胃腺癌及正常胃黏膜组织蛋白质组双向电泳图谱分析

目的:对胃腺癌与配对正常胃黏膜双向电泳(2-DE)图谱进行比较分析,筛选差异表达蛋白质.方法:采用2-DE技术分离胃腺癌和正常胃黏膜组织总蛋白.银染后采用magicscan图像获取仪扫描2-DE凝胶图像,采用Imagemaster6.0凝胶图像分析软件进行分析.结果:胃腺癌和正常胃黏膜2-DE图谱上蛋白质斑点主要分布在等电点p14～7、相对分子质量20 000～90 000范围内,共有28个差异表达蛋白质,其中15个在胃腺癌组织中表达上调,13个表达下调.结论:胃腺癌与正常胃黏膜组织蛋白质组表达有差异.     

人宫颈癌组织及癌旁正常组织蛋白质组的比较研究

目的:构建人宫颈癌组织及癌旁组织双向凝胶电泳图谱,筛选差异表达的蛋白质.方法: 人宫颈癌组织及癌旁正常组织标本10例,应用固相pH梯度双向凝胶电泳后,ImageScanner扫描图像、ImageMaster 2-DE Elite4.01软件分析识别差异表达的蛋白质;应用质谱仪得到相应的肽质指纹图谱,联网ExPASY网站查询相关数据库鉴定获得的差异蛋白质点.结果: 癌组织和癌旁正常组织凝胶的平均蛋白质点数分别为1285±85和1063±76,凝胶上蛋白质点位置上有较好的重复性,不同胶间蛋白质点在等电聚焦方向的偏差是(1.78±0.18) mm,在SDS-PAGE方向上的偏差为(1.89±0.21) mm;比较10例宫颈癌组织及癌旁正常组织的双向凝胶电泳图谱,未匹配总点数为156±18,对31个差异蛋白质点进行肽质指纹图分析,初步鉴定了8个与肿瘤基因、细胞周期调控、信号转导等有关的蛋白质.结论: 建立了分辨率高且重复性较好的人宫颈癌组织与癌旁正常组织的双向凝胶电泳图谱,并成功鉴定出部分与宫颈癌癌变相关的差异表达的蛋白质,为筛选宫颈癌特异性的诊断分子标志物奠定了的基础.     

人支气管上皮蛋白质样品制备方法及其癌变各阶段组织2-DE图谱的建立

目的： 改良、优化支气管上皮组织的蛋白质样品制备方法,建立人支气管上皮癌变过程各阶段组织的2-DE图谱,为识别鉴定肺鳞癌癌变相关蛋白质奠定基础。方法：收集、筛选人支气管正常上皮、鳞状化生、不典型增生和上皮浸润癌组织标本。改良的脱氧胆酸-三氯醋酸(deoxycholate-trichloroaetic acid,DOC-TCA)法提纯支气管上皮总蛋白质,应用固相pH梯度双向凝胶电泳分离各阶段组织的总蛋白质,凝胶经银染显色后,用ImageMaster 2D软件分析双向电泳图谱。结果：应用改良的DOC-TCA法提纯的支气管上皮总蛋白质进行双向电泳,可获得分辨率高、重复性好的人支气管上皮组织的双向电泳图谱。正常上皮、鳞状化生、不典型增生和浸润癌4种组织凝胶的蛋白质点数依次为1 190±63,1 227±69,1 272±71,1 326±82。选取同例鳞状上皮化生组织进行3次重复性检测,3块凝胶的平均蛋白质点数为1 216 ±75 ,平均匹配点数为1 082±67,匹配率达 89.3% , 且3块胶在蛋白质点位置上有较好的重复性,不同胶间蛋白质点在IEF方向的偏差为（0.835±0.247） mm ,在SDS-PAGE方向上的偏差为（0.921±0.104） mm。结论：改良的DOC-TCA沉淀法是一种较好的支气管上皮组织蛋白质样品制备法,初步建立了分辨率较高且重复性好的人支气管上皮癌变各阶段组织的双向凝胶电泳图谱。     

双向电泳分析失神经支配大鼠腓肠肌蛋白的表达变化

目的:建立骨骼肌蛋白分离的双向电泳(2-DE)技术体系,寻找失神经支配骨骼肌与正常骨骼肌蛋白的表达差异。方法:建立失神经腓肠肌模型,提取腓肠肌总蛋白,进行第一向等电聚焦(IEF)和第二向十二烷基硫酸钠聚丙烯酰胺凝胶电泳(SDS-PAGE)技术分离总蛋白,使用PDQuest软件对凝胶图像进行分析,并测量了凝胶蛋白斑点在IEF和SDS-PAGE方向上的位置偏差。结果:失神经组和正常组腓肠肌组织3块凝胶的平均蛋白质点数分别为560±16和545±13,平均匹配点数分别为471±19和447±17,匹配率达84.1%和82.1%。对比分析了两种腓肠肌组织的双向电泳图谱发现,平均匹配点数为445±8;发现在失神经支配后有80个点发生了明显而稳定的质和量(大于10倍或小于0.1倍)的改变。不同凝胶间蛋白质点在IEF方向的偏差为(1.203±0.763)mm,在SDS-PAGE方向上的偏差为(1.062±0.529)mm。结论:失神经组和正常组腓肠肌的双向电泳图谱存在明显差异,失神经后表达下调的蛋白可能与维持肌肉正常功能相关,而表达上调的蛋白则可能与肌萎缩的发生过程相关。     

侵袭性与非侵袭性垂体腺瘤的蛋白质差异表达谱的建立

目的：建立分辨率高和重复性好的侵袭性与非侵袭性垂体腺瘤的双向凝胶电泳图谱 ,并识别鉴定出其差异表达的蛋白质。方法： 利用固相 pH梯度双向凝胶电泳分离侵袭性与非侵袭性垂体腺瘤的总蛋白质,凝胶经银染显色后,运用ImageMaster 2D图像分析软件进行比较分析、识别差异表达的蛋白质,应用基质辅助激光解吸电离飞行时间质谱 (MALDI-TOF-MS)及数据库搜索鉴定部分差异蛋白质点。结果：得到了分辨率较高、重复性较好的侵袭性与非侵袭性垂体腺瘤的双向凝胶电泳图谱,将同一侵袭性垂体腺瘤组织和非侵袭性肿瘤组织分别进行了3次双向凝胶电泳,3块凝胶的蛋白质点数分别为1 080±24和1 035±28,匹配点数分别为975±45和918±56,匹配率分别达90.3％和88.7％。同一组织的3块胶在蛋白质点位置上有较好的重复性 ,不同胶间蛋白质点在等电聚焦(IEF)方向的偏差是(1.563±0.259) mm,在十二烷基磺酸钠聚丙烯酰胺凝胶电泳(SDS-PAGE)方向上的偏差为(1.088±0.206) mm。比较分析侵袭性垂体腺瘤和非侵袭性垂体腺瘤凝的双向凝胶电泳图谱,发现两者间的99个表达有明显差异的蛋白质点。初步对其中30个差异蛋白质点进行了肽质量指纹图分析,鉴定出一些与细胞周期调控、信号转导等有关的蛋白质。 结论：建立了分辨率较高且重复性较好的侵袭性与非侵袭性垂体腺瘤的双向凝胶电泳图谱,并识别鉴定出一些侵袭性与非侵袭性垂体腺瘤之间差异表达的蛋白质,为进一步筛选侵袭性垂体腺瘤的蛋白质表达数据库提供了基础。     

RASSF1A表达或缺失的两类早期肺腺癌组织差显蛋白的筛选与鉴定

目的:建立RASSFlA表达或表达缺失的两类不同早期肺腺癌组织蛋白质的双向电泳凝胶图谱.筛选并鉴定存在明显表达差异的蛋白质.方法:采用Western印迹技术,从RASSFlA转录缺失和RASSFIA转录正常的早期肺腺癌标本中筛选出RASSFlA表达和表达沉默的癌组织各5例.提取组织可溶性总蛋白,采用固相pH梯度双向凝胶电泳技术分离总蛋白,建立两类不同早期肺腺癌组织蛋白质的双向电泳凝胶图谱;PDQuest凝胶图像分析软件比较分析,筛选出差异表达的蛋白质点;基质辅助激光解吸电离飞行时间质谱获得相应蛋白质点的肽质量指纹图谱;搜索蛋白质数据库鉴定差异表达的蛋白质.结果:建立了重复性较好的RASSFIA表达或表达缺失的两类不同早期肺腺癌组织蛋白质双向电泳凝胶图谱;筛选出存在明显表达差异的蛋白质点17个,挖取其中的9个进行质谱分析,9个蛋白质点均得到了满意的肽质量指纹图谱;搜索蛋白质数据库鉴定出5种蛋白质,分别是:细胞色素b5(cytochrome b5),60S磷酸核楮体蛋白P2(60S acidic ribosomal protein P2),碳酸酐酶1(carbonic anhydrase-1),5-吡咯啉羧酸还原酶1(pyrroline-5-carboxylate reductase-1)和载脂蛋白A-I前体蛋白(apolipoprorein A-I precursor).结论:成功建立了RASSFlA表达或表达缺失的两类不同早期肺腺癌组织蛋白质的双向电泳凝胶图谱,从中鉴定出5种存在明显表达差异的蛋白质,为进一步研究RASSFIA作用的信号转导途径奠定了初步的基础.     

肺癌组织蛋白质组双向凝胶电泳方法的建立和优化

目的建立和优化肺癌组织蛋白质组双向凝胶电泳(2-DE)技术,获取高分辨率、重复性好的蛋白质2-DE图谱。方法提取肺癌组织蛋白,以固相pH梯度胶条做第一向等电聚焦电泳,十二烷基磺酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)为第二向,在上述过程中分别对蛋白提取、等电聚焦电泳和凝胶染色进行控制和优化,利用ImageMaster 2Dplatinum 6.0分析软件获得二维凝胶图像。结果实验重复3次,共获15幅二维凝胶图像,平均蛋白质点数为1 137±57;随机选取1例标本的2幅图像,利用软件对2幅图像中蛋白点进行匹配,匹配率为90.4%。结论优化肺癌组织蛋白质组2-DE技术可获得高分辨率、重复性好的蛋白质2-DE图谱,为开展肺癌组织蛋白质组学研究打下基础。     

盘状红斑狼疮皮损角质形成细胞的蛋白组学初步分析

 【目的】建立盘状红斑狼疮皮损和正常皮肤角质形成细胞的双向电泳图谱，分析二者蛋白质的表达差异及特点。【方法】运用固相pH梯度双向凝胶电泳分离6例盘状红斑狼疮患者皮损及正常皮肤角质形成细胞总蛋白，考马斯亮蓝染色后用ImageMaster 2D图象软件比较分析，识别差异蛋白。【结果】盘状红斑狼疮皮损和正常对照电泳图谱平均蛋白点分别为1566±32和1682±38，平均匹配点数为1286±14 和1314±21，匹配率为82.12%和78.12%。分辨差异蛋白点共22个，11个在皮损高表达，2个在皮损组低表达，有5个点仅在病例组表达，4个点仅在正常对照组表达。【结论】成功获得分辨率高且重复性较好的蛋白质组双向电泳图谱，识别重复表达的差异蛋白，为后续蛋白功能研究奠定基础。     

先天性巨结肠组织蛋白质组成分的双向凝胶电泳分析

目的 分析先天性巨结肠(HD)狭窄段及正常段肠管组织双向凝胶电泳的蛋白质表达图谱,寻找差异表达的蛋白质.方法 采用固相pH梯度双向聚丙烯酰胺凝胶电泳(2-DE)分离20例HD患者配对狭窄段及正常段肠管组织的总蛋白,银染显色,ImageMaster 2D Platinum 6.0软件分析差异表达的蛋白质点.结果 获得了重复性较好的2-DE银染图谱.图像分析显示,狭窄段和正常段肠管组织凝胶的平均匹配率分别为78.1%和86.7%.差异分析共获得103个差异表达的蛋白质点.结论 采用2-DE分离HD肠管组织的总蛋白可获得重复性较好的结果.获得的差异蛋白质点为寻找HD早期诊断的分子标记物提供了基础资料.     

鼻息肉的2-DE图谱建立和蛋白质组学分析

目的:建立鼻息肉及鼻黏膜双向凝胶电泳(two-dimensional polyacrylamide gel electrophoresis,2-DE)图谱,鉴定差异表达蛋白质。方法:收集鼻息肉及鼻黏膜样本各7例,采用固相pH梯度2-DE,凝胶银染,扫描图像,ImageMaster2-DE软件比较分析等方法,识别差异表达蛋白质,通过质谱分析得到相应肽质指纹图谱(peptide mass fingerprint,PMF),采用PeptIdent软件查询Swiss-Protand TreMBL数据库,鉴定差异表达蛋白质。结果:建立了鼻息肉和鼻黏膜蛋白质的2-DE图谱。鼻息肉和鼻黏膜3块凝胶的平均蛋白质点数分别为825±78和936±62;平均匹配点数为682±96和821±78,匹配百分率为82.7%和87.7%;同一鼻息肉的3块胶在蛋白质点位置上有较好的重复性,不同胶间蛋白质点在等电聚焦(IEF)方向偏差为(1.06±0.14)mm,在SDS-PAGE方向偏差为(1.45±0.21)mm。比较分析两种组织各7例样本的2-DE图谱,鼻息肉和鼻黏膜蛋白质点数为1458个和1617个,平均匹配点数为1026个。质谱分析差异蛋白质点40个,获取34张PMF,查询数据库鉴定出蛋白质24个。结论:建立了分辨率高和重复性好的鼻息肉及鼻黏膜2-DE图谱,识别鉴定出一些与鼻息肉病变相关的蛋白质。     

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.