黄芪甲苷孵育脂肪源性干细胞对顺铂诱导肾小管上皮细胞凋亡的影响

背景：干细胞移植用于治疗急性肾损伤的有效性已经被多个研究证实,但其对肾小管上皮细胞损伤的修复机制尚不明确。 目的：观察黄芪甲苷孵育后的脂肪源性干细胞对顺铂诱导的肾小管上皮细胞凋亡的保护作用及机制。 方法：实验分为4组。2.5 μmol/L顺铂诱导肾小管上皮细胞 24 h,建立肾小管细胞损伤模型(顺铂损伤组);将脂肪源性干细胞与损伤肾小管上皮细胞共培养(脂肪源性干细胞+损伤肾小管上皮细胞组);利用Transwell小室将20 mg/L黄芪甲苷孵育脂肪源性干细胞48 h后与损伤肾小管上皮细胞共培养(黄芪甲苷孵育脂肪源性干细胞+损伤肾小管上皮细胞组);以正常肾小管上皮细胞做对照(正常对照组)。 结果与结论：与肾小管上皮细胞损伤组相比,AV/PI和TUNEL结果均显示脂肪源性干细胞+肾小管上皮细胞组和20 mg/L 黄芪甲苷脂肪源性干细胞+肾小管上皮细胞组肾小管上皮细胞发生凋亡的比例和数量明显减少;ELISA结果表明20 mg/L黄芪甲苷脂肪源性干细胞+肾小管上皮细胞组胰岛素样生长因子1分泌显著提高(P < 0.05);Western blot进一步显示20 mg/L 黄芪甲苷脂肪源性干细胞+肾小管上皮细胞组caspase-3蛋白水平明显下降,而Bcl-2的表达量明显增加(P < 0.05)。表明黄芪甲苷孵育的人脂肪源性干细胞对顺铂诱导的肾小管上皮细胞凋亡具有抑制作用,从而有利于肾小管损伤的早期恢复,其保护机制可能与增加胰岛素样生长因子1分泌,抑制caspase-3表达、上调Bcl-2水平有关。中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程全文链接：     

硫酸镁预防瑞芬太尼术后痛觉过敏

目的观察术中静脉注射硫酸镁预防瑞芬太尼麻醉患者术后痛觉过敏的效果。方法择期行腹腔镜胆囊切除术患者60例,随机分为硫酸镁治疗组（M组,n=30）和对照组（C组,n=30）。常规诱导插管后,以0.4μg·kg-1.min-1速率维持瑞芬太尼输注,调节七氟醚吸入浓度维持BIS至40~60之间。M组麻醉诱导后15 min内静脉注射硫酸镁50 mg/kg后以15 mg·kg-1.h-1速率维持输注至手术结束;C组输注等质量生理盐水。记录术后30 min、1 h、2 h、4 h、8 h、12 h、24 h VAS评分,测定患者术前1 d及术后2 h、24 h静脉血Mg2＋浓度;手术前1 d及术后24 h进行2 min步行测验（2MWT）。观察并记录患者恶心呕吐、镇静、低血压等不良反应发生率。结果与C组比较,M组患者术后2 h及以内VAS评分有统计学差异（P〈0.05）,术后4~24 hVAS评分差异无统计学意义（P〉0.05）。术后2 h,M组血浆中Mg2＋浓度较C组显著增加（P〈0.05）,术后24 h恢复正常。M组患者术后24 h 2MWT距离恢复较C组快（P〈0.05）,M组患者恶心呕吐发生率较C组显著降低（P〈0.05）。结论术中静脉注射硫酸镁可有效预防瑞芬太尼术后痛觉过敏,促进患者术后功能恢复且不增加不良反应。     

Ligation of CD8alpha on human natural killer cells prevents activation-induced apoptosis and enhances cytolytic activity

It has been previously shown that the subset of human natural killer (NK) cells which express CD8 in a homodimeric alpha/alpha form are more cytotoxic than their CD8- counterparts but the mechanisms behind this differential cytolytic activity remained unknown. Target cell lysis by CD8- NK cells is associated with high levels of effector cell apoptosis, which is in contrast to the significantly lower levels found in the CD8alpha+ cells after lysis of the same targets. We report that cross-linking of the CD8alpha chains on NK cells induces rapid rises in intracellular Ca2+ and increased expression of CD69 at the cell surface by initiating the influx of extracellular Ca2+ ions. We demonstrate that secretion of cytolytic enzymes initiates NK-cell apoptosis from which CD8alpha+ NK cells are protected by an influx of exogenous calcium following ligation of CD8 on the NK-cell surface. This ligation is through interaction with fellow NK cells in the cell conjugate and can occur when the target cells lack major histocompatibility complex (MHC) Class I expression. Protection from apoptosis is blocked by preincubation of the NK cells with anti-MHC Class I antibody. Thus, in contrast to the CD8- subset, CD8alpha+ NK cells are capable of sequential lysis of multiple target cells.     

胰岛素对滋养层细胞凋亡的调节作用及机制

目的 :探讨胰岛素对培养的滋养细胞凋亡的影响及可能的机制。方法 :将培养的妊娠早期滋养层细胞 ,分为正常对照组(细胞 +培养液 )、H2 O2 组 (细胞 +培养液 +H2 O2 )和胰岛素 +H2 O2 组 (细胞 +H2 O2 +胰岛素 )。采用透射电镜观察及流式细胞术 ,观察H2 O2 诱导的细胞凋亡及胰岛素对H2 O2 诱导的细胞凋亡的抑制作用。并检测胰岛素对滋养细胞caspase 3的活性及Bcl 2蛋白表达的影响。结果 :H2 O2 可诱导培养的滋养细胞凋亡 ,透射电镜下可见特征性的细胞核改变。胰岛素可显著抑制H2 O2 诱导的细胞凋亡 ,流式细胞仪检测其凋亡率较H2 O2 组显著下降 (P <0 .0 1)。H2 O2 组滋养细胞中caspase 3的活性较对照组显著增高 (P <0 .0 1) ,而Bcl 2蛋白的表达则较对照组显著下降 (P <0 .0 1)。结论 :胰岛素可明显抑制H2 O2诱导的滋养细胞凋亡 ,其机制可能与降低caspase 3的活性和促进Bcl 2蛋白的表达有关     

己糖激酶2通过抑制线粒体凋亡通路减少LPS引起的人肺上皮BEAS-2B细胞凋亡

目的:探讨脂多糖(LPS)诱导人正常肺上皮BEAS-2B细胞凋亡的分子机制,并对己糖激酶2(HK2)在该效应中的作用进行分析。方法:采用不同浓度的LPS作用于BEAS-2B细胞建立损伤模型,CCK-8实验检测细胞存活率; Hoechst 33342染色及Annexin V/PI双染法分析细胞凋亡水平;通过使用线粒体凋亡通路抑制剂或者外在凋亡通路抑制剂鉴定细胞凋亡通路;在BEAS-2B细胞中转染HK2过表达质粒以验证HK2对上述效应的影响。Western blot法确认HK2过表达效果;免疫荧光实验检测HK2亚细胞定位。结果:CCK-8实验结果显示,LPS以时间和剂量依赖性方式降低BEAS-2B细胞活力; Hoechst 33342染色结果表明,给予LPS处理后的BEAS-2B细胞核出现固缩和碎裂;同时Annexin V/PI双染实验结果表明,处于凋亡状态的细胞由2. 89%增加至42. 4%,细胞凋亡率明显升高(P 0. 05)。线粒体凋亡通路执行蛋白caspase-9特异性抑制剂可显著抑制细胞凋亡,而caspase-8抑制剂却无此效应。在BEAS-2B细胞的凋亡过程中伴随着HK2的表达下调,而HK2过表达可以有效阻止以上事件的发生。结论:己糖激酶2可通过抑制线粒体凋亡通路减少LPS引起的人肺上皮细胞凋亡。     

Effects of acrylonitrile on apoptosis of rat cerebral nerve cells

The brain, especially the hippocampus, is sensitive to damage caused by anoxic chemicals. In this study, we established a rat model of acrylonitrile poisoning with administration by gavage, aiming to determine the influence of acrylonitrile on rat cerebral nerve cells. Transmission electron microscopy observation and TdT-mediated dUTP nick-end labelling (TUNEL) staining were used to explore preliminarily the apoptotic changes of cerebral nerve cells. The pathogenesis revealed by transmission electron microscopy indicated that apoptosis in the control group was more serious than that of the exposure groups. The results of TUNEL staining showed the apoptotic rate was significantly higher in the control group than that of other exposure groups. All the results indicated that acrylonitrile can inhibit the apoptosis of rat cerebral nerve cells, which is closely related to its animal carcinogenicity.     

Establishment and chemical transformation of human skin epithelial cells in vitro

Summary This paper describes the in vitro establishment and chemical treatment of human foreskin epithelial cells which transform the cells to an anchorage-independent state as demonstrated by growth in soft agar. The procedures described include (a) production of primary cultures of human epithelial cells, (b) cytotoxicity determination of putative chemical carcinogens, (c) chemical transformation protocol, and (d) evaluation of chemical transformation as indicated by anchorage-independent growth.     

硫酸镁联合低分子肝素治疗早发型子痫前期的病例对照研究

目的探讨硫酸镁联合低分子肝素治疗早发型子痫前期的疗效及其妊娠结局。方法将50例早发型子痫前期患者分为硫酸镁治疗组（n=22）和硫酸镁联合低分子肝素治疗组（n=28）,并进行病例对照分析,观察治疗前后的凝血功能、肾功能变化及妊娠结局。结果两组患者治疗后收缩压及舒张压均明显下降,但下降幅度差异无统计学意义（P〉0.05）。排除治疗前凝血功能或肾功能基础值的影响,硫酸镁联合低分子肝素治疗使凝血酶原时间（PT）明显延长、凝血酶原时间国际标准化比值（PT-INR）明显增大、凝血酶时间（TT）明显延长,差异有统计学意义（P〈0.05）,但上述各指标改变后仍在正常范围以内。硫酸镁联合低分子肝素治疗对尿蛋白及血尿酸的减少影响更显著,差异有统计学意义（P〈0.05）。两个治疗组之间的血清尿素氮和血清肌酐的变化差异无统计学意义（P〉0.05）。两组之间的分娩孕周、期待治疗时间、新生儿体重及新生儿出生Apgar评分及母儿并发症发生率差异均无统计学意义（P〉0.05）。结论在传统治疗的基础上加用低分子肝素辅助治疗早发型子痫前期是安全的,且可以对肾功能起到保护作用,明显减少尿蛋白量。尚未观察到低分子肝素对妊娠结局的影响。     

硫酸右旋糖苷对人胃癌MKN1细胞的形态学影响

目的探讨硫酸右旋糖苷（DS）对人胃癌MKN1细胞株的形态学影响。方法用位相差显微镜、荧光免疫细胞染色法和共聚焦显微镜观察固定细胞和活细胞在培养盘上的附着过程及形态变化。结果IKN1细胞通过变形,形成伪足黏附到培养盘,并与其他细胞接触形成单细胞层。DS抑制了MKN1细胞的黏附,培养48h后仍有部分细胞处于游离状态。结论DS阻止人胃癌细胞株MKN1的黏附过程。     

Replication of measles virus in cultured human thymic epithelial cells

Measles virus can replicate in cultures of both infantile and fetal human thymic epithelial cells. Virus-induced cytopathology including syncytium formation was first evident around 24 hr after viral inoculation of these cultures. At the same time, the cultures began to lose their characteristic thymus-like organizational structure. Viral antigens were detected in infected cells by indirect immunofluorescence, and the presence of progeny virions was demonstrated in culture fluids.     

Nerve growth factor prevents apoptosis of cord blood-derived human cultured mast cells synergistically with stem cell factor

BACKGROUND: Stem cell factor (SCF) has been identified as a critical survival factor of human mast cells. Other cytokines which possess survival promotion activity on human mast cells are less known. OBJECTIVE: We examined the survival promotion activity of nerve growth factor (NGF) on cord blood-derived human cultured mast cells. METHODS: Expression and function of NGF receptors on the mast cells were examined by RT PCR, flowcytometric analysis, immunoprecipitaion and western blotting. The survival promotion activity of NGF to the mast cells was examined. To evaluate the proliferating activity of NGF on the human cultured mast cells, flow cytometric analysis with propidium iodide staining was applied. To confirm whether the human mast cell growth activity of NGF was caused by a suppression of apoptosis, the proportion of the cells containing in situ DNA fragmentation was counted. RESULTS: The human cultured mast cells expressed the high affinity receptor p140trk but not the low affinity receptor p75LNGFR. NGF induced the phosphorylation of p140trk. NGF alone could not support the survival of the mast cells, however, the addition of NGF to the culture medium containing recombinant SCF led to a significant increase of the number of survival mast cells. No significant changes of the cell cycle from G0/G1 phase to the S/G2 + M phases were observed by NGF. In contrast, the addition of NGF to the medium with SCF showed a significant inhibitory effect on the apoptosis of the mast cells. CONCLUSION: NGF may act as a key factor to promote the survival of human mast cells synergistically with SCF through the prevention of apoptosis.     

幽门螺杆菌对体外培养的胃上皮细胞增殖与凋亡的影响

目的　研究H .pylori对体外培养的胃上皮细胞增殖与凋亡的影响。方法　以SGC 790 1细胞作为H .pylori感染的体外细胞模型 ,用Ki 6 7抗原的免疫组化分析检测了H .pylori标准菌株NCTC 116 37活菌对胃上皮细胞增殖的影响 ,同时用流式细胞术、荧光染色技术检测了细胞凋亡率。结果　H .pylori在较低浓度 (≤ 1.6× 10 5CFU/ml)时对细胞增殖有促进作用 ,而在较高浓度 (≥ 8× 10 5CFU/ml)时抑制细胞增殖。H .pylori以浓度依赖方式诱导胃上皮细胞凋亡 ,Hoechst 332 5 8荧光染色和流式细胞术两种方法所得结果一致。结论　细胞凋亡与增殖间的不平衡亦可部分解释人体感染H .pylori后所表现的多样化结局     

Adherence of Helicobacter pylori to cultured human gastric epithelial cells.

Experiments were performed to demonstrate that adherence of Helicobacter pylori to gastric epithelial cells causes alterations in the cell cytoskeleton. H. pylori intimately attached to cultured human gastric epithelial cells on small cellular projections, while there was no intimate association of H. pylori with cultured human esophageal epithelial cells. Fluorescein-conjugated phalloidin staining of gastric epithelial cells showed that H. pylori adherence stimulated actin polymerization; this stimulation was not observed with esophageal cells. Also, this organism's selectivity for gastric mucosa was supported by rare binding of bacteria to esophageal epithelial cells and gastric fibroblasts.     

Aspergillus fumigatus conidia inhibit tumour necrosis factor- or staurosporine-induced apoptosis in epithelial cells

A major innate immune response to inhaled conidia of the opportunistic pathogen Aspergillus fumigatus (Af) is the synthesis of pro-inflammatory cytokines, which include tumour necrosis factor (TNF)-alpha, a known inducer of apoptosis. Modulation of host cell apoptosis has been reported to be one of the mechanisms whereby pathogens overcome host cell defences. Our study was designed to investigate whether or not Af conidia could modulate apoptosis induced by TNF-alpha or staurosporine (STS). Exposure of epithelial cells treated by these inducers and exposed to Af conidia decreased the number of apoptotic cells detected by Annexin V staining, analysis of nuclear morphology, terminal deoxynucleotidyl transferase-mediated fluorescein-dUTP nick end-labelling reaction and immunoblotting. Inhibition of apoptosis by Af conidia was seen in cells of the A549 pneumocyte II line, human tracheal epithelial 16HBE and primary human respiratory cells. Inhibition of apoptosis by Af conidia was also observed when apoptosis was induced by co-cultivating A549 cells with activated human alveolar macrophages. Unlike Af conidia, conidia of Cladosporium cladosporioides as well as latex beads or killed Af conidia have no inhibitory effect on TNF-alpha or STS-induced apoptosis. For TNF-induced apoptosis, the observed anti-apoptotic effect of Af conidia was found to be associated with a significant reduction of caspase-3.     

C-反应蛋白诱导人肾小管上皮细胞凋亡

目的:探讨与微炎症状态相应的C-反应蛋白(CRP)水平是否诱导肾小管上皮细胞凋亡。方法:以微炎症状态相应的CRP浓度刺激HK-2细胞。采用AnnexinⅤ-FITC、PI染色和流式细胞术检测凋亡细胞的百分率。采用Hoechst 33258染色观察肾小管上皮细胞凋亡的形态学改变。比色法检测细胞caspase-3活性。Real-time PCR检测促凋亡基因bax、抗凋亡基因bcl-2的mRNA表达。结果:CRP呈剂量和时间依赖性地诱导HK-2细胞凋亡,细胞凋亡在CRP浓度为10 mg/L时达高峰,在20 mg/L时则以晚期凋亡和坏死为主。Hoechst 33258细胞核染色显示CRP作用的HK-2细胞呈现染色质浓缩、碎裂或染色质边集等细胞凋亡的特点。CRP增高细胞caspase-3的酶活性、上调促凋亡基因bax的表达和下调抗凋亡基因bcl-2的表达。结论:CRP轻度增高可诱导肾小管上皮细胞凋亡。     

体外培养人羊膜上皮细胞的表型及分化能力

背景：人羊膜上皮细胞具有多系分化能力,是再生医学中重要的细胞来源。目前的研究多集中于对其分化能力的考察,而体外培养过程中羊膜上皮细胞的生物学特征如何变化尚不清楚。 目的：分析体外培养对人羊膜上皮细胞生长、表型及向心肌样细胞分化的能力等生物学特性的影响,探讨原代人羊膜上皮细胞干性标志物SSEA-4的表达水平与人羊膜上皮细胞生物学特性变化之间的关联性。 方法：使用统一分离方法获得原代羊膜上皮细胞并进行体外培养。利用CCK-8、流式细胞仪及real-time PCR等手段检测不同培养阶段人羊膜上皮细胞的增殖、表型以及向心肌样细胞分化的能力。 结果与结论：不同胎儿样本来源的原代人羊膜上皮细胞的SSEA-4表达在26.7%-97%,存在很大的个体差异。并且,随着传代次数的增加,人羊膜上皮细胞的SSEA-4表达水平显著降低,其下降程度与原代SSEA-4的表达水平无关。另外,培养后人羊膜上皮细胞的心肌分化潜能也存在很大个体差异,且其差异与原代人羊膜上皮细胞的SSEA-4表达水平的高低无关。结果提示,不同胎儿样本来源的原代人羊膜上皮细胞的SSEA-4表达水平受到个体差异的影响,需要建立更准确的临床样本筛选指标来稳定获得原代高表达SSEA-4的胎儿样本,以实现对人羊膜上皮细胞的质量监控。另外,体外培养过程中SSEA-4的表达水平受到培养条件的影响,需要继续优化培养条件以维持其高表达。此外,人羊膜上皮细胞向心肌样细胞分化的能力受到样本个体差异以及培养条件的影响,在今后还需要进一步研究。 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程全文链接：     

Accumulation of heparan sulfate in the culture of human melanoma cells with different metastatic ability

Glycosaminoglycans were metabolically labeled in subconfluent cultures of highly metastatic 7Gp122 and poorly metastatic IC8 variants and of the low metastatic parental M4Be human melanoma cell line. Proteoglycans were separated by DEAE Trisacryl chromatography from the culture medium, from the heparin extract of the cell layer and from the heparin-extracted cell residue lyzed with detergents. Glycosaminoglycans were released from the proteoglycans by reductive alkaline hydrolysis and heparan sulfate (HS) was detected by deaminative cleavage with nitrous acid. Expressed on cell protein basis, the labeled HS content in the medium and in the cell layer decreased with increasing metastatic ability. The extraction of HS with heparin from the 7Gp122 cells indicated that this variant was enriched in (polypeptide bound) HS non inserted into the plasma membrane, compared with the low metastatic IC8 and M4Be cells. The HS fraction in heparin extract and in the heparin-extracted cell residue exhibited molecular mass heterogeneity on gel permeation chromatography and it contained HS fragments. Scission with nitrous acid followed by molecular sieve chromatography of the degradation products indicated that the tetra- and disaccharide repeats separated by the N-sulfated glucosamine residues were present in about equal amounts and constituted 60% of the HS chains in the IC8 and M4Be cells. HS from 7Gp122, IC8 and M4Be cells did not bind antithrombin III with high affinity but it was capable of binding bFGF in in vitro assay.     

下调HIPK2基因表达促进顺铂诱导的肾小管上皮细胞HKC凋亡

目的探讨HIPK2对顺铂诱导的人肾小管上皮细胞(HKC)凋亡的影响。方法构建顺铂诱导的HKC细胞凋亡模型,实时荧光定量PCR和Western blot检测HIPK2的表达;设计合成2条HIPK2 siRNA干扰片段,通过脂质体转染HKC细胞,建立HIPK2干扰的细胞株;实时荧光定量PCR和Western blot分别检测HIPK2 mRNA和蛋白的表达;再用顺铂处理细胞,Annexin V/PI检测细胞凋亡;Western blot检测Bax表达的影响。结果顺铂呈剂量依赖性诱导的HKC细胞凋亡过程中,HIPK2 mRNA和蛋白表达水平均明显下调(P0.05);转染siRNA后可显著降低HIPK2在HKC细胞内mRNA和蛋白的表达(P0.05),且HIPK2表达降低后会促进顺铂诱导的HKC细胞凋亡。结论HIPK2可抑制顺铂诱导的HKC细胞凋亡。     

参附注射液和硫酸镁佐治老年人慢性充血性心力衰竭的临床分析

目的 观察参附注射液和硫酸镁佐治老年人慢性充血性心力衰竭的临床疗效。方法 慢性充血性心力衰竭老年患者63例，随机分为对照组31例，治疗组32例。对照组常规给予降压药、强心药和利尿药治疗，治疗组在此基础上静脉滴注参附注射液和硫酸镁注射液，14d为1疗程，比较两组的疗效。结果 对照组总有效率为71％，治疗组为97％，疗效相差显著（P〈0．05）；治疗组的心率、左心室射血分数较对照组改善明显（P〈0．05）。结论 参附注射液和硫酸镁佐治老年人慢性充血性心力衰竭疗效较好，能较好地改善心功能。     

miR-24通过SIRT1调控晶状体上皮细胞凋亡的研究

目的探讨氧化应激情况下miR-24对晶状体细胞凋亡的调控。方法采用实时定量PCR检测40例白内障患者晶状体上皮组织及临近晶状体上皮组织中miR-24的表达水平,并在氧化应激情况下检测miR-24的表达变化。通过miR-24 mimics、miR-24 inhibitor转染晶状体上皮细胞SAR01/04以过表达和敲低miR-24,利用pcDNA3.1-SIRT1转染SAR01/04以过表达SIRT1,FITC/PI流式细胞术检测晶状体细胞凋亡情况,Western blot检测凋亡相关蛋白Bcl-2、Bax表达情况,CCK-8检测细胞活性状态。结果miR-24在白内障晶状体上皮组织中的表达高于临近组织,氧化应激情况下促进miR-24表达增高,差异有统计学意义(P<0.05);氧化应激情况下,敲低miR-24抑制晶状体上皮细胞的凋亡;过表达或敲低miR-24可以分别降低或促进SIRT1的表达;过表达miR-24和过表达SIRT1后抑制了晶状体细胞的凋亡。结论氧化应激促进晶状体上皮细胞miR-24表达上调,miR-24通过下调SIRT1促进晶状体上皮细胞的凋亡,为白内障的靶向治疗提供一定的策略。