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
2011年
12期
925-929
,共5页
陈晨%杨沫%张遵真%吴媚%邓雯文
陳晨%楊沫%張遵真%吳媚%鄧雯文
진신%양말%장준진%오미%산문문
DNA聚合酶β%氢醌类%膜电位%过氧化物酶类
DNA聚閤酶β%氫醌類%膜電位%過氧化物酶類
DNA취합매β%경곤류%막전위%과양화물매류
DNA polymerase beta%Hydroquinones%Membrane potentials%Peroxidases
目的 探讨DNA聚合酶β(DNA polymerase beta,polβ)对氢醌(HQ)所致细胞凋亡的影响及其可能的分子机制.方法 以polβ野生型(polβ+/+)、polβ缺陷型(polβ-/-)及polβ高表达型(polβoe)小鼠胚胎成纤维细胞为研究对象,以不同浓度的HQ溶液染毒后,采用流式细胞术分别检测HQ对细胞凋亡和线粒体膜电位( △Ψm)的影响,试剂盒法测定细胞内活性氧(ROS)和羟自由基(·OH)含量;化学发光法分析细胞内超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)活力.结果 与对照组相比,各染毒组凋亡细胞率和△Ψm降低的细胞比例明显增加,差异均有统计学意义(P＜0.05).与polβ+/+细胞比较,20.00、40.00、80.00 μmol/L染毒组polβ-/-细胞凋亡率明显增高,10.00、20.00、40.00、80.00μmol/L染毒组polβ oe细胞凋亡率明显降低,差异均有统计学意义(P＜0.05).与polβ+/+细胞(20.60％±0.57％、37.95％+0.64％、44.50％±1.27％、57.55％+1.06％)比较,10.00、20.00、40.00、80.00 μmol/L染毒组polβ-/-细胞△Ψm降低的细胞比例(33.60％±1.55％ 、46.05％±1.77％、52.75％±2.05％、75.20％±0.56％)明显增高,polβ oe细胞△Ψm降低的细胞比例(16.05％±1.20％、29.80％±1.21％、35.15％±1.06％、53.80％±0.85％)明显降低,差异均有统计学意义(P＜0.05).与polβ+/+细胞比较,各染毒组polβ-/-细胞产生的荧光强度明显增高,polβ oe细胞的荧光强度明显降低,差异均有统计学意义(P＜0.05).与polβ+/+细胞相比,20.00、40.00 μmol/L染毒组polβ-/-细胞内·OH含量明显增高,polβ oe细胞内·OH含量明显降低,差异有统计学意义(P＜0.05).在相同浓度HQ染毒剂量下,polβ-/-细胞内SOD和GSH-Px消耗最快,polβ oe细胞内的SOD和GSH-Px消耗速度缓慢.结论 HQ可诱导细胞产生ROS,降低△Ψm,从而介导凋亡的发生;polβ可以帮助细胞对抗ROS的产生,降低线粒体发生去极化的几率,从而对HQ介导的凋亡起到一定的保护作用.
目的 探討DNA聚閤酶β(DNA polymerase beta,polβ)對氫醌(HQ)所緻細胞凋亡的影響及其可能的分子機製.方法 以polβ野生型(polβ+/+)、polβ缺陷型(polβ-/-)及polβ高錶達型(polβoe)小鼠胚胎成纖維細胞為研究對象,以不同濃度的HQ溶液染毒後,採用流式細胞術分彆檢測HQ對細胞凋亡和線粒體膜電位( △Ψm)的影響,試劑盒法測定細胞內活性氧(ROS)和羥自由基(·OH)含量;化學髮光法分析細胞內超氧化物歧化酶(SOD)和穀胱甘肽過氧化物酶(GSH-Px)活力.結果 與對照組相比,各染毒組凋亡細胞率和△Ψm降低的細胞比例明顯增加,差異均有統計學意義(P＜0.05).與polβ+/+細胞比較,20.00、40.00、80.00 μmol/L染毒組polβ-/-細胞凋亡率明顯增高,10.00、20.00、40.00、80.00μmol/L染毒組polβ oe細胞凋亡率明顯降低,差異均有統計學意義(P＜0.05).與polβ+/+細胞(20.60％±0.57％、37.95％+0.64％、44.50％±1.27％、57.55％+1.06％)比較,10.00、20.00、40.00、80.00 μmol/L染毒組polβ-/-細胞△Ψm降低的細胞比例(33.60％±1.55％ 、46.05％±1.77％、52.75％±2.05％、75.20％±0.56％)明顯增高,polβ oe細胞△Ψm降低的細胞比例(16.05％±1.20％、29.80％±1.21％、35.15％±1.06％、53.80％±0.85％)明顯降低,差異均有統計學意義(P＜0.05).與polβ+/+細胞比較,各染毒組polβ-/-細胞產生的熒光彊度明顯增高,polβ oe細胞的熒光彊度明顯降低,差異均有統計學意義(P＜0.05).與polβ+/+細胞相比,20.00、40.00 μmol/L染毒組polβ-/-細胞內·OH含量明顯增高,polβ oe細胞內·OH含量明顯降低,差異有統計學意義(P＜0.05).在相同濃度HQ染毒劑量下,polβ-/-細胞內SOD和GSH-Px消耗最快,polβ oe細胞內的SOD和GSH-Px消耗速度緩慢.結論 HQ可誘導細胞產生ROS,降低△Ψm,從而介導凋亡的髮生;polβ可以幫助細胞對抗ROS的產生,降低線粒體髮生去極化的幾率,從而對HQ介導的凋亡起到一定的保護作用.
목적 탐토DNA취합매β(DNA polymerase beta,polβ)대경곤(HQ)소치세포조망적영향급기가능적분자궤제.방법 이polβ야생형(polβ+/+)、polβ결함형(polβ-/-)급polβ고표체형(polβoe)소서배태성섬유세포위연구대상,이불동농도적HQ용액염독후,채용류식세포술분별검측HQ대세포조망화선립체막전위( △Ψm)적영향,시제합법측정세포내활성양(ROS)화간자유기(·OH)함량;화학발광법분석세포내초양화물기화매(SOD)화곡광감태과양화물매(GSH-Px)활력.결과 여대조조상비,각염독조조망세포솔화△Ψm강저적세포비례명현증가,차이균유통계학의의(P＜0.05).여polβ+/+세포비교,20.00、40.00、80.00 μmol/L염독조polβ-/-세포조망솔명현증고,10.00、20.00、40.00、80.00μmol/L염독조polβ oe세포조망솔명현강저,차이균유통계학의의(P＜0.05).여polβ+/+세포(20.60％±0.57％、37.95％+0.64％、44.50％±1.27％、57.55％+1.06％)비교,10.00、20.00、40.00、80.00 μmol/L염독조polβ-/-세포△Ψm강저적세포비례(33.60％±1.55％ 、46.05％±1.77％、52.75％±2.05％、75.20％±0.56％)명현증고,polβ oe세포△Ψm강저적세포비례(16.05％±1.20％、29.80％±1.21％、35.15％±1.06％、53.80％±0.85％)명현강저,차이균유통계학의의(P＜0.05).여polβ+/+세포비교,각염독조polβ-/-세포산생적형광강도명현증고,polβ oe세포적형광강도명현강저,차이균유통계학의의(P＜0.05).여polβ+/+세포상비,20.00、40.00 μmol/L염독조polβ-/-세포내·OH함량명현증고,polβ oe세포내·OH함량명현강저,차이유통계학의의(P＜0.05).재상동농도HQ염독제량하,polβ-/-세포내SOD화GSH-Px소모최쾌,polβ oe세포내적SOD화GSH-Px소모속도완만.결론 HQ가유도세포산생ROS,강저△Ψm,종이개도조망적발생;polβ가이방조세포대항ROS적산생,강저선립체발생거겁화적궤솔,종이대HQ개도적조망기도일정적보호작용.
Objective To explore the effect and mechanism of DNA polymerase β expression level on cell apoptosis and mitochondrial membrane potential induced by hydroquinone.Methods Polβ wild-type cells (polβ +/+),polβ overexpressed cells (polβ oe) and polβ null cells (polβ -/-) were applied as a model cell system,The effect of cell apoptosis and mitochondrial membrane potential induced by different doses of hydroquinone were analyzed by flow cytometry.The ROS and ·OH assay kit were used to examine the cellular ROS and ·OH level.The activity of cellular SOD and GSH-Px were tested by Chemiluminescence method after exposed to different concentrations of hydroquinone.Results With the dose of hydroquinone increased,the rate of apoptosis and falling of mitochondrial membrane potential (△Ψm) in cells were increased compared with the control.When compared with polβ +/+ cells,the rate of apoptosis in polβ -/- cells exposed to 20.00,40.00,80.00μmol/L hydroquinone increased and the rate of apoptosis in polβ oe cells exposed to 10.00,20.00,40.00,80.00μ mol/L hydroquinone decreased (P＜0.05).Compared with polβ +/+ cells (20.60％±0.57％,37.95％±0.64％,44.50％±1.27％,57.55％±1.06％ ),the rate of cell which undergone mitochondrial depolarization in polβ -/-cells treated with 10.00,20.00,40.00,80.00 μmol/L hydroquinone (33.60％±1.55％,46.05％±1.77％,52.75％±2.05％,75.20％±0.56％) increased.The rate of cell which undergone mitochondrial depolarization in polβ oe cells exposed to 10.00,20.00,40.00,80.00 μmol/L hydroquinone ( 16.05％+ 1.20％,29.80％± 1.21％,35.15％±1.06％,53.80％±0.85％ ) decreased (P＜0.05).When compared with polβ +/+ cells,fluorescent intensity of polβ -/- cells treated with different dosages of hydroquinone increased,while which of polβ oe cells decreased (P＜0.05).Compared with polβ +/+ cells,.OH level of polβ -/- cells treated with 20.00,40.00 μmol/L hydroquinone significantly enhanced,while which of polβ oe cells decreased sharply (P＜0.05).Under the same concentrations of hydroquinone,the activity of SOD and GSH-Px were decreased most rapidly in polβ -/- cells.The activity of SOD and GSH-Px in polβ oe cells decreased slower than in the polβ-/- cells.Conclusion Hydroquinone could induced apoptosis by the generation of ROS and decrease of △Ψm; polβ could protect cells from apoptosis induced by hydroquinone through decrease of ROS level and depolarization of mitochondria.
