姜黄油诱导人肝癌细胞凋亡及其机制

探讨姜黄油的抗肝癌作用及其作用机制,采用二甲基噻唑蓝比色法(MTT)检测肝癌细胞的活力,流式细胞仪检测细胞凋亡率以及线粒体膜电位的变化,Western blot方法检测凋亡相关蛋白Bax、Bcl-2、caspase-3、caspase-9以及细胞色素C的表达。结果发现,姜黄油能时间依赖性地抑制肝癌细胞的生长,降低线粒体膜电位并诱导其凋亡;升高细胞内Bax与Bcl-2的比值;诱导细胞色素C从线粒体中的释放并激活caspase-9和caspase-3。结果提示姜黄油可能通过线粒体途径诱导肝癌细胞的凋亡。     

活性氧激活线粒体凋亡通路是亚硒酸钠诱导NB4细胞凋亡的重要机制

目的 探讨亚硒酸钠诱导NB4细胞凋亡过程中,活性氧引起的线粒体膜电位丧失及凋亡作用的相关机制。方法 用活性氧荧光探针DCFH-DA探测经亚硒酸钠诱导的NB4细胞内活性氧的含量。流式细胞术检测细胞凋亡率及线粒体膜电位变化。Westem blot检测胞浆中细胞色素C含量的变化及细胞总蛋白中Bcl-xl、Bax和Bid的含量变化及切割行为。同时应用活性氧清除剂MnTmPy及活性氧激活剂BSO预处理细胞,观察Bid及凋亡的变化。结果 活性氧清除剂MnTmPy可有效抑制亚硒酸钠诱导的NIM细胞凋亡、线粒体膜电位丧失及Bid的切割;而BSO可促进线粒体膜电位丧失。结论 亚硒酸钠诱导NIM细胞凋亡的可能机制是通过刺激细胞产生活性氧,一方面下调抗凋亡蛋白Bcl-xl,另一方面激活线粒体打孔蛋白Bax和Bid,损伤线粒体使膜电位降低,促使线粒体释放细胞色素C。     

2-甲基-3-羟基蒽醌诱导人乳腺癌MCF-7细胞凋亡的机制

为探讨2-甲基-3-羟基蒽醌抗肿瘤作用及其机制,本研究采用锥虫蓝法检测细胞活力,流式细胞仪检测细胞周期变化、细胞凋亡率、线粒体膜电位及细胞内游离钙的变化,Western blot方法检测凋亡相关蛋白caspase-4、caspase-7、caspase-9、Bcl-2、Bax、JNK、细胞色素C的表达。结果发现：2-甲基-3-羟基蒽醌时间依赖性地抑制乳腺癌细胞的生长,升高细胞内游离钙含量,降低线粒体膜电位并诱导其凋亡;药物上调Bax并下调Bcl-2蛋白的表达;诱导caspase-4、caspase-7、caspase-9、calpain的活化及细胞色素C的释放。结果提示2-甲基-3-羟基蒽醌可能通过Ca²⁺/calpain/caspase-4途径诱导人乳腺癌MCF-7细胞凋亡。     

冬凌草甲素通过线粒体途径诱导人胆囊癌GBC-SD细胞凋亡

目的 研究冬凌草甲素诱导人胆囊癌GBC-SD细胞的凋亡作用及其机制.方法 MTS法检测冬凌草甲素对GBC-SD细胞的生长抑制作用;瑞氏染色法观察细胞的形态学变化;流式细胞仪检测细胞凋亡率、线粒体膜电位改变、caspase-3活性变化;Western blot检测caspase-9活化情况.结果 冬凌草甲素能显著抑制GBC-SD细胞增殖,呈时间剂量依赖性(P＜0.05).细胞形态学观察可见冬凌草甲素可诱导细胞发生凋亡.流式细胞仪检测结果显示28 μmol/L药物处理GBC-SD细胞24h后,细胞凋亡率为51.28％±2.65％.随着药物浓度增加,GBC-SD细胞线粒体膜电位逐渐下降而caspase-3活性逐渐增强,56μmol/L组caspase-3活性是对照组的11倍.Western blot分析结果显示caspase-9酶原被激活,且活化条带随药物浓度的增加而增强.结论 冬凌草甲素可能通过细胞线粒体膜电位下降激活caspase-3并最终诱导GBC-SD细胞凋亡.     

AAVC-1诱导A549细胞凋亡的线粒体机制研究

目的:探讨皖南五步蛇蛇毒抑瘤组分Ⅰ(AAVC-1)诱导人非小细胞型肺癌A549细胞凋亡的作用,并分析其可能的线粒体机制。方法:以不同作用浓度的AAVC-1处理A549细胞,采用Annexin V/PI双染流式细胞仪分析法检测A549细胞凋亡百分率;JC-1检测细胞线粒体的膜电位变化,免疫组化检测细胞色素C在线粒体内的分布。结果:不同浓度梯度AAVC-1 1.0、3.0、9.0、20.0μg/m L处理24 h后,与对照组相比,A549细胞早期凋亡率明显增加(P<0.05);线粒体膜电位绿色荧光百分比由(23.01±6.07)%上升至(87.26±9.54)%(P<0.01),荧光显微镜观察到绿色荧光增强;免疫组化显示,随着AAVC-1浓度增加,胞质内细胞色素C平均光密度值明显增加(P<0.05)。结论:AAVC-1可以通过降低线粒体膜电位和促进细胞色素C的释放,激活线粒体通路诱导人非小细胞型肺癌A549的凋亡。     

线粒体途径在丹参酮ⅡA诱导人宫颈癌细胞凋亡中作用

目的探讨丹参酮ⅡA诱导宫颈癌HeLa细胞凋亡的可能途径。方法体外培养人宫颈癌HeLa细胞,用不同浓度丹参酮ⅡA处理后,流式细胞仪检测HeLa细胞凋亡,JC-1染色后流式细胞仪检测线粒体膜电位的改变,Western印迹法检测细胞色素C(Cyto C)从线粒体释放情况。结果不同浓度丹参酮ⅡA作用48h后,与空白对照组比较,HeLa细胞凋亡率均增加;线粒体Cyto C表达减少,细胞质Cyto C表达增加;线粒体膜电位电压较高细胞(正常细胞)数量减少,线粒体膜电位降低的细胞数量增加,差异均有统计学意义(F=116.5～1 064.6,q=4.11～75.38,P<0.01)。结论丹参酮ⅡA对HeLa细胞的凋亡诱导作用与线粒体膜电位降低及线粒体释放Cyto C有关。     

鲎素诱导HL-60细胞凋亡的机制探讨

目的 研究鲎素诱导HL-60细胞凋亡时,半胱氨酸蛋白酶(Caspases)活性的变化,探讨鲎素诱导HL-60细胞凋亡的可能机制.方法 以鲎素20 μg/ml处理HL-60细胞0～24 h.流式细胞仪检测线粒体膜电位.Western blot法和发色底物法检测Caspase-3、Caspase-6、Caspase-8和Caspase-9活性变化.Western blot法检测胞质中细胞色素C的变化.结果 鲎素处理HL-60细胞后,线粒体膜电位下降.Caspase-3、Caspase-6、Caspase-8和Caspase-9被激活,呈时间依赖性改变.鲎素处理HL-60细胞6 h Caspase-8活性最高,而Caspase-9在12 h活性最高,Caspase-6活性在18 h达峰值,Caspase-3活性峰值时间在24 h后.胞质中细胞色素C含量呈时间依赖性增加.结论 鲎素诱导HL-60细胞凋亡可能是多途径的,至少与线粒体途径和死亡受体途径相关.     

Survivin反义寡核苷酸经线粒体途径诱导肺癌细胞凋亡研究

目的: 探讨survivin反义寡核苷酸(antisense oligonucleotides,ASODN)诱导肺癌细胞凋亡的作用及survivin抗凋亡的分子机制。方法: 脂质体介导下,分别以100 nmol/L、300 nmol/L、500 nmol/L survivin ASODN作用于肺癌细胞株NCI-H446后,在不同时间用流式细胞仪(flow cytometry,FCM)检测凋亡;Rh123染色法检测细胞线粒体膜电位(mitochondrial potentialmembrane,△Ψm)变化,EL ISA法检测胞质细胞色素C(cytochrome C,Cyt C)浓度,比色法检测胞质内天冬氨酸特异性半胱氨酸蛋白酶-9(cystenyl aspartate specific proteinase-9,caspase-9)活性变化;加入caspase-9抑制剂Z-LEHD-FMK后FCM检测细胞凋亡。结果: Survivin ASODN 500 nmol/L作用72 h效果最佳,细胞凋亡指数(AI)达48.35%,增殖指数(PI)为24.38%;Survivin ASODN导致细胞△Ψm逐渐下降,并相继引起Cyt C的释放和caspase-9的激活。三者变化具有时间依赖性和差异性;Z-LEHD-FM K显著抑制了survivin ASODN诱导的细胞凋亡。结论: Survivin主要通过调控线粒体凋亡途径发挥抗肺癌细胞凋亡作用;Survivin ASODN能够显著诱导肺癌细胞凋亡,抑制细胞增殖。     

EGCG活化线粒体途径诱导人胃癌细胞凋亡

目的 探讨表没食子儿茶素没食子酸酯(EGCG)通过活化线粒体途径诱导人胃癌BGC823细胞凋亡的作用机制.方法 MTT法检测EGCG对BGC823细胞生长的影响;PI染色流式细胞术(FCM)分析测定细胞凋亡率;细胞线粒体跨膜电位(△(Ψ)m)用荧光染料罗丹明123染色FCM分析测定;Westem blot检测线粒体凋亡信号传导通路相关蛋白的表达.结果 EGCG对BGC823细胞生长有显著抑制作用,呈时间和剂量依赖性.EGCG以剂量依赖方式诱导BGC823细胞凋亡.EGCG(20μg/mL、40 μg/mL、80μg/mL)作用48 h后,BGC823细胞中△(Ψ)m降低,细胞色素c(Cyt c)释放增加,caspase-9蛋白表达增加,呈剂量依赖性.结论 EGCG通过活化线粒体信号传导途径诱导BGC823细胞凋亡.     

线粒体凋亡途径参与TRAIL诱导胃腺癌细胞凋亡

目的研究TRAIL诱导胃腺癌细胞凋亡的途径。方法PI染色、流式细胞仪检测细胞凋亡;广谱caspase抑制剂(z-VAD-fmk)、caspase-8抑制剂(z-IETD-fmk)和caspase-9抑制剂(z-LEHD-fmk)分别与TRAIL共同作用后PI染色、流式细胞仪检测细胞凋亡率的变化;Western blot检测caspase-3的活化及PARP的裂解;JC-1染色、流式细胞仪分析线粒体膜电位的变化。结果TRAIL诱导胃腺癌细胞凋亡具有剂量和时间依赖性,BGC-823较SGC-7901对TRAIL诱导的凋亡更敏感,TRAIL(100μg/L)作用24h细胞凋亡率分别是59.9%、24.3%;3种caspase抑制剂都能很大程度上阻止TRAIL诱导胃腺癌细胞凋亡;caspase-3、PARP在TRAIL作用早期即活化、裂解,且BGC-823比SGC-7901发生得更快;线粒体膜电位随TRAIL处理时间延长而不断下降。结论TRAIL能诱导胃腺癌细胞凋亡,且这种凋亡依赖于caspases;除死亡受体途径外,线粒体途径也参与了凋亡信号通路。     

Value of D-Dimers in patients with acute aortic dissection

Objective: To evaluatel the value of D-dimers in patients with acute aortic dissection (AAD). Methods: This study consisted of 16 patients with AAD and 27 non-AAD patients. Serum D-dimets were measured by Sta-Liatest D-DI immunoturbidimetric assay. Results: D-dimer level was higher (P ＜ 0.001) in patients with AAD(7.91 ± 5.52 μg/ml) than that in non- AAD group(1.57±1.24 μg/ml). D-dimer was positive (＞0.4 μg/ml) in all patients with AAD and in 10 control group patients (37%). Among patients with acute AAD, D-dimers tended to be higher in Stanford A than in Stanford B (8.67 ± 4.31 μg/ml vs. 3.24±1.27 μg/ml, P ＜0.01). D-dimer values tended to be higher in more extended disease(3.84 ± 1.65 μg/ml, 8.57 ± 3.58 μg/ml and 11.87 ± 5.69 μg/ml in thoracic aorta, thoracic and abdominal aorta, thoracic and abdominal aorta and iliacal arteries, respectively, P ＜ 0.05 for both 8.57 ± 3.58 and 11.87 ± 5.69 vs. 3.84 ± 1.65 ). Including the control group into the analysis, we found a sensitivity of 100%, a negative predictive value of 100%, and a specificity of 66% and a positive predictive value of 64% for D-dimer in diagnosis of AAD in our patients with suspected AAD. Conclusion: D-dimer was elevated in patients with AAD. A negative D-dimer test result could be useful in excluding AAD.     

Research of combined liver-kidney transplantation model in rats

Objective： To set up a simple and reliable rat model of combined liver-kidney transplantation. Methods： SD rats served as both donors and recipients. 4℃ sodium lactate Ringer＇s was infused from portal veins to donated livers,and from abdominal aorta to donated kidneys, respectively. Anastomosis of the portal vein and the inferior vena cava （IVC） inferior to the right kidney between the graft and the recipient was performed by a double cuff method, then the superior hepatic vena cava with suture. A patch of donated renal artery was anastomosed to the recipient abdominal aorta. The urethra and bile duct were reconstructed with a simple inside bracket. Results： Among 65 cases of combined liver-kidney transplantation, the success rate in the late 40 cases was 77.5%. The function of the grafted liver and kidney remained normal. Conclusion： This rat model of combined liver-kidney transplantation can be established in common laboratory conditions with high success rate and meet the needs of renal transplantation experiment.     

BLOOD PRESSURE CHANGE WITH AGE IN SALT-SENSITIVE TEENAGERS

Objective To observe blood pressure change with age in salt-sensitive teenagers whose salt sensitivity were determined by repeated testing.Methods Salt sensitivity was determined through intravenous infusion of normal saline combined with volume-depletion by oral diuretic furosemide in 55 teenagers. After five years, salt sensitivity was re-examined and subject blood pressure was followed up. Blood pressure changes in salt-sensitive teenagers were compared to that of non-salt sensitive teenagers over five years.Results After 5 years, the repetition rate of salt sensitivity determined by intravenous saline loading is 92.7%. In teenagers with salt sensitivity on the baseline, both the systolic blood pressure increments and increment rates were much higher than non-salt sensitive teenagers (12.7±12.1 mmHg vs. 2.8±5.2 mmHg, P＜ 0.01; 12.2%± 12.0% vs. 2.5% ±4.4%, P＜ 0.001,respectively). There was a similar trend for diastolic blood pressure (8.4 ± 6.4 mmHg vs. 3.7 ± 6.4 mmHg, P = 0.052; 13.2% ±10.6 % vs. 6.8%± 10.1%, P = 0.053, respectively).Conclusions Salt sensitivity determined by intravenous saline loading showed good reproducibility. Blood pressure increments with age were much higher in salt-sensitive teenagers than non-salt sensitive teenagers, especially in terms of systolic blood pressure.     

安心颗粒对急诊经皮冠状动脉介入术后生活质量影响的研究

目的：评价使用安心颗粒对急诊经皮冠状动脉介入术（PPCI）术后生活质量的影响.方法：将160例接受PPCI的急性ST段抬高型心肌梗死患者随机分为安心颗粒组（术前顿服安心颗粒8.8g,术后安心颗粒4.4 g/次,每日2次）和对照组（仅接受基础药物治疗）.所有患者均服用阿司匹林、氯吡格雷和阿托伐他汀.分别在入院时、出院前1d、出院后180 d时,应用心肌梗死多维度量表（MIDAS）、中文版SF-36评价量表对患者生活质量评分.并观察术后30 d以内的出血并发症、血小板减少症发生情况.结果：入院时和出院前1d,两组患者的心肌梗死MIDAS、SF-36量表评分比较无差异（P＞0.05）;出院后180 d时,与对照组比较,安心颗粒组MIDAS、SF-36评分明显减低（P＜0.05）;组内与入院时比较,两组出院前1d、出院后180 d时,MIDAS、SF-36评分均降低（P＜0.05）.两组患者在随访期间均无大量出血、少量出血、重度和极重度血小板减少症发生,安心颗粒组有4例、对照组有7例发生不明显出血（P＞0.05）.两组发生轻度血小板减少症的患者数比较无差异（P＞0.05）.结论：PPCI使用安心颗粒,能改善急性ST段抬高型心肌梗死患者的生活质量,且不增加出血风险.     

The comparative studies of the influences of Urapidil and Nicardipine on sino-atrial node function, atrio-ventricular node function and hemodynamics

Objective:To investigate the influences of urapidil and nicardipine on rabbit sinus function,atrio-ventricular node function and hemodynamics.Methods:Thirty-two Angora's rabbits were selected and randomly divided into four groups.U1 group:urapidil 0.25 mg/kg;U2 group:urapidil 0.5 mg/kg;N1 group:nicardipine 10 μg/kg;N2 group:nicardipine 20 μg/kg.All these medicine were administrated within 30 seconds.Measurements were taken before and after the administration of urapidil or nicardipine for the following data:mean blood pressure(MAP),heart rate(HR),sino-atrial conduction time(SACT),maximal sinoatrial recovery time(SNRTmax)corrected sinus node recovery time(CSNRT),index of sinus node recovery time(SNRTI),Wenckebach A-V conduction frequency (WB),and P-R interval.Results:Significant MAP and HR changes were identified in all of the four groups before and after administration of both urapidil and nicardipine.No significant changes could be found in the rest of the parameters.Intergroup analysis showed that SACT and CSNRT of N1 and N2 groups were shorter than those of the U2 group(P＜0.01);the MAP decreased(P＜0.01)and the HR increased drastically(P＜0.01).Conclusions:Neither urapidil(0.25 mg/kg,0.5 mg/kg)nor nicardipine(10μg/kg,20μg/kg)has any significant influence on rabbit sinus function or rabbit atrio-ventricular node function.Nicardipine could be a better choice than urapidil for parafunctional sinus node patients.     

Expression of OPGmRNA and ODFmRNA in the patients of hip fracture due to osteoporosis

Objective：To investigate the gene expression of osteoprotegerin（OPG） and osteoclast differentiation factor（ODF） in the bone tissue of patients with hip fracture due to osteoporosis. Methods：OPGmRNA and ODFmRNA in the bone tissue in 50 cases of osteoporosis sufferers（over 50 years old） with hip fracture（Observer Group） and 30 cases of hip facture sufferers with no osteoporosis（Control group） were analyzed with the Semi-Quantitative RT-PCR method. Results：The mRNA expressed of ODF, OPG were both high in the patients with hip fracture. In the control group, the expression of OPG mRNA was observed, while the expression of ODF mRNA was very slight. Conclusion：Aged patients contained all signals including OPG, ODF that are essential for inducing osteoclastogenesis and promoting bone resorption.     

The clinicopathological and immuohistochemical analysis of solid-pseudopapillary tumor of the pancreas： report of 9 cases

Objective：To investigate the clinical features, pathological characteristics and immunophenotype of solid-pseudopapillary tumor of the pancreas（SPTP）. Methods：Nine surgically treated cases of SPTP were retrospectively reviewed. Hematoxylin and Eosin（HE） staining and immunohistochemical staining were used to analyze all cases, and the general clinical data was collected. Results：Six patients were asymptomatic except for a palpable mass. Two patients complained of vague-epigastric pain. One patient appeared jaundice. The tumor was encapsulated and solid tissues alternately with cystic tissues. Histologically, the histological structure of solid portion was pseudopapillary with a fibrovascular core. Tumor cells were uniform and medium-sized which were arranged in sheets ets or nests or pseudopapillary patterns. Immunohistochemical studies demonstrated that SPTP proved positive in vimentin（9/9 cases）, AAT（9/9 cases）, NSE（9/9 cases）, ACT（7/9 cases）, CK20（2/9 cases）, CgA（1/9 cases）, S-100（3/gcases）, PR（4/gcases）, Syn（3/9 cases） and CD56（5/9cases）, negative in CEA and ER. Conclusion：SPTP is a tumor predominantly occurring in young women frequently without special symptoms. This tumor has various characteristical histological patterns with different immunophenotype.     

Dynamic Changes in Serum Estradiol and Progesterone levels in Patients of Premenstrual Syndrome with Adverse Flow of Liver-qi

Objective:To probe into the influence of changes of ovarian hormones on the pathogenesis of the specific sub-type premenstrual syndrome(PMS)and reveal partial microcosmic mechanisms of adverse flow of liver-qi.Methods:Estradiol(E2)and progesterone(P)levels in serum were determined at different phases of menstrual cycle by radioimmunoassay.Results:In the group of PMS with adverse flow of liver-qi.the secretive peak value Of E2 and P at the follicular phase significantly decreased,and the secretive peak value at the luteal phase did not come into being.Conclusions:Low E2 and P secretive peak at the follicular phase and absence of secretive peak at the luteal phase is one of the microcosmic mechanisms of PMS with adverse flow of liver-qi.One of the pathophysiologic mechanisms of specific sub-type PMS is probably the continuous low level of E2and P.     

Real-time Three-dimensional Echocardiography in Assessment of Left Ventricular and Right Ventricular Volumes

Real-time three-dimensional echocardiography (RT3DE)is a new ultrasound technique that enables dynamic threedimensional visualization and quantification of the heart in real time. Investigation of feasibility and methodology of RT3DE in determining left ventricular (LV) and right ventricular (RV) volumes, RT3DE was performed in 35 normal adults using Philips SONOS 7500 system with a 2-4 MHz matrix array transducer. The 60°×60° "pyramid" volume database was obtained and analyzed on a TomTec echo workstation. Both LV and RV volumes were calculated with four 3DE methods (i.e. apical 2, 4, 8, and 16-plane) through manually tracing ventricular endocardial borders in end diastole and end systole. Stroke volumes were then calculated. LV volume was also measured by 2DE Simpson's rule using GE VIVID 7 ultrasound machine.     

SCREENING FOR A 21-CHROMOSOME ABNORMALITY IN PREIMPLANTED EMBRYOS OF ELDERLY WOMEN

Increasing maternal age is the only etiological factor unequivocally linked to Down's syndrome in humans. The occurrence rate of newborns with Down's syndrome is about 1/220 in women over 35 years old. However, the occurrence rate in embryos fertilized in vitro, of the elder woman is unclear. Using FISH we screened the number of chromosome 21 in preimplanted embryos of 5 elderly women (average age, 38.4 years) to study the feasibility and necessity of screening trisomy 21 in embryos in patients over 35 years old at the in vitro fertilization (IVF) center.