β联蛋白对体外过氧化氢所致人皮肤成纤维细胞衰老表型的影响

目的 观察高表达的β联蛋白对体外过氧化氢（H2O2）所致正常人皮肤成纤维细胞（NHSF）衰老表型的影响。方法 NHSF分别转染pcDNA3.1-β联蛋白或空载体pcDNA3.1,然后150 μmol/L H2O2处理2 h。实验分为转染H2O2处理组（NHSF + β联蛋白 + H2O2）、转空载体H2O2处理组（NHSF + 空载体 + H2O2）以及转空载体组（NHSF + 空载体）。用RT-PCR和Western印迹分析β联蛋白的表达,显微镜观察细胞形态,试剂盒检测衰老相关的β半乳糖苷酶（SA-β-gal）、细胞活性氧（ROS）以及超氧化物歧化酶（SOD）的活性。所有数据均用SPSS 13.0软件进行统计分析,多组间比较采用ANOVA检验。结果 pcDNA3.1-β联蛋白明显上调了NHSF中β联蛋白的表达。NHSF + 空载体 + H2O2组β联蛋白mRNA和蛋白水平（与GAPDH mRNA和蛋白的比值分别为0.2900 ± 0.0195和0.3130 ± 0.0171）明显低于NHSF + 空载体组（分别为0.5963 ± 0.0400和0.6190 ± 0.0090）,两组比较,均P ＜ 0.05;NHSF + β联蛋白 + H2O2组β联蛋白表达水平（0.7953 ± 0.0074）高于NHSF + 空载体 + H2O2组（P ＜ 0.05）。SA-β-gal染色率在NHSF + 空载体组、NHSF + 空载体 + H2O2组、NHSF + β联蛋白 + H2O2组分别为（2.9667 ± 0.2517）%、（37.70 ± 0.9539）%、（29.330 ± 0.6359）%,各组比较,差异有统计学意义（P ＜ 0.05）。NHSF + 空载体、NHSF + 空载体 + H2O2、NHSF + β联蛋白 + H2O2组ROS活性分别为（50.9963 ± 9.2688）%、（109.9190 ± 11.5215）%、（75.1063 ± 3.0138）%,SOD水平分别为（88.0856 ± 3.9181）、（35.5585 ± 3.4438）、（61.7029 ± 3.1716） U/mg,各组比较,差异均有统计学意义（P ＜ 0.05）。结论 高表达的β联蛋白能够抑制细胞衰老相关β-半乳糖甘酶（SA-β-gal）和细胞活性氧（ROS）的表达,同时上调了SOD的活性。 【关键词】 β连环素; 过氧化氢; 成纤维细胞; 氧化性应激; 细胞衰老     

没食子酸对H2O2诱导的SH-SY5Y神经细胞损伤保护作用的研究

目的：以H₂O₂诱导SH-SY5Y细胞氧化损伤为模型,探讨没食子酸对H₂O₂诱导SH-SY5Y细胞氧化损伤的影响及其机制。方法：筛选没食子酸(gallic acid,GA)对SH-SY5Y细胞无毒剂量范围,以此剂量范围研究没食子酸对H₂O₂诱导SH-SY5Y细胞氧化损伤的保护作用。将细胞分为5组:正常对照组(Control),H₂O₂模型组(Model),GA 5µmol•L^-1组(GA1)、GA 10µmol•L^-1组(GA2) 和GA25µmol•L^-1组(GA3)。GA组先加入GA预处理1 h后再加入终浓度为200 µmol•L^-1 H₂O₂。Hoechst 33258 染色观察凋亡细胞的形态学特征;碘化丙啶染色流式细胞仪检测细胞周期构成比;diaphorase催化的INT显色反应检测乳酸脱氢酶(lactate dehydrogenase, LDH)释放量;DCFH-DA检测细胞内活性氧(reactive oxygen species, ROS);利用天冬氨酸蛋白水解酶-3(caspase-3)可以催化底物Ac-DEVD-pNA的反应检测caspase-3活性。结果：过氧化氢组与正常对照组相比,终浓度为200 μmol•L^-1 H₂O₂作用于细胞24 h后,细胞活力下降为65%,差异具有统计学意义(P<0.01),细胞凋亡率上升(P<0.01);明显出现了细胞凋亡的形态学特征;增殖指数(proliferation index, PI)降低(P<0.05); LDH释放量和细胞内ROS增加(P<0.01),caspase-3活性增强(P<0.01)。与H₂O₂损伤组相比,终浓度为5～25 µmol•L^-1 GA能显著改善上述指标的变化(P<0.05)。结论：GA在一定剂量范围内对H₂O₂诱导的 SH-SYSY神经细胞损伤具有明显的保护作用,其保护效应可能与清除ROS,减轻DNA氧化损伤,抑制线粒体通路介导的细胞凋亡有关。     

Inhalation of hydrogen gas reduces liver injury during major hepatotectomy in swine

AIM:To study the effect of H2 gas on liver injury in massive hepatectomy using the Intermittent Pringle maneuver in swine. METHODS:Male Bama pigs (n = 14) treated with ketamine hydrochloride and Sumianxin Ⅱ as induction drugs followed by inhalation anesthesia with 2% isoflurane, underwent 70% hepatotectomy with loss of bleeding less than 50 mL, and with hepatic pedicle occlusion for 20 min, were divided into two groups:Hydrogen-group (n = 7), the pigs with inhalation of 2% hydrogen by the tracheal intubation during major hepatotectomy; Contrast-group (n = 7), underwent 70% hepatotectomy without inhalation of hydrogen. Hemo-dynamic changes and plasma concentrations of alanine aminotransferase (ALT), aspartate aminotransferase (AST), hyaluronic acid (HA), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and malondialdehyde (MDA) in liver tissue were measured at pre-operation, post-hepatotectomy (PH) 1 h and 3 h. The apoptosis and proliferating cell nuclear antigen (PCNA) expression in liver remnant were evaluated at PH 3 h. Thenwe compared the two groups by these marks to evaluate the effect of the hydrogen in the liver injury during major hepatotectomy with the Pringle Maneuver in the swine. RESULTS: There were no significant differences in body weight, blood loss and removal liver weight between the two groups. There was no significant difference in changes of portal vein pressure between two groups at pre-operation, PH 30 min, but in hydrogen gas treated-group it slightly decrease and lower than its in Contrast-group at PH 3 h, although there were no significant difference (P = 0.655). ALT and AST in Hydrogen-group was significantly lower comparing to Contrast-group (P = 0.036, P = 0.011, vs P = 0.032, P = 0.013) at PH 1 h and 3 h, although the two groups all increased. The MDA level increased between the two group at PH 1 h and 3 h. In the hydrogen gas treated-group, the MDA level was not significantly significant at pre-operation and significantly low at PH 1 h and 3 h comparing to Contrast-group (P = 0.0005, P = 0.0004). In Hydrogen-group, the HA level was also significantly low to Contrast-group (P = 0.0005, P = 0.0005) although the two groups all increased at PH 1 h and 3 h. The expression of cluster of differentiation molecule 31 molecules Hydrogen-group was low to Contrast-group. However, PCNA index (%) was not statistically significant between the two groups (P = 0.802). Microphotometric evaluation of apoptotic index (AI) in terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-stained tissue after hepatotectomy for 3h, the AI% level in the hydrogen was significantly low to Contrast-group (P = 0.012). There were no significant difference between Hydrogen-group and Contrastgroup at pre-operation (P = 0.653, P = 0.423), but after massive hepatotectomy, the TNF-α and IL-6 levels increase, and its in Hydrogen-group was significantly low compared with Contrast-group (P = 0.022, P = 0.013, vs P = 0.016, P = 0.012), respectively. Hydrogen-gas inhalation reduce levels of these markers and relieved morphological liver injury and apoptosis.CONCLUSION: H2 gas attenuates markedly ischemia and portal hyperperfusion injury in pigs with massive hepatotectomy, possibly by the reduction of inflammation and oxidative stress, maybe a potential agent for treatment in clinic.     

Impact of different surface treatment methods on bond strength between fiber post and composite core material

PurposeTo assess the impact of different surface treatments on the push-out bond strength between fiber post and a composite resin core material.Material and methodsSeventy-two glass-fiber posts were randomly assigned into six groups according to the method of surface treatment: Control (no treatment), silane, sandblasting, hydrofluoric acid, hydrogen peroxide, and hydrogen peroxide with sandblasting. Two posts from each group were inspected under a scanning electron microscope to assess the surface modifications and 10 posts were employed for the push-out test. Each post was placed vertically in the middle of a cylindrical putty matrix and a dual-cure composite resin material was applied for core build-up. Two discs of each specimen were cut using a low-speed diamond saw (total 120 discs). The push-out test was executed using a universal testing machine at a crosshead speed of 0.5 mm/min. Statistical analysis was performed using one-way ANOVA and Tukey’s test (p ≤ 0.05). The mode of failure of each disc was evaluated under SEM.ResultsThe sandblasting and hydrofluoric acid groups presented significantly higher bond strength than control and hydrogen peroxide groups. The hydrogen peroxide groups exhibited significantly the lowest bond strength of all groups. There was no significant difference between the control and silane groups. All groups showed predominantly adhesive failure except the hydrogen peroxide with sandblasting, where the cohesive failure of the post was predominant.ConclusionsSandblasting and hydrofluoric acid surface treatments demonstrated superior results to silane and hydrogen peroxide. The combined method of hydrogen peroxide and sandblasting could weaken the fiber post and lead to clinical fractures.     

应用口腔粘膜细胞微核试验评价职业接触硫化氢工人的遗传损伤初探

目的　探讨口腔粘膜细胞微核试验在长期接触低浓度硫化氢 (H2 S)工人的遗传损伤监护中的应用。方法　以 2 7名硫化氢作业工人为接触组 ,选择基本条件与接触组均衡而不接触硫化氢的 31名工人为对照组 ,分析其外周血淋巴细胞及口腔粘膜细胞微核率的变化。结果　外周血淋巴细胞及口腔粘膜细胞微核率有增高趋势 ,多元回归统计结果显示 ,接触硫化氢的时间长短和吸烟量与微核率呈正相关 (P <0 0 5 )。相关分析显示 ,对照组和接触组的外周血淋巴细胞微核率与口腔粘膜细胞微核率存在相关关系 (P <0 0 1 )。结论　长期接触低浓度硫化氢可能会对工人造成一定的遗传损伤。在硫化氢接触人群的遗传损伤监护中可用口腔粘膜细胞微核试验替代外周血淋巴细胞微核试验     

过氧化氢与肿瘤发生发展的关系

H2O2与肿瘤的发生发展关系密切。传统观点认为上调肿瘤细胞内H2O2可导致氧胁迫并诱导细胞凋亡,但近年来陆续有研究者发现大量的具有抗氧化活性的药物也能诱导肿瘤细胞凋亡,因此H2O2与肿瘤细胞的关系成为急需探讨的问题。文章综述了近年来关于H2O2与肿瘤发生、发展、凋亡关系的研究进展,为肿瘤的有效化学预防及新的治疗策略提供思路。     

1H MRS在神经肿瘤临床中的应用

1H-MRS是一种无创性检测活体组织能量代谢、生化改变及化合物定量分析的新技术,今年来广泛应用于脑肿瘤的诊断、鉴别诊断、术前分级及预后分析中.本文重点总结<'1>H-MRS的临床应用现状及分析方法,并简要对其发展进行综述.     

二烯丙基硫醚拮抗微囊藻毒素-LR致细胞凋亡

目的 探讨二烯丙基硫醚( DAS)对微囊藻毒素-LR (MCLR)诱导的细胞凋亡及活性氧产生的影响.方法 体外培养人肝癌细胞株HT17,用1μmol/L MCLR染毒的同时给予DAS(0.02～2 mmol/L)作用4、24h,同时设立溶剂对照组、DAS对照组和单纯MCLR染毒组.应用四甲基偶氮噻唑蓝法检测细胞存活率,流式细胞术检测细胞凋亡,采用荧光探针2-[6-(4'-羟基)苯氧基-3H-占吨醇-3-on-9-yl] -苯甲酸检测细胞内活性氧水平.结果 0.2和2mmol/L DAS干预组细胞存活率分别为(80.91±5.25)％,(90.13±4.47)％,高于MCLR染毒组的(69.50±3.03)％,差异有统计学意义(P＜0.05);而0.2和2 mmol/L DAS干预组细胞凋亡率分别为(10.87±0.52)％,(8.52±0.55)％,低于MCLR染毒组的(13.29±1.16)％,差异有统计学意义(P＜0.05);0.2和2 mmol/L DAS干预组的活性氧水平明显下降(P＜0.05).结论 DAS可拮抗MCLR诱导的活性氧产生及细胞凋亡.     

海康灵各组分抗神经细胞损伤有效部位和有效剂量的研究

目的:通过比较海康灵复方中单味药对过氧化氢(H2O2)诱导SH-SY5Y神经细胞损伤的保护作用,筛选更为理想的量效关系。方法:采用体外细胞培养的方法,复制神经母细胞瘤(SH-SY5Y)细胞H2O2损伤模型。通过MTT法测定细胞存活率,检测海康灵复方中单味药与复方对SH-SY5Y细胞损伤的保护作用。结果:根据最佳有效剂量组建的海康灵新复方(浓度为0.159g.mL-1)同单味药与原复方比较,能减轻H2O2引起的SH-SY5Y细胞的损伤,明显提高细胞的存活率。结论:海康灵新复方对H2O2诱导的SH-SY5Y细胞损伤具有明显的保护作用。     

Hydrogen inhalation ameliorates lipopolysaccharide-induced acute lung injury in mice

Acute lung injury (ALI) is a serious illness, the incidence and mortality of which are very high. Free radicals, such as hydroxyl radicals (OH) and peroxynitrite (ONOO⁻), are considered to be the final causative molecules in the pathogenesis of ALI. Hydrogen, a new antioxidant, can selectively reduce OH and ONOO⁻. In the present study, we investigated the hypothesis that hydrogen inhalation could ameliorate ALI induced by intra-tracheal lipopolysaccharide (LPS, 5 mg/kg body weight). Mice were randomized into three groups: sham group (physiological saline + 2% hydrogen mixed gas), control group (LPS + normal air) and experiment group (LPS + 2% hydrogen mixed gas). Bronchoalveolar lavage fluid (BALF) was performed to determine the total protein concentrations and pro-inflammatory cytokines. Lung tissues were assayed for oxidative stress variables, wet/dry (W/D) ratio, histological, immunohistochemistry and Western blotting examinations. Our experiments exhibited that hydrogen improved the survival rate of mice and induced a decrease in lung W/D ratio. In addition, hydrogen decreased malonaldehyde and nitrotyrosine content, inhibited myeloperoxidase and maintained superoxide dismutase activity in lung tissues and associated with a decrease in the expression of TNF-α, IL-1β, IL-6 and total protein concentrations in the BALF. Hydrogen further attenuated histopathological alterations and mitigated lung cell apoptosis. Importantly, hydrogen inhibited the activation of P-JNK, and also reversed changes in Bax, Bcl-xl and caspase-3. In conclusion, our data demonstrated that hydrogen inhalation ameliorated LPS-induced ALI and it may be exerting its protective role by preventing the activation of ROS–JNK–caspase-3 pathway.