左旋精氨酸/一氧化氮通路对小鼠胰岛β细胞凋亡的影响

目的研究左旋精氨酸／一氧化氮通路对胰岛β细胞凋亡的影响，初步探讨一氧化氮（NO）诱导胰岛β细胞凋亡的作用。方法体外培养的NIT-1小鼠胰岛β细胞，用不同浓度的L-精氨酸（L-arg）处理，6h后采用Griess化学显色法检测细胞培养液内的NO水平，Annexin-V／PI双染流式细胞术检测细胞的凋亡。而后联合应用hemoglobin处理细胞，分为3组：对照组；L-arg组；L-arg＋hemoglobin组；6h后检测NO的水平和细胞的凋亡，并用Western Blot检测P53蛋白的表达。结果与对照组比较，L-arg 4mg／mL处理组的NO水平最高（P〈0．05），细胞凋亡最明显（P〈0．05）；hemoglobin干预后，NO水平和细胞凋亡率均较L-arg单独处理组明显降低（P〈0．05），与正常对照组之间差异没有显著性（P〉0．05）；Western Blot提示L—arg组的P53蛋白的表达水平明显高于对照组和hemoglobin处理组（P〈0．05）。结论L-arg／NO通路可诱导NIT-1小鼠胰岛β细胞凋亡，并呈现一定程度的剂量依赖性；NO可能通过P53的活化诱导NIT-1胰岛细胞凋亡。     

大黄素对化疗药诱导肝癌细胞凋亡的增强作用

目的 观察大黄素对化疗药5-氟尿嘧啶（5-Fu）诱导肝癌细胞HepG2凋亡的影响。方法 不同浓度大黄素单用或与5-Fu联合作用HepG2细胞，以流式细胞仪（FCM）及末端脱氧核苷酸转移酶介导的dUTP缺口末端标记（TUNEL）法检测细胞凋亡；免疫组织化学（sP）法检测Bax、Bcl-2、凋亡诱导因子（AIF）蛋白表达的改变。结果 2、4μg／m1大黄素分别与5-Fu联用，作用细胞16、24、48h后，时间依赖性地诱导HepG2细胞凋亡，且随大黄素浓度的增加细胞凋亡率有增加的趋势。联用组细胞凋亡率及Bax、Bcl-2、AIF蛋白表达与5-Fu单用组相比差异均有极显著性意义（均P〈0．01）。各组细胞凋亡率与Bax蛋白表达呈正相关（r=0．881，P〈0．05），与Bcl-2蛋白表达呈负相关（r=-0．942，P〈0．01），与AIF蛋白表达呈正相关（r=0．900，P〈0．05）。结论 大黄素可显著增强5-Fu诱导HepG2细胞凋亡的作用，可能与调控Bax、Bcl-2、AIF蛋白表达有关。     

大黄素联合5-氟尿嘧啶对人胃癌细胞MKN45增殖抑制及诱导凋亡

目的研究大黄素联合5-氟尿嘧啶（5-Fu）对人胃癌细胞MKN45增殖及诱导凋亡的作用。方法不同浓度大黄素单用或与5-Fu联合作用于人胃癌MKN45细胞,甲氮唑蓝法观察对MKN45的生长抑制作用;荧光染色法、流式细胞仪分析MKN45的凋亡。结果10、20μg／mL大黄素分别与5-Fu联用对人胃癌MKN45细胞有明显的生长抑制作用,其抑制作用呈时间及浓度依赖性（P〈0．05）;流式细胞仪分析大黄素联合5-Fu能浓度依赖性诱导MKN45细胞凋亡和G2／M期阻滞,大黄素与5-Fu联用细胞凋亡率与5-Fu单用组相比差异有统计学意义（P〈0．051;吖啶橙染色观察联合作用组细胞核内可见浓染致密的颗粒荧光、新月型改变、核固缩或片段化及凋亡小体的形成。结论大黄素联合5-Fu可显著增强诱导人胃癌MKN45细胞凋亡的作用。     

吡格列酮对炎性因子所致胰岛细胞凋亡的保护作用

目的：探讨Ppar-γ激动剂吡格列酮（PGZ）对炎性因子所诱导的胰岛细胞凋亡的保护作用，初步研究其作用机制。方法：Western—blotting检测NIT-1细胞的Ppar-γ受体蛋白表达；联合IL-1β及IFN-γ作用于体外培养的胰岛细胞株NIT-1，建立胰岛细胞炎症模型，观察（IL-1β＋IFN-γ）与PGZ预孵育NIT-1细胞的凋亡；MTT法、流式细胞术检测细胞生长抑制率及凋亡率，比色法检测凋亡细胞caspase-3比活性。结果：NIT-1细胞表达Ppat-γ蛋白；（IL—1β＋IFN-γ）组作用24h对NIT-1细胞生长抑制率、凋亡率、caspase-3比活性分别为49．8％、28．0％、3．5，对照组3项指标分别为0％、4．3％、1，两组比较均有十分显著差异（P〈0．01）；PGZ组3项指标分别为2．7％、7．1％、1.3，与对照组比较，均无明显差异（P〉0．05）；（PGZ＋IL-1β＋IFN-γ）组的3项指标分别为18．6％、14．8％、1．5，与（IL-1β＋IFN-γ）组比较均有差异（P〈0．05），与对照组比较分别为P〉0．05，P〈0．05，P〈0．05。结论：NIT-1细胞表达Ppar-γ受体蛋白；联合IL—1β及IFN-γ明显诱导NIT-1细胞凋亡，PGZ显著抑制IL-1β及IFN-γ诱导的NIT-1细胞凋亡，其机制与降低caspase-3活性有关。     

大黄素抑制食管癌EC-109细胞增殖的机制探讨

目的：探讨大黄素对人食管癌细胞株EC-109增殖和凋亡的影响,并初步探讨其作用机制。方法：不同浓度（0．0,2．5、5．0、10．0、20．0μg/mL）大黄索作用食管癌细胞EC-109,应用噻唑蓝比色法检测大黄素对食管癌细胞增殖的影响;用Annexin V-FTTC碘化丙啶双染流式细胞术检测细胞凋亡情况;流式细胞术检测细胞内活性氧变化情况。结果：不同浓度大黄素可明显抑制EC．109细胞增殖,并且呈浓度-时间依赖性;在作用EC-109细胞2h后,细胞内活性氧明显增加;12h后可见细胞凋亡率分别为8．1％、15．6％、24．0％、36．5％。结论：大黄素能明显抑制EC-109细胞的增殖,通过诱导细胞内活性氧的产生引起食管癌细胞EC-109凋亡。     

大黄素诱导白血病K562细胞凋亡及对Caspase-3基因表达的影响

目的：探讨大黄素对人白血病K562细胞的增殖抑制与诱导凋亡作用．方法：采用四唑蓝比色试验（MTT）检测大黄素对K562细胞的增殖抑制作用;通过电子显微镜观察细胞的形态学变化;运用原位缺口末端标记技术检测大黄素对K562细胞的凋亡效应;采用流式细胞技术检测细胞周期变化与凋亡情况;通过比色法检测Caspase-3的相对活性,RT—PCR检测细胞内凋亡相关基因Caspase-3的转录水平．结果：大黄素能显著抑制K562细胞的增殖,作用24,72h后的半数抑制率浓度分别为80,40μmol／L;处理组细胞可见典型的凋亡形态学改变;TUNEL法检测到K562细胞在大黄素的诱导下出现凋亡;流式细胞仪结果分析发现,处理组的K562细胞被阻滞于G0／G1期,与对照组相比,凋亡率明显升高并呈现时间-剂量依赖性（P〈0．01）;大黄素可触发Caspase-3的活性增高（P〈0．01）,并呈现剂量依赖性;RT—PCR结果显示,Caspase-3 mRNA表达上调．结论：大黄素能有效地抑制K562细胞的增殖并诱导其凋亡,可能与Caspase-3活化有关．     

游离脂肪酸对大鼠胰岛分泌功能和胰岛细胞凋亡的影响研究

目的：研究游离脂肪酸对体外培养SD大鼠胰岛功能和胰岛细胞凋亡的影响并探讨可能机制。方法：取健康雄性SD大鼠胰岛原代分离培养,分为6组,培养1d：NC1组（5．6mmol／L葡萄糖）、FFA1组（0．25mmol／L）、FFA2组（0．5mmol／L）;培养3d：NC2组（5．6mmol／L葡萄糖）、FFA3组（0．25mmol／L）、FFA4组（0．5mmol／L）;每组6个样本,每个样本20个胰岛。采用RT—PCR扩增胰岛素、PDX-1、Bax、Caspase-3基因mRNA表达,TUNEL法检测胰岛细胞凋亡率,放射免疫法检测胰岛素水平。结果：①FFA1、FFA2组Insulin、PDX-1基因表达量、胰岛素分泌较NC1组明显下降,FFA1、FFA2组凋亡基因Bax、Caspase-3表达量及胰岛细胞凋亡率较NC1组明显升高,差异有统计学意义（P〈0．05）;②FFA3、FFA4组Insulin、PDX-1基因表达量、胰岛素分泌较NC2组明显下降,FFA3、FFA4组凋亡基因Bax、Caspase-3表达量及胰岛细胞凋亡率较NC2组明显升高,差异有统计学意义（P〈0．05）。结论：游离脂肪酸抑制体外培养SD大鼠胰岛细胞的胰岛素分泌功能,其可能机制是FFA抑制胰岛素基因、PDX-1基因的合成,刺激胰岛细胞凋亡基因Bax、Caspase-3表达,并导致胰岛细胞凋亡,最终导致胰岛细胞分泌功能下降。     

高糖诱导胰岛细胞凋亡机制的研究

目的：探索不同浓度葡萄糖引起胰岛细胞凋亡的机制。方法：用胶原酶P消化Ficol1400纯化的成年SD大鼠胰岛细胞，经RPMI1640培养7天后铺成单层，分别设立对照组和不同浓度的葡萄糖组，用放免法测定加葡萄糖后1、3、5、7、9和11天胰岛素浓度。用流式细胞术测定加处理因素后11天的胰岛细胞凋亡率和免疫组化法测定bax及bcl-2蛋白的表达。另取单层培养的胰岛细胞，亦设立对照组和加入不同浓度甘露醇组，培养11天后，用免疫组化法测定bax和bcl-2蛋白的表达，流式细胞术测定胰岛细胞凋亡率。结果：①葡萄糖为11．1mmol／L时，胰岛素浓度开始升高；当葡萄糖升至22．2mmol／L时达高峰，且随时间延长而下降。②葡萄糖为11．1和22．2mmol／L时，胰岛细胞凋亡率最高，5．5mmol／L组与对照组的凋亡率差异无显著性意义。甘露醇组胰岛细胞的bax及bcl-2蛋白表达与对照组比较无显著差异，胰岛细胞凋亡率与对照组比较亦无著著差异。③葡萄糖在11．1-22．2mmol／L时，bax蛋白表达阳性，bcl-2蛋白表达阴性，葡萄糖在33．3mmol／L时，bax和bcl-2蛋白的表达匀为阴性，5．5mmol／L组bax和bcl-2蛋白的表达与对照组无显著差异。结论：高血糖（葡萄糖为11．1和22．2mmol／L）可引起胰岛细胞凋亡和胰岛素释放增加，当葡萄糖浓度升至33．3mmol／L时，胰岛细胞凋亡率开始下降，胰岛素释放也减少。表明高渗状态与高血糖诱导的胰岛细胞凋亡无关。     

姜酚肟对谷氨酸诱导PC12细胞损伤的保护作用

【目的】探讨姜酚肟对谷氨酸（glutamate，Glu）诱导PC12细胞损伤的保护作用。【方法】体外培养PCI2细胞，建立谷氨酸诱导PC12细胞损伤的模型；采用MTT法检测细胞活力，流式细胞术和Hoechst 33258 DNA染色法检测细胞凋亡。【结果】经5mmol／L的谷氨酸处理24h后，PC12细胞活力比对照组降低，仅为对照组的58．3％。在一定浓度范围内，姜酚肟能保护PC12细胞免受谷氨酸（5mmol／L）的影响，但较高浓度时对细胞有一定的毒性。经不同浓度姜酚肟（3.125、6．25、12．5、25μg／mL）预处理后，细胞活力明显提高，其保护作用随浓度降低而升高，当浓度为6．25μg／mL时效果最好，使PC12细胞的存活率达到75．2％（P〈0．01），浓度为3．125μg／mL时，细胞存活率降为70．2％（P〈0．01）。谷氨酸（5mmol／L）能诱导PCI2细胞凋亡，处理24h后细胞凋亡率为20．1％，经姜酚肟（3．125、6．25、12．5、25μg／mL）预处理后，细胞凋亡率明显降低，分别为3．5％（P〈0．01），2．8％（P〈O．01），9．6％（P〈0．01），17．7％（P〈0．05）。【结论】谷氨酸能诱导PCI2细胞凋亡，较低浓度的姜酚肟能有效保护PC12细胞免受谷氨酸诱导的细胞损伤。其中6．25μg／mL的姜酚肟效果最好。     

软脂酸对胰岛细胞凋亡作用的观察

目的通过观察软脂酸对体外培养的原代胰岛细胞的凋亡作用，探讨游离脂肪酸在糖尿病发病中的作用。方法体外培养Wistar大鼠胰岛细胞，用不同浓度软脂酸(分别为0，0．125，0．25，0．5mmol／L)共同孵育48h，用Tunel法计算胰岛细胞凋亡率。AO／EB染色观察细胞形态变化。结果0．125，0．25，0．5mmol／L的软脂酸均有抑制高糖刺激下胰岛细胞的胰岛素分泌，促进胰岛细胞凋亡的作用。结论软脂酸可抑制胰岛细胞的分泌功能，促进胰岛细胞凋亡，其凋亡程度与软脂酸的浓度有关。     

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.