PCNA在小鼠胚胎着床过程中的作用

目的:探讨增殖细胞核抗原(Proliferating cell nuclear antigen,PCNA)在小鼠胚胎着床过程中的作用.方法:实时荧光定量聚合酶链反应(Real-time Fluorescence Quantitative PCR,Real-time FQ-PCR)技术检测未孕及孕2、3、4、5、6、7 d(分别记为d0、d2、d3、d4、d5、d6、d7)小鼠子宫内膜PCNA mRNA的表达;子宫角注射PCNA单克隆抗体观察胚泡着床数.结果:FQ-PCR测得随着小鼠妊娠天数的增加,PCNA mRNA表达量也逐渐增高,在孕d4、d5达到峰值,孕d6、d7逐渐下降.子宫角注射PCNA单克隆抗体后胚泡着床数明显减少.结论:PCNA在妊娠早期子宫内膜持续表达,可能参与了胚泡着床.     

小鼠胚泡着床前后子宫内膜细胞间粘附分子-1 mRNA表达水平的变化

目的　研究小鼠胚泡着床前后子宫内膜细胞间粘附分子1(ICAM1)在转录水平的表达变化。方法　小鼠雌雄合笼后建立小鼠早孕模型,用逆转录聚合酶链式反应(RTPCR),测定ICAM1mRNA的表达。结果　小鼠正常动情期子宫内膜有一定ICAM1mRNA的表达,着床前后其表达显著升高,并在着床前一天即孕3d达到高峰,着床后逐渐出现下降趋势。结论　ICAM1在胚泡着床前后明显上调,和胚泡着床密切相关。     

整合素β3在围植入期小鼠子宫内膜中的表达

目的 检测整合素β3在围植入期小鼠子宫内膜中的表达,明确整合素β3表达规律.方法 将清洁级性成熟期昆明小鼠44只,雌性24只,雄性20只,雌性小鼠阴道涂片检查为动情前期,在此期注射PMSG和HCG,4只雌鼠不合笼,用颈椎脱臼法处死,剪取子宫组织(孕D0,对照组),直接液氮保存和福尔马林固定保存,其余的雌:雄小鼠1∶1合笼,次日检查阴栓,发现阴栓计为妊娠第1天.分别于妊娠第4、5、6、7和8天用颈椎脱臼法处死,迅速剖腹,剪取子宫组织予以保存(实验组),采用免疫组化方法和Western-blotting技术分析围植入期小鼠子宫内膜中的整合素β3表达规律,采用RT-PCR技术半定量分析围植入期小鼠子宫内膜整合素β3 mRNA表达特征.结果 小鼠整个围植入期子宫内膜中均有整合素β3表达,但表达的高峰在着床窗口期,即孕D5.孕D0(非孕)子宫内膜的整合素β3表达较弱,孕D4时表达增强,孕D5时表达最强,孕D6时表达突然回落,孕D7、孕D8时表达减弱,降至非孕水平.结论 整合素β3在围植入期小鼠子宫内膜中的表达随着孕龄的增加逐渐增强,但表达的高峰在孕D5,与植入窗的开放相吻合,是子宫内膜接受性的客观标志,提示整合素β3在胚泡着床过程中起重要作用,该研究为阐明胚泡着床机制提供实验依据.     

骨桥蛋白(OPN)在围植入期小鼠子宫内膜中的表达

目的 检测骨桥蛋白在围植入期小鼠子宫内膜中的表达,明确OPN表达规律.方法 将清洁级性成熟期昆明小鼠44只,雌性24只,雄性20只,雌性小鼠阴道涂片检查为动情前期,在此期注射PMSG和HCG,4只雌鼠不合笼(孕D0,对照组),用颈椎脱臼法处死,剪取子宫组织,直接液氮保存和福尔马林固定保存,其余的雌:雄小鼠1∶1合笼,次日检查阴栓,发现阴栓计为妊娠第1天.分别于妊娠第4、5、6、7和8天用颈椎脱臼法处死,迅速剖腹,剪取子宫组织予以保存(实验组),采用免疫组化方法和Western-blotting技术分析围植入期小鼠子宫内膜中的骨桥蛋白表达规律,采用RT-PCR技术半定量分析围植入期小鼠子宫内膜骨桥蛋白mRNA表达特征.结果 小鼠整个围植入期子宫内膜中均有OPN表达,但表达的高峰在着床窗口期,即孕D5.孕D0(非孕)子宫内膜的OPN表达较弱,孕D4时表达增强,孕D5时表达最强,孕D6时表达突然回落,孕D7、孕D8时表达减弱,降至非孕水平.结论 骨桥蛋白在围植入期小鼠子宫内膜中的表达随着孕龄的增加逐渐增强,但表达的高峰在孕D5,与植入窗的开放相吻合,是子宫内膜接受性的客观标志,提示骨桥蛋白在胚泡着床过程中起重要作用,该研究为阐明胚泡着床机制提供了实验依据.     

PTEN基因在小鼠胚胎着床过程中作用的实验研究

目的探讨PTEN基因小鼠胚胎着床过程中的作用。方法实时荧光定量聚合酶链反应(real-time fluores-cence quantitativePCR,FQ-PCR)分别检测未孕及孕1、3、4、5、7d(分别记为d0、d1、d3、d4、d5、d7)小鼠子宫内膜PTEN基因mRNA的表达和子宫角注射反义寡核苷酸观察胚泡着床数。结果FQ-PCR测得妊娠的子宫内膜组织PTEN的表达高于未妊娠的子宫内膜组织,且随着妊娠天数的增加呈现逐渐增加的趋势,到妊娠第5天达到最高。子宫角注射PTEN反义寡核苷酸后胚泡着床数明显减少。结论PTEN在妊娠早期子宫内膜持续表达,可能参与了胚泡着床。     

TACE在着床窗口期小鼠子宫内膜上皮细胞中的表达及其抗体对胚泡粘附的影响

目的:通过检测肿瘤坏死因子α转换酶(Tumor necrosis factor α convening enzyme,TACE)在小鼠子宫内膜上皮细胞中的表达及其在体外着床模型中对胚泡粘附的影响,初步探讨TACE在小鼠生殖中的作用以及其与胚胎着床的关系.方法:体外培养孕4d(d4)小鼠子宫内膜上皮细胞和胚泡,构建胚泡体外着床模型;通过细胞免疫组化鉴定小鼠子宫内膜上皮细胞;运用RT-PCR和细胞免疫组化分别检测小鼠子宫内膜上皮细胞中TACE mRNA和蛋白的表达;向体外着床模型中添加不同浓度的TACE抗体,观察对胚泡体外着床的影响,并用细胞免疫化学检测抗体干预后各培养体系中TACE蛋白质的表达情况.结果:原代培养小鼠子宫内膜上皮细胞生长旺盛,鉴定结果显示:上皮细胞对角蛋白抗体有棕黄色阳性表达,对波形蛋白抗体为阴性表达.PCR结果显示TACE mRNA在共培养体系中有高表达,免疫组化结果与RT-PCR结果相一致;小鼠胚泡体外共培养具有较高的粘附率,添加TACE抗体后胚泡帖附率均显著低于对照组帖附率(P<0.05),且随着抗体浓度的降低胚泡粘附率随之升高,免疫化学检测结果也显示随着抗体浓度的降低TACE的表达量随之升高.结论:TACE可能参与了胚泡着床过程,有利于小鼠胚胎粘附、扩展,为胚胎的着床做准备.     

小鼠子宫内膜胚泡着床点及旁组织白血病抑制因子基因表达研究

目的:探讨白血病抑制因子(LIF)基因在小鼠胚泡着床窗口期了宫内膜表达的特定部位.方法:采用半定量逆转录聚合酶链反应(RT-PCR),对20只孕期第4天的小鼠子宫内膜着床点和旁组织的LIF mRNA进行检测.结果:20只孕期第4天的小鼠着床点LIF mRNA表达平均水平为0.5316,旁组织LIF mRNA表达平均水平为0.3711.统计处理t检验差异具有显著性(P＜0.05).结论:在胚泡着床窗口期,小鼠子宫内膜胚泡着床点和旁组织LIF基因表达存在差异.     

GRB7基因在早孕小鼠子宫内膜中的表达

目的：研究生长因子结合受体7（growth factor receptor bound 7,GRB7）在早孕小鼠子宫内膜中的表达规律,探讨其在胚胎着床过程中的作用。方法：分别应用实时荧光定量PCR、免疫组化和Western blot方法检测未妊娠及妊娠早期d1～7小鼠子宫内膜GRB7基因及蛋白的表达;孕3 d下午8点子宫角注射GRB7反义寡核苷酸,孕7 d观察胚泡着床数较正常组有无变化。结果：实时荧光定量PCR测得未妊娠（d0）和妊娠早期（d1～7）小鼠子宫内膜组织均有GRB7 mRNA表达,并在孕5 d达到峰值;免疫组化及Western blot分析显示GRB7蛋白在子宫内膜的表达规律与实时荧光定量PCR基本一致,孕1 d到孕4 d,GRB7蛋白主要表达部位为腔上皮、腺上皮,基质细胞少量表达,孕5 d、孕6 d,GRB7蛋白表达较之前增强,并且在基质细胞开始明显表达,孕5 d最强;孕6 d着床点GRB7蛋白高表达于初级蜕膜区,着床旁GRB7蛋白高表达于基质细胞;子宫角注射GRB7反义寡核苷酸后胚泡着床数较正常组明显减少。结论：GRB7在早孕小鼠子宫内膜中呈动态表达,提示它参与了胚胎着床的发育过程。     

健胎液对模型小鼠着床期子宫内膜容受性的影响

目的:研究健胎液对胚泡着床障碍小鼠着床期子宫内膜容受性的影响。方法:于妊娠第4天9∶00利用米非司酮建立胚泡着床障碍小鼠模型,予健胎液进行干预,于妊娠第4天21∶00处死小鼠,观察小鼠胚泡着床率及平均着床胚泡数,采用放免法检测血清及子宫组织雌二醇、孕酮的含量,采用逆转录聚合酶链式反应(RT-PCR)检测子宫ERmRNA、PRmRNA表达。结果:与模型组相比,中药组着床率和平均着床胚胎数均显著升高,与对照组无显著差异;模型组子宫内膜发育不良,中药组子宫内膜的发育与正常组相似;3组血清及子宫组织的雌二醇和孕酮的含量无显著改变;中药组子宫ER mRNA、PR mRNA表达均较模型组显著提高,与正常组比较差异无显著性。结论:健胎液通过促进子宫内膜发育,调节胚泡着床障碍小鼠子宫组织ER、PR基因的表达,改善子宫内膜容受性,利于胚泡着床。     

瘦素及其受体系统在小鼠胚泡着床过程中子宫内膜的表达

目的:探讨瘦素(leptin)及其受体(Ob-Rb)在小鼠胚泡着床过程中子宫内膜的表达规律,以明确其在胚泡着床过程中的可能作用.方法:采用半定量逆转录聚合酶链反应(RT-PCR),检测小鼠胚泡着床过程中子宫内膜leptin及其受体(Ob-Rb)的表达.结果:leptin在胚泡着床过程中在小鼠子宫内膜也都有表达,各时期的表达水平差异不明显(P＞0.05);Ob-Rb在胚泡着床过程中在小鼠子宫内膜也都有表达,各时期的表达水平差异(P＜0.01),且在孕期第4天表达水平最高,在5到7天仍然维持在一定水平.结论:瘦素及其受体(Ob-Rb)可能在胚泡着床过程中发挥作用,同时也可能参与着床后胚胎的生长发育.     

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.