TNFα、Leptin-Rb、NF-κB和PPAR-γ在酒精性肝病大鼠肝细胞中的表达

目的 探讨TNFα、NF-κB、Leptin-Rb和PPAR-γ等在大鼠酒精性肝病中与肝组织炎症、坏死和纤维化的关系.方法 胃造瘘法建立长期大鼠酒精性肝病模型,分别在第4、8、12、24和52周用免疫组化、原位杂交和RT-PCR技术检测不同阶段大鼠肝组织中TNFα、NF-κB、PPAR-γ和Leptin-Rb等的表达.结果 TNFα、NF-κB和Leptin-Rb水平在模型组大鼠肝细胞中,第8周表达逐渐增强,第24周起高水平表达一直持续至52周(P<0.01).PPAR-γ在对照组和第4、8周模型组肝细胞中表达最强,第12周表达开始减弱.TNFα和NF-κB在肝细胞中的表达呈明显正相关关系,与PPAR-γ则呈负相关关系(P<0.01).结论 TNFα和NF-κB与肝细胞病变程度有很好的一致性关系,Leptin-Rb在肝细胞的表达与肝纤维化发展相一致,PPAR-γ则刚好相反.     

体外大鼠成骨细胞对破骨细胞形成的影响

目的通过将鼠成骨细胞与破骨细胞直接共培养的实验方法,研究成骨细胞对破骨细胞分化及功能成熟的影响。方法按照M-CSF 30μg/L、RANKL 50μg/L的浓度对鼠骨髓单个核细胞诱导培养6 d,与原代培养3 d的鼠成骨细胞按细胞数量1∶1直接共培养,加入1,25-(OH)2D31×10-8mol/L和PGE21×10-6mol/L,利用形态学观察、TRAP染色、骨吸收陷窝检测等方法对共培养细胞进行鉴定。结果成骨细胞与破骨细胞共培养时,成骨细胞有明显的生长优势;染色后显微镜下可见大量呈单层排列的成骨细胞,偶见TRAP( )的破骨细胞。结论成骨细胞对破骨细胞分化及功能成熟的影响与两者的相对数量有关。本实验中,由于成骨细胞快速生长,当其数量多于破骨细胞时,可能使1,25-(OH)2D3对成骨细胞RANKL表达的上调作用不足,且PGE2对成骨细胞OPG表达的下调作用不足,从而主要表现为对破骨细胞形成及分化的抑制作用。     

快速老化小鼠的学习记忆缺陷及其生化机制

目的用快速老化小鼠(SAMP8)模拟老年痴呆,观察其学习记忆能力,并探讨其学习记忆能力下降的可能机制。方法采用10月龄SAMP8,并用同龄抗快速老化小鼠(SAMR1)作为正常对照。用水迷路检测近记忆以及空间学习记忆能力。用生物化学方法检测大脑皮层线粒体膜电位和ATP酶活力,观察线粒体功能;检测皮层和海马胆碱乙酰基转移酶(ChAT)和乙酰胆碱酯酶(AChE)的活力,观察中枢胆碱能神经功能;检测血清超氧化物歧化酶(SOD)活力和丙二醛(MDA)含量,观察其全身性氧化应激状况。结果与SAMR1相比,SAMP8出现明显的衰老体征,其近记忆和空间学习记忆能力明显减弱,具有衰老和痴呆特征;皮层线粒体膜电位和ATP酶活力显著降低,海马ChAT活力显著下降,AChE活力显著升高,血清SOD活力稍有增高。结论SAMP8可以很好地模拟老年痴呆,其机制包括大脑皮层线粒体功能降低、海马胆碱能神经功能下降以及氧化应激等。     

早期自然流产患者绒毛组织中核转录因子-kappa B的表达

目的 探讨核转录因子-κB(NF-κB)在绒毛组织中的表达及活化与早期自然流产的关系.方法 采用免疫组化方法检测早期自然流产患者和同期妊娠的健康妇女绒毛组织中NF-κB p65的表达;激光共聚焦显微镜(CLSM)检测NF-κB p65的核区与胞浆区比值(FN/FC)以观察其活化情况.结果 早期自然流产患者绒毛合体滋养层细胞NF-κB表达强度低于正常早期妊娠妇女,其差异具有统计学意义(P<0.001);早期自然流产患者绒毛间质细胞NF-κB表达的FN/FC(0.7602)明显高于正常早期妊娠妇女(0.6977)(t=3.511,P<0.001).结论 NF-κB在早期自然流产患者绒毛组织合体滋养细胞中的低表达以及其在间质细胞中的过度活化可能与早期自然流产的发生有关.     

生存素在5-氟尿嘧啶诱导人绒癌细胞株JAR凋亡中的作用

目的探讨生存素(survivin)在化疗药物5-氟尿嘧啶(5-Fu)诱导的人绒癌细胞株(JAR)凋亡中的作用。方法用JAR细胞株进行培养传代,以不同浓度5-Fu作用于JAR细胞,MTT方法观察5-Fu对JAR细胞生长的抑制情况,流式细胞仪检测细胞凋亡率,用逆转录聚合酶链(RT-PCR)和免疫印迹技术(Western-blot)方法检测survivin基因和蛋白的表达。结果①5-Fu对JAR细胞的生长具有抑制作用,并有浓度和时间的依赖性;②5-Fu诱导JAR细胞凋亡,凋亡率随浓度而增加;③5-Fu作用后survivin基因和蛋白表达减少,随浓度增加而减少。结论5-Fu可能是通过降低survivin表达水平,诱导JAR细胞凋亡。     

降钙素基因相关肽与内皮素受体A在兔实验性蛛网膜下腔出血后基底动脉的表达

目的探讨蛛网膜下腔出血(SAH)后脑血管内皮细胞降钙素基因相关肽(CGRP)及内皮素受体A(ETRA)的表达与迟发性脑血管痉挛(DCVS)之间的关系。方法日本大耳白家兔60只,随机分为SAH模型组25只,盐水组25只,穿刺组5只,正常组5只。采用枕大池二次注血法制作SAH模型,灌注处死后取基底动脉制成切片。进行CGRP及ETRA的免疫组化和HE染色,观察CGRP与ETRA的免疫表达和病理结构变化,并用图像分析方法测量血管周长,使用方差分析进行统计学检验。结果基底动脉周长在SAH后均发生明显缩窄,其中在5d时缩小最强烈。CGRP与ETRA免疫阳性标记颗粒在SAH组的基底动脉组织中均有表达,其动态变化规律为CGRP的表达在1h时最明显,在3、5d依次表达减少,5d达到最低点,7d后开始逐渐升高,10d时表达相对比较明显,但未能达到正常水平,CGRP在穿刺组、正常对照组和盐水组也有散在不规则表达。在正常组、穿刺组和盐水组的基底动脉内ETRA有表达;SAH组基底动脉内ETRA在3d组和5d组表现为高表达,以3d组为强,而其中血管内皮细胞表达最强烈,平滑肌也有表达,10d组的表达仍然较强。结论在SAH后脑血管内皮细胞CGRP和ETRA的表达呈现明显的动态改变,但二者的变化趋势并不同步;CGRP和ETRA在DCVS的发生发展中起重要的作用。     

NT4-Ant-Shepherdin[79-87]融合基因重组载体的构建

目的设计构建携带NT4-Ant-Shepherdin[79-87]的cDNA融合基因的重组载体,为针对Survivin的靶向治疗奠定基础。方法应用非对称引物/模板法、PCR技术制备NT4-Ant-Shepherdin[79-87]cDNA片断,连接于pGEM-T-Easy载体,经克隆测序、酶切后与PBV220/NT4质粒连接;转化感受态细胞E.coliDH5α,亚克隆获得NT4-Ant-Shepherdin[79-87]融合基因。结果克隆出Ant-Shepherdin[79-87]基因,经酶切及测序证实结果正确;连接PBV220/NT4,经克隆、酶切,琼脂糖凝胶电泳证实获得321bp的NT4-Ant-Shepherdin[79-87]目的基因片断。结论通过分子生物学技术成功构建了携带NT4-Ant-Shepherdin[79-87]融合基因的重组载体,为进一步研究针对Survivin的靶向抗肿瘤作用奠定了基础。     

Role of candesartan against cerebral ischemia and oxidative damage in normotensive rats

Objective Angiotensin Ⅱ (Ang Ⅱ ) contributes to modulating blood pressure by stimulation of Ang Ⅱ AT1 receptors. We devised a rat transient middle cerebral artery occlusion (MCAO) model to assess whether oxidative damage is decreased after pretreatment with Angiotensin Ⅱ AT1 receptor blocker (ARB). Methods After 2 weeks pretreatment with ARB 0. 5 and 1 mg/kg, the male Wister rats were subjected to 2 h middle cerebral artery occlusion (MCAO). At 24 h, the lumen diameter of middle cerebral artery, the plasma level of 8-hydroxy-2'-deoxyguanosine (8-OHdG), and HIF-1 a levels were recorded and compared. Results After pretrcatment with ARB 0.5 and 1 mg/kg, blood pressure did not significantly change compared with that of controls. In the group of candesartan at 1 mg/(kg· day), the lumen diameter was significantly increased compared to that in control group [(86.0±5.0) μm vs. (69.0± 2.1) μm; P<0. 01, n = 6- 8]. The plasma 8-OHdG levels of ARB pretreatment groups were decreased. In immunohistochemical findings, 8-OHdG- and HIF-1α-containing cells in ARB pretreatment groups were decreased. Conclusion Brain ischemia and oxidative damage can be reversed by AT1 receptor blockade in normotensive rats after transient cerebral artery occlusion.     

The changes of neuronal activity of the zone incerta in a rodent model of Parkinson＇s disease

Objective To study the changes in neuronal activity of the zone incerta (ZI) following the unilateral lesion of the nigrostriatal pathway. Methods Eiectrophysiological recordings of ZI neurons were made in normal rats and in two groups of rats at different time intervals after injection of 6-hydrodopamine (6-OHDA) into the pars compacta of snbstantia nigra by extracellular recording in vivo. Results The results showed a significant increase in the mean firing rate of ZI during the second and fourth weeks after 6-OHDA lesion [-n=32, (3.6±2. 2)Hz, P<0.001; n=35, (9.3±6.6)Hz, P<0. 001, respectively] compared to that of normal rats [n=39, (9.2±5.2)Hz]. However, no significant change was observed between two groups of 6-OHDA-lesioned rats. With regard to firing pattern, 7.7%(3/39) of ZI neurons discharged regularly, 82.1% (32/39) irregularly and 10. 3% (4/39) in bursts in normal rats. During the second week after 6-OHDA lesion, the regular, irregular and bursting firing neurons recorded in the ZI were 9.4% (3/32), 59. 4% (19/32) and 31.3% (10/32), and during fourth week, the regular, irregular and bursting firing neurons were 14.3% (5/35), 57.1% (20/35) and 28. 6% (10/35), respectively. The firing pattern of the neurons in the three different groups did not change significantly. Conclusion These results suggest that the firing rate of ZI neurons in 6-OHDA-lesioned rats is increased significantly, which may contribute to the pathophysiology of Parkinson's disease.     

Effect of ABCA1-V771M polymorphism on plasma lipid levels and its relationship with coronary atherosclerotic heart disease

Objective To explore the risk association of ABCA1-V771M polymorphism with coronary heart disease (CHD) in Hart nationality in Northwest of China. Methods With case-control study, ABCA1-V771M polymorphism was detected in 204 unrelated Hart nationality people in Northwest of China, and all the subjects by coronary angiography were grouped into 106 cases and 98 controls. The genotypes and alleles frequency distribution of ABCA1-V771M polymorphisms were analyzed by PCR-RFLP analysis, and the clinical statistics of serum lipids were compared and its effects of ABCA1-V771M polymorphism on the plasma lipid levels and coronary atherosclerotic heart disease were analyzed. Results The genotypic frequencies of ABCA1-V771M polymorphism matched well under Hardy-Weinberg equilibrium (P>0.05), V and M allelic frequencies were 33.3% and 66.7%. In comparison with VV VM genotype carriers, MM genotypes carriers had much lower plasma levels of HDL-C (P<0. 001) and much higher plasma levels of TG (P<0. 05). M allelic frequency in CHD group was significantly higher than V allelic frequency (P<0. 05). M allele was related with more severity of atherosclerosis in the coronary artery than V allele (P<0.05). However, there was no obvious difference in the incidence of AMI among carriers with three genotypes of ABCA1-V771M polymorphism (P>0.05). Conclusion ABCA1-V771M polymorphism was not only associated with the plasma levels of HDL-C and TG, but also related to the susceptibility and severity of coronary atheroselerotic heart disease. Moreover, M771 allele appeared to be atherogenie among Han population in Northwest of China.