大鼠肝再生终止阶段肝细胞CCAAT增强子结合蛋白α mRNA、微小RNA-144-3p和3种环状RNA的表达和作用

Expression and role of CCAAT enhancer binding protein α mRNA,microRNA-144-3p and three kinds of circular RNAs of hepatocytes during the rat liver regeneration termination

目的了解大鼠肝再生终止阶段CCAAT增强子结合蛋白α(CEBPα)mRNA、miR-144-3p、rnoLOC100365958_0001、rno-Usp3_0010和rno-AC241873_0010调节肝细胞进入G_(1)期的途径和方法。方法按Higgins等方法制备大鼠2/3肝切除(PH)模型,按Smedsrod等方法分离肝细胞,用大规模定量分析技术检测大鼠肝再生中肝细胞竞争性内源RNA(ceRNA)表达的变化,用Cytoscape 3.2软件构建ceRNA的相互作用网络,用ceRNA综合分析等方法解析它们表达的相关性和作用相关性。结果PH后6 h和72 h时,CEBPαmRNA的比值为1.12±0.27和2.52±0.15,miR-144-3p为2.97±0.81和0.99±0.36,rno-LOC100365958_0001为0.16±0.05和6.78±0.62,rnoUsp3_0010为0.13±0.01和7.61±0.69,rno-AC241873_0010为0.09±0.02和7.53±0.69。CEBPα促进的G_(1)期相关基因细胞周期蛋白G2(CCNG2)为0.25±0.06和1.10±0.22,抑制的G_(1)期相关基因ETS变异转录因子6(ETV6)为2.22±0.68和0.99±0.27,MAPK14为2.01±0.32和0.85±0.11,信号转导子和转录激活因子3(STAT3)为2.88±0.24和1.08±0.07,肿瘤坏死因子(TNF)为2.29±0.51和0.86±0.51。CEBPα促进的G_(0)期相关基因G_(0)/G_(2)期开关2(G_(0)S2)为0.64±0.08和4.71±0.91,磷脂酶A2-IVA(PLA2G4A)为0.42±0.13和2.83±0.62。结论PH后6 h时,rno-LOC100365958_0001、rno-Usp3_0010、rno-AC241873_0010和miR-144-3p的相互作用加强了后者对CEBPαmRNA的抑制,不利于CEBPα形成,有利于CEBPα抑制的G_(1)期相关基因表达和肝细胞处于G_(1)期。相反,PH后72 h时,CEBPαmRNA上调,有利于CEBPα促进G_(0)期相关基因表达和肝细胞处于G_(0)期。

Objective To explore the pathways and patterns which CCAAT enhancer binding proteinα(CEBPα)mRNA,miR-144-3 p,rno-LOC100365958_0001,rno-Usp3_0010 and rno-AC241873_0010 regulate the hepatocytes to enter G_(1) phase during rat liver regeneration(LR).Methods The rat 2/3 hepatectomy(partial hepatectomy,PH)model was prepared as described by Higgins,the hepatocytes of rat liver right lobes were isolated in 9∶00-11∶00 am according to the method of Smedsrod et al,the large-scale quantitative detection of ceRNAs was processed by the high-throughput biotechnology,the interaction network of competitive endogenous RNAs(ceRNAs)was constructed by Cytoscape 3.2 software,and the correlation in expression and role of ceRNAs was analyzed by ceRNA comprehensive analysis.Results It was found that at 6 hours and 72 hours after PH,the ratio value of CEBPα mRNA shows 1.12±0.27 and 2.52±0.15,miR-144-3 p displays 2.97±0.81 and 0.99±0.36,rno-LOC100365958_0001 indicates 0.16±0.05 and 6.78±0.62,rnoUsp3_0010 shows 0.13±0.01 and 7.61±0.69,rno-AC241873_0010 indicates 0.09±0.02 and 7.53±0.69.The G_(1) phaserelated gene cylin G2(CCNG2)promoted by CEBPα0.25±0.06 and 1.10±0.22,but the G_(1) phase-related genes inbibited by CEBPαwere following,ETS variant transcription factor 6(ETV6)2.22±0.68 and 0.99±0.27,MAPK142.01±0.32 and 0.85±0.11,the signal transducer and activator of transcription 3(STAT3)为2.88±0.24 and 1.08±0.07,tumor necrosis factor(TNF)2.29±0.51 and 0.86±0.51.The G_(0) phase-related genes promoted by CEBPαwere following,G_(0)/G_(1) witch 2(G_(0)S2)0.64±0.08 and 4.71±0.91,phospholipase A2 group IVA(PLA2G4A)0.42±0.13 and 2.83±0.62.Conclusion The interaction of miR-144-3 p and rno-LOC100365958_0001,rno-Usp3_0010 and rno-AC241873_0010 leads to CEBPαmRNA to bind with miR-144-3 p,to CEBPαbeing formed difficultly,and to the expression of the G_(1) phase-related genes inbibited by CEBPαprobablely,and to the hepatocytes being in G_(1) phase at 6 hours after PH.On the contrary,CEBPαmRNA is up-regulated at 72 hours after PH,that is helpful for the expression of the G_(0) phase-related genes promoted by CEBPαand for the hepatocytes to be in G_(0) phase.