may misfold, activate cellular stress pathways, and be toxic to the podocytes. Modulation of protein misfolding and stress pathway activation may be a potential therapeutic approach for certain forms of FSGS.
Protein kinases in the kidney
The Ste20-like protein kinase, SLK, is essential for embryonic development, and may play a key role in cytoskeletal structure and cell membrane assembly, cell cycle progression, wound healing, tumor growth and metastasis. Our studies have demonstrated increased expression and activation of SLK in the developing kidney and recovery from ischemic acute renal failure, which recapitulates developmental mechanisms. When SLK expression increases, its actions are anti-proliferative or pro-apoptotic, both in renal cell culture models and in podocytes in transgenic mice. SLK is expressed primarily in renal tubular and glomerular epithelial cells, in both developing and adult kidney. The regulation of SLK activity is complex and involve changes in mRNA stability, dimerization, phosphorylation, as well as protein-protein interactions. Signaling by SLK occurs via activation of stress protein kinase pathways. Our studies have employed cell culture and in vivo models (transgenic mice, knockout mice, zebrafish), and have been directed at characterizing the regulation, signaling, and functions of SLK in the kidney. The overall aim of these studies is to obtain a better understanding of the role of protein kinases, including SLK in the mediation of injury, and determining if modulating SLK activity can repair renal cell injury.