中华结核和呼吸杂志
中華結覈和呼吸雜誌
중화결핵화호흡잡지
Chinese Journal of Tuberculosis and Respiratory Diseases
2011年
6期
442-446
,共5页
杜艳%胡晓芸%韩葆芬%杜永成
杜豔%鬍曉蕓%韓葆芬%杜永成
두염%호효예%한보분%두영성
尼古丁%蛋白激酶C%细胞外信号调节MAP激酶类%人脐静脉内皮细胞
尼古丁%蛋白激酶C%細胞外信號調節MAP激酶類%人臍靜脈內皮細胞
니고정%단백격매C%세포외신호조절MAP격매류%인제정맥내피세포
Nicotine%Protein kinase C%Extracellular signal-regulated kinase%Human umbilical vascular endothelial cells (HUVECs)
目的 探讨蛋白激酶C(PKC)-胞外信号调节激酶(ERK)1/2信号通路在尼古丁诱导人脐静脉内皮细胞(HUVECs)表达纤维溶解酶原激活物抑制物-1(PAI-1)中的作用.方法 体外培养HUVECs,采用不同实验条件尼古丁进行干预,ELISA法测定细胞上清液中PAI-1的浓度,观察尼古丁作用的最佳浓度和时间.进一步分别用PKC的抑制剂星型胞菌素staurosporine (STS)和ERK的抑制剂PD98059干预HUVECs,观察PKC或ERK被阻断后对尼古丁诱导的HUVECs 表达PAI-1的影响,ELISA测定各组细胞上清液中PAI-1蛋白的表达水平,RT-PCR检测各组细胞PAI-1 mRNA的表达.结果 100 μmol/L尼古丁组PAI-1蛋白水平[(22.6 ± 1.1)μg/L]明显高于对照组[(14.2± 2.8)μg/L,q=5.64,P＜0.05];以100 μmol/L 尼古丁分别与HUVECs孵育0、4、6、8、12及24 h,各组PAI-1蛋白表达呈时间依赖性升高,并在12 h达到高峰(F=32.063,P＜0.05);尼古丁组PAI-1 mRNA及蛋白含量[(1.32±0.20),(21.08±0.83)μg/L]明显高于对照组[(0.73±0.10),(13.39± 0.93)μg/L,q=8.43、11.97,均P＜0.05];尼古丁+STS组PAI-1 mRNA及蛋白含量[(1.07±0.10),(16.19±2.15)μg/L]较尼古丁组降低(q=5.61、7.61,均P＜0.05),但仍高于对照组(q=7.84、4.36,均P＜0.05);尼古丁+ PD98059组PAI-1 mRNA及蛋白表达[(1.12±0.11),(17.52±1.72)μg/L]低于尼古丁组(q=4.68、5.54,均P＜0.05),仍高于对照组(q=8.77、6.43,均P＜0.05).结论 PKC-ERK1/2信号通路在尼古丁诱导的血管内皮细胞PAI-1表达上调中发挥一定作用.
目的 探討蛋白激酶C(PKC)-胞外信號調節激酶(ERK)1/2信號通路在尼古丁誘導人臍靜脈內皮細胞(HUVECs)錶達纖維溶解酶原激活物抑製物-1(PAI-1)中的作用.方法 體外培養HUVECs,採用不同實驗條件尼古丁進行榦預,ELISA法測定細胞上清液中PAI-1的濃度,觀察尼古丁作用的最佳濃度和時間.進一步分彆用PKC的抑製劑星型胞菌素staurosporine (STS)和ERK的抑製劑PD98059榦預HUVECs,觀察PKC或ERK被阻斷後對尼古丁誘導的HUVECs 錶達PAI-1的影響,ELISA測定各組細胞上清液中PAI-1蛋白的錶達水平,RT-PCR檢測各組細胞PAI-1 mRNA的錶達.結果 100 μmol/L尼古丁組PAI-1蛋白水平[(22.6 ± 1.1)μg/L]明顯高于對照組[(14.2± 2.8)μg/L,q=5.64,P＜0.05];以100 μmol/L 尼古丁分彆與HUVECs孵育0、4、6、8、12及24 h,各組PAI-1蛋白錶達呈時間依賴性升高,併在12 h達到高峰(F=32.063,P＜0.05);尼古丁組PAI-1 mRNA及蛋白含量[(1.32±0.20),(21.08±0.83)μg/L]明顯高于對照組[(0.73±0.10),(13.39± 0.93)μg/L,q=8.43、11.97,均P＜0.05];尼古丁+STS組PAI-1 mRNA及蛋白含量[(1.07±0.10),(16.19±2.15)μg/L]較尼古丁組降低(q=5.61、7.61,均P＜0.05),但仍高于對照組(q=7.84、4.36,均P＜0.05);尼古丁+ PD98059組PAI-1 mRNA及蛋白錶達[(1.12±0.11),(17.52±1.72)μg/L]低于尼古丁組(q=4.68、5.54,均P＜0.05),仍高于對照組(q=8.77、6.43,均P＜0.05).結論 PKC-ERK1/2信號通路在尼古丁誘導的血管內皮細胞PAI-1錶達上調中髮揮一定作用.
목적 탐토단백격매C(PKC)-포외신호조절격매(ERK)1/2신호통로재니고정유도인제정맥내피세포(HUVECs)표체섬유용해매원격활물억제물-1(PAI-1)중적작용.방법 체외배양HUVECs,채용불동실험조건니고정진행간예,ELISA법측정세포상청액중PAI-1적농도,관찰니고정작용적최가농도화시간.진일보분별용PKC적억제제성형포균소staurosporine (STS)화ERK적억제제PD98059간예HUVECs,관찰PKC혹ERK피조단후대니고정유도적HUVECs 표체PAI-1적영향,ELISA측정각조세포상청액중PAI-1단백적표체수평,RT-PCR검측각조세포PAI-1 mRNA적표체.결과 100 μmol/L니고정조PAI-1단백수평[(22.6 ± 1.1)μg/L]명현고우대조조[(14.2± 2.8)μg/L,q=5.64,P＜0.05];이100 μmol/L 니고정분별여HUVECs부육0、4、6、8、12급24 h,각조PAI-1단백표체정시간의뢰성승고,병재12 h체도고봉(F=32.063,P＜0.05);니고정조PAI-1 mRNA급단백함량[(1.32±0.20),(21.08±0.83)μg/L]명현고우대조조[(0.73±0.10),(13.39± 0.93)μg/L,q=8.43、11.97,균P＜0.05];니고정+STS조PAI-1 mRNA급단백함량[(1.07±0.10),(16.19±2.15)μg/L]교니고정조강저(q=5.61、7.61,균P＜0.05),단잉고우대조조(q=7.84、4.36,균P＜0.05);니고정+ PD98059조PAI-1 mRNA급단백표체[(1.12±0.11),(17.52±1.72)μg/L]저우니고정조(q=4.68、5.54,균P＜0.05),잉고우대조조(q=8.77、6.43,균P＜0.05).결론 PKC-ERK1/2신호통로재니고정유도적혈관내피세포PAI-1표체상조중발휘일정작용.
Objective To explore the role of protein kinase C (PKC)- extracellular signal-regulated kinase (ERK)1/2 signal pathway in the process of plasminogen activator inhibitor-1(PAI-1) protein and mRNA expression in cultured human umbilical vein endothelial cells(HUVECs) induced by nicotine. Methods HUVECs were cultured to examine the effect of nicotine on the expression of secreting PAI-1 in HUVECs on different experimental conditions. The expression of PAI-1 protein was measured by ELISA. PKC inhibitor staurosporine (STS)and ERK1/2 inhibitor PD98059 were used to detect PKC or ERK1/2 function on the expression of PAI-1 in HUVECs induced by nicotine. The PAI-1 mRNA expression was determined by RT-PCR. Results The expression level of PAI-1 protein in 100 μmol/L nicotine treated group [(22.6±1.1) μg/L] increased significantly compared to the control group [(14.2±2.8) μg/L; q=5.64,P<0.05]. After stimulation with 100 μmol/L nicotine for 0,4,6,8,12 and 24 h, the levels of PAI-1 protein increased over time and reached the peak at 12 h (F=32.063,P<0.05).The PAI-1 mRNA and protein expression in nicotine treated group [(1.32±0.20), (21.08 ± 0.83) μg/L] increased significantly compared to the control group [(0.73±0.10), (13.39±0.93) μg/L; q=8.43,11.97,all P<0.05].Compared with nicotine treated group , the PAI-1 mRNA and protein expression in nicotine and STS treated group [(1.07±0.10),(16.19±2.15) μg/L] decreased significantly(q=5.61,7.61, all P<0.05), but still higher than the control group (q=7.84,4.36, all P<0.05). In nicotine and PD98059 treated group, the PAI-1 mRNA and protein expression [(1.12±0.11),(17.52±1.72) μg/L] decreased significantly compared to the nicotine treated group(q= 4.68,5.54, all P<0.05), still higher than the control group (q=8.77,6.43, all P<0.05). Conclusion PKC-ERK1/2 signal pathway may play a partial role in the up-regulation of PAI-1 induced by nicotine in HUVECs.
