Knock in p38g and knock out p38d effect on gene expression in TLR4 signalling in bone marrow derived macrophages using LPS.

Evidence implicating p38? and p38d (p38?/p38d) in inflammation are mainly based on experiments using p38?/p38d deficient (p38?/d-/-) mice, which show low levels of TPL2, the kinase upstream of MKK1-ERK1/2 in myeloid cells. This could obscure p38?/p38d roles, since TPL2 is essential for regulating inflammation. Here we generated a p38?D171A/D171A/p38d-/- (p38?/dKIKO) mouse, expressing kinase-inactive p38? and lacking p38d. This mouse exhibited normal TPL2 levels, making it an excellent tool to elucidate specific p38?/p38d functions. p38?/dKIKO mice showed a reduced inflammatory response and less susceptibility to LPS-induced septic shock and Candida albicans infection than wild-type mice. Gene expression analyses in LPS-activated WT and p38?/dKIKO macrophages revealed that p38?/p38d regulated numerous genes implicated in innate immune response. Overall design: WT and p38?/dKIKO bone marrow derived macrophages were stimulated with 100 ng/ml LPS for 0, 30 and 60 min, and gene expression was analysed by RNA-sequencing (RNA-Seq) to identify target genes specifically affected by the lack of p38? kinase activity and p38d expression.