基因沉默整合素连接激酶对滋养细胞生物学功能的影响

目的：研究基因沉默整合素连接激酶（ILK）对人滋养细胞TEV-1细胞增殖、迁移和侵袭等的影响。方法：构建ILK-siRNA慢病毒载体后,转染人TEV-1细胞,分ILK基因沉默组（KD组）和空白质粒转染组（NC组）进行;qRT-PCR和Western blot检测mRNA和蛋白表达,MTT法、Transwell法和划痕实验等检测细胞的增殖、迁移和侵袭能力。结果：基因沉默ILK后,TEV-1细胞的ILK基因和蛋白的表达水平显著降低（P＜0.05）,证明干扰ILK表达成功。沉默ILK后,TEV-1细胞的增殖能力、侵袭能力、迁移能力均显著下降（P＜0.05或P＜0.01)。ILK-siRNA转染TEV-1细胞后,MMP-9和p-GSK3β mRNA和蛋白水平显著下降（P＜0.01）。结论：沉默ILK通过下调GSK3β和MMP-9的表达抑制人滋养细胞的增殖、迁移和侵袭。     

siRNA沉默PTP4A1表达对人舌鳞癌细胞系TCA8113体内外生物学行为的影响

目的 探讨特异性沉默人舌鳞癌细胞系TCA8113中PTP4A1基因的表达,并研究沉默该基因表达后对TCA8113细胞体内外生物学行为的影响.方法 采用siRNA的慢病毒载体siRNA-PTP4A1作为阳性干扰组(KD),空病毒载体作为阴性对照组(NC),未作任何处理的TCA8113细胞作为空白对照组(CON).Real-time PCR和Westem blot检测PTP4A1基因的干扰效果,MTT法检测3组细胞的增殖能力,流式细胞技术分析3组细胞凋亡和细胞周期情况,平板克隆形成实验检测体外克隆形成能力,Western blot分别检测3组细胞抑制凋亡蛋白Bcl-2、促凋亡蛋白Bax的表达情况,3组细胞分别裸鼠成瘤后观察移植瘤的生长情况.结果 siRNA-PTPA1转染TCA8113细胞成功,Real-time PCR和Western blot 检测PTP4A1基因的抑制率达到55％和60％以上(P ＜0.05);MTT结果显示KD组细胞增殖能力小于NC、CON组;细胞凋亡率也明显增加,KD组早期凋亡为(17.48 ±0.70)％,而NC、CON组的早期细胞凋亡率为(7.34±0.70)％和(4.86±0.25)％ (P ＜0.01);KD组细胞周期阻滞在G1/G0和S期(P＜0.05);Western blot结果显示Bcl-2蛋白表达降低(P＜0.01),Bax蛋白表达增高(P＜0.01);KD组体外克隆形成数小于NC、CON组(P＜0.01);KD组裸鼠移植瘤的生长速度缓慢,质量和体积均小于NC、CON组(P＜0.0l).结论 siRNA PTP4A1在体内外均能有效抑制舌鳞癌TCA8113细胞的恶性生物学行为,PTP4A1基因可能与舌鳞癌的发生、发展相关.     

特异性ILK siRNA对膀胱癌移植瘤生长及p-Akt、p-GSK3β表达的影响

目的 探讨应用整合素连接激酶（integrin-linked kinase,ILK）特异siRNA沉默ILK基因的表达对膀胱癌裸鼠移植瘤生长及p-Akt、p-GSK3β表达的影响。方法实验分3组：BIU-87细胞组（正常BIU-87细胞）、BIU-87-NC组（转染阴性对照质粒的BIU-87细胞）、BIU-87 siR...     

慢病毒介导shRNA特异性沉默促肝细胞再生磷酸酶1基因表达抑制舌癌细胞迁移侵袭能力

目的：探讨特异性沉默促肝细胞再生磷酸酶1（phosphatase of regenerating liver cell-1,PRL-1）基因表达对舌癌细胞迁移、侵袭能力的影响。方法：设计、合成5条针对PRL-1基因的短发夹RNA（short hairpin RNA,shRNA）干扰序列,构建于慢病毒载体质粒中,PCR电泳及基因测序验证。转染293T细胞,Western blot筛选最佳干扰序列,包装产生慢病毒。转染TCA8113细胞,筛选、建立稳定转染细胞株;real-time PCR检测PRL-1基因沉默效率;划痕实验和Transwell实验分别比较慢病毒转染细胞（KD组）、空病毒转染细胞（NC组）及TCA8113细胞（CON组）迁移、侵袭能力变化。结果：PCR电泳及基因测序证实5条shRNA序列定向插入慢病毒载体质粒中。Western blot筛选出PRL-1-shRNA-1基因沉默效率最高,包装获得慢病毒滴度为 7×108 TU/ml。通过转染、筛选,建立了TCA8113细胞稳定转染细胞株;real-time PCR测得PRL-1基因mRNA表达明显下降（F=809.120,P=0.000）;PRL-1基因沉默后细胞迁移能力降低,穿过人工基底膜的细胞数KD组（25.5±0.4）明显少于NC组（81.5±2.0）和CON组（88.5±2.3）（F=1 092.970,P=0.000）。结论：沉默PRL-1基因表达能有效抑制舌癌细胞迁移、侵袭能力。     

整合素连接激酶在子宫内膜癌组织中的表达及其对细胞迁移和侵袭能力的影响

目的：观察子宫内膜癌患者癌组织中整合素连接激酶（ILK）的表达水平,通过体外细胞实验探讨其对子宫内膜癌细胞迁移和侵袭能力的影响,并阐明其作用机制。方法：采用免疫组织化学SP法检测29例子宫内膜癌患者癌组织和癌旁组织细胞中ILK阳性表达率。分别将ILK siRNA和siRNA control转染至子宫内膜癌HEC-1B细胞作为干扰组和空载组,以不作处理的细胞作为对照组;采用Western blotting法检测HEC-1B细胞中ILK、蛋白激酶B（Akt）、磷酸化Akt（p-Akt）、糖原合成酶激酶3β（GSK-3β）和磷酸化GSK-3β（p-GSK-3β）蛋白表达水平,细胞划痕实验检测细胞迁移距离,Transwell小室法检测侵袭细胞数。结果：29例子宫内膜癌患者癌组织中ILK阳性表达率为79.3%,癌旁组织中ILK阳性表达率为10.3%,子宫内膜癌组织中ILK阳性表达率明显高于癌旁组织（P<0.05）。与对照组和空载组比较,干扰组HEC-1B细胞中ILK、p-Akt和p-GSK-3β蛋白表达水平明显降低（P<0.05）,Akt和GSK-3β蛋白表达水平差异无统计学意义（P>0.05）;细胞划痕实验,与对照组和空载组比较,干扰组HEC-1B细胞迁移距离明显缩短（P<0.05）,侵袭细胞数明显减少（P<0.05）。结论：ILK在子宫内膜癌组织中呈高水平表达,干扰ILK表达能够降低子宫内膜癌细胞的迁移和侵袭能力,其作用机制可能与其抑制Akt和GSK-3β蛋白磷酸化有关。     

异土木香内酯对人舌鳞癌Tca8113细胞的增殖、迁移和侵袭的影响

目的 研究异土木香内酯对舌鳞癌细胞Tca8113的增殖、迁移和侵袭能力的影响.方法 用CCK-8法检测异土木香内酯作用于舌鳞癌Tca8113细胞的半数抑制浓度(IC50),EDU实验方法检测异土木香内酯抑制Tca8113细胞增值能力.检测土木香内酯对Tca8113细胞凋亡影响实验.Western Blot检测异土木香内酯对Tca8113细胞凋亡蛋白表达的影响.结果 实验结果表明,异土木香内酯作用于舌鳞癌Tca8113细胞的半数抑制浓度(IC50)为20μM,20μM异土木香内酯可以抑制舌鳞癌Tca8113细胞活性(P<0.01),细胞增值能力(P<0.01);进一步结果表明,异土木香内酯还可以抑制对Tca8113 P迁移,同时降低Tca8113细胞侵袭能力(P<0.01);促进Tca8113细胞凋亡的影响(P<0.01).Western Blot实验结果表明,异土木香内酯可以下调舌癌Tca8113细胞的Bcl-2蛋白,上调caspase3蛋白的表达.结论 实验结果表明异土木香内酯对舌癌Tca8113细胞有抑制作用.     

增强DNA聚合酶beta基因表达对人舌癌细胞生物学行为的影响

目的探讨DNA聚合酶beta（polβ）对人舌癌Tca8113细胞增殖和侵袭力的影响。方法运用野生型人polβ重组绿色荧光蛋白表达载体pEGFP-C3-polβ转染Tca8113细胞,分别用RT-PCR、Western blot检测polβ转录及蛋白表达水平;细胞生长实验检测舌癌细胞增殖的变化;Boyden室细胞侵袭实验检测舌癌细胞侵袭能力的变化。结果 pEGFP-C3-polβ转染Tca8113细胞获得稳定细胞株,与空白对照组及空载体组比较,polβmRNA及蛋白表达水平均明显增高,野生型polβ导入的舌癌细胞增殖和侵袭能力均明显减弱（P〈0.05）。结论 DNA polβ表达增强能够降低人舌癌细胞的增殖和侵袭能力。     

干扰素调节因子1对舌鳞癌细胞增殖侵袭和迁移的影响

目的 研究干扰素调节因子对舌鳞癌细胞的侵袭和迁移的影响.方法 采用实时定量PCR和免疫组化检测IRF1在舌鳞癌组织中的表达水平,通过构建IRF1的过表达和IRF1沉默的质粒,在舌鳞癌细胞Tca8113中过表达IRF1或降低IRF1的表达水平后,检测其对Tca8113细胞增殖,侵袭和迁移的影响.结果 IRF1在舌鳞癌组织...     

FAK基因沉默促进舌癌细胞失巢凋亡及抑制转移的实验研究

目的:研究黏着斑激酶(FAK)基因沉默是否可促进舌癌细胞系Tca8113失巢凋亡,进而逆转其失巢凋亡抗性,抑制舌癌细胞侵袭转移。方法:以舌癌细胞株Tca8113为研究对象,首先利用RNA干扰方法抑制FAK基因表达。采用poly-HEMA诱导悬浮培养,进而采用流式细胞仪观察体外培养过程中,FAK基因沉默对舌癌细胞株Tca8113失巢凋亡抗性的影响。最后以划痕实验、Transwell小室侵袭模型研究舌癌细胞迁移及侵袭能力。结果 :在体外培养过程中,Tca8113细胞具有很强的失巢凋亡抗性。针对FAK的siRNA可显著下调FAK基因的表达。当FAK基因下调后,Tca8113细胞失巢凋亡抗性显著下降(P<0.01),并且细胞的侵袭及迁移能力明显下降(P<0.01)。结论:FAK基因沉默可促进失巢凋亡,逆转舌癌细胞失巢凋亡抗性,进而抑制其转移,是一个潜在的抗舌癌转移分子治疗的靶点。     

斑蝥素抑制舌癌细胞生长及转移能力

摘 要:【目的】 观察斑蝥素(Cantharidin)对舌癌细胞株Tca8113?CAL-27生长及转移能力的抑制作用?【方法】 舌癌细胞株Tca8113?CAL-27处理组采用斑蝥素处理,通过噻唑蓝(MTT)比色法检测细胞生长能力;通过流式细胞术检测细胞的细胞周期分布;通过实时定量PCR和Western-blot检测p21?细胞周期蛋白依赖激酶1(CDK1)?基质金属蛋白酶-2?9(MMP-2,9)基因表达的变化;通过Transwell小室及划痕实验检测斑蝥素对细胞侵袭和迁移能力的影响?【结果】 斑蝥素抑制舌癌细胞生长,呈剂量和时间依赖性(P﹤0.01)?斑蝥素处理后G2/M期细胞比例上升(P﹤0.01),呈G2/M期阻滞?斑蝥素处理后,抑制G2/M周期转换的p21表达上调(P﹤0.01),而促进G2/M周期转换的CDK1表达下调(P﹤0.01)?斑蝥素可显著抑制舌癌细胞的侵袭能力,处理24 h使Tca8113?CAL-27细胞侵袭力分别下降(93.50 ± 3.32)%(P﹤0.01),(61.33 ± 4.11)%(P﹤0.01)?斑蝥素处理后MMP-2?9 mRNA表达量显著降低?划痕实验显示斑蝥素可显著抑制舌癌细胞的迁移能力(P﹤0.01),处理12 h使Tca8113?CAL-27细胞迁移能力分别下降(57.60 ± 8.98)%(P﹤0.01),(49.94 ± 5.63)%(P﹤0.01)?【结论】 斑蝥素通过阻滞细胞周期抑制舌癌细胞的生长,并通过抑制舌癌细胞迁移和对基质的降解,抑制舌癌细胞转移?     

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.