中国病理生理杂志
中國病理生理雜誌
중국병리생리잡지
CHINESE JOURNAL OF PATHOPHYSIOLOGY
2009年
11期
2113-2117
,共5页
万磊%李菊香%洪葵%丁浩%夏子荣%苏海%颜素娟%吴延庆%吴清华%程晓曙
萬磊%李菊香%洪葵%丁浩%夏子榮%囌海%顏素娟%吳延慶%吳清華%程曉曙
만뢰%리국향%홍규%정호%하자영%소해%안소연%오연경%오청화%정효서
Nox-1%心肌细胞%缺氧复氧%心肌营养素-1
Nox-1%心肌細胞%缺氧複氧%心肌營養素-1
Nox-1%심기세포%결양복양%심기영양소-1
Nox -1%Cardiomyocytes%Hypoxia - reoxygenation%Cardiotrophin -1
目的:探讨心肌细胞急性缺氧复氧损伤时NADPH氧化酶亚单位nox-1的变化及心肌营养素-1的作用.方法:用改良的方法培养出生1-3 d的乳鼠心肌细胞,分为6组:(1)对照组;(2)缺氧复氧组;(3)缺氧复氧+CT-1组;(4)缺氧复氧+CT-1+LY294002组 (PIK3/Akt 阻断剂);(5)缺氧复氧+CT-1+PD98059组(ERK 阻断剂);(6)缺氧复氧+CT-1+助溶剂DMSO组. CT-1 的浓度为10 μg/L.MTS法测定心肌细胞的存活率,四氯四乙基苯丙咪唑基羰化青碘化物(JC1)检测心肌细胞线粒体膜电位(Δψm),二氯荧光黄双乙酸盐(DCFH-CA)检测细胞活性氧(ROS),流式细胞仪检测心肌细胞凋亡率.Nox-1蛋白采用Western blotting检测.结果:缺氧复氧培养后心肌细胞凋亡率及细胞内ROS较对照组明显增加,分别是(19.7%±1.4% vs 2.1%±0.5%, 14.07%±1.25% vs 3.54%±0.86%, P<0.05),而心肌细胞存活率显著降低,线粒体膜电位(Δψm)下降;nox-1表达明显升高.CT-1处理的心肌细胞,较缺氧复氧组心肌细胞存活率明显上升 (87.0%±7.3%),而心肌细胞凋亡率及细胞内ROS 显著减少,Δψm水平增加,nox-1蛋白表达下调.而CT-1的这些作用能被PIK3/Akt和ERK阻断剂抑制.结论:心肌细胞急性缺氧复氧损伤时NADPH氧化酶亚单位nox-1表达上调,而心肌营养素-1能通过下调nox-1表达,发挥对心肌细胞保护作用.
目的:探討心肌細胞急性缺氧複氧損傷時NADPH氧化酶亞單位nox-1的變化及心肌營養素-1的作用.方法:用改良的方法培養齣生1-3 d的乳鼠心肌細胞,分為6組:(1)對照組;(2)缺氧複氧組;(3)缺氧複氧+CT-1組;(4)缺氧複氧+CT-1+LY294002組 (PIK3/Akt 阻斷劑);(5)缺氧複氧+CT-1+PD98059組(ERK 阻斷劑);(6)缺氧複氧+CT-1+助溶劑DMSO組. CT-1 的濃度為10 μg/L.MTS法測定心肌細胞的存活率,四氯四乙基苯丙咪唑基羰化青碘化物(JC1)檢測心肌細胞線粒體膜電位(Δψm),二氯熒光黃雙乙痠鹽(DCFH-CA)檢測細胞活性氧(ROS),流式細胞儀檢測心肌細胞凋亡率.Nox-1蛋白採用Western blotting檢測.結果:缺氧複氧培養後心肌細胞凋亡率及細胞內ROS較對照組明顯增加,分彆是(19.7%±1.4% vs 2.1%±0.5%, 14.07%±1.25% vs 3.54%±0.86%, P<0.05),而心肌細胞存活率顯著降低,線粒體膜電位(Δψm)下降;nox-1錶達明顯升高.CT-1處理的心肌細胞,較缺氧複氧組心肌細胞存活率明顯上升 (87.0%±7.3%),而心肌細胞凋亡率及細胞內ROS 顯著減少,Δψm水平增加,nox-1蛋白錶達下調.而CT-1的這些作用能被PIK3/Akt和ERK阻斷劑抑製.結論:心肌細胞急性缺氧複氧損傷時NADPH氧化酶亞單位nox-1錶達上調,而心肌營養素-1能通過下調nox-1錶達,髮揮對心肌細胞保護作用.
목적:탐토심기세포급성결양복양손상시NADPH양화매아단위nox-1적변화급심기영양소-1적작용.방법:용개량적방법배양출생1-3 d적유서심기세포,분위6조:(1)대조조;(2)결양복양조;(3)결양복양+CT-1조;(4)결양복양+CT-1+LY294002조 (PIK3/Akt 조단제);(5)결양복양+CT-1+PD98059조(ERK 조단제);(6)결양복양+CT-1+조용제DMSO조. CT-1 적농도위10 μg/L.MTS법측정심기세포적존활솔,사록사을기분병미서기탄화청전화물(JC1)검측심기세포선립체막전위(Δψm),이록형광황쌍을산염(DCFH-CA)검측세포활성양(ROS),류식세포의검측심기세포조망솔.Nox-1단백채용Western blotting검측.결과:결양복양배양후심기세포조망솔급세포내ROS교대조조명현증가,분별시(19.7%±1.4% vs 2.1%±0.5%, 14.07%±1.25% vs 3.54%±0.86%, P<0.05),이심기세포존활솔현저강저,선립체막전위(Δψm)하강;nox-1표체명현승고.CT-1처리적심기세포,교결양복양조심기세포존활솔명현상승 (87.0%±7.3%),이심기세포조망솔급세포내ROS 현저감소,Δψm수평증가,nox-1단백표체하조.이CT-1적저사작용능피PIK3/Akt화ERK조단제억제.결론:심기세포급성결양복양손상시NADPH양화매아단위nox-1표체상조,이심기영양소-1능통과하조nox-1표체,발휘대심기세포보호작용.
AIM: To observe the change of subunit of NADPH oxidation enzyme complex nox - 1 protein in cardiocyte hypoxia - reoxygenation injury and the role of cardiotrophin -1.METHODS: Cardiomyocytes from the hearts of 1 -3 d old neonatal rats were prepared by a modified method. Five groups were included in the study: control; hypoxia/ reoxygenation; hypoxia/reoxygenation + CT - 1; CT - 1 + hypoxia/reoxygenation + LY294002 (PIK3/Akt inhibitor) ; CT -1 + hypoxia/reoxygenation + PD98059 (ERK inhibitor) ; CT - 1 + hypoxia/reoxygenation + DMSO. The concentration of CT -1 was 10 μg/L. The survival rate of myocytes was evaluated by MTS method. Apoptosis, mitochondrial permeability transition pore ( △ψm) and reactive oxygen species ( ROS) were detected by flow cytometry. Nox - 1 protein was determined by Western blotting. RESULTS: Apoptosis of cardiomyocytes and the level of ROS (19.7% ±1.4% vs 2.1% ± 0.5% , 14.07% ± 1.25% vs 3.54% ± 0.86% , P < 0.05 ) increased markedly after hypoxia/reoxygenation, but cardio-myocyte survival rate and the level of △ψm (40.55% ±4.25% vs 86.28% ±7.15% , P <0.01) decreased significantly. The expression of nox - 1 protein was upregulated markedly. With CT - 1 intervention, cardiomyocyte survival rate increased markedly, apoptosis, both ROS and expression of nox - 1 protein reduced significantly. The level of △ψm increased obviously. The effect of CT - 1 was inhibited by LY294002.No significant effect was observed on cells survival in DMSO group, which confirmed that LY294002 was specifically involved in blocking the protective effect of CT - 1.CONCLUSION : The expression of subunit of NADPH oxidation enzyme complex nox - 1 protein is upregulated markedly in cardiocyte hypoxia - reoxygenation injury. CT - 1 protects cardiac cells against hypoxia - reoxygenation injury by downregulating the expression of nox -1 protein to decrease the level of ROS.
