Mcart1基因敲除对小鼠巨噬细胞代谢及表型的影响OA

Objective To investigate the effects of Mcart1 gene knockout on macrophage metabolism and phenotype.Methods Two types of bone marrow-derived macrophages(BMDMs)were obtained from Mcart1flox/flox and Mcart1Lyz2-cre mice.Basal respiration of both cell types was measured using a Seahorse analyzer.The activity of α-ketoglutarate dehydrogenase(OGDH)was assessed in Mcart1 stably knockout RAW264.7 cells(Mcart1-/--RAW264.7)and wild-type RAW264.7 cells(RAW264.7).The two types of BMDMs were induced into M1 mac-rophages using LPS,and whole-cell reactive oxygen species(ROS)levels were measured by flow cytometry,while dynamic expression of inflammatory mediators was monitored by qPCR.The two types of BMDMs were also induced into M2 macrophages using IL-4,and expression of M2 anti-inflammatory and tissue repair mediators was measured by qPCR,while protein levels of classical M2 activation markers were detected using Western blot.Results Com-pared with Mcart1flox/flox BMDMs,Mcart1 knockout significantly reduced basal respiration in BMDMs,and Mcart1 stable knockout decreased OGDH activity in macrophages(P＜0.01).Mcart1 knockout significantly enhanced ROS production in M1 macrophages and upregulated early inflammatory mediators Il1b,Tnf,and Nos2(P＜0.05),while having no significant effect on the expression of M2 anti-inflammatory and tissue repair mediators Arg1,Mrc1,Ret-nla,Cd163,Fn1,or classical activation proteins p-Stat6 and p-Stat3 in M2 macrophages.Conclusions Mcart1 knockout significantly suppresses basal respiration in macrophages,inhibits OGDH enzyme activity,increases oxi-dative stress and inflammatory mediator levels in M1 macrophages,and does not significantly affect functional medi-ator levels in M2 macrophages.