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BACKGROUND:M2 macrophages have the function of reducing inflammatory factors and promoting tissue healing.Therefore,how to regulate M2 polarization of macrophages has been a hot research topic in recent years,and some miRNAs have been found to have this function. OBJECTIVE:To investigate the effects of miR-378a on the polarization of the Raw264.7 macrophage cell line. METHODS:The M1 polarization of macrophages was induced by lipopolysaccharide and interferon-γ.Interleukin-4 induced M2 polarization and the expression of endogenous miR-378a in each cell type was detected using qRT-PCR to verify whether miR-378a was involved in the polarization of macrophages.By transfection with lentivirus as the vector of overexpression of miR-378a,the stable expression of miR-378a cell lines was screened.Macrophage M1 polarization was induced synergically by lipopolysaccharide and interferon-γ.Macrophage M2 polarization was induced by interleukin-4.The levels of M1/M2 polarization-related cytokines in the supernatant of the macrophage culture medium were determined by enzyme-linked immunosorbent assay.qRT-PCR was used to detect the polarization characteristics of M1/M2-type macrophages and the mRNA expression levels of related cytokines. RESULTS AND CONCLUSION:(1)The expression level of endogenous miR-378a in Raw264.7 cells of each group increased after macrophage polarization.(2)Compared with the non-transfected group,the expressions of proinflammatory cytokine-induced nitric oxide synthase,tumor necrosis factor-α,interleukin-6 and interleukin-1β in macrophage M1 induced polarization were significantly decreased in the miR-378a transfection group(P<0.05);the levels of inducible nitric oxide synthase,tumor necrosis factor-α and interleukin-6 in cell supernatant were also significantly decreased(P<0.05).(3)Compared with the non-transfected group,the expressions of CD206,interleukin-10 and arginase-I in macrophage M2 induced polarization were significantly increased(P<0.05);the levels of CD206 and interleukin-10 in cell supernatant were also significantly increased(P<0.05)in the miR-378a transfection group.(4)It is indicated that overexpression of miR-378a promotes the M2 polarization of macrophages and inhibits the M1 polarization of macrophages.
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ObjectiveThe type 2C protein phosphatases （PP2C） are involved in numerous plant signal transduction pathways. They mainly participate in plant stress response and regulate second metabolites biosynthesis via negatively regulating MAPK signaling pathway. Herein，we were to identify and analyze PP2C （CsPP2C） gene family from hemp genome，in hope of providing comprehensive insights for studying CsPP2C function during the development of hemp. MethodMolecular Evolutionary Genetics Analysis （MAGA）-X was used to construct phylogenetic tree. Expert Protein Analysis System （ExPASy），WoLF PSORT，Multiple EM for Motif Elicitation （MEME），Batch Conserved Domain Search （Batch-CD-Search），PlantCare，and TBtools were used，respectively，to predict CsPP2C physicochemical properties，subcellular localization，conserved motifs，protein structure，cis-element in promoter and collinearity with Arabidopsis PP2C. Cannabis sativa transcriptome and Real-time polymerase chain reaction（Real-time PCR） were used to analyze and verify gene expressions，respectively. ResultFifty-two CsPP2C with conserved domains were identified from the entire genome of hemp，encoding proteins ranging from 244 to 1 089 aa in length and with molecular weights ranging from 26.76 to 122.53 kDa. Those genes were mainly distributed in the nucleus，cytoplasm and chloroplast. The 47 CsPP2C were divided into 10 subfamilies，and the remaining 5 were not clustered. Seven pairs of homologous genes between hemp and Arabidopsis thaliana were identified according to collinear analysis. The light-responsive elements and abscisic acid elements are most abundant in the prediction. The gene expression heat map showed varied expression pattern of CsPP2C in different tissues. Real-time PCR results of three CsPP2C were consistent with transcriptome data. Moreover，alternative splicing analysis showed that some CsPP2C had alternative-splicing genes during evolution. ConclusionWe predicted and analyzed CsPP2C gene family in genomic scale and showed that CsPP2C are involved in many biological processes，whereby provides foundation for CsPP2C functional study.
RESUMO
ObjectiveThe type 2C protein phosphatases （PP2C） are involved in numerous plant signal transduction pathways. They mainly participate in plant stress response and regulate second metabolites biosynthesis via negatively regulating MAPK signaling pathway. Herein，we were to identify and analyze PP2C （CsPP2C） gene family from hemp genome，in hope of providing comprehensive insights for studying CsPP2C function during the development of hemp. MethodMolecular Evolutionary Genetics Analysis （MAGA）-X was used to construct phylogenetic tree. Expert Protein Analysis System （ExPASy），WoLF PSORT，Multiple EM for Motif Elicitation （MEME），Batch Conserved Domain Search （Batch-CD-Search），PlantCare，and TBtools were used，respectively，to predict CsPP2C physicochemical properties，subcellular localization，conserved motifs，protein structure，cis-element in promoter and collinearity with Arabidopsis PP2C. Cannabis sativa transcriptome and Real-time polymerase chain reaction（Real-time PCR） were used to analyze and verify gene expressions，respectively. ResultFifty-two CsPP2C with conserved domains were identified from the entire genome of hemp，encoding proteins ranging from 244 to 1 089 aa in length and with molecular weights ranging from 26.76 to 122.53 kDa. Those genes were mainly distributed in the nucleus，cytoplasm and chloroplast. The 47 CsPP2C were divided into 10 subfamilies，and the remaining 5 were not clustered. Seven pairs of homologous genes between hemp and Arabidopsis thaliana were identified according to collinear analysis. The light-responsive elements and abscisic acid elements are most abundant in the prediction. The gene expression heat map showed varied expression pattern of CsPP2C in different tissues. Real-time PCR results of three CsPP2C were consistent with transcriptome data. Moreover，alternative splicing analysis showed that some CsPP2C had alternative-splicing genes during evolution. ConclusionWe predicted and analyzed CsPP2C gene family in genomic scale and showed that CsPP2C are involved in many biological processes，whereby provides foundation for CsPP2C functional study.