子痫前期血清诱发血管内皮细胞凋亡机制的实验研究

目的：探讨子痫前期患者血清对体外生长血管内皮细胞凋亡情况的影响及发生机制.方法：对血管内皮细胞进行体外培养,分别加入正常孕妇血清和重度子痫前期患者血清,采用流式细胞术检测血管内皮细胞凋亡率、半胱天冬氨酸蛋白酶-3（Caspase-3）活性,并在培养体系中加入阳离子染料JC-1后检测血管内皮细胞线粒体跨膜电位变化.结果：①子痫前期血清组诱导血管内皮细胞早期凋亡阳性率与晚期细胞凋亡阳性率分别为（10.8±3.2）％与（6.8±1.8）％,明显高于正常孕妇血清组的早期凋亡阳性率与晚期细胞凋亡阳性率（4.2±1.7）％与（3.3±1.1）％（P＜0.05）;②子痫前期血清组诱导血管内皮细胞Caspase-3阳性率为（8.3±1.2）％,明显高于正常孕妇血清组诱导血管内皮细胞Caspase-3阳性率（3.6±1.1）％（P＜0.05）：③子痫前期血清组诱导血管内皮细胞其线粒体损伤率为（8.9±2.1）％,明显高于正常孕妇血清组诱导血管内皮细胞的线粒体损伤率（3.0±0.8）％（P＜0.05）.结论：子痫前期患者血清可促进体外生长血管内皮细胞的凋亡,Caspase-3和线粒体凋亡在子痫前期患者血清促血管内皮细胞凋亡中可能起重要作用.     

Effect of resveratrol on TNF-α-induced vascular endothelial inflammation in vitro

目的 观察白藜芦醇对TNF-α刺激引起的内皮细胞炎症反应的影响,并围绕TNFR1探索相关作用机制.方法 不同浓度(0.01、0.1、1、2.5、5、10、20、50、100、200、400 ng/ml)TNF-α处理人血管内皮细胞株EA.hy926,24h后MTT法和ELISA法分别检测细胞增殖活力和细胞培养液中slCAM-1释放水平;白藜芦醇(浓度为0.1、1、10 μmol/L)预处理24h后,再用TNF-α( 10 ng/ml)处理24h,利用ELISA法检测细胞培养液中sICAM-1释放水平,qRT-PCR法检测ICAM-1mRNA和TNFR1 mRNA表达水平.结果 高浓度TNF-α(≥100 ng/ml)刺激EA.hy926细胞后,细胞增殖活力显著下降(P＜0.05),当TNF-浓度为200 ng/ml时,与对照组比较细胞增殖活力下降约59.7％;浓度为10 ng/ml的TNF-α刺激内皮细胞后,细胞的ICAM-1及TNFR1 mRNA表达明显上调(P＜0.05);白藜芦醇预处理内皮细胞后可明显下调TNFR1 mRNA表达水平(P＜0.05),并显著抑制TNF-α诱导的ICAM-1的表达(P＜0.05),且抑制作用随着浓度的增加而增强.结论 白藜芦醇可能通过抑制TNFR1表达,进而抑制TNF-α刺激引起的炎性因子释放,减轻内皮炎性损伤.     

中药尿酸利仙含药血清对TNF-a诱导的人血管内皮细胞VCAM-1mRNA表达的影响

目的:观察中药尿酸利仙(含药血清)对肿瘤坏死因子-α(TNF-α)诱导的人血管内皮细胞(ECV)血管细胞黏附分子-1(VCAM-1)mRNA表达的影响。方法:①体外培养人血管内皮细胞,分别用不同浓度(12.5、25、50μg/ml)的TNF-α刺激;②体外培养人血管内皮细胞,培养液中加入TNF-α至终浓度为25μg/ml,分别孵育细胞8、16、24、48小时。③用25μg/ml的TNF-α孵育细胞16小时,同时用含不同浓度(2.5%、5%、10%)中药尿酸利仙含药血清的培养液进行干预,用逆转录-聚合酶链式反应(RT-PCR)技术测定血管内皮细胞VCAM-1mRNA表达。结果:低、中、高浓度TNF-α刺激均可显著上调血管内皮细胞VCAM-1mRNA表达(P0.01),各刺激组间差异无统计学意义(P0.05)。25μg/ml的TNF-α在16、24和48小时均可显著上调血管内皮细胞VCAM-1mRNA表达(P0.01),各干预组间差异无统计学意义(P0.05)。低、中、高浓度的中药尿酸利仙含药血清干预均可显著降低VCAM-1mRNA表达(P0.05,P0.01)。结论:中药尿酸利仙含药血清可抑制VCAM-1mRNA过度表达,可能是减轻TNF-α诱导的人血管内皮细胞炎症损伤及抑制炎症反应的机制之一。     

子痫前期血清对血管内皮细胞凋亡及细胞因子表达的影响

目的 探讨子痫前期患者血清诱导血管内皮细胞核因子-κB(NF-κB)活性及血管内皮细胞黏附分子-1(VCAM-1)表达的结果.方法 对血管内皮细胞株进行体外培养,分别加入正常妊娠及子痫前期患者血清.2 h后Western blot法测定细胞胞浆NF-κB抑制因子(I-κB)和胞核NF-κB活性;48 h后流式细胞仪检测细胞凋亡,酶联免疫法测定VCAM-1的表达.结果 子痫前期患者血清培养的血管内皮细胞胞浆I-κB含量明显低于正常妊娠血清培养组(P＜0.05),胞核NF-κB 含量以及VCAM-1的表达明显高于正常妊娠血清培养组(P＜0.05).结论 子痫前期患者血清可促进血管内皮细胞VCAM-1的表达、NF-κB的活性以及血管内皮细胞的凋亡;NF-κB在子痫前期患者血清促血管内皮细胞凋亡及VCAM-1的表达过程中可能起重要作用.     

乌司他丁对大鼠多脏器功能障碍综合征炎性因子的影响及意义

目的 通过乌司他丁（UTI）干预多脏器功能障碍综合征（MODS）大鼠,检测外周血炎性因子表达,研究UTI对MODS的可能保护作用及机制.方法 将90只Wistar大鼠随机分为对照组、模型组与UTI组,模型组与UTI组大鼠分别通过尾静脉以15 g/kg注射内毒素（LPS）,UTI组建模前24 h和建模后连续3d,每日2次以1＆#215;105 U/kg腹腔注射UTI进行干预和治疗,观察各时间点各组的体征表现并收集血清用ELISA方法检测TNF-α、IL-1α、IL-4和IL-10的表达水平.结果 ①对照组、模型组和UTI组的死亡率分别为：0％、33％和16％.②TNF-α、IL-1α的表达水平UTI组均低于相对应的模型组.③模型组IL-4的表达水平在6h与24 h与对照组相比明显的升高（P＜0.05）,UTI组与对照组相比在0h、12 h（P＜0.05）,24 h（P＜0.01）显著升高;模型组IL-10的表达水平在12h与对照组相比明显升高（P＜0.05）,UTI组在6 h～12 h时IL-10的表达水平与对照组相比明显升高（P＜0.05）.结论 UTI能降低LPS介导的大鼠MODS的死亡率,可抑制TNF-α、IL-1α释放和促进IL-4、IL-10释放,有效改善MODS的发生发展.     

α干扰素对TNF-α诱导宫颈癌Hela细胞凋亡及NF-κB表达的研究

目的:探讨应用α干扰素与宫颈癌细胞核因子κB(NF-κB)表达的关系以及对肿瘤细胞凋亡的影响.方法:采用TUNEL法检测细胞凋亡,Western blot分析方法检测NF-κB表达.结果:100 U/ml的α干扰素与10 ng/ml肿瘤坏死因子α(TNF-α)联合作用24 h后,细胞凋亡率达到92%;与15 ng/ml TNF-α联合作用12 h后,凋亡率上升至92.6%.α干扰素不影响NF-κB的表达,TNF-α可显著增加NF-κB的表达(P＜0.01),α干扰素预先作用后加用TNF-α可以显著抑制NF-κB的表达(P＜0.01).结论:α干扰素能够显著抑制TNF-α诱导宫颈癌Hela细胞NF-κB的表达,增强TNF-α诱导Hela细胞凋亡的作用.     

肿瘤坏死因子α对人脐静脉内皮样细胞增殖及syndecan-4蛋白表达的影响

目的 观察肿瘤坏死因子α(TNF-α)对体外培养的人脐静脉内皮样细胞株ECV304增殖及syndecan-4蛋白表达的影响.方法 体外培养人脐静脉内皮样细胞株ECV304,分别应用1、10、20、100 ng/ml的TNF-α作用24 h及36 h并设立对照组进行比较,采用MTS/PES法确定人脐静脉内皮样细胞的增殖状态.利用Western blotting蛋白免疫印迹法测定细胞syndecan-4蛋白的表达情况.结果 24 h各组细胞增殖率分别为对照组(1.956±0.214),TNF-α100 ng/ml组(2.154±0.250),TNF-α20 ng/ml组(2.26±0.151),TNF-α10 ng/ml组(2.118±0.205),TNF-α1 ng/ml组(2.106±0.136).统计分析显示低至1 ng/ml的TNF-α仍能明显刺激人脐静脉内皮样细胞的增殖(P＜0.05),其中以TNF-α 20 ng/ml组细胞增殖最显著P＜0.05).36 h各剂量组的细胞增殖率均明显低于24 h的细胞增殖率(P＜0.05).TNF-α对人脐静脉内皮样细胞的syndecan-4蛋白表达有显著的增强作用(P＜0.05).结论 TNF-α对体外培养的人脐静脉内皮样细胞的增殖及syndecan-4蛋白表达均有明显的促进作用,但随着时间的改变,这种作用有所变化.     

子痫前期患者血清12-羟基二十碳四烯酸浓度变化及其对血管内皮细胞凋亡的影响

目的观测子痫前期患者血清12-羟基二十碳四烯酸（12-HETE）浓度变化及对绒毛膜外滋养细胞诱导血管内皮细胞凋亡的影响,探讨12-HETE参与在子痫前期发生、发展的可能机制。方法收集2019年6月至2020年6月复旦大学附属中山医院吴淞医院子痫前期患者临床症状出现前26例和出现后23例,以及与之孕周、年龄及BMI相匹配的正常孕妇26例及正常未孕妇女14例血清样本,利用超高压液相色谱串联质谱仪（UPLC-MS）检测各组血清12-HETE浓度;建立人血管内皮细胞的单培养及其与胎盘滋养细胞HTR-8/Snveo的共培养模型,分为正常对照组、12-HETE处理组、共培养对照组、共培养12-HETE处理组。采用流式细胞术检测各组细胞血管内皮细胞的凋亡情况,计算凋亡率。结果12-HETE的保留时间为3.33min,母离子碎片为319.33m/z,离子碎片为179.22m/z,在20~2000pg/ml的线性范围内,相关系数r2=0.99,能够满足样本检测需求。子痫前期患者临床症状出现前、后12-HETE浓度均高于正常孕妇组和正常未孕妇女组（均P＜0.01）。血管内皮细胞单培养中,12-HETE处理组细胞凋亡率显著低于正常对照组（P＜0.01）;共培养对照组血管内皮细胞的凋亡率显著高于正常对照组（P＜0.01）;共培养12-HETE处理组血管内皮细胞的凋亡率显著低于共培养对照组（P＜0.01）。结论子痫前期患者临床症状出现前后外周血中12-HETE浓度均显著升高,其可能通过抑制血管内皮细胞凋亡参与子痫前期的发生、发展。     

重度子痫前期患者的血清作用下NK细胞对血管内皮细胞分泌内皮素-1及前列环素的影响

目的 研究在重度子痫前期患者的血清作用下,NK细胞对血管内皮细胞分泌内皮素-1(ET-1)及前列环素的影响.方法 重度子痫前期组、正常妊娠组各18例,采用1∶1配比的病历对照研究.用20％重度子痫前期及正常妊娠妇女的血清孵育NK细胞,加入脐带血管内皮细胞(HUVEC)培养体系中培养24 h.采用放射免疫方法检测NK细胞与HUVEC培养上清液中ET-1及前列环素的稳定代谢产物6-keto-PGF1α的水平.结果 重度子痫前期组血管内皮细胞分泌ET-1的水平为(40.35±8.94)pg/mL,与正常妊娠组(33.06±7.88)pg/mL相比显著增高(P＜0.01),差异具有统计学意义;重度子痫前期组血管内皮细胞分泌6-ke-to-PGF1α的水平为(527.23±102.59)pg/mL,与正常妊娠组(629.64±114.45)pg/mL相比显著降低(P＜0.01),差异具有统计学意义.结论 在重度子痫前期患者的血清作用下,NK细胞导致血管内皮细胞分泌血管收缩因子内皮素-1增多,分泌血管舒张因子前列环素减少.     

乌司他丁对大鼠肝脏冷缺血-再灌注肺损伤的影响

目的 研究乌司他丁（UTI）预处理对肝缺血-再灌注肺损伤（IRI）的影响.方法 将SPF级SD大鼠30只随机分为3组：假手术组（S组,n=10）大鼠仅进行单纯开关腹手术,游离相应血管;模型对照组（M组,n=10）大鼠采用肝脏冷缺血-再灌注模型手术,不给予任何干预措施;UTI组（U组,n=10）大鼠采用肝脏冷缺血-再灌注模型,在切皮时经尾静脉给予UTI,10万U/kg.各组于缺血再灌注后12 h收集血清和肺组织标本,利用ELISA检测TNF-α和IL-6表达水平;切取肺组织,测定肺组织丙二醛（MDA）和超氧化物歧化酶（SOD）含量,光学显微镜下观察肺组织病理学结果.结果 与S组相比,M组和U组大鼠血清TNF-α和IL-6浓度及肺组织MDA含量均升高（P＜0.05）,肺组织SOD活性均降低（P＜0.05）;与M组相比,U组血清TNF-α和IL-6浓度及肺组织MDA的含量均降低（P＜0.05）,肺组织SOD活性均升高（P＜0.05）.结论 UTI预处理可明显减轻大鼠肝脏冷缺血-再灌注后肺损伤;肝脏移植.     

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.     

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.     

Scientific principles and rigorous processes should be followed in developing clinical guidelines for therapeutic interventions of integrative medicine

A clinical guideline for the therapeutic interventions of integrative medicine may be defined as a written document which states a series of recommendations on therapeutic interventions of integrative medicine for a special disease or condition. The guideline may provide assistance to medical professionals in making clinical decisions aimed at improving the clinical outcome of patients and reducing the costs of medical care（~＇4~. Recommendations issued by a guideline should be based on the best available evidence in both Western and Chinese medicine. For fulfilling this purpose, the development of clinical guidelines for therapeutic interventions in the field of integrative medicine should follow scientific principles and undergo a rigorous processes.