Effect Of Coronin-1 On Macrophage Polarization By Regulating The Expression Of CD14

Research background:Macrophage Coronin-1 is related to its mediated inflammatory response,and the phenotypic polarization state of macrophages determines its pro-inflammatory or anti-inflammatory functions.Studying the correlation between Coronin-1 and macrophage polarization and its molecular mechanism can provide new ideas for tumor prevention and treatment and inflammation control.Objective:To investigate the correlation between Coronin-1 and macrophage polarization and its possible mechanism.Methods:1.The RAW264.7 macrophage models of M1 and M2 were established.The polarization indexes of i NOS and Arg-1 were detected by Real-time PCR and Western blot,then the expression level of Coronin-1 in macrophages with different polarization phenotypes was detected.2.Over-expressed(Coronin-1 Plus)and knockdown(Coronin-1 KO)cell lines were constructed,and the expression of Coronin-1 was detected by Real-time PCR and Western blot to verify whether the cell lines were successfully constructed.The expression level of i NOS in over-expressed,knockdown and normal Coronin-1 cells was detected.3.The interaction and enrichment of Coronin-1 and CD14-related proteins were analyzed by PPI、GO、KEGG,and the expression levels of CD14 in the over-expressed,knockdown and normal-expressed Coronin-1 cells were detected by real-time PCR and Western blot.4.Over-expressed,knockdown and normal-expressed Coronin-1 cells were treated with 100ng/m L LPS for 48 hours,and the expression level of i NOS was detected by Real-time PCR and Western blot.Then,the CD14 knockdown cell lines(Control-CD14Minus)and CD14 knockdown Coronin-1 Plus cell lines(Coronin-1plus-CD14Minus)were constructed.After LPS treatment(100ng/m L),the expression of i NOS was detected by real-time PCR and Western blot.Results:1.M1 and M2 macrophage models were successfully constructed,and the expression of Coronin-1 in M1 type was significantly higher than that in the Control group(P<0.05)。2.Over-expression(Coronin-1 Plus)and knockdown expression(Coronin-1 KO)cell lines were successfully constructed.The over-expression rate of Coronin-1 Plus was 50.2%,and the knockout rate of Coronin-1 KO was 93%.The m RNA expression of i NOS in Coronin-1 Plus group was significantly lower than that in Control group(P<0.001).3.The expression of CD14 in Coronin-1 Plus group was significantly higher than that in Control group(P<0.05).4.After LPS treatment,the expression of i NOS in Coronin-1 Plus group was significantly higher than that in Control group(P<0.01).CD14 Minus and Coronin-1plus-CD14 Minus cell lines were successfully constructed.CD14 Minus silencing efficiency was 54.3% and Coronin-1 plus-CD14 Minus silencing efficiency was 62.1%.The i NOS level of CD14 Minus and Coronin-1 plus-CD14 Minus cells treated with LPS was not significantly different from that of the Control group(P>0.05).Conclusion:1.The M1 polarization of macrophages leads to the high expression of Coronin-1.2.Coronin-1 over-expression does not directly lead to M1 polarization of macrophages.3.Coronin-1 over-expression can lead to up-regulation of CD14 expression,and Coronin-1 can promote the LPS-induced M1 polarization of macrophages through the CD14 pathway.