Master of Science
Microbiology and Immunology
Dr. Anthony M. Jevnikar
Ischemia-reperfusion injury (IRI) contributes to reduced long term organ transplant survival. IRI is a complex process related to multiple mechanisms including regulated forms of cell death such as apoptosis and necroptosis. We have previously shown that preventing necroptosis by genetic elimination of the key molecule (RIPK3) or apoptosis inhibition through caspase-8 silencing are of benefit in kidney IRI. As these pathways of regulated cell death are unique, we tested the hypothesis that deletion of both RIPK3 and caspase-8 activity in kidneys would synergistically improve renal IRI, compared to inhibition of individual pathways. Interestingly we found that IRI was not improved in vivo using RIPK3/caspase-8 null (DKO) mice. In vitro testing of renal tubular epithelial cells (TEC) from DKO kidneys showed enhanced death following exposure to IFN-γ + IL-1β suggesting enhanced activation of an intrinsic apoptosis pathway. Indeed enhanced caspase-9 and -3 activation was observed, as well as death of DKO TEC was inhibited by the BAX inhibiting peptide V5 (BIP). Finally, murine cytomegalovirus (MCMV) infection, which can block intrinsic apoptosis, was able to abrogate death in DKO TEC. In the absence of cell death, cytokine exposed DKO TEC showed increased expression of pro-inflammatory chemokines. Collectively these results indicate that cell death pathways are complex and counterbalanced. In the absence of RIPK3 and caspase-8, intrinsic apoptosis and chemokine expression is up-regulated in TEC which may account for the absence of the expected benefit from the loss of both apoptosis and necroptosis. These results highlight the complex biology that results from targeted perturbations of death pathways in TEC and suggest that the inhibition of multiple forms of cell death will likely be required to maximize clinical benefit in IRI and transplants.
Sung, Ron, "Contribution of enhanced intrinsic apoptosis to renal ischemia reperfusion injury in the absence of RIPK3 and Caspase-8 dependent regulated cell death" (2015). Electronic Thesis and Dissertation Repository. 3080.