Receptor‐interacting protein kinase 3 mediates macrophage/monocyte activation in autoimmune hepatitis and regulates interleukin‐6 production

Abstract

Background: The mechanisms of macrophages/monocytes in autoimmune hepatitis (AIH) remain unclear. We investigated the role of receptor-interacting protein kinase 3 (RIP3), a key inflammatory signal adapter, in macrophage/monocyte activation in AIH. Methods: Liver tissues and monocytes from patients were collected to evaluate the relationship between macrophage activation and RIP3 by double-immunofluorescence and Western blotting. RAW264.7 macrophages were used to study the regulation of RIP3 signaling on inflammatory cytokines. Results: Compared to the hepatic cyst, the majority of accumulated macrophages expressed RIP3 in AIH liver tissues. Moreover, RIP3 expression of monocytes was correlated with the levels of serum hepatic enzyme in AIH. Furthermore, RIP3 signaling was activated by lipopolysaccharide in RAW264.7 macrophages, which was accompanied with upregulated interleukin (IL)-1b, IL-6, and IL-10 and downregulated IL-4 and transforming growth factor-b. Notably, necrostatin-1, the specific inhibitor of the RIP3 signaling pathway, and 6-thioguanine (6-TG), the active metabolite of azathioprine, predominantly reduced IL-6 production compared to other cytokines. Moreover, the gene level of IL-6 was dramatically increased in AIH liver tissues. Conclusions: RIP3 signaling is involved in macrophage/monocyte activation in AIH and mediates IL-6 production, and is a novel molecular mechanism of 6-TG, indicating that it might be a promising therapeutic target for AIH treatment. KeywordsAutoimmune hepatitis, macrophages, receptor-interacting protein kinase 3, interleukin-6Received: 4 October 2017; accepted: 2 January 2018 Key summaryOur study determined a novel molecular mechanism of macrophage/monocyte activation and inflammatory response in autoimmune hepatitis (AIH). Moreover, the role of receptor-interacting protein kinase 3 (RIP3) in human AIH disease was reported for the first time. In addition, this study points out the relationship between interleukin (IL)-6 and AIH, and IL-6 can be downregulated through inhibiting the RIP3 signaling of macrophages. Finally, our investigation provided a novel mechanism for the effect of thiopurines on the immune system.

Bifidobacterium animalis ssp. Lactis 420 Mitigates Autoimmune Hepatitis Through Regulating Intestinal Barrier and Liver Immune Cells

“…RIP3 has been increasingly recognized as a key inflammatory signal adapter, which mediates inflammation through necroptosis as well as non-necroptosis function ( 30 , 48 ). Our previous studies have found that RIP3 signaling was involved in macrophage/monocyte activation in the liver tissues of AIH patients and is correlated with the levels of serum hepatic enzyme ( 29 ). Then, we examined the activation of RIP3 and MLKL (the direct downstream effector of RIP3) in liver macrophages of the mice.…”

“…It is commonly accepted that macrophages are implicated in the pathological inflammation and fibrosis of liver diseases and activated macrophages are present in the portal area of AIH ( 26 – 28 ). Our previous studies have found that receptor-interacting protein kinase 3 (RIP3) signaling was involved in LPS-induced macrophage/monocyte activation in AIH ( 29 ). RIP3 kinase activity supports the recruitment of the mixed lineage kinase domain-like (MLKL) to trigger membrane leakage with the consequent production of pro-inflammatory cytokines ( 1 , 30 , 31 ).…”

RIP3 blockade prevents immune-mediated hepatitis through a myeloid-derived suppressor cell dependent mechanism

“…The upregulation of RIP3 expression has been described in AIH and other immune disorders and supposed to contribute to the progression of the pathology [45,46]. Zhang et al [31] showed that the accumulated macrophages express RIP3 in AIH liver tissues. And RIP3 deletion has been reported to protect against ConA-induced immune hepatitis [47].…”

“…Receptor interacting protein kinase 3 (RIP3) is a molecular switch between tumor necrosis factor (TNF)-induced apoptosis and necrosis [30]. We previously showed that RIP3 signaling is highly correlated with serum transaminase levels in patients with AIH, indicating that RIP3 signal plays important roles in the pathogenesis of the disease [31]. On the other hand, RIP3 also targets and delivers negative signals to MDSCs recruitment in tumor research [32,33].…”

RIPK3 promoter hypermethylation in hepatocytes protects from bile acid induced inflammation and necroptosis

“…1A). [47][48][49] To analyze the purity of the isolated primary hepatocytes and macrophages, a western blot against CD68 and albumin was performed from the same lysates.…”