目的观察线粒体未折叠蛋白反应(mitochondrial unfolded protein response,mtUPR)在氯化锂-匹鲁卡品(pilocarpine,PILO)致痫大鼠海马神经中的变化及线粒体特异性抗氧化剂Mito-TEMPO对其影响。方法采用PILO诱导癫痫大鼠模型,并进一步采...目的观察线粒体未折叠蛋白反应(mitochondrial unfolded protein response,mtUPR)在氯化锂-匹鲁卡品(pilocarpine,PILO)致痫大鼠海马神经中的变化及线粒体特异性抗氧化剂Mito-TEMPO对其影响。方法采用PILO诱导癫痫大鼠模型,并进一步采用线粒体特异性抗氧化剂Mito-TEMPO进行干预;将成年雄性Wistar大鼠随机分为空白对照组(CON组)、致痫组(PILO组)、Mito-TEMPO组和PILO+Mito-TEMPO组;采用Nissl染色观察海马神经元损伤,电子透射显微镜观察线粒体超微结构,活性氧荧光探针(DCFDA)检测线粒体ROS生成,Rhodamine123染色法检测线粒体膜电位变化,Western blot法检测线粒体热休克蛋白HSP60、蛋白酶LONP1、线粒体蛋白酶CLpP的表达。结果(1)与CON组相比,PILO组海马神经线粒体超微结构破坏严重,线粒体ROS生成增多,线粒体膜电位降低;(2)与CON组相比,PILO组海马神经HSP60、LONP1和CLpP表达增加;(3)与PILO组相比,PILO+Mito-TEMPO组线粒体超微结构破坏减轻,线粒体ROS生成明显减少,线粒体膜电位增高;(4)与PILO组相比,PILO+Mito-TEMPO组海马神经HSP60、LONP1和CLpP表达降低。结论mtUPR在癫痫海马神经损伤中明显激活,Mito-TEMPO可能通过调控mtUPR对癫痫海马线粒体损伤发挥保护作用。展开更多
The serological hallmark of primary biliary cirrhosis (PBC) is the presence of pyruvate dehydrogenase complex E2 subunit (PDC-E2) antimitochondrial antibodies (AMAs). Anti- PDC-E2 antibodies cross-react specifically w...The serological hallmark of primary biliary cirrhosis (PBC) is the presence of pyruvate dehydrogenase complex E2 subunit (PDC-E2) antimitochondrial antibodies (AMAs). Anti- PDC-E2 antibodies cross-react specifically with mycobacterial hsp65, and we have demonstrated that the motif SxGDL[ILV]- AE shared by PDC- E2212- 226 and hsp’s is a cross-reactive target. Having found that this same motif is present only in β -galactosidase of Lactobacillus delbrueckii (BGAL LACDE), we hypothesized that this homology would also lead to cross-reactivity. The mimics were tested via ELISA for reactivity and competitive cross-reactivity using sera from100 AMA-positive and 23 AMA-negative PBC patients and 190 controls. An Escherichia colt (ECOLI) PDC-E2 mimic that has been pathogenetically linked to PBC but lacks this motif has been also tested. Anti-BGAL266-280 LACDE antibodies were restricted to AMA-positive patients (54 of 95, 57% ) and belonged to immunoglobulin (Ig) G3. Of the 190 controls, 22 (12% ; P < .001) had anti-BGAL266-280 antibodies, mainly of the IgG4 subclass. ECOLI PDC-E2 reactivity was virtually absent. BGAL266- 280/PDC- E2212- 226 reactivity of the IgG3 isotype was found in 52 (52% ) AMA-positive PBC patients but in only 1 of the controls (P < .001). LACDE BGAL266- 280/PDCXE2212- 226 reactivity was due to cross-reactivity as confirmed via competition ELISA.Antibody affinity for BGAL266- 280 was greater than for PDC- E2 mimics. Preincubation of a multireactive serum with BGAL266- 280 reduced the inhibition of enzymatic activity by 40% , while marginal effect (12% ) or no effect (2% ) was observed in human or ECOLIPDC-E2 mimics. In conclusion, IgG3 antibodies to BGAL LACDE cross-react with the major mitochondrial autoepitope and are characteristic of PBC.展开更多
文摘目的观察线粒体未折叠蛋白反应(mitochondrial unfolded protein response,mtUPR)在氯化锂-匹鲁卡品(pilocarpine,PILO)致痫大鼠海马神经中的变化及线粒体特异性抗氧化剂Mito-TEMPO对其影响。方法采用PILO诱导癫痫大鼠模型,并进一步采用线粒体特异性抗氧化剂Mito-TEMPO进行干预;将成年雄性Wistar大鼠随机分为空白对照组(CON组)、致痫组(PILO组)、Mito-TEMPO组和PILO+Mito-TEMPO组;采用Nissl染色观察海马神经元损伤,电子透射显微镜观察线粒体超微结构,活性氧荧光探针(DCFDA)检测线粒体ROS生成,Rhodamine123染色法检测线粒体膜电位变化,Western blot法检测线粒体热休克蛋白HSP60、蛋白酶LONP1、线粒体蛋白酶CLpP的表达。结果(1)与CON组相比,PILO组海马神经线粒体超微结构破坏严重,线粒体ROS生成增多,线粒体膜电位降低;(2)与CON组相比,PILO组海马神经HSP60、LONP1和CLpP表达增加;(3)与PILO组相比,PILO+Mito-TEMPO组线粒体超微结构破坏减轻,线粒体ROS生成明显减少,线粒体膜电位增高;(4)与PILO组相比,PILO+Mito-TEMPO组海马神经HSP60、LONP1和CLpP表达降低。结论mtUPR在癫痫海马神经损伤中明显激活,Mito-TEMPO可能通过调控mtUPR对癫痫海马线粒体损伤发挥保护作用。
文摘The serological hallmark of primary biliary cirrhosis (PBC) is the presence of pyruvate dehydrogenase complex E2 subunit (PDC-E2) antimitochondrial antibodies (AMAs). Anti- PDC-E2 antibodies cross-react specifically with mycobacterial hsp65, and we have demonstrated that the motif SxGDL[ILV]- AE shared by PDC- E2212- 226 and hsp’s is a cross-reactive target. Having found that this same motif is present only in β -galactosidase of Lactobacillus delbrueckii (BGAL LACDE), we hypothesized that this homology would also lead to cross-reactivity. The mimics were tested via ELISA for reactivity and competitive cross-reactivity using sera from100 AMA-positive and 23 AMA-negative PBC patients and 190 controls. An Escherichia colt (ECOLI) PDC-E2 mimic that has been pathogenetically linked to PBC but lacks this motif has been also tested. Anti-BGAL266-280 LACDE antibodies were restricted to AMA-positive patients (54 of 95, 57% ) and belonged to immunoglobulin (Ig) G3. Of the 190 controls, 22 (12% ; P < .001) had anti-BGAL266-280 antibodies, mainly of the IgG4 subclass. ECOLI PDC-E2 reactivity was virtually absent. BGAL266- 280/PDC- E2212- 226 reactivity of the IgG3 isotype was found in 52 (52% ) AMA-positive PBC patients but in only 1 of the controls (P < .001). LACDE BGAL266- 280/PDCXE2212- 226 reactivity was due to cross-reactivity as confirmed via competition ELISA.Antibody affinity for BGAL266- 280 was greater than for PDC- E2 mimics. Preincubation of a multireactive serum with BGAL266- 280 reduced the inhibition of enzymatic activity by 40% , while marginal effect (12% ) or no effect (2% ) was observed in human or ECOLIPDC-E2 mimics. In conclusion, IgG3 antibodies to BGAL LACDE cross-react with the major mitochondrial autoepitope and are characteristic of PBC.
