大黄素诱导人肾上皮HK-2细胞凋亡及内质网应激的介导作用

目的 研究大黄素诱导人肾小管上皮HK-2细胞凋亡的作用及其机制是否与内质网应激有关。方法 不同浓度大黄素处理HK-2细胞不同时间。MTT法检测细胞存活率,实时荧光定量PCR（RT-PCR）检测葡萄糖调节蛋白78（GRP78）、CCAAT/增强子结合蛋白同源蛋白（CHOP）、转录激活因子3（ATF3）和X盒结合蛋白1（XBP1）的基因表达,Western blotting法检测caspase-3、GRP78、CHOP、真核生物启动因子2α（eIF2α）的蛋白表达。结果 大黄素以浓度相关方式降低HK-2细胞存活率,诱导caspase-3剪切。RT-PCR检测表明,大黄素能诱导内质网相关基因GRP78、CHOP、ATF3基因表达,Western blottinh检测结果进一步证实大黄素能诱导GRP78、CHOP的蛋白表达。大黄素还明显诱导eIF2α磷酸化和XBP1 mRNA剪切。在大黄素给药之前给予内质网应激干扰药4-苯基丁酸和salubrinal,能增加HK-2细胞的存活率。结论 大黄素能诱导HK-2凋亡,内质网应激参与了大黄素诱导HK-2细胞凋亡的作用。     

内质网应激介导滋养细胞凋亡在妊娠期肝内胆汁淤积症中的作用及机制

目的探讨内质网应激介导滋养细胞凋亡在妊娠期肝内胆汁淤积症（ICP）中的作用及机制。方法收集2015 年12 月~ 2016年12月在南方医科大学南方医院住院分娩的ICP孕妇20例为病例组,另以同期因社会因素行择期剖宫产术的20例无合 并症的正常孕妇为正常对照组。利用HE染色计量观察正常对照孕妇及ICP孕妇胎盘合体细胞结节数量;RT-PCR法检测正常 孕妇及ICP孕妇胎盘组织中GRP78、CHOP、caspase-3及caspase-7的mRNA表达;建立不同浓度胆酸（0、10、50、100 μmol/L）体 外刺激HTR-8/SVneo人早孕滋养细胞株模型,利用RT-PCR法检测HTR-8/SVneo细胞GRP78、CHOP、caspase-3及caspase-7的 mRNA表达及利用蛋白印迹法检测GRP78、CHOP蛋白的表达;caspase-3、caspase-7活性检测试剂盒分别检测其活化程度;电镜 观察细胞凋亡形态。结果ICP 组较对照组孕妇胎盘组织中合体细胞结节数明显增多（P<0.01）;GRP78、CHOP、caspase-3 及 caspase-7的mRNA表达明显上调（P<0.05）;不同浓度（0、10、50、100 μmol/L）去氧胆酸作用细胞24 h后,GRP78、CHOP、caspase-3 及caspase-7 mRNA的表达水平较对照组显著增高（P<0.05）,GRP78、CHOP蛋白的表达水平较对照组也显著增高,且均存在浓 度依赖效应;50 μmol/L去氧胆酸作用细胞24 h后检测caspase-3及caspase-7活性增高（P<0.05）;50 μmol/L去氧胆酸作用细胞 12 h后电镜观察HTR-8/SVneo细胞呈现凋亡小体。结论ICP孕妇胎盘组织中存在滋养细胞过度凋亡及内质网应激激活的现 象。去氧胆酸可以呈浓度依赖性地促进滋养细胞内质网应激相关基因mRNA和蛋白的表达,引起细胞凋亡,提示内质网应激介 导的滋养细胞凋亡在ICP的发病机制中可能起着重要的作用。     

全反式维甲酸通过内质网应激诱导N-乙酰氨基葡萄糖基转移酶V受阻的人肝癌SMMC-7721细胞凋亡

目的初步探讨全反式维甲酸（all-trans-retinoic acid，ATRA）诱导N-乙酰氨基葡萄糖基转移酶V（N—acetylglucosaminyltransferase V，GnT—V）表达受阻的人肝癌SMMC-7721细胞（AsGnT—V／SMMC-7721）发生凋亡与细胞中内质网应激的关系。方法利用RT-PCR检测ATRA处理前后AsGnT-V／SMMC-7721细胞中内质网应激关键分子GRP78（glucose regulated protein 78）、XBP1（X box binding protein 1）等的变化。并用Western blot检测ATRA处理前后AsGnT—V／SMMC-7721细胞中内质网应激相关凋亡途径中的重要分子CHOP（C／EBP homologus protein）、caspase-9、caspase-12以及caspase-3的变化。结果GRP78和XBP1的变化表明经ATRA处理后AsGnT-V／SMMC-7721细胞的内质网应激加剧了，而与内质网应激相关的凋亡分子CHOP、caspase-9、caspase-12以及caspase-3都发生了激活。结论ATRA诱导AsGnT—V／SMMC-7721细胞发生的凋亡与内质网应激有关。     

1-脱氧甘露糖野尻霉素诱导人肝癌SMMC7721细胞发生内质网应激

目的探讨人肝癌SMMC7721细胞中1-脱氧甘露糖野尻霉素（1-deoxymannojirimycin，DMJ）对高尔基体中α-甘露糖苷酶Ⅰ活性的抑制作用与内质网应激的关系。方法人肝癌SMMC7721细胞经DMJ诱导后．用RT-PCR和Western blot检测内质网应激中的重要分子葡萄糖调节蛋白78（glucose regulated protein 78，GRP78）、X盒结合蛋白1（X box binding protein 1,XBP1）、C／EBP同源蛋白（C／EBP homologus protein,CHOP）及caspase-12的表达变化。结果GRP78的mRNA和蛋白质表达量升高；XBP1的mRNA发生剪接．蛋白相对分子质量从33000变为54000；CHOP蛋白表达量上调；caspase-12剪切激活。结论DMJ可诱导SMMC7721细胞发生内质网应激。     

大黄酸通过非活性氧依赖性内质网应激通路诱导L-02细胞凋亡

研究大黄酸对人正常肝细胞L-02的凋亡作用及探讨其机制。应用MTT法评价大黄酸对L-02细胞的活性抑制作用;流式细胞术检测大黄酸诱导L-02细胞的凋亡;DCFH-DA荧光探针检测细胞内活性氧(ROS)含量;实时荧光定量PCR(RT-qPCR)和Western blot技术检测细胞内质网应激相关基因和蛋白的表达变化;并探讨抗氧化剂N-乙酰半胱氨酸(NAC)对内质网应激蛋白及caspase-4、caspase-3的影响。实验结果显示:大黄酸能够呈剂量和时间依赖性抑制L-02细胞活性,增加L-02细胞凋亡率;诱导细胞ROS水平增高,NAC预给药可显著降低其水平;大黄酸能显著上调L-02细胞内的葡萄糖调节蛋白78(GRP 78)、活化转录因子4(ATF-4)和C/EBP同源蛋白(CHOP)基因的mRNA水平;显著增加GRP 78,磷酸化c-jun 氨基末端激酶(p-JNK),CHOP蛋白表达;NAC不能抑制大黄酸诱导的GRP 78,p-JNK,CHOP蛋白表达上调,亦不能抑制大黄酸诱导的caspase-4及caspase-3活性增加。表明大黄酸能够诱导L-02细胞凋亡,其作用机制与非活性氧依赖性的内质网应激通路有关。     

大鼠压疮局部不同干预温度对内质网应激及细胞凋亡的影响

目的比较大鼠压疮局部不同干预温度对内质网应激(ERS)及细胞凋亡的影响,为临床防治压疮提供实验依据。方法使用缺血再灌注法建立压疮模型,将40只SPF级成年雄性SD大鼠随机分成假手术组(sham组)、模型组、热干预组和冷干预组4组。在实验终点于冰上剪取各组大鼠受压部位肌肉组织,用HE染色观察骨骼肌病理学变化;Western blot检测内质网应激相关蛋白GRP78、caspase-12与CHOP的表达水平;免疫荧光观察caspase-12与CHOP的表达情况;TUNEL染色观察细胞凋亡情况。结果与模型组相比,热干预组的骨骼肌细胞损伤程度加重,凋亡细胞增多,免疫荧光结果显示caspase-12与CHOP的阳性表达增加,Western blot结果显示GRP78、caspase-12与CHOP的表达较模型组均上升(P0.05);而冷干预组骨骼肌细胞损伤程度减轻,凋亡细胞减少,免疫荧光结果显示caspase-12与CHOP的阳性表达减少,Western blot结果显示GRP78、caspase-12与CHOP的表达较模型组均下降(P0.05)。结论局部冷干预通过抑制内质网应激介导的凋亡通路而减轻大鼠压疮损伤;局部热干预加剧ERS而促进细胞凋亡的发生,加重大鼠压疮损伤。     

N-乙酰半胱氨酸对脂多糖诱导大鼠肝细胞内质网应激的抑制作用

目的:探讨N-乙酰半胱氨酸(N-acetyl-L-cysteine,NAC)对脂多糖(lipopolysaccharide,LPS)诱导大鼠肝细胞内质网应激的影响.方法:培养大鼠肝细胞系BRL细胞,用NAC或LPS分别或共同处理BRL细胞,其中共同处理时把NAC加入培养液中预处理1h再给予LPS处理肝细胞.用锥虫蓝拒染法检测细胞存活率;Hoechst33258染色荧光显微技术检测肝细胞凋亡的形态学变化;免疫印迹法检测内质网应激标志蛋白GRP78及凋亡相关蛋白CHOP、Caspase-12和Caspase-3的表达.结果:与溶剂对照组比较,LPS能时间依赖性地引起肝细胞细胞活力显著降低和细胞凋亡增多,诱导内质网应激标志蛋白GRP78表达明显上调;NAC预处理抑制LPS引起细胞存活率降低和细胞凋亡增多,并且能显著抑制LPS诱导的GRP78及凋亡相关蛋白CHOP、Caspase-12和Caspase-3表达上调.结论:NAC通过阻断LPS诱导的肝细胞内质网应激而减轻肝细胞损伤.     

大黄素对糖尿病大鼠肠平滑肌内质网应激及调控机制的研究

目的 探讨大黄素对糖尿病大鼠肠平滑肌细胞凋亡机制及调控机制.方法 SD大鼠分为对照组、糖尿病组与大黄素组,成模10周时,检测小肠推进率;TUNEL法测肠平滑肌细胞凋亡;免疫组织化学测细胞葡萄糖调节蛋白78 (GRP78)及Caspase-12蛋白表达.结果 糖尿病组与大黄素组大鼠于成模后出现多尿、多饮、多食、体质量减轻症状;血糖水平显著增高(P＜0.05).成模10周时,与对照组比较,糖尿病组大鼠小肠推进率显著下降(P＜0.05),肠平滑肌细胞凋亡率、GRP78、Caspase-12蛋白表达显著增加(P＜0.05).与糖尿病组比较,大黄素组小肠推进率显著增高(P＜0.05),肠平滑肌细胞凋亡率、GRP78、Caspase-12蛋白表达显著降低(P＜0.05).结论 内质网应激途径介导了糖尿病大鼠肠平滑肌细胞凋亡;大黄素可能降低糖尿病大鼠肠平滑肌内质网应激介导的细胞凋亡,从而改善肠动力.     

LBH589对胃癌细胞增殖的影响及其机制的研究

目的 探讨组蛋白去乙酰化酶抑制剂LBH589对胃癌细胞系BGC-823增殖的影响.方法 体外培养、胃癌BGC-823细胞,MTT检测LBH589对BGC-823细胞增殖率的影响;Western Blotting检测内质网应激相关蛋白GRP78,内质网凋亡相关蛋白CHOP、caspase-4表达水平的改变.结果 与对照组...     

异氟醚对β淀粉样蛋白诱导的大鼠PC12细胞凋亡和内质网应激的影响及其机制

目的：探讨异氟醚对β淀粉样蛋白（Aβ）25-35诱导的大鼠PC12细胞凋亡和内质网应激（ERS）的影响,并阐明其作用机制。方法：将PC12细胞随机分为正常对照组、10 μmol•L-1Aβ25-35组（Aβ组）、2%异氟醚组（Iso组）和2%异氟醚联合10 μmol•L^-1 Aβ25-35组（Iso+Aβ组）。MTT法检测各组PC12细胞存活率;Hoechst 33342核染色法检测各组PC12细胞凋亡形态;Western blotting法检测各组PC12细胞内质网分子伴侣葡萄糖调节蛋白78(GRP78)、ERS相关凋亡信号蛋白C/EBP同源蛋白(CHOP)、磷酸化氨基末端蛋白激酶(p-JNK)和caspase-12表达量。结果：与正常对照组比较,Aβ组和Iso组PC12细胞存活率明显降低（P<0.05）,细胞凋亡率明显增加（P<0.05）,GRP78表达量明显上调（P<0.05）,ERS相关凋亡信号蛋白CHOP、p-JNK和caspase-12表达量均明显增加（P<0.05）;与Aβ组比较,Iso + Aβ组PC12细胞存活率明显降低（P<0.05）,细胞凋亡率明显增加（P<0.05）,GRP78、CHOP、p-JNK和caspase-12表达量均明显增加（P<0.05）。结论：异氟醚能够促进Aβ25-35诱导的大鼠PC12细胞凋亡,其机制与激活ERS及其相关的凋亡信号通路有关联。     

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.