Netrin-1对肺泡上皮细胞钠离子通道α亚基的调控及其机制  

Regulatory effect of Netrin-1 on expression of epithelial sodium channel α-subunit in alveolar epithelia cells and potential mechanism

作　　者：何婧[1] 王导新[1] 

机构地区：[1]重庆医科大学附属第二医院呼吸内科,重庆400010

出　　处：《中国生物制品学杂志》2013年第6期817-821,共5页Chinese Journal of Biologicals

基　　金：国家自然科学基金(30971303)

摘　　要：目的研究Netrin-1对肺泡上皮细胞钠离子通道α亚基(epithelial sodium channelα-subunit,α-ENaC)表达的调控及其可能的机制,并探讨其在急性肺损伤(acute lung injury,ALI)及急性呼吸窘迫综合征(acute respiratory distresssyndrome,ARDS)中的作用。方法取对数生长期的A549细胞,用相同浓度的Netrin-1(500μg/L),分别作用0、3、6、12、24和48 h,用不同浓度的Netrin-1(0、0.5、5、50和500μg/L),作用12 h。将A549细胞分为药物组(500μg/LNetrin-1)、抑制组(1μmol/L PSB1115+500μg/L Netrin-1)和对照组(仅加RPMI1640培养基),培养12 h。RT-PCR法分别检测各组α-ENaC基因mRNA的转录水平;Western blot法检测各组α-ENaC蛋白的表达水平。结果相同浓度下,Netrin-1作用A549细胞6 h后,α-ENaC基因mRNA相对转录水平及蛋白相对表达量显著高于对照组(P<0.05),作用12 h时达最高;相同时间内,Netrin-1浓度大于5μg/L时,α-ENaC基因mRNA相对转录水平及蛋白的相对表达量均明显高于对照组(P<0.05),且呈剂量依赖性,浓度为500μg/L时达最高;药物组α-ENaC基因mRNA相对转录水平及蛋白相对表达量均明显高于对照组及抑制组(P<0.05),PSB1115可明显抑制α-ENaC基因mRNA的转录和蛋白表达。结论 Netrin-1可通过腺苷A2b受体途径,从基因水平上调α-ENaC的表达,有益于ALI及ARDS的预后。Objective To investigate the regulatory effect of Netrin-1 on expression of epithelial sodium channel α-subunit（α-ENaC）in alveolar epithelia A549 cells and the potential mechanism,as well as role of Netrin-1 in acute lung injury（ALI）and acute respiratory distress syndrome（ARDS）.Methods A549 cells at logarithmic growth phase were incubated with 500 μg/L Netrin-1 for 0,3,6,12,24 and 48 h respectively,or Netrin-1 at various concentrations（0,0.5,5,50 and 500 μg/L）for 12 h,then divided into control,drug and inhibition groups,and added with RPMI1640 medium,500 μg/L Netrin-1 and 1 μmol/L PSB1115＋500 μg/L Netrin-1 respectively,and further incubated for 12 h.The transcription levels of α-ENaC mRNAs in various groups were determined by RT-PCR,while the expression level of αENaC protein by Western blot.Results Both the expressions of α-ENaC at mRNA and protein levels in A549 cells after incubation with 500 μg/L Netrin-1 for 6 h were significantly higher than those in control group（P ＆lt; 0.05）and reached peak values 12 h after incubation.However,both the expression levels in A549 cells after incubation with Netrin-1 at concentrations of more than 5 μg/L for 12 h were significantly higher than those in control group（P ＆lt; 0.05）,which were dose-dependent and reached the peak values after incubation with 500 μg/L Netrin-1.Both the expression levels were significantly higher in drug group than in control and inhibition groups（P ＆lt; 0.01）,indicating that PSB1115 inhibited the mRNA transcription and protein expression of α-ENaC significantly.Conclusion Netrin-1 up-regulated the expression of α-ENaC at gene level via adenosine A2b receptor pathway,which was beneficial to the prognosis of ALI and ARDS.

关 键 词：神经轴突生长导向因子 腺苷A2b受体 上皮细胞钢离子通道α亚基 呼吸窘迫综合征 急性肺损伤 

分 类 号：Q256[生物学—细胞生物学]