Abstract:目的:研究志贺菌中Ⅱ类整合子的流行现状、与耐药的相关性及其携带的耐药基因.方法:K-B法测定26株志贺菌的药敏情况;PCR-限制片段长度多态性(RFLP)筛查整合子并分类;长片段PCR扩增Ⅱ类整合子可变区并进行序列测定.结果:65.4％(17/26例)的志贺菌中检出整合子,PCR-RFLP结果显示均含有Ⅱ类整合子,其中有12例还同时含有Ⅰ类整合子;17例Ⅱ类整合子保守区阳性的标本,有16例可变区扩增亦阳性,大小约为2 200bp,对其测序结果显示含有编码对磺胺类抗生素耐药的基因dfrA1、对链丝菌素耐药的基因sat1和对氨基糖苷类抗生素耐药的基因aadA1.结论:在国内率先报道了Ⅱ类整合子广泛存在于志贺菌中.Ⅱ类整合子与志贺菌的耐药具有相关性.

Abstract:目的:研究亚溶解剂量补体C5b-9(SC5b-9)复合物刺激大鼠肾小球系膜细胞(GMCs)增生和细胞外基质(ECM)分泌的作用,并探讨NF-κB核转录因子是否介入SC5b-9的促增生效应.方法:体外纯培养大鼠GMCs,并行光镜、电镜及功能鉴定.质控SC5b-9后,将GMCs分为5组,即SC5b-9刺激组、SC5b-9+PDTC组(吡咯啉烷二甲基硫脲)、ATS组、灭活人血清组及完全营养液组.应用RT-PCR检测40 min PCNA和FN mRNA水平;同时应用免疫组化检测18 h GMCs增殖细胞核抗原(PCNA)、平滑肌肌动蛋白(α-SMA)和纤维粘连蛋白(FN)及细胞核内NF-κB p65表达情况.结果:实验40 min,SC5b-9刺激组PCNA和FN mRNA表达上调,PDTC处理后表达则明显下降,其余3组PCNA均呈阴性,另3组FN则均有一定基础表达.SC5b-9刺激组,GMCs免疫组化显示:PCNA、α-SMA、FN、NF-κB p65表达增加,PDTC处理后则4者表达明显下调,其余各组,PCNA、α-SMA、NF-κBp65均呈阴性,FN亦有一定的基础表达.结论:SC5b-9可能通过激活NF-κB促进GMCs增生和ECM分泌.

Abstract:目的:观察脂联素基因mRNA在3T3-L1脂肪前体细胞诱导分化过程中表达水平的变化.方法:体外培养3T3-L1脂肪前体细胞,应用MIX、地塞米松、胰岛素诱导其分化,采用RT-PCR技术检测诱导分化不同时间(0～10天)脂肪细胞中脂联素基因mRNA的表达水平.结果:脂联素基因低表达于3T3-L1脂肪前体细胞中,并随脂肪前体细胞分化成熟,该基因表达水平呈逐渐上调趋势,除在诱导剂作用后第0天与第2天、第1～2天、第3～4天、第6天与第8～10天、第7～10天各时段内差异无显著性(P＞0.05)外,其余各时段之间表达水平差异均有显著性(P＜0.05).结论:在3T3-L1脂肪前体细胞分化过程中脂联素基因表达逐渐上调,可能有利于脂肪细胞的分化成熟.

Abstract:目的:探讨MyoD基因诱导大鼠心脏成纤维细胞分化为成肌细胞的可能性.方法:将MyoD基因重组腺病毒载体转染大鼠心脏成纤维细胞,观察转染后大鼠心脏成纤维细胞形态的变化;用RT-PCR和Western blot方法检测MyoD和肌细胞生成素的表达;免疫组化检测骨骼肌肌球蛋白和结蛋白的表达.结果:MyoD基因转染后的大鼠心脏成纤维细胞形态和排列方式发生明显变化;RT-PCR可检测出转染后细胞表达MyoD;Western blot结果表明转染后细胞表达MyoD和肌细胞生成素;免疫组化检测骨骼肌肌球蛋白和结蛋白表达均为阳性.结论:MyoD基因可诱导体外培养的大鼠心脏成纤维细胞分化为成肌细胞,为进一步研究MyoD对心肌损伤的修复作用奠定了基础.

Abstract:目的:探讨三七皂甙Rg1对心肌细胞缺血/再灌注(I/R)损伤的作用.方法:分离培养SD乳鼠心肌细胞,建立I/R模型.分组:①正常对照组;②I/R组:缺氧2 h,复氧1 h;③Rg1治疗组:在缺氧前24 h予三七皂甙Rg-1;④ST+Rg1治疗组:在缺氧前24h予Rg-1及缺氧前1 h予蛋白激酶C特异性阻断剂Staurosporine(ST).计算心肌细胞锥虫蓝摄取率、测定细胞内游离钙、细胞凋亡率(流式细胞法).结果:锥虫蓝摄取率、细胞内游离钙、细胞凋亡率分别为:正常对照组为(2.3±0.3)％、(117.1±14.3)％、(2.6±0.4)％;I/R组为(21.5±2.0)％、(355.3±31.9)％、(20.1±2.3)％;Rg1治疗组为(9.8±0.9)％,(179.5±11.3),(10.4±0.94)％;ST+Rg1治疗组为(16.2±1.7)％、(294.7±18.5)％、(15.9±2.8)％.3项检测数据I/R组明显高于各组(P＜0.05);Rg1治疗组明显低于I/R组(P＜0.05)和高于正常对照组(P＜0.05);ST+Rg1治疗组明显高于Rg1组(P＜0.05).结论:细胞死亡、细胞凋亡、钙超载参与了心肌I/R损伤;三七总甙Rg1可减少SD乳鼠心肌细胞I/R的损伤;三七总甙Rg1的抗I/R损伤的作用可能通过激活蛋白激酶C途径.

Abstract:目的:研究姜黄素对双侧颈总动脉永久性结扎(2VO)所致血管性痴呆大鼠学习记忆功能减退的作用.方法:以2VO来制备大鼠血管性痴呆模型,将大鼠分为3组(假手术组、溶剂对照组、给药组),一部分于术后2周检测脑组织氧自由基(O2-)和羟自由基(OH),另一部分于术后4周和8周分别进行水迷宫实验,来检测各组大鼠学习记忆能力的差异,通过HE染色和免疫组化来观察细胞数量和形态的差异.结果:姜黄素能明显降低因2VO所造成的氧自由基(O2-)和羟自由基(OH)水平的升高(P＜0.05),改善由2VO所致的学习记忆功能的减退(P＜0.05和P＜0.01).HE染色和免疫组化证实,姜黄素减少了2VO所导致的神经细胞死亡.结论:姜黄素能改善2VO所致的学习记忆减退,其机制与减少自由基的产生以及减少神经细胞死亡有关.

Abstract:目的:探讨超声介导SonoVue微泡破裂法促进外源基因在肝脏中表达的作用,评估其转染效率、安全性.方法:经SD大鼠门静脉置管输入SonoVue微泡建立动物模型,以虫荧光素酶基因为报告基因,观察不同浓度的微泡促进虫荧光素酶基因在肝脏中的表达强度与持续时间,实验结束后采下腔静脉血检测肝肾功能并取肝、肾组织常规HE染色行组织学检查.结果:随着微泡浓度的增加,虫荧光素酶基因表达逐渐增加,在微泡浓度为100％(V/V)时,转染效率最高,持续到21天仍有表达.转染前后的肝肾功能(AST、ALT、ALB、BUN、Cr)无明显变化,HE切片未发现肝肾组织结构的改变.结论:超声介导微泡破裂法能有效地促进外源基因在肝脏中的表达,为肝脏疾病的基因治疗提供了新的途径.

Abstract:目的:探讨不同表型幽门螺杆菌(Hp)对Hp相关性胃炎和胃癌病变及其p53表达的影响.方法:对137例Hp(+)慢性胃炎(CG)、消化性溃疡及胃癌患者和34例Hp(-)患者用Western blot检测Hp细胞毒素相关蛋白(CagA)、空泡毒素(VacA)、尿素酶(Urease)及其亚型.评价Hp(+)CG中慢性炎症(CI)、多形核活动性(PA)、Hp定植密度(DH)的组织学等级.用免疫组化检测p53表达,以H-score方法评价.结果:①Hp 128 ku CagA、116 ku CagA、95 ku VacA、91 ku VacA、30 ku UreA表达在轻度CI组的比例低于中、重度CI组(P均＜0.05),95 ku VacA、91 ku VacA和30 ku UreA表达在无PA组的比例低于有PA组(P均＜0.05).②Hp(+)组P53 Hscore均高于Hp(-)CG组(P=0.002).在Hp+CG中,中重度CI组P53 Hscore均高于轻度组(P=0.025),而在Hp(-)CG组中,差异无显著性.③Hp+胃癌中,未发现任何独立表型与CG和胃癌的53 Hscore有关(P＞0.05).结论:Hp多种毒力因子对胃黏膜CI程度加重、PA的发生和发展起促进作用,Hp感染及DH增加均促进p53表达无关;未见Hp任何独立表型与53表达有关,提示Hp可能通过多种毒力因素及此研究未提及的其他因素的综合作用促进p53过度表达.

Abstract:目的:研究应用白三烯受体拮抗剂(孟鲁司特钠)对哮喘小鼠肺组织中细胞周期蛋白D1(Cyclin D1)的表达影响及对支气管哮喘气道重塑的作用,探讨白三烯受体拮抗剂在哮喘治疗中的作用.方法:将30只BALB/c小鼠随机分为正常对照组、哮喘组、孟鲁司特钠组3组,每组10只;卵蛋白致敏和激发建立哮喘小鼠模型;HE染色观察各组气道炎症发生及气道结构改变情况;免疫组化观察肺组织中Cyclin D1的表达及定位;RT-PCR及Western-blot测定各组小鼠肺组织中Cyclin D1的mRNA和蛋白表达变化.结果:HE染色提示哮喘组与对照组相比出现嗜酸性粒细胞浸润增多、纤毛脱失、平滑肌细胞层增厚等改变,而治疗组上述改变较哮喘组为轻;免疫组化显示Cyclin D1在哮喘小鼠气道平滑肌细胞、内皮细胞、成纤维细胞中皆有表达而在对照组中表达减弱(P＜0.05),而治疗组表达量较哮喘组低(P＜0.05);RT-PCR及Western blot检测发现Cyclin D1在哮喘组表达较对照组为高(P＜0.01),而治疗组表达量低于哮喘组(P＜0.01).结论:哮喘小鼠肺组织中Cyclin D1表达量较正常组为高;白三烯受体拈抗剂能够抑制Cyclin D1表达,减轻气道炎症反应、延缓气道重塑进程.

Abstract:目的:构建以GFP做标志基因的放射敏感基因Egr-1调控单纯疱疹病毒胸苷激酶基因(herps simplex virus thymidine kinase,TK)的腺病毒载体.方法:以PCl-neo-Egr-1-TK(pET)为模板扩增Egr-1,插入到pAdTrack的XbaI和XhoI位点,构建重组质粒pAdTrack/Egr-1;与pET酶切获得TKcDNA相连,构成pAdTrack/Egr-1-TK;用PmeI酶切后与pAdEasy-1混合,应用细菌内同源重组法获得重组表达质粒pAdEgr-1-TK.结果:重组表达质粒pAdEgr-1-TK的酶切鉴定符合预期结果,感染肿瘤细胞可见绿色荧光表达.结论:通过细菌内同源重组成功构建含GFP做标志基因的pAdEgr-1-TK重组表达质粒,为研究Egr-1调控自杀基因及对肿瘤细胞进行灭活打下了基础.

Abstract:目的:观察氯胺酮雾化吸入对哮喘大鼠肺泡灌洗液及血清白细胞介素-13(IL-13)的影响.方法:雄性无特殊病原体(SPF级)SD大鼠40只,随机分成对照组(C组)、哮喘模型组(A组)、0.1 g/L地塞米松组(D组)及25g/L、50 g/L的氯胺酮预处理组(K1和K2组).A组用鸡卵清蛋白(OVA)吸附氢氧化铝佐剂辅以灭活的百日咳杆菌菌苗注射致敏2次,第2次致敏1周后雾化吸入OVA激发,D组,K1组和K2组用同样的方法致敏,只是在每次激发前给于地塞米松或氯胺酮预处理.C组分别在相同时间注射和吸入生理盐水.所有的大鼠在最后1次激发后24 h检测血清和支气管肺泡灌洗液(BALF)中IL-13浓度(ELISA法).结果:无论在血清或BALF中A组的IL-13浓度均显著高于C组(P＜0.05);D组,K1组和K2组的两种IL-13的浓度均显著低于A组(P＜0.01或P＜0.05),各组分别与C组相应的值比较均没有统计学差异(P＞0.05);3组相对应值之间没有显著差异(P＞0.05).结论:两种浓度的氯胺酮雾化吸入均可以降低哮喘大鼠血清及肺泡灌洗液中IL-13的浓度,提示临床应用中不必追求较大剂量.

Abstract:目的:观察精囊腺摘除后SD大鼠的性反应,分析精囊分泌功能对性功能的影响.方法:实验组采用普通的外科手术将雄性大鼠精囊腺摘除,对照组暴露两侧精囊腺后即行缝合.在对雌鼠给予药物催情的情况下,分两组进行交配,观察两组雄鼠爬高潜伏期、爬高总次数、插入潜伏期等性功能相关指标,并进行统计学分析.结果:与对照组相比,SD大鼠在摘除精囊腺后,爬高潜伏期、插入潜伏期延长,2 h内爬高总次数、2 h内插入总次数及有射精活动鼠数减少,两组间的差异性具有极显著性;两组的命中率比较,P＜0.05,其差异具有显著性.结论:精囊腺摘除后,大鼠的性功能显著降低,说明精囊腺对维持大鼠的正常性功能具有重要影响.

Abstract:目的:研究As2O3磁性纳米微球的制备及其联合磁流体热疗(MFH)对宫颈癌Sih细胞株的治疗作用.方法:采用改良的乳化冷冻凝聚法制备As2O3磁性纳米微球,能谱仪、原子荧光光谱仪对其进行表征;高频交变磁场中进行体外升温实验.四甲基偶氮唑蓝(MTT)比色法检测As2O3磁性纳米微球联合MFH对宫颈癌Siha细胞株生长的影响,流式细胞仪(FCM)检测凋亡.结果:所制备As2O3磁性纳米微球近似球形,粒径约为120 nm,含砷量为0.61％,在输出电流I=300A的高频交变磁场中具有升温能力,且能达到肿瘤治疗的有效温度(41℃～46℃);As2O3磁性纳米微球联合MFH能抑制宫颈癌Siha细胞株的生长,促进其凋亡,且均较单纯As2O3液及单纯MFH明显.结论:As2O3磁性纳米微球可同时发挥As2O3的细胞毒性作用和磁感应加热的联合定向治疗作用,效果优于单一治疗,为临床治疗宫颈癌提供新的方法.

Abstract:目的:评价心肌特异性脂肪酸结合蛋白(hFABP)在不停跳冠状动脉旁路移植术(OPCABG)围术期诊断心肌缺血损伤的效率.方法:随机择期OPCABG患者68例(ASAⅡ～Ⅲ),在麻醉诱导前(T0)、术中吻合血管(T1)、手术结束(T2)、术后6 h(T3)、18 h(T4)、36 h(T5)时间位点测定肌钙蛋白Ⅰ(cTnI)、肌酸磷酸激酶同工酶(CK-MB)及血清hFABP浓度,并观察血流动力学的变化.结果:有12例患者(17.6％)在手术中出现ECG的ST段上升＞0.1 mV或肺毛细血管楔压(PCWP)＞18 mmHg的心肌缺血损伤,其血清hFABP、cTnI、CK-MB系列浓度均保持在较高的水平,血清hFABP浓度的峰值比cTnI、CK-MB浓度的峰值均高并提前数小时出现;而56例患者(82.4％)血清hFABP、cTnI、CK-MB系列浓度均保持在较低的水平.结论:与cTnI和CK-MB相比,hFABP对OPCABG围术期心肌缺血损伤具有更快速、更易于诊断的价值.

Abstract:目的:通过对心肌致密化不全心肌病家系的分析,了解该病在中国患者中的遗传规律及临床特点,以实现早期诊断与治疗.方法:对患者及其亲属采用彩色多普勒超声心动图检查,通过对左心室长轴、左心室短轴、心尖四腔等多切面扫查,观察心肌、心内膜的组织结构和血流分布等特征性超声心动图表现,诊断心肌致密化不全心肌病.结果:所有先证者的超声心动图检查均发现受累心腔内有异常隆突、粗大的肌小梁,其间存在深陷的隐窝,心腔内血液与之相通,并伴有不同类型的心律失常,1例发生肺栓塞.在调查的4个家系中除1例为散发外,其余家系内均发现心肌病患者,其中1名亲属为肥厚型心肌病.结论:通过家系调查提示心肌致密化不全心肌病有遗传性,但遗传规律尚不能明确,且本研究首次发现该病与肥厚型心肌病同时出现在一个家系中,其联系有待进一步研究.

Abstract:目的:了解系统性红斑狼疮(SLE)患者白细胞介素(IL)-10、IL-12异常表达的相互关系,探讨IL-10对IL-12分泌的影响.方法:取26例SLE患者及正常对照18例志愿者的外周血(PBMCs),用ELISA法测定其血清、单个核细胞(PBMCs)培养上清、植物血凝素(PHA)刺激培养上清中IL-10、IL-12水平;在SLE患者及正常对照PBMCs中分别加入正常人血清、SLE血清、SLE血清+IL-10抗体,PHA刺激培养24 h,用ELISA法测培养上清中IL-12的水平.结果:SLE患者血清、PBMCs培养上清中自发产生和PHA刺激诱生的IL-10及IL-12P40水平均异常增高.然而仅血清IL-10水平与SLEDAI呈正相关,但血清IL-12P40水平、PBMCs培养上清自发产生的及PHA刺激诱生的IL-12P40水平均与SLEDAI呈正相关.尽管SLE患者血清IL-10水平与IL-12P40血清及PBMCs培养上清水平呈正相关,但体外实验时,在PBMCs培养中,加入SLE血清,可显著抑制IL-12的产生,采用IL-10抗体中和掉SLE血清中的IL-10后,正常人或SLE患者的PBMCs培养上清中IL-12可明显增多.结论:SLE患者IL-10及IL-12表达水平异常增高,且血清水平与SLE活动性正相关;体外细胞培养实验中SLE血清中IL-10可明显抑制IL-12的分泌,但在细胞因子网络环境中两者之间相互作用更为复杂.

Abstract:目的:探讨应用超声对麻醉雄性Balb/c小鼠心功能进行评价的价值.方法:采集27只麻醉后雄性Balb/c小鼠的二维、M型、多普勒心脏超声图像,分别测量评价心脏结构和功能的各项参数.结果:所有小鼠均可获得反映心脏结构和功能的二维及M型超声心动图,约89％的小鼠能获得反映心脏血流动力学的多普勒超声心动图,统计学分析得到心脏功能正常的雄性Balb/c小鼠心脏结构和功能各项参数的参考值.结论:通过超声可以动态研究Balb/c小鼠心脏结构,准确评价小鼠心脏舒张收缩功能,为今后评价Balb/c小鼠心脏疾病模型提供定性定量指标.

Abstract:目的:探讨肝癌患者肝移植术前、术后使用希罗达+表阿霉素的可行性、安全性及疗效.方法:回顾性研究16例原发性肝癌患者肝移植前后辅助化疗的临床资料.化疗方案为希罗达2 500mg/(m2·d),口服,第1～14天;表阿霉素20 mg,/m2,静脉滴注,第1～2天,分析化疗时机、化疗方案、化疗药物的不良反应及近期疗效.结果:16例接受化疗患者存活时间均超过8个月,最长1例24个月;死亡3例,死亡原因为移植肝肿瘤复发和肝内转移及远处转移,化疗不良反应中,12例出现消化道反应,8例发生骨髓抑制,6例出现手足综合征,4例出现肝功能损害,均为轻中度.结论:肝癌患者肝移植术前、术后化疗联用希罗达+表阿霉素是可行的,不良反应可以耐受,可望延长患者存活时间.选择化疗时机可能对肿瘤复发和患者的生存率有影响.

Abstract:Objective: To observe the effects of Bushen Zhuanggu Formul on interleukin 6 (IL-6) level and bone metabolism of the ovariectomized rats, and to explore the possible mechanism of preventing and treating postmenopausal osteo porosis by traditional Chinese Formul. Methods: SD rats were randomly divided into 4 groups: normal control group (N),ovariectomized control group (OVX), estrogen-treated group (OVX+E) traditional and Chinese medicine-treated group (OVX+CM). After treatment, IL-6 levels in rat bone tissue media and serum were examined by ELISA. The proximal tibias(PTM) of rats were undecalcified for quantitative bone histomorphometry. Results: Compared with OVX group, both Bushen Zhuanggu medicine and estrogen decreased IL-6 level in rat bone tissue media. However, the effects of Bushen Zhuanggu Formul were weaker than those of estrogen. Furthermore, Bushen Zhuanggu Formul and estrogen produced a reduction in bone turnover by inhibiting bone formation and bone resorption at the same time. Conclusion: Bushen Zhuanggu Formul prevents ovariectomic bone loss by inhibiting IL-6 production in rat bone tissues.

Abstract:Objective: To construct green fluorescent protein (GFP) retroviral vector (pLgXSN), and to investigate the expression of GFP in primary rat myoblast. Methods: GFP cDNA was subcloned into the plasmid pLgXSN, and the recombinant vector was transfected into packaging cell PT67. G418 was used to select positive colony. Myoblasts were infected by a high-titer viral supernatant. The recombinant retroviral plasmid vector was identified by restriction endonuclease analysis and DNA sequence analysis. Confocal microscopy and flow cytometry were used to detect the expression of GFP. Results: The GFP cDNA sequence was identical to that of GenBank. Recombinant retroviral plasmid vector pLgGFPSN was constructed successfully. The titer of the packaged recombinant retrovirus was 1 × 106 cfu/ml. Bright green fluorescence of the transfected cells was observed under confocal microscope 48 h after transfection. The transfection rate was 33﹪. The effective expression of GFP in myoblast infected by recombinant retrovirus lasted for 6 weeks. Conclusion: GFP gene could be effectively and stably expressed in myoblast, which suggests that GFP could act as a marker for studies on myoblast.

Abstract:Objective: To study the amelioration effect of Adenovirus5-mediated human hepatocyte growth factor (Ad5-HGF) on postinfarction heart failure in the swine myocardial infarction model. Methods: Twelve SuZhong young swine were randomly divided into 2 groups with 6 swine in each group: Ad5-HGF-treated group and null-Ad5 group. Four weeks after ligation at left anterior descending coronary artery in swine hearts, Ad5-HGF was transferred to the swine myocardium. Simultaneously,Gated myocardial perfusion imaging was performed to evaluate cardiac perfusion and heart function. After three weeks, Gated myocardial perfusion imaging was performed again, then the hearts were harvested and sectioned to examine the expression of HGF through ELISA. Results: High expression of human HGF was observed in the myocardium of Ad5-HGF-treated group. From 4 weeks to 7 weeks after operation, Left ventricular ejection fraction was increased in Ad5-HGF-treated group. The improvement in LVEF was greater in Ad5-HGF-treated group than that in null-Ad5 group at 7 weeks after operation. Cardiac perfusion was significantly improved in the Ad5-HGF-treated group. Conclusion: High expression of human HGF was observed in the myocardium through intracoronary transfection, which suggests that HGF can ameliorate heart function in swine with postinfarction heart failure.

Abstract:Objective: To investigate whether the method of bridgy duct established between the recipient's spleen artery and the donor's gastroduodenal artery could inhibit the apoptosis of liver and bile duct cells caused by hepatic artery ischemia.Methods: Twenty-four mongrel dogs from Xi'an area were used to establish simplified models of dog orthotopic liver transplantation and divided into three groups randomly: HAI group (hepatic artery ischemia group),BBB group (bypassing the blood by a bridgy duct) and control group. After cold perfusion, The samples were collected from liver and bile duct in each group at different time and fixed in glutaraldehyde and 4﹪ polyformaldehyde respectively. At last, the apoptosis of liver and bile duct cells were observed and the apoptotic indexes were calculated. Results: Two hours after cold perfusion, apoptotic phenomenon was common in HAI group, rare in BBB group, while no apoptotic phenomenon was observed in control group. TUNEL staining showed that there was no significant difference in apoptotic index among the three groups immediately after cold perfusion. However, with time going, the apoptotic cells were increased in three groups, and the difference in apoptotic index was significant among three groups (P ＜ 0.01 ). Conclusion: Bridgy duct of hepatic artery can inhibit the apoptosis of liver and bile duct cells caused by HAI significantly.

Abstract:Objective: To investigate the clinical significance of survivin in endometrial carcinoma and to investigate the relationship between the expression of survivin and Ki-67. Methods: Immunohistochemical S-P (streptavidin-biotin-peroxidase complex)method was performed to detect the expression of survivin and Ki-67 antigen in 15 cases of normal endometrium, 21 cases of endometrial simple and complex hyperplasia, 22 cases of endometrial atypical hyperplasia, and 61 cases of endometrial carcinoma. Results: Survivin was hardly detected in some normal endometrium in the proliferative phase and in the secretory phase. However, the level of survivin expression in atypical hyperplasia endometrium(72.73﹪)was higher than that in normal endometrium (7.14﹪)(P ＜ 0.05), including simple and complex hyperplasia (42.38﹪)(P ＜ 0.01), and was lower than that in endometrial carcinoma (90.17﹪)(P ＜ 0.05). Moreover, significant correlation was present between the expression of survivin and the characteristics of endometrial carcinoma, including clinical stage, histological grade and the presence of invasion to myometrium (P ＜ 0.05). In addition, Ki-67 antigen expression was positively correlated with survivin expression in all specimen. Ki-67 labeled indexes (LIs)in hyperplasia endometrium were significantly lower than those in atypical hyperplasia endometrium and endometrial carcinoma (P ＜ 0.01 ), while there was no significant difference in Ki-67 LIs between atypical hyperplasia endometrium and endometrial carcinoma(P ＞ 0.05). There was no significant relationship between Ki-67 LIs and the characteristics of endometrial carcinoma, including histological grade, clinical stage or the invasion to myometrium(P ＞ 0.05). Conclusion: Survivin may participate in the onset and progression of endometrial carcinoma through inhibiting apoptosis and promoting proliferation. Survivin expression is correlated with the malignant degree and prognosis of tumor. Ki-67 is also associated with carcinogenesis and progression of endometrial carcinoma. The results suggest that survivin could be a diagnostic and prognostic marker for endometrial carcinoma and might provide pathways to treat the patients with recurrent or refractory or rudimental endometrial carcinoma.

Abstract:Objective: To determine the encephalic region correlated with epilepsy by manganese-enhanced MRI (MEMRI) and determine the correlation of epilepsy with calcium overloading. Methods: The cats were divided into two groups.The first group underwent EEG examination and ethological observation. The second group underwent MEMRI measurement. Signal enhanced encephalic regions were sectioned. Results: The achievement ratio of convulsive cats intramusclelarly injected with PTZ was 80﹪. MEMRI showed diffuse signal enhancement in the cerebral cortex of the cats with generalized tonic-clonic convulsive seizures compared with control animals. The enhancement rate of frontal-parietal-occipital lobe was 34.6﹪ and 22.9﹪ in temporal lobe compared with the control groups. Signal enhancement on frontal-parietal lobe persisted for 24 h after epileptic seizures were induced. The neurons of enhanced encephalic regions showed obvious degeneration and necrosis. Conclusion: Seizures can be induced in cats by intramuscular injection of PTZ (55 mg/kg). Frontal-parietal lobe is the correlated encephalic regions of epilepsy. MEMRI plays an important role in localizing and revealing pathogenesis of epileptic seizures.

Abstract:Objective: To study the biologic effects of various concentrations of thrombin on aquaporin 4 (AQP4) expression in rat primary cultured astrocytes, and to explore the regulation mechanism of transmembrane water transportation in astrocytes after intracerebral hemorrhage (ICH). Methods: Primary cultured astrocytes were incubated in culture mediums containing various concentrations of thrombin for 24 h and harvested. AQP4 mRNA and AQP4 protein expression were determined by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemical technique. Cell apoptosis was detected by TdT-mediated dUTP nick end labeling (TUNEL) technique. Cell morphology was observed by phase contrast microscope, and cell viability was assayed by MTT. Results: AQP4 mRNA and AQP4 protein showed a low expression in normal astrocytes. The expression of AQP4 mRNA and AQP4 protein significantly increased in the astrocytes treated with 100 U/ml or 200 U/ml thrombin (P ＜ 0.01),and these astrocytes swelled. The number of TUNEL positive cells significantly increased. On the other hand, AQP4 mRNA and AQP4 protein expression were down-regulated in the astrocytes treated with 0.5 U/ml or l U/ml thrombin (P ＜ 0.05),and the cell morphology did not change. Few TUNEL positive cells were observed. Conclusion: AQP4 over-expression induced by high concentrations of thrombin causes an increased permeability of water in astrocytic membrane. On the contrary, the decreased AQP4 expression prevents the astrocytes from swelling and apoptosis.

Abstract:Objective: To investigate the distribution of TRPA1 (one kind of the TRP-like ion channel family) channel in the hippocampus and cerebral cortex of rat. Methods: RT-PCR was used to amplify the fragment of TRPA1 in the DRG (dorsal root ganglion), hippocampus and cerebral cortex of adult SD rat. In situ hybridization staining was used to show the distribution of TRPA1 mRNA in the hippocampus and cerebral cortex of adult rat brain. Results: Both RT-PCR and in situ hybridization staining showed that TRPA1 mRNA was expressed in hippocampus and cerebral cortex of the adult rat brain. Conclusion: Ourresults suggest that there is expression of TRPA1 mRNA both in the hippocampus and cerebral cortex of the adult rat brain.

Abstract:Objective: To explore the mechanism of endoplasmic reticulum stress (ERS) response and related apoptosis in dopaminergic neurons death. Methods: Nerve growth factor (NGF)-treatedPC12 cells were treated with 6-OHDA, MPP+ and rotenone. MTr assay and flow cytometry were used to measure the cell viability and the rate of celluar apoptosis induced by those neurotoxins. The expression of ERS-related gene XBP1, Grp78, CHOP, caspase-12 in drug-treated group and reserpine preincubation group was determined with RT-polymerase chain reaction (RT-PCR) and immunohistochemistry. Results: After the exposure to different toxins, the viability of PC12 cells were decreased by 52﹪, 44﹪, 40﹪ at 100μM6-OHDA, 75 μM MPP+, 20 nM rotenone for 24 h respectively. FCM assay confirmed time-dependent cell apoptosis (P ＜ 0.01 ). The gene and protein expression of XBP1, Grp78 in drug-treated group were significantly increased and reached their peaks 8 h after the treatment(P ＜ 0.05).The expression levels of CHOP and caspase-12 gene were increased 16-24 h after the treatment(P ＜ 0.01 ), but the expression level of caspase-12 was inhibited by reserpine preincubayion (P ＜ 0.05). Conclusion: The excessive ERS and relative activated cell apoptosis pathway may be associated with selective death of dopaminergic neurons.

Abstract:Objective: To study the effects of simulated microgravity induced by tail-suspension on histological structures and antioxidant system in testis of adult rats. Methods: Forty Spraque-Dawley adult male rats were randomly divided into four groups, two experimental groups and two synchronic control groups. Rats in the two experimental groups were tail-suspended for 14 d and 28 d respectively, then serum hormone, testicular morphology and biochemical changes were evaluated. Results:Compared with the synchronic controls, serum testosterone (T) levels and testicular superoxide dismutase (SOD) significantly decreased(P ＜ 0.05), while testicular malondialdehyde(MDA) significantly increased (P ＜ 0.05) in tail-suspended rats. The changes were more prominent in the 28 d tail-suspended rats. In addition, hematoxylin and eosin (HE) staining showed that seminiferous tubules atrophied, spermatogenic cells decreased, and seminiferous epithelial cells disintegrated and shed, abnormal multinucleated giant cells appeared and the lumina were azoospermic in tail-suspended rats. Conclusion: Simulated microgravity induced by tailsuspension has a harmful effect on male reproduction of adult rats by decreasing the ability of testicular antioxidant defense.

Abstract:Objective: To establish a stable and high efficient method for collection of CD4+CD25+ regulatory T cells from rats in vitro. Methods: CD4+CD25+ regulatory T cells were isolated from the rat splenic cells through two steps by magic cell sorting (MACS) system. The first step was negative selection of CD4+ T cells by cocktail antibodies and anti-IgG magic microbeads, and the second step was positive selection of CD25+ T cells by anti-CD25 PE and anti-PE magic microbeads. The purity and viability of separated cells were measured by flow cytometry (FACS) and Trypan blue staining. The suppressive ability of seperated cells on the proliferation of CD4+CD25- T cells was assessed by cell proliferation assay. Results: The purity of negatively enriched CD4+ T cells was 79﹪-87﹪ (83.6﹪ ± 2.5﹪) , and the purity of positively enriched CD4+CD25+ T cells was 86﹪-93﹪ ( 90.2 ± 1.8﹪) with the viability of 92﹪-95﹪ (92.8﹪ ± 3.4﹪). The enriched cells significantly suppressed the proliferation of CD4+CD25- T cells in mixed lymphocyte culture (P ＜ 0.05). Conclusion: An effective method can be established for enrichment of CD4+CD25+ regulatory T cells in two steps by MACS, with satisfied cell purity, viability and function.

Abstract:Objective: To develop an appropriate mass screening system to evaluate hearing loss for primary school children, especially in rural areas. Methods: In Jiangsu Province, at a rural primary school-Longdu Primary School, 317 of 479 rural students (from Grade 1 to Grade 6, over 50 students for each grade) were screened by, using the Chinese Hearing Questionnaire for School Children (CHQS). Two weeks later, they accepted transiently evoked otoacoustic emissions (TEOAEs) test. At the same time, all participants were given a 'gold standard' audiological assessment that comprised of otoscopy, pure tone, and tympanometry. After eight weeks, 103 out of the 317 students accepted the second TEOAEs and 'gold standard' test to confirm the results. Results: There was no statistical difference in the results of two 'gold standard' test (χ2 = 0.14, 0.5 ＜ P ＜ 0.75). Compared with the 'gold standard' results, the hit rate, sensitivity, specificity of the CHQS was 57﹪ ± 2.78﹪, 61﹪ ± 10.18﹪, 57﹪± 2.89﹪ respectively(α = 0.43, β = 0.39). Youden Index J, LR+, LR-, PV+ and PV- was 0.18 ± 0.11, 1.42, 0.68, 0.10, 0.95(Az ＜ 0.54, EF = 0.75-0.95). While a higher system accuracy was obtained by TEOAEs [HR = 97 ± 0.71﹪, Sen = 69 ± 7.8﹪,Spe = 98 ± 0.52﹪(α = 0.02, β = 0.31), Youden Index J = 0.67 ± 0.08, LR+ = 34.5, LR- = 0.32, PV+ = 0.71, PV- = 0.99(Az = 0.85, EF = 0.97)]. Conclusion: TEOAEs got a high efficiency and specificity values with reasonable sensitivity. However,CHQS needs substantial modifications to improve its sensitivity in evaluating hearing loss.

Abstract:Objective: To study the method of cryopreserving rat hepatocytes and double collagen gel culture measurement after its cryopreservation. Methods: Rat hepatocytes, isolated by two-step perfusion with collagenase using an extra corporeal perfusion apparatus, were cryopreserved in double collagen gel with culture medium added by epidermal growth factor(EGF).The expression of cell function and cellular morphology were examined during culture. Results: The hepatocytes cryopreserved in double collagen gel concluding EGF showed good morphology and biological characteristics. After thawing, the MTT metabolism and protein synthesis of hepatocytes in sandwich ± EGF groups were better than those in control group. And the morphology and function of hepatocytes in sandwich group was better than that in EGF group(P ＜ 0.05). Conclusion: Double collagen gel culture can keep hepatocyte's activities. Thawed hepatocytes can be cultivated with collagenous matrix, which provides an environment that more closely resembles that in vivo and maintain the expression of certain liver-specific function of hepatocytes.

Abstract:Objective: To investigate etiological facts and treatment of biliary sludge, cast and stone following orthotopic liver transplantation(OLT). Methods: A review was made using data collected from 81 cases with OLTs performed in our center from February 2003 to January 2004, and confirmed by retrospective study. Etiological factors of biliary sludge, cast and stone following OLT were analyzed, and treatment of biliary sludge, cast and stone following OLT were discussed. Result: Nine cases of biliary sludge, cast and stone were diagnosed and the incidence rate was 11.1﹪. Of these, five were biliary sludge and cast, 2 were bile stone and 2 were necrotic debris. Two cases with hepatic artery embolism received retransplantation and survived. The other one with hepatic artery embolism was ameliorated with nasobiliary drainage by ERCP. Two cases with biliary sludge and cast were resolved by non-operative treatment. Four cases were reoperated, 2 resolved and 2 cases died. Conclusion: Biliary injury and ischemia reperfusion injury, reject reaction, infection and changes of bile kinetics are the important factors causing biliary sludge, cast and stone following OLT. Shortening the time of cold and heat ischemia reperfusion injury of liver, reducing the injury of the blood supply of donor bile duct, actively preventing and early treating of infection and rejection reaction might reduce the incidence rate of biliary sludge, cast and stone following OLT.