溴莫尼定治疗大鼠视神经夹挫伤机制探讨

目的观察0.2%酒石酸溴莫尼定对视神经夹挫伤大鼠视网膜形态及bcl-2/bax表达的影响,探讨其作用机制。方法雌性SD大鼠90只随机分为正常组、模型组、治疗组,每组30只。60只大鼠用视神经钳夹法制作SD大鼠视神经夹挫伤模型,治疗组30只鼠给予0.2%酒石酸溴莫尼定点眼,于实验的l、3、5、7、14、21d处死大鼠,用苏木精-伊红染色计数各组鼠视网膜神经节细胞（RGCs）的数量,用透射电镜观察和比较各组鼠视网膜的超微结构改变,免疫组织化学染色检测鼠视网膜中bcl-2及bax的表达。结果正常组视网膜结构正常,治疗组较模型组视网膜形态损伤减轻。造模后3～21d,模型组和治疗组较正常对照组RGCs数量明显减少（P〈0.05）,但治疗组较模型组RGCs数量明显增加（P〈0.01）。造模后5～7d,治疗组bcl-2在鼠视网膜中的表达量较模型组明显增加（P〈0.01）,但bax表达量明显减少（P〈0.01）。模型组和治疗组的bcl-2在鼠视网膜中的表达量较正常组增加但bax表达量明显减少（P〈0.05）。结论0.2%酒石酸溴莫尼定对大鼠视神经夹挫伤有一定的治疗作用,其机制与抑制凋亡有关。     

DNA-fragmentation and apoptosis-related proteins of muscle cells in motor neuron disorders

Apoptosis has been decribed as one of the mechanisms of muscle fiber loss in infantile spinal muscular atrophy. In order to investigate if muscle fiber-apoptosis plays a role in other denervating disorders as well, we studied DNA-fragmentation, a hallmark of apoptosis, by the TUNEL-method and, moreover, the expression patterns of apoptosis-related proteins in 2 patients suffering from ALS and in 6 patients with polyneuropathy. We identified DNA-cleavage in muscle fibers of all these patients. Furthermore, we found strong expression of bax and ICE promoting apoptosis in muscle fibers. However, also strong expression of the anti-apoptotic factor bcl-2 was found. Our findings indicate that defective innervation may prompt muscle fibers to activate an intrinsic "suicide" programme which is promoted by the pro-apoptotic factors bax and ICE, which seems to induce formation of apoptotic bodies by cleavage of actin. Nevertheless, there are also anti-apoptotic strategies in muscle fibers manifested by expression of the bax-antagonist bcl-2 which is able to neutralize high bax levels.     

ISCHEMIC PRECONDITIONING RELIEVES ISCHEMIA/REPERFUSION INJURY OF HIPPOCAMPUS NEURONS IN RAT BY INHIBITING p53 AND BAX EXPRESSIONSA

Objective To examine whether ischemic preconditioning (IPC) can protect neuron against delayed death in CA1 subfield of hippocampus following reperfusion of a lethal ischemia in rats, and explore the role of p53 and bax in this process.Methods We examined the effect of IPC on delayed neuron death, neuron apoptosis, expressions of p53 and bax gene in the CA1 area of hippocampus in the rats using HE staining, flow cytometry, RT-PCR, and immunohistochemis-try technique.Results IPC enhanced the quantity of survival cells in the CA1 region of hippocampus (216 ±9 cells/0. 72 mm2 vs. 30 ±5 cells/0. 72 mm2, P＜0. 01), decreased the percentages of apoptotic neurons of hippocampus caused by is-chemia/reperfusion (2. 06% ±0.21% vs. 4.27% ±0. 08% , P＜0. 01), and weakened the expressions of p53 and bax gene of hippocampus compared with ischemia/reperrusion without IPC.Conclusion IPC can protect the neurons in the CA1 region of hippocampus against apoptosis caused by ischemia/reperfusion, and this process may be related to the reduced expressions of p53 and bax.     

bcl-2、bax在砷中毒大鼠肾近端小管表达

目的 探讨砷中毒对大鼠肾近端小管上皮细胞凋亡调控基因bc l-2、bax影响。方法 清洁级SD大鼠40只,体重为120～150g,高、低剂量染砷组各15只,对照组10只。高、低剂量染砷组分别给予三氧化二砷(AS₂O₃)10、0.4 mg/kg水溶液自由饮用,对照组饮用蒸馏水。分笼喂养4个月,取肾脏标本,采用免疫组织化学二步法、细胞计数和图像分析方法测定bc l-2、bax表达。结果 高、低剂量染砷组肾近端小管上皮bc l-2阳性细胞计数分别为(1.85±1.22)与(5.47±1.62)个,明显低于对照组(8.03±2.42)个,平均灰度值逐渐增高,差异具有统计学意义(P<0.05);高、低剂量染砷组肾近端小管上皮bax阳性细胞数分别为(14.88±3.02)与(6.89±1.86)个,明显高于对照组(2.18±1.52)个,平均灰度值随着染砷剂量的增加而下降,差异具有统计学意义(P<0.05)。结论 bc l-2、bax参与了砷中毒大鼠近端小管上皮细胞凋亡过程。     

切除部分Ranvier区对骺板软骨细胞bcl-2和bax表达的影响

目的:探讨凋亡相关基因bax、bcl-2在R anv ier区部分切除后骺板软骨组织中的表达。方法:切除12mm长的新西兰大耳白兔的胫骨近端R anv ier区,于术后1、4周处死实验兔,从免疫组化方面观察凋亡相关基因bax、bcl-2在R anv ier区部分切除后不同周龄的兔胫骨近端骺板软骨组织中表达的变化特征。结果:随着R anv ier区部分切除后,骨桥形成前骺板软骨组织中的软骨细胞凋亡率明显低于对照组,骨桥形成后软骨细胞凋亡率明显高于对照组。结论:R anv ier区和骨桥对骺板软骨细胞的凋亡率有促进作用,骨桥的促进作用更明显。     

DNA‐fragmentation and expression of apoptosis‐related proteins in experimentally denervated and reinnervated rat facial muscle

Muscle fibres may undergo apoptotic cell death in several neuromuscular disorders such as denervated muscle fibres in spinal muscular atrophies. We investigated DNA‐fragmentation ( in situ by the TUNEL‐method) and expression of apoptosis‐associated proteins in experimentally denervated and reinnervated rat facial muscle up to 24 weeks after surgery to evaluate the rate and time lapse of apoptotic muscle fibre loss. While denervated muscle displayed constantly high rates of DNA‐fragmentation, denervated and immediately reinnervated muscle showed a distinct decrease of primarily elevated DNA‐cleavage, finally resembling rates of normal controls. Denervated muscle fibres revealed strong immunoreactivity of the anti‐apoptotic proteins bcl‐2 and bcl‐xL, and the pro‐apoptotic factor bax. In reinnervated muscle fibres, only bcl‐2 was constantly up‐regulated while bcl‐xL and bax diminished after the 7th week. The present findings indicate that denervation may prompt muscle fibres to activate an intrinsic 'suicide' programme to undergo apoptosis. High levels of bcl‐2 after denervation may sustain cell survival until reinnervation, e.g. after accidental nerve damage or in neurodegenerative disorders. Furthermore, increasing levels of bcl‐2 are able to neutralize high apoptosis‐promoting bax levels. Interventions modifying DNA‐fragmentation and the expression of apoptosis‐related proteins may lead to new therapeutic concepts in denervating disorders of muscle in the absence of other primary therapies.     

Expression of Bcl-2 and bax in the frontoparietal cortex of the rat following cardiac arrest

Following 10-min cardiac arrest and resuscitation, male Sprague-Dawley rats developed posthypoxic myoclonus. This phenomenon peaked at 14 days and disappeared by 45 days after cardiac arrest. The mechanisms for the initial dysfunction and later restoration of motor function are not completely known. In the present study, involvement of Bcl-2 and Bax in these phenomena was investigated. In the frontoparietal cortex, both bcl-2 and bax mRNA levels were significantly increased 1, 3, 7, 14, and 28 days postresuscitation. bax mRNA levels continued to be high 45 days postcardiac arrest, whereas bcl-2 mRNA levels were returned to control levels. The apoptotic cells were found in layers IV to VI of the frontoparietal cortex of rats 3 days postcardiac arrest. These results indicate that after cardiac arrest, the initial rise of Bax levels may mediate apoptosis and neurodegeneration in the rat brain. At later time points, increased levels of Bcl-2 may contribute to recovery of motor function in posthypoxic rats.     

低温保存不同时间大鼠带瓣管道活性与bax、bcl-2基因表达的关系

目的 观察低温保存不同时间大鼠带瓣血管活性及bax和bcl-2基因的表达,探讨基因检测是否能成为带瓣血管活性指标.方法 Wistar大鼠80只,体质量250～350 g,分为新鲜组A组,-196 ℃液氮冻存3、6、9个月为B、C、D 3组.检测细胞结构、糖代谢及bax、bcl-2基因的变化.结果 大鼠带瓣血管的各组葡萄糖代谢率测定,A组(4.365±0.784)与B(4.383±0.548)、C组(4.446±0.608)、D组(4.090±0.657)差异无统计学意义(P＞0.05);bax基因的表达分别为20%、45%、60%和85%,差异有统计学意义(P＜0.05);bcl-2基因表达分别25%、30%、20%和35%,差异无统计学意义(P＞0.05).结论 经冻存后的大鼠带瓣血管活性良好,提示bax基因可作为检测带瓣血管活性的指标,bcl-2的表达或高表达可能是细胞能耐受冻存打击的原因之一.

Abstract：

Objective To study the expression of bax and bcl-2 genes and activity of valved conduit in rats by cryopreservation for different lengths, and to probe whether genetic detection will become one of the indexes of valved conduit activity. Methods Eighty Wistar rats were divided into four groups: 20cases of fresh blood vessels ( group A ), 20 cases of frozen vessels for 3 months ( group B ), 20 for 6months ( group C), and 20 for 9 months ( group D). The changes in cellular structure, gluoce metabolism and the expression of bax and bcl-2 proteins were observed. Results The glucose metabolism rate of the rate valved conduit in groups A, B, C and D was 4. 365 ± 0. 784, 4. 383 ± 0. 548, 4. 446 ± 0. 608, and 4. 090 ± 0. 657 respectively, with the difference being not significant ( P ＞ 0. 05 ). The positive expression rate of bax protein in groups A, B, C and D was 20%, 45%, 60% and 85% respectively (P＜0. 05).The positive expression rate of bcl-2 in groups A, B, C and D was 25%, 30%, 20% and 35% respectively ( P ＞ 0. 05 ). Conclusion The frozen rat valved vascular showed good activity. bax gene can be regarded as one of the indicators for detecting valved vessel activity. The expression or over-expression of bcl-2gene may be one of the reasons why the cells can tolerate the frozen preservation.     

量子点荧光技术标记口腔鳞癌细胞中bax的应用研究

目的利用量子点(QD)优良的光学性质对人舌癌Tca8113细胞内bax进行特异性荧光标记。方法利用QD545nm通过间接免疫荧光法,在激光共聚焦显微镜下观测人舌癌Tca8113细胞内bax的表达,并激光连续照射1 h,用激光共聚焦显微镜自带软件Leica Confocal Software测量量子点QD545nm的荧光信号强度。结果激光共聚焦显微镜下可见人舌癌Tca8113细胞内bax明显表达,且激光连续照射1 h,QD545nm标记的荧光未见明显衰减,而FITC标记的荧光1 h内已基本淬灭。结论量子点荧光标记技术能对细胞内bax进行标记。