目的利用生物信息学的方法分析细粒棘球绦虫Fis1(EgFis1)蛋白的抗原表位,为分子肽疫苗的研发奠定理论基础。方法在NCBI数据库中检所并下载EgFis1蛋白的氨基酸序列;利用EXpasy软件预测蛋白的理化性质;采用SignalP5.0和TMHMM sever 2.0软...目的利用生物信息学的方法分析细粒棘球绦虫Fis1(EgFis1)蛋白的抗原表位,为分子肽疫苗的研发奠定理论基础。方法在NCBI数据库中检所并下载EgFis1蛋白的氨基酸序列;利用EXpasy软件预测蛋白的理化性质;采用SignalP5.0和TMHMM sever 2.0软件预测EgFis1蛋白的信号肽和跨膜结构域;利用SOPMA和SWISS-MODLE预测EgFis1蛋白的二级结构和三级结构;采用IEDB、ABCpred和SYFPEITHI数据库预测EgFis1蛋白的T、B细胞表位。结果细粒棘球绦虫EgFis1是由157个氨基酸组成的等电点为5.86,分子质量单位为16.93ku的蛋白质;不含有信号肽序列,但具有一个跨膜结构域;其二级结构中α-螺旋占比例为48.41%,延伸连占19.11%,β-转角占7.01%,无规则卷曲占25.48%。亲水性较强的区域主要位于12aa-24aa,42aa-52aa,64aa-69aa,81aa-90aa,97aa-105aa,126aa-150aa。Eg-Fis1含有3个优势B细胞表位,7个T细胞表位以及2个T、B联合表位。结论生物信息学方法预测EgFis1蛋白含有4个优势B细胞表位、7个T细胞表位以及2个T、B联合表位,可作为免疫治疗和药物治疗的靶点。展开更多
Selective brain hypothermia is considered an effective treatment for neuronal injury after stroke,and avoids the complications of general hypothermia.However,the mechanisms by which selective brain hypothermia affects...Selective brain hypothermia is considered an effective treatment for neuronal injury after stroke,and avoids the complications of general hypothermia.However,the mechanisms by which selective brain hypothermia affects mitochondrial fission remain unknown.In this study,we investigated the effect of selective brain hypothermia on the expression of fission 1 (Fis1) protein,a key factor in the mitochondrial fission system,during focal cerebral ischemia/reperfusion injury.Sprague-Dawley rats were divided into four groups.In the sham group,the carotid arteries were exposed only.In the other three groups,middle cerebral artery occlusion was performed using the intraluminal filament technique.After 2 hours of occlusion,the filament was slowly removed to allow blood reperfusion in the ischemia/reperfusion group.Saline,at 4℃ and 37℃,were perfused through the carotid artery in the hypothermia and normothermia groups,respectively,followed by restoration of blood flow.Neurological function was assessed with the Zea Longa 5-point scoring method.Cerebral infarct volume was assessed by 2,3,5-triphenyltetrazolium chloride staining,and apoptosis was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining.Fis1 and cytosolic cytochrome c levels were assessed by western blot assay.Fis1 mRNA expression was assessed by quantitative reverse transcription-polymerase chain reaction.Mitochondrial ultrastructure was evaluated by transmission electron microscopy.Compared with the sham group,apoptosis,Fis1 protein and mRNA expression and cytosolic cytochrome c levels in the cortical ischemic penumbra and cerebral infarct volume were increased after reperfusion in the other three groups.These changes caused by cerebral ischemia/reperfusion were inhibited in the hypothermia group compared with the normothermia group.These findings show that selective brain hypothermia inhibits Fis1 expression and reduces apoptosis,thereby ameliorating focal cerebral ischemia/reperfusion injury in rats.Experiments were authorized by the Ethics Committee of Qingdao Municipal Hospital of China (approval No.2019008).展开更多
目的:探讨线粒体分裂蛋白1(mitochondrial fission protein 1,FIS1)在顺铂(cisplatin,CDDP)耐药的骨肉瘤(osteosarcoma,OS)组织中的表达及对CDDP耐药OS细胞凋亡的作用。方法:采用免疫组织化学法(immunohistochemistry,IHC)和蛋白质印迹...目的:探讨线粒体分裂蛋白1(mitochondrial fission protein 1,FIS1)在顺铂(cisplatin,CDDP)耐药的骨肉瘤(osteosarcoma,OS)组织中的表达及对CDDP耐药OS细胞凋亡的作用。方法:采用免疫组织化学法(immunohistochemistry,IHC)和蛋白质印迹法检测CDDP耐药的OS组织(CDDP-resistance组)和CDDP敏感的OS组织(CDDP-sensity组)中FIS1的表达水平,生物信息学方法分析FIS1在OS中的相关作用通路,流式细胞术检测CDDP耐药细胞凋亡及过表达FIS1对细胞活性氧(reactive oxygen species,ROS)的作用,蛋白质印迹法检测FIS1对P53信号通路的作用,OS公共数据分析FIS1与P53的相关性。结果:FIS1在18例CDDP-sensity组中表达明显比其在15例CDDP-resistance组中表达增高;FIS1在OS耐CDDP细胞系MG63/Cis-R细胞中的蛋白质表达低于普通MG63组;人OS组织中与FIS1相关的前10位KEGG(Kyoto Encyclopedia of Genes and Genomes)通路,凋亡通路排在首位。在OS耐CDDP细胞系MG63/Cis-R细胞中,FIS1高表达组细胞与对照组相比,细胞凋亡比例增高(P<0.05),ROS水平降低(P<0.05),P53、BAX和cleaved Caspase3蛋白质表达上调,BCL-2分子表达降低;在OS组织中同样发现FIS1与P53、BAX、cleaved Caspase3表达正相关,与BCL-2表达负相关。结论:FIS1通过激活P53通路进而调控CDDP耐药OS细胞的凋亡,而FIS1可能在OS CDDP耐药中发挥重要作用,有望成为OS化疗耐药的新靶点。展开更多
[目的]研究饲料钙对母鼠饮水型氟染毒后子代大鼠肾细胞线粒体损伤的影响。[方法]选用健康初断乳SD雌性大鼠100只,随机分为对照组、染氟组(100 mg/L Na F)、低钙组(0.063%CaCO_3)、低钙染氟组(100 mg/L Na F+0.063%CaCO_3)和高钙染氟组(1...[目的]研究饲料钙对母鼠饮水型氟染毒后子代大鼠肾细胞线粒体损伤的影响。[方法]选用健康初断乳SD雌性大鼠100只,随机分为对照组、染氟组(100 mg/L Na F)、低钙组(0.063%CaCO_3)、低钙染氟组(100 mg/L Na F+0.063%CaCO_3)和高钙染氟组(100 mg/L Na F+7%CaCO_3);饲养3个月后,雌雄鼠合笼交配。取日龄14 d及28 d仔鼠雌雄各10只,以其肾脏细胞线粒体标志酶琥珀酸脱氢酶(SDHase)活性及脂质过氧化指标丙二醛(MDA)水平,肾脏细胞凋亡状况,线粒体分裂/融合蛋白Fis1、Drp1和Mfn2表达水平为观察指标。[结果]与染氟组相比,高钙染氟组SDHase活性升高(P<0.05),低钙染氟组SDHase活性降低(P<0.05)。与对照组相比,雌鼠各组肾脏线粒体MDA含量均升高(P<0.05)。与对照组相比,染氟各组仔鼠凋亡细胞增多;与染氟组相比,低钙染氟组凋亡细胞增多,而高钙染氟组凋亡细胞减少。与对照组相比,低钙染氟组14 d雄鼠的分裂蛋白Fis1表达升高(P<0.05);低钙染氟组和高钙染氟组28 d雄鼠的分裂蛋白Drp1升高(P<0.05)。[结论]氟中毒能够造成大鼠肾脏细胞线粒体内分裂/融合蛋白Fis1、Drp1和Mfn2表达异常,引起肾脏细胞线粒体损伤。高钙饲料摄入能降低线粒体内脂质过氧化反应,减轻高氟对子代肾脏细胞的毒性作用,而低钙饲料摄入会加剧高氟的毒性作用。展开更多
文摘目的利用生物信息学的方法分析细粒棘球绦虫Fis1(EgFis1)蛋白的抗原表位,为分子肽疫苗的研发奠定理论基础。方法在NCBI数据库中检所并下载EgFis1蛋白的氨基酸序列;利用EXpasy软件预测蛋白的理化性质;采用SignalP5.0和TMHMM sever 2.0软件预测EgFis1蛋白的信号肽和跨膜结构域;利用SOPMA和SWISS-MODLE预测EgFis1蛋白的二级结构和三级结构;采用IEDB、ABCpred和SYFPEITHI数据库预测EgFis1蛋白的T、B细胞表位。结果细粒棘球绦虫EgFis1是由157个氨基酸组成的等电点为5.86,分子质量单位为16.93ku的蛋白质;不含有信号肽序列,但具有一个跨膜结构域;其二级结构中α-螺旋占比例为48.41%,延伸连占19.11%,β-转角占7.01%,无规则卷曲占25.48%。亲水性较强的区域主要位于12aa-24aa,42aa-52aa,64aa-69aa,81aa-90aa,97aa-105aa,126aa-150aa。Eg-Fis1含有3个优势B细胞表位,7个T细胞表位以及2个T、B联合表位。结论生物信息学方法预测EgFis1蛋白含有4个优势B细胞表位、7个T细胞表位以及2个T、B联合表位,可作为免疫治疗和药物治疗的靶点。
基金supported by the Natural Science Foundation of Shandong Province of China,No.ZR2015HM023(to MSW)the Science and Technology Plan Project of Qingdao City of China,No.19-6-1-50-nsh(to MSW)
文摘Selective brain hypothermia is considered an effective treatment for neuronal injury after stroke,and avoids the complications of general hypothermia.However,the mechanisms by which selective brain hypothermia affects mitochondrial fission remain unknown.In this study,we investigated the effect of selective brain hypothermia on the expression of fission 1 (Fis1) protein,a key factor in the mitochondrial fission system,during focal cerebral ischemia/reperfusion injury.Sprague-Dawley rats were divided into four groups.In the sham group,the carotid arteries were exposed only.In the other three groups,middle cerebral artery occlusion was performed using the intraluminal filament technique.After 2 hours of occlusion,the filament was slowly removed to allow blood reperfusion in the ischemia/reperfusion group.Saline,at 4℃ and 37℃,were perfused through the carotid artery in the hypothermia and normothermia groups,respectively,followed by restoration of blood flow.Neurological function was assessed with the Zea Longa 5-point scoring method.Cerebral infarct volume was assessed by 2,3,5-triphenyltetrazolium chloride staining,and apoptosis was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining.Fis1 and cytosolic cytochrome c levels were assessed by western blot assay.Fis1 mRNA expression was assessed by quantitative reverse transcription-polymerase chain reaction.Mitochondrial ultrastructure was evaluated by transmission electron microscopy.Compared with the sham group,apoptosis,Fis1 protein and mRNA expression and cytosolic cytochrome c levels in the cortical ischemic penumbra and cerebral infarct volume were increased after reperfusion in the other three groups.These changes caused by cerebral ischemia/reperfusion were inhibited in the hypothermia group compared with the normothermia group.These findings show that selective brain hypothermia inhibits Fis1 expression and reduces apoptosis,thereby ameliorating focal cerebral ischemia/reperfusion injury in rats.Experiments were authorized by the Ethics Committee of Qingdao Municipal Hospital of China (approval No.2019008).
文摘目的:探讨线粒体分裂蛋白1(mitochondrial fission protein 1,FIS1)在顺铂(cisplatin,CDDP)耐药的骨肉瘤(osteosarcoma,OS)组织中的表达及对CDDP耐药OS细胞凋亡的作用。方法:采用免疫组织化学法(immunohistochemistry,IHC)和蛋白质印迹法检测CDDP耐药的OS组织(CDDP-resistance组)和CDDP敏感的OS组织(CDDP-sensity组)中FIS1的表达水平,生物信息学方法分析FIS1在OS中的相关作用通路,流式细胞术检测CDDP耐药细胞凋亡及过表达FIS1对细胞活性氧(reactive oxygen species,ROS)的作用,蛋白质印迹法检测FIS1对P53信号通路的作用,OS公共数据分析FIS1与P53的相关性。结果:FIS1在18例CDDP-sensity组中表达明显比其在15例CDDP-resistance组中表达增高;FIS1在OS耐CDDP细胞系MG63/Cis-R细胞中的蛋白质表达低于普通MG63组;人OS组织中与FIS1相关的前10位KEGG(Kyoto Encyclopedia of Genes and Genomes)通路,凋亡通路排在首位。在OS耐CDDP细胞系MG63/Cis-R细胞中,FIS1高表达组细胞与对照组相比,细胞凋亡比例增高(P<0.05),ROS水平降低(P<0.05),P53、BAX和cleaved Caspase3蛋白质表达上调,BCL-2分子表达降低;在OS组织中同样发现FIS1与P53、BAX、cleaved Caspase3表达正相关,与BCL-2表达负相关。结论:FIS1通过激活P53通路进而调控CDDP耐药OS细胞的凋亡,而FIS1可能在OS CDDP耐药中发挥重要作用,有望成为OS化疗耐药的新靶点。
文摘[目的]研究饲料钙对母鼠饮水型氟染毒后子代大鼠肾细胞线粒体损伤的影响。[方法]选用健康初断乳SD雌性大鼠100只,随机分为对照组、染氟组(100 mg/L Na F)、低钙组(0.063%CaCO_3)、低钙染氟组(100 mg/L Na F+0.063%CaCO_3)和高钙染氟组(100 mg/L Na F+7%CaCO_3);饲养3个月后,雌雄鼠合笼交配。取日龄14 d及28 d仔鼠雌雄各10只,以其肾脏细胞线粒体标志酶琥珀酸脱氢酶(SDHase)活性及脂质过氧化指标丙二醛(MDA)水平,肾脏细胞凋亡状况,线粒体分裂/融合蛋白Fis1、Drp1和Mfn2表达水平为观察指标。[结果]与染氟组相比,高钙染氟组SDHase活性升高(P<0.05),低钙染氟组SDHase活性降低(P<0.05)。与对照组相比,雌鼠各组肾脏线粒体MDA含量均升高(P<0.05)。与对照组相比,染氟各组仔鼠凋亡细胞增多;与染氟组相比,低钙染氟组凋亡细胞增多,而高钙染氟组凋亡细胞减少。与对照组相比,低钙染氟组14 d雄鼠的分裂蛋白Fis1表达升高(P<0.05);低钙染氟组和高钙染氟组28 d雄鼠的分裂蛋白Drp1升高(P<0.05)。[结论]氟中毒能够造成大鼠肾脏细胞线粒体内分裂/融合蛋白Fis1、Drp1和Mfn2表达异常,引起肾脏细胞线粒体损伤。高钙饲料摄入能降低线粒体内脂质过氧化反应,减轻高氟对子代肾脏细胞的毒性作用,而低钙饲料摄入会加剧高氟的毒性作用。
