PINK1敲除对氟化钠处理小鼠肺脏线粒体损伤、氧化应激和炎症反应的影响

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中文摘要：以6周龄雄性野生型(WT)C57BL/6J小鼠和磷酸酶与张力蛋白同源物诱导激酶1(PINK1)基因敲除(PINK1-/-)小鼠为对象,将小鼠随机分为 WT对照组,WT模型组和PINK1-/-模型组.WT模型组和PINK1-模型组小鼠饮用含100 mg/L氟化钠(NaF)的蒸馏水,WT对照组小鼠饮用蒸馏水,处理时间为12周.通过免疫印迹(Western blot)研究线粒体自噬相关蛋白表达水平;用透射电镜观察肺脏线粒体超微结构损伤;ATP通过检测试剂盒和RT-qPCR分别测定ATP水平和mtDNA拷贝数;DHE染色检测肺脏活性氧(ROS)水平;比色法测定超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活力以及丙二醛(MDA)的含量;TUNEL染色检测肺脏细胞凋亡;酶联免疫吸附法(ELISA)测定肺泡灌洗液中肿瘤坏死因子(TNF-α)和白介素6(IL-6)的水平.结果显示,与 WT模型组比较,PINK1-/-模型组小鼠肺脏中帕金森病蛋白2(Parkin)和微管相关蛋白1轻链3Ⅱ(LC3-Ⅱ)的蛋白表达水平显著降低(P＜0.05),p62/SQSTM1和线粒体外膜转运孔蛋白20(TOM20)蛋白表达水平显著增加(P＜0.05);与WT模型组比较,PINK1-模型组小鼠肺脏受损线粒体百分比增加(P＜0.05),ATP水平和mtDNA拷贝数减少(P＜0.05).PINK1-/-模型组小鼠肺脏ROS水平、MDA含量和TUNEL阳性细胞数升高,SOD和CAT活力降低,较 WT模型组有显著性差异(P＜0.05).此外,与 WT模型组比较,PINK1-/-模型组小鼠肺泡灌洗液中TNF-α和IL-6的水平显著增加(P＜0.05).结果表明,PINK1介导的线粒体自噬可减弱NaF引起的小鼠肺脏损伤.

外文标题：Effects of PINK1 knockout on sodium fluoride(NaF)-induced mitochondrial inju-ry,oxidative stress,and inflammation in mice lungs

外文摘要：Six-week-old male wild-type(WT)and PTEN-induced kinase 1(PINK1)knockout mice at C57BL/6J background were adopted and were randomly divided into the WT control group,WT model group,and PINK1-/-model group.Mice in the WT model group and PINK1-/-model group were given distilled water containing 100 mg/L sodium fluoride(NaF),mice in the WT control group were given distilled water for 12 weeks.The expressions of mitophagy-related pro-teins were investigated by Western blot.Ultrastructural damages of lung mitochondria were ob-served by transmission electron microscopy.ATP levels and mtDNA copy numbers were deter-mined by ATP detection kit and RT-qPCR,respectively.Reactive oxygen species(ROS)were de-tected by DHE staining.The activities of superoxide dismutase(SOD),catalase(CAT),and malon-dialdehyde(MDA)were determined by colorimetric method.The apoptosis of lung cells was detec-ted by TUNEL staining.The levels of tumor necrosis factor(TNF-α)and interleukin-6(IL-6)in alveolar lavage fluid were determined by enzyme linked immunosorbent assay(ELISA).The results showed that compared with the WT model group,the protein changes of Parkinson disease protein 2(Parkin)and microtubule-associated protein 1 light chain 3 Ⅱ(LC3-Ⅱ)were markedly decreased(P＜0.05),while the protein expressions of p62/SQSTM1 and mitochondrial 20 kDa outer mem-brane protein(TOM20)were significantly decreased(P＜0.05).The percentages of damaged mito-chondria in lung cells were increased,while the ATP levels and mtDNA copy number were de-creased in the PINK 1-/-model group when compared with those in the WT model group(P＜0.05).The ROS level,MDA content,and TUNEL-positive cell numbers were significantly in-creased,while the activities of SOD and CAT were decreased in the lungs of PINK1-/-model group,which were significantly different from those in WT model group(P＜0.05).Additionally,compared with the WT model group,the levels of TNF-α and IL-6 in alveolar lavage fluid of PINK1-/-model group were significantly increased by PINK1 knockout(P＜0.05).The results in-dicated that PINK1-mediated mitophagy could attenuate NaF-induced lung injury in mice.

外文关键词：

sodium fluoridelungPTEN-induced kinase 1mitophagyoxidative stressinflamma-tion

作者：

宋超、张爱国、王坤丽、宋予震、董青、靳双星、米俊宪、王宏魁、石冬梅、王俊东、彭巍、罗琴

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作者单位：

河南牧业经济学院动物医药学院郑州市动物营养代谢病与中毒病重点实验室,河南郑州 450000

华农(肇庆)生物产业技术研究院有限公司,广东肇庆 526000

青海省畜牧兽医科学院,青海西宁 810000

关键词：

氟化钠肺脏磷酸酶与张力蛋白同源物诱导激酶1 线粒体自噬氧化应激炎症

基金：

国家自然科学基金国家自然科学基金青海省科技计划河南省自然科学基金青年科学基金河南省高等学校重点科研项目河南省科技攻关计划

项目编号：

31902285321026312021-ZJ-73623230042021821A230005222102110155

出版年：

2024

DOI：

10.16303/j.cnki.1005-4545.2024.04.10

中国兽医学报

吉林大学

中国兽医学报

CSTPCD北大核心

影响因子：0.702

ISSN：1005-4545

年,卷(期)：2024.44(4)

参考文献量21