Combined expression of A1 and A20 achieves optimal protection of renal proximal tubular epithelial cells

Background. Apoptotic death of renal proximal tubular epithelial cells (RPTECs) is a feature of acute and chronic renal failure. RPTECs are directly damaged by ischemia, inflammatory, and cytotoxic mediators but also contribute to their own demise by up-regulating proinflammatory nuclear factor-kappaB (NF-kappa B)-dependent proteins. In endothelial cells, the Bcl family member A1 and the zinc finger protein A20 have redundant and dual antiapoptotic and anti-inflammatory effects. We studied the function(s) of A1 and A20 in human RPTECs in vitro. Methods. Expression of A1 [reverse transcription-polymerase chain reaction (RT-PCR) and A20 (Northern and Western blot analysis)] in RPTECs was evaluated. A1 and A20 were overexpressed in RPTECs by recombinant adenoviral-mediated gene transfer. Their effect upon inhibitor of NF kappa B alpha (I kappa B alpha) degradation (Western blot), NF-kappa B nuclear translocation [electrophoretic mobility shift assay (EMSA)], up-regulation of intercellular adhesion molecule-1 (ICAM-1) [fluorescence-activated cell sorter (FACS)] and monocyte chemoattractant protein-1 (MCP-1) (Northern blot) and apoptosis [terminal deoxynucleotiddyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) nick-end labeling (TUNEL)] and FACS analysis of DNA content) was determined. Results. A1 and A20 were induced in RPTECs as part of the physiologic response to tumor necrosis factor (TNF). A20, but not A1, inhibited TNF-induced NF-kappa B activation by preventing I kappa B alpha degradation, hence subsequent up-regulation of the proinflammatory molecules ICAM-1 and MCP-1. Unexpectedly, A20 did not protect RPTECs from TNF and Fas-mediated apoptosis while A1 protected against both stimuli. Coexpression of A1 and A20 in RPTECs achieved additive anti-inflammatory and antiapoptotic cytoprotection. Conclusion. A1 and A20 exert differential cytoprotective effects in RPTECs. A1 is antiapoptotic. A20 is anti-inflammatory via blockade of NF-kappa B. We propose that A1 and A20 are both required for optimal protection of RPTECs from apoptosis (A1) and inflammation (A20) in conditions leading to renal damage.