Analyzing necroptosis using an RIPK1 kinase inactive mouse model of TNF shock

Abstract

The serine/threonine kinase RIPK1 has numerous biological and pathological functions, mediating prosurvival as well as prodeath apoptotic and necroptotic signaling pathways downstream of various receptors, including death receptors and Toll-like receptors (TLRs). RIPK1 has been implicated in various diseases, including ischemia–reperfusion injury and inflammatory bowel disease (IBD). The recent generation of RIPK1 kinase inactive mice has enabled us to genetically interrogate the role of RIPK1 kinase-mediated necroptosis in disease models. Here, we describe procedures utilizing kinase inactive Ripk1D138N/D138N mice to analyze necroptosis induction in vitro in bone-marrow derived macrophages (BMDMs) and in vivo in a murine model of TNF-induced shock.