Medical Biology - Theses
Permanent URI for this collection
Search Results
1 - 1 of 1
-
ItemSHARPIN and RIPK1 are key regulators of cell death and inflammation in vivo( 2015)Therapeutic tumour necrosis factor (TNF) inhibition has been remarkably successful in the treatment of inflammatory and autoimmune diseases such as rheumatoid arthritis, inflammatory bowel disease and psoriasis. Receptor interacting serine/threonine protein kinase 1 (RIPK1) and SHANK-associated RH domain interacting protein (SHARPIN) are proteins that regulate TNF and other immune signalling pathways. In mice, RIPK1 deficiency causes perinatal lethality whilst SHARPIN deficiency due to the inactivating chronic proliferative dermatitis gene mutation results in multi-organ disease including severe dermatitis. In both cases the underlying mechanism for the pathology has been unknown. TNF is best known for upregulating pro-survival and inflammatory transcriptional pathways although cell death in the form of caspase-8-dependent apoptosis or Receptor interacting serine/threonine protein kinase 3- (RIPK3) and Mixed lineage kinase domain-like- (MLKL) dependent programmed necrosis can also result. Whilst apoptosis is generally viewed as being immunologically inert and non-inflammatory, programmed necrosis, known as necroptosis, results in the release of cellular contents into the extracellular matrix and this can drive inflammation. In vitro, SHARPIN and RIPK1 regulate the transcriptional arm of TNF signalling and RIPK1 is also required for TNF-induced necroptosis. Both SHARPIN and RIPK1 also regulate transcriptional and cell death signalling from multiple other immune receptors. We therefore hypothesised that deregulated cell death was causative for the phenotypes caused by RIPK1 and SHARPIN deficiency. This work shows that SHARPIN deficient keratinocytes and dermal fibroblasts are sensitive to TNF-induced cell death. The SHARPIN deficient skin phenotype was completely prevented by deletion of the gene encoding TNF or its death receptor Tumour necrosis factor receptor 1 (TNFR1). SHARPIN deficient mice with caspase-8 heterozygosity and RIPK3 deficiency were almost fully protected from multi-organ inflammation and dermatitis, whilst unexpectedly, combined SHARPIN, RIPK3 and caspase-8 deficiency resulted in lethality. We also show that RIPK1 deficient mice on a C57BL/6 background die perinatally due to TNFR1-independent RIPK3- and MLKL-dependent systemic inflammatory disease, providing strong evidence that RIPK1 is both dispensable for, but required to inhibit, necroptosis. Mice doubly deficient for RIPK1 and RIPK3 or RIPK1 and MLKL were overtly normal at birth but after three to five days became runted and appeared to succumb to an intestinal phenotype marked by excessive apoptosis. Caspase-8 deficiency did not prevent the RIPK1 deficient perinatal lethality but prevented the intestinal phenotype. Finally, analogous to the deleterious effects of caspase-8 and RIPK3 in SHARPIN deficient mice, mice triply deficient for RIPK1, RIPK3 and caspase-8 were viable well into adulthood, and fertile, but eventually developed severe lymphoproliferative disease. These data show that in mice, RIPK1 or SHARPIN deficiency results in widespread inflammation and pathology due to unrestrained caspase-8-dependent apoptosis and RIPK3- and MLKL-dependent necroptosis, in a tissue specific manner. The results highlight that excessive apoptosis can be inflammatory, and that TNF can cause inflammation indirectly by causing excessive cell death and not only by upregulating inflammatory cytokines. RIPK1 activates necroptosis and has been successfully therapeutically targeted in pre-clinical trials. This research shows that RIPK1 also inhibits necroptosis in some tissue types, indicating a protective function for RIPK1 that must be preserved when therapeutically targeting it. These findings may have implications for the treatment of human disease states including stroke, heart attack and inflammatory bowel disease thought to involve excessive and pathological cell death.