RNAseqencing of wild type (WT) and Ripk3-/- (KO) AML-12 cells in the presence (PA) of absence (C) of 125 µM palmitate-bovine serum albumin for 24 h

Purpose: Necroptosis as been implicated in various deseases. The goal of this study is to invastigate the impact of RIPK3 in the lipid metabolism of hepatic cells. Methods: AML-12 cells invalidated or not for RIPK3 were exposed to free fatty acid and the mRNA profiles of wild type (WT) or knockout (RIPK3-KO) AML-12 cells control (C) or exposed to palmitic acid (PA) were generated by deep sequencing, in 5 copies, using Illumina NOVAseq 6000 plateform. Differential expression analysis between two conditions/groups (five biological replicates per condition) was performed using DESeq2 R package. Genes with an adjusted P value < 0.05 found by DESeq2 were assigned as differentially expressed. qRT–PCR validation was performed using SYBR Green assays Results: The DEGs were clustered using a hierarchical clustering algorithm, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis unveiled that transcripts associated with OXPHOS and thermogenesis were markedly upregulated in unchallenged Ripk3-/- hepatocytes compared with WT cells an effect that was aggravated by the PA treatment. Similarly, gene ontology enrichment analysis showed that mitochondria respiration-related proteins were overrepresented among the cellular components that were upregulated in Ripk3-/- hepatocytes compared with WT cells Conclusions: The absence of Ripk3 impacts on hepatocyte transcriptomic profile, increasing the expression of genes implicated in mitochondria bioenergetics and function, rescuing mitochondria respiration after free fatty acid overload. Overall design: RNAseqencing of wild type (WT) and Ripk3-/- (KO) AML-12 cells in the presence (PA) of absence (C) of 125 µM palmitate-bovine serum albumin for 24 h