TMEM88 Modulates The Secretion Of Inflammatory Cytokine In Alcoholic Fatty Liver Disease

Objective: Alcoholic fatty liver disease(AFLD)is a progressive disease,which is caused by long-term heavy drinking.The formation mechanism of AFLD is comprehensive and multifactorial,involving the interaction and feedback between many components and signaling molecules,including a variety of cells,cytokines,complement components and transcription factors.At present,it is considered that the occurrence and development of AFLD is a complex process with multiple mechanisms and channels.Intracellular signals not only appear in hepatic parenchymal cells,but also Kupffer cells and hepatic stellate cells.In AFLD,Kupffer cells are activated and release a large number of cytokines,chemokines and oxidative active substances,which stimulate liver parenchymal cells and lead to inflammatory injury.Therefore,the research of AFLD mainly focuses on the secretion of inflammatory factors by Kupffer cells,which is of great significance.Transmembrane protein 88(TMEM88)is a new type of membrane protein that has attracted more and more attention in recent years.At present,the understanding of the expression,subcellular localization and potential molecular mechanism of TMEM88 is limited,and its potential application value is gradually being concerned.Recent studies have found that TMEM88 can bind to PDZ domain of dishevelled-1(DVL)in the Wnt signaling pathway,cut off the connection between DVL and frizzle and LRP5 / 6,and then block the dissociation of degradation complex,resulting in the loss of β-catenin in cytoplasm It is recognized by the phosphorylation site of GSK3 β and degraded by phosphorylation,thus inhibiting the classical Wnt/β-catenin signaling pathway.Recent studies have shown that TMEM88 inhibits the accumulation of collagen and the production of extracellular matrix in liver fibrosis.The relationship between TMEM88 and AFLD in liver diseases is unclear.In this study,we investigated the effects of TMEM88 on the secretion of inflammatory cytokines in Kupffer cells and RAW264.7 cells stimulated by ethanol.In addition,we also detected the mechanism of YAP signaling pathway in RAW264.7 cells stimulated by ethanol.Methods: In vivo,the mice model of AFLD was established by gaobin method,then the localization of TMEM88 in macrophages of AFLD was detected,and then the expression of TMEM88 in AFLD was detected.Adenovirus with silencing effect(ADV-TMEM88)and its control were injected into C57 BL / 6J mice via tail vein injection,and primary Kupffer cells were extracted by in situ perfusion technique.The protein and m RNA expressions of IL-6,TNF-α and IL-1β were detected by Western blotting and RT-q PCR.In vitro,mouse monocyte macrophages(RAW264.7)were stimulated by alcohol.p EGFP-C1-TMEM88 and TMEM88 siRNA were used to overexpress and silence TMEM88,respectively.The protein and mRNA expressions of IL-6,TNF-α and IL-1 β were detected by Western blotting and RT-qPCR.Flow cytometry was used to detect the effect of TMEM88 on the apoptosis of RAW264.7cells;Edu staining was used to detect the effect of TMEM88 on the proliferation of RAW264.7 cells;finally,the specific regulatory mechanism of TMEM88 on AFLD was observed.Results: The overexpression of TMEM88 leads to the up regulation of IL-6,TNF-α and IL-1β,suggesting that it may be related to the occurrence,development and end of inflammation.In addition,TMEM88 silencing can reduce the secretion of IL-6,TNF-α and IL-1β in RAW264.7 cells.In addition,flow cytometry showed that overexpression of TMEM88 promoted the apoptosis of RAW264.7 cells,while silencing TMEM88 inhibited the apoptosis of RAW264.7 cells.Edu staining showed that overexpression of TMEM88 inhibited the proliferation of RAW264.7 cells,while silencing TMEM88 inhibited the proliferation of RAW264.7 cells.Finally,we demonstrated that TMEM88 may regulate the secretion of inflammatory factors secreted by macrophages in AFLD through YAP/P-YAP signaling pathway.Conclusions: All in all,these experimental outcomes indicated that TMEM88 plays a proinflammatory effect on EtOH-stimulated RAW264.7 cells through the YAP signaling pathway.