中华劳动卫生职业病杂志
中華勞動衛生職業病雜誌
중화노동위생직업병잡지
CHINESE JOURNAL OF INDUSTRIAL HYGIENE AND OCCUPATIONAL DISEASES
2012年
12期
937-940
,共4页
李爱萍%侯志国%范晶晶%吉晓明%王婷%倪春辉
李愛萍%侯誌國%範晶晶%吉曉明%王婷%倪春輝
리애평%후지국%범정정%길효명%왕정%예춘휘
1-磷脂酰肌醇3-激酶%信号通路%二氯化硅%肌动蛋白类%信号通路
1-燐脂酰肌醇3-激酶%信號通路%二氯化硅%肌動蛋白類%信號通路
1-린지선기순3-격매%신호통로%이록화규%기동단백류%신호통로
1-Phosphatidylinositol 3-kinase%Signaling system%Silicon olioxide%Actins%PI3K/Akt pathway
目的 探讨磷脂酰肌醇3激酶(PI3K)/丝氨酸/苏氨酸蛋白激酶(Akt)信号通路在二氧化硅(SiO2)粉尘诱导的人胚胎支气管上皮细胞(HBE)表达转化生长因子(TGF)-β和α-平滑肌肌动蛋白(α-SMA)中的作用.方法 将体外培养的HBE细胞分成5组:(1)空白对照组；(2)SiO2处理组；(3)Akt磷酸化抑制剂组；(4)同时用Akt磷酸化抑制剂和SiO2处理组；(5)Akt磷酸化抑制剂处理细胞24 h再用SiO2处理组；SiO2暴露浓度为100 μg/ml,磷酸化抑制剂Ly294002浓度为10 μmol/L;用蛋白免疫印迹法(Western blot)检测Akt、磷酸化Akt(p-Akt)、TGF-β 、α-SMA表达水平.用免疫荧光技术检测上述处理过程中α-SMA蛋白在细胞中的定位和表达情况.结果 SiO2可以明显促使Akt磷酸化,48 h p-Akt表达增加了近l倍,72 h表达最高；TGF-β在12h明显增高,48 h达高峰；α-SMA表达在24 h开始增高,72h表达上调了1倍.Akt磷酸化抑制剂Ly294002可以有效抑制Akt的磷酸化,明显降低α-SMA的表达,尤其是先加抑制剂与细胞作用24 h后再加SiO2,p-Akt和α-SMA表达均明显下调,分别下调了1.5倍和7.6倍,对TGF-p表达则无明显影响.免疫荧光结果显示,SiO2可明显诱导HBE细胞内α-SMA的表达,而Akt磷酸化抑制剂Ly294002可以使HBE细胞内α-SMA蛋白的荧光强度明显减弱.结论 SiO2诱导HBE细胞表达α-SMA可能是通过PI3K/Akt信号通路实现的,Akt磷酸化抑制剂Ly294002可以有效抑制SiO2诱导的α-SMA表达.
目的 探討燐脂酰肌醇3激酶(PI3K)/絲氨痠/囌氨痠蛋白激酶(Akt)信號通路在二氧化硅(SiO2)粉塵誘導的人胚胎支氣管上皮細胞(HBE)錶達轉化生長因子(TGF)-β和α-平滑肌肌動蛋白(α-SMA)中的作用.方法 將體外培養的HBE細胞分成5組:(1)空白對照組；(2)SiO2處理組；(3)Akt燐痠化抑製劑組；(4)同時用Akt燐痠化抑製劑和SiO2處理組；(5)Akt燐痠化抑製劑處理細胞24 h再用SiO2處理組；SiO2暴露濃度為100 μg/ml,燐痠化抑製劑Ly294002濃度為10 μmol/L;用蛋白免疫印跡法(Western blot)檢測Akt、燐痠化Akt(p-Akt)、TGF-β 、α-SMA錶達水平.用免疫熒光技術檢測上述處理過程中α-SMA蛋白在細胞中的定位和錶達情況.結果 SiO2可以明顯促使Akt燐痠化,48 h p-Akt錶達增加瞭近l倍,72 h錶達最高；TGF-β在12h明顯增高,48 h達高峰；α-SMA錶達在24 h開始增高,72h錶達上調瞭1倍.Akt燐痠化抑製劑Ly294002可以有效抑製Akt的燐痠化,明顯降低α-SMA的錶達,尤其是先加抑製劑與細胞作用24 h後再加SiO2,p-Akt和α-SMA錶達均明顯下調,分彆下調瞭1.5倍和7.6倍,對TGF-p錶達則無明顯影響.免疫熒光結果顯示,SiO2可明顯誘導HBE細胞內α-SMA的錶達,而Akt燐痠化抑製劑Ly294002可以使HBE細胞內α-SMA蛋白的熒光彊度明顯減弱.結論 SiO2誘導HBE細胞錶達α-SMA可能是通過PI3K/Akt信號通路實現的,Akt燐痠化抑製劑Ly294002可以有效抑製SiO2誘導的α-SMA錶達.
목적 탐토린지선기순3격매(PI3K)/사안산/소안산단백격매(Akt)신호통로재이양화규(SiO2)분진유도적인배태지기관상피세포(HBE)표체전화생장인자(TGF)-β화α-평활기기동단백(α-SMA)중적작용.방법 장체외배양적HBE세포분성5조:(1)공백대조조；(2)SiO2처리조；(3)Akt린산화억제제조；(4)동시용Akt린산화억제제화SiO2처리조；(5)Akt린산화억제제처리세포24 h재용SiO2처리조；SiO2폭로농도위100 μg/ml,린산화억제제Ly294002농도위10 μmol/L;용단백면역인적법(Western blot)검측Akt、린산화Akt(p-Akt)、TGF-β 、α-SMA표체수평.용면역형광기술검측상술처리과정중α-SMA단백재세포중적정위화표체정황.결과 SiO2가이명현촉사Akt린산화,48 h p-Akt표체증가료근l배,72 h표체최고；TGF-β재12h명현증고,48 h체고봉；α-SMA표체재24 h개시증고,72h표체상조료1배.Akt린산화억제제Ly294002가이유효억제Akt적린산화,명현강저α-SMA적표체,우기시선가억제제여세포작용24 h후재가SiO2,p-Akt화α-SMA표체균명현하조,분별하조료1.5배화7.6배,대TGF-p표체칙무명현영향.면역형광결과현시,SiO2가명현유도HBE세포내α-SMA적표체,이Akt린산화억제제Ly294002가이사HBE세포내α-SMA단백적형광강도명현감약.결론 SiO2유도HBE세포표체α-SMA가능시통과PI3K/Akt신호통로실현적,Akt린산화억제제Ly294002가이유효억제SiO2유도적α-SMA표체.
Objective To explore the role of the phosphotidylinositol 3 kinase/protein kinase B (PI3K/Akt) pathway in silica-induced α-SMA (c smooth muscle actin) expression in HEB (human bronchial epithelial)cell.Methods The cultured HBE cells were divided into 5 groups:control,silica,PI3K inhibitor (Ly294002),both PI3K inhibitor (Ly294002) and silica at the same time and the inhibitor 24h ahead of silica.The final concentrations of PI3K inhibitor and silica were 10 μmol/L and 100 μg/ml,respectively.Western blots were used to detect protein expressions of Akt,p-Akt,TGF-β and α-SMA.The location and expression of α-SMA were measured by immunofluorescence assay.Results HBE cell line exposed to silica can induce Akt phosphorylation,in which expressions of p-Akt were up regulated 1 times at 48 and the highest at 72 h.The expressions of TGFβ increased remarkably at 12 h and the peak at 48 h after silica exposure,while the expressions of α-SMA increased at 24 h and the highest at 72 h.However,the PI3K inhibitor (Ly294002) significantly down regulated α-SMA expression.When the cell line exposed to the PI3K inhibitor ahead of silica 24 h,the expressions of p-Akt and α-SMA were more remarkably down regulated which were decreased 1.5 times and 7.6 times respectively compare to silica exposure group.But no significant changes werc found for TGFβexpressions.The immunofluorescence assay showed that silica can induce α-SMA expression,which located in cytoplasma,and PI3K inhibitor can decrease the expression.Conclusions Silica induced α-SMA expression in HBE cell line is by targeting the PI3K/Akt pathway and PI3K inhibitor can repress α-SMA expression.
