白藜芦醇诱导肺癌细胞凋亡和细胞周期阻滞的分子机制

目的探讨白藜芦醇对肺癌细胞系A549和H1299细胞增殖与细胞周期阻滞及细胞凋亡之间的联系。方法采用MTT法检测白藜芦醇对A549和H1299细胞增殖的影响;用Western印迹法检测白藜芦醇作用后A549和H1299中凋亡相关蛋白cleaved-caspase-3、cleaved-caspase-7、cleaved-caspase-9、cleaved-PARP及细胞周期相关蛋白cyclin D1、cyclin D3和CDK2、CDK4、CDK6、p53、p18、p21、p27的表达情况;通过流式细胞仪技术检测白藜芦醇诱导A549和H1299细胞凋亡率以及细胞周期阻滞的影响。结果白藜芦醇对A549和H1299的生长均有抑制作用,呈浓度依赖性;经白藜芦醇处理后,A549和H1299的周期蛋白cyclin D1、cyclin D3和CDK2蛋白的表达水平上调,而周期蛋白CDK4、CDK6、p53、p18、p21、p27的表达水平下调;A549和H1299的凋亡相关蛋白cleaved-caspase-3、cleaved-caspase-7、cleaved-caspase-9、cleaved-PARP的表达水平上调;流式细胞仪检测A549和H1299的细胞凋亡率随白藜芦醇的浓度上升而增加,流式细胞仪检测细胞周期即可观察到随白藜芦醇的浓度的增加,细胞周期明显阻滞于G1/S期。结论白藜芦醇可能是通过促进caspases通路的活化,从而促进细胞凋亡;周期蛋白cyclin D1、cyclin D3和CDK2蛋白的表达水平的上调和CDK4、CDK6、p53、p18、p21、p27的表达水平的下调可能是白藜芦醇诱导人肺癌细胞发生G1/S期阻滞的关键机制。     

葛根素对人小细胞肺癌H446细胞周期和相关周期蛋白表达的影响

目的观察葛根素诱导人小细胞肺癌H446细胞周期的阻滞作用,进一步探讨其作用机制。方法采用MTT法测定葛根素对H446细胞生长的影响;采用流式细胞术检测处理后H446细胞周期分布和周期蛋白cyclinD1、CDK4和P27表达水平。结果MTT法表明葛根素对H446细胞生长有较强的抑制作用;流式细胞仪检测结果发现细胞周期阻滞在G0/G1期,量效关系良好,并检测到细胞周期蛋白cyclin D1、CDK4表达减弱,P27表达增强。结论葛根素可通过诱导细胞周期阻滞而发挥体外抗肺癌作用,其机制可能涉及细胞周期相关蛋白cyclin D1、CDK4和P27表达的调控。     

靶向抑制Fbxw8对前列腺癌细胞增殖能力的影响

目的 探讨Fbxw8对人前列腺癌DU145细胞增殖及对增殖相关基因cyclinA、cyclinB、CDK1、p21和p27表达的影响.方法 RT-PCR和Western blot法验证Fbxw8 siRNAs的干扰效率;利用CCK-8检测法检测细胞的增殖能力;流式细胞仪检测细胞周期分布情况;Western blot法检测细胞增殖相关蛋白CDK1、CDK2、cyclinA、cyclinB、P21、P27的表达.结果 Fbxw8 siRNAs显著下调DU145细胞中Fbxw8mRNA及蛋白水平的表达(P＜0.01);Fbxw8 siRNAs显著抑制DU145细胞增殖活力,其中72 h抑制最明显(P＜0.05);流式细胞术分析显示,Fbxw8 siRNAs可阻滞细胞于G2/M期,G2/M期细胞百分数由15.32％ (control siRNA)增加到29.03％(Fbxw8 siRNA 1)和28.39％(Fbxw8 siRNA 2);Western blot结果显示,Fbxw8 siRNAs可促进周期蛋白依赖性激酶抑制剂p21、p27的表达,同时下调G2/M期调控蛋白CDK1、CDK2、cyclin A及cyclin B1的表达.结论 靶向抑制DU145细胞中Fbxw8的表达可显著抑制细胞的增殖,使阻滞细胞于G2/M期,并且可能是通过下调CDK1、CDK2、cyclin A及cyclin B1的表达,上调P21、P27的表达实现的.     

TRPM8抑制剂BCTC对前列腺癌DU145细胞的抑癌作用

目的:探讨TRPM8抑制剂BCTC对前列腺癌DU145细胞的抑癌作用。方法:PCR和Western blot检测TRPM8的表达;MTT法检测DU145细胞增殖能力;划痕实验、transwell实验检测细胞迁移和侵袭能力;流式细胞术检测细胞周期和凋亡;Western blot检测周期相关蛋白(p-AKT,p-GSK-3β,cyclin B1,cyclin D1,CDK2/4/6)、凋亡相关蛋白(Bcl-2,Bax,caspase-3)、迁移相关蛋白(pFAK,MMP2)表达水平的变化。结果:PCR和Western blot提示TRPM8在前列腺癌DU145细胞中表达显著高于正常前列腺上皮PNT1A细胞;BCTC处理后,DU145细胞增殖能力下降(P<0.05),细胞周期表现为G0/G1阻滞期(P<0.05),迁移、侵袭能力也明显下降(P<0.05),但凋亡相关实验显示BCTC并不引起细胞凋亡(P>0.05)。Western blot显示BCTC能下调p-AKT,cyclin D1,CDK2,CDK6,MMP2和pFAK等的表达,而上调p-GSK-3β的表达。结论:TRPM8抑制剂BCTC能抑制前列腺癌DU145细胞增殖、迁移和侵袭能力,是一个极具潜力的抗前列腺癌药物。     

环孢素A对体外人系膜细胞cyclin E,cyclin D1,p27kip1蛋白表达的影响

目的探讨环孢素A(CsA)对人系膜细胞增殖的抑制作用及细胞周期蛋白D1、E、p27kip1表达影响。方法人系膜细胞常规体外培养,分为实验组和对照组,实验组加入不同浓度的CsA(0.1,1,5μmol/L)干预。药物作用24,48,72 h后MTT比色法测定细胞增殖情况。药物作用48 h后采用流式细胞仪检测细胞周期。药物作用48 h后Western blot法检测细胞中cyc-lin D1、cyclin E、p27kip1蛋白质表达情况。结果 CsA对人系膜细胞具有时间、浓度依赖性抑制作用;且浓度依赖性阻滞细胞周期,使G0/G1期细胞增多,S期细胞减少(P<0.05)。CsA浓度依赖性下调cyclin D1、cyclin E蛋白,上调p27kip1蛋白的表达(P<0.05)。结论 CsA通过下调cyclin D1、cyclin E蛋白,上调p27kip1蛋白的表达,调节细胞周期调控通路,阻滞细胞于G0/G1期,有效抑制人系膜细胞的增殖。     

辛基酚对MDA-MB-231乳腺癌细胞周期及周期蛋白表达的影响

目的:观察辛基酚(OP)对细胞周期及周期蛋白表达的影响,探讨OP抑制MDA-MB-231乳腺癌细胞增殖的可能分子机制.方法:以MTT试验、流式细胞分析、免疫细胞化学和RT-PCR等方法观察OP对MDA-MB-231乳腺癌细胞增殖、细胞周期、CDK2,CDK4,cyclin D1和cyclin D3表达的影响.结果:4, 8 μmol/L OP作用MDA-MB-231乳腺癌细胞48h时,其抑制率为(17.1±4.1)%, (40.3±8.7)%,cyclin D1表达量荧光值为55.1±10.2,41.4±11.2,比对照组(89.9±11.2)明显降低(P＜0.05). 8 μmol/L OP作用MDA-MB-231乳腺癌细胞24 h和72 h,G0/G1期细胞为(67.0±17.6)%和(44.4±11.9)%,G0/G1期细胞比对照组[24 h,(46.6±12.8)%;72 h,(15.3±3.1)%]明显增高(P＜0.05). OP导致MDA-MB-231乳腺癌细胞中CDK2,CDK4,cyclin D1和cyclin D3 mRNA的表达降低,且cyclin D1蛋白的表达也降低.结论:OP能抑制MDA-MB-231乳腺癌细胞增殖,其机制可能与OP能降低乳腺癌细胞中CDK2,CDK4,cyclin D1和cyclin D3 mRNA及其蛋白的表达有关.     

网脊衣酸上调p21CIP1蛋白诱导前列腺癌细胞周期阻滞的作用分析

目的 探讨天然化合物网脊衣酸(RB)对前列腺癌LNCaP细胞的周期阻滞作用及分子机制。方法 细胞经RB处理后,流式细胞术检测细胞周期的变化;溴脱氧尿嘧啶核苷(BrdU)标记法检测细胞增殖情况;Western blotting检测p53、p21 CIP1(p21)及细胞周期相关蛋白的表达;实时定量PCR检测p21的 mRNA水平的变化;荧光素酶报告基因检测RB对p21启动子活性的影响;转染p53突变体质粒检测p53对p21表达的影响;基因敲除p21检测其对细胞周期的作用。结果 RB可使LNCaP细胞阻滞于G0/G1期,抑制周期相关的Cyclin D、Cyclin E、Cdk 4和p-Rb表达。同时,RB可时间依赖性地诱导LNCaP细胞中野生型p53表达、促进其入核,同时上调p21的mRNA和蛋白水平。阻断p53的活性后,可抑制RB介导的p21表达及其启动子活性。而敲除p21基因,可降低RB所介导的G0/G1期阻滞,同时G2/M期细胞数量增加。结论 RB通过激活p53,在转录水平上调靶基因p21的表达,导致前列腺癌LNCaP细胞阻滞于G0/G1期,从而抑制细胞的增殖。     

PTEN基因影响K562细胞周期的分子机制探讨

【目的】探讨抑癌基因PTEN对人慢性粒细胞白血病细胞系K562细胞周期的影响。【方法】将携带有野生型PTEN和绿色荧光蛋白的腺病毒（Ad-PTEN-GFP）及对照载体腺病毒（Ad-GFP）转染人慢性粒细胞白血病细胞系K562。MTT检测细胞增殖抑制率;流式细胞仪检测转染效率、细胞凋亡率和细胞周期的变化;荧光定量PCR（FQ-PCR）检测PTEN、CyclinD1、CyclinD2、CDK4、P27Kip1 mRNA水平变化,Western blot检测上述蛋白表达水平的变化。【结果】与Ad-GFP组比较,Ad-PTEN-GFP转染K562细胞后,细胞凋亡率最高为30%,细胞周期显示：G0/G1期细胞比例由54.9%增加至78.5%,G2/M期比例由30.2%降至13.6%,Cyclin D1、Cyclin D2、CDK4 mRNA及Cyclin D1蛋白表达降低,P27Kip1 mRNA及蛋白表达增加。【结论】PTEN基因高表达可以促进K562细胞凋亡,并通过抑制Cyclin D1、Cyclin D2、CDK4及增加P27Kip1表达使细胞周期阻滞在G0/G1期。     

双黄升白颗粒对化疗所致骨髓抑制Lewis肺癌荷瘤小鼠细胞周期的双重调控作用及其机制

目的：探讨双黄升白颗粒对化疗所致骨髓抑制荷瘤小鼠骨髓和肿瘤细胞周期双重调控的作用机制。 方法：本研究采用Lewis肺癌荷瘤小鼠,30只小鼠随机分为空白组、模型组和治疗组。除空白组外,腹腔注射环磷酰胺制作骨髓抑制模型。治疗组用40g/（kg·d）双黄升白颗粒治疗6d后,流式细胞仪检测细胞周期情况,并计算增殖指数（proliferationindex,PI）;蛋白印迹法和免疫组织化学法检测骨髓及肿瘤组织中细胞周期蛋白依赖激酶4（cyclin-dependent kinase4,CDK4）、细胞周期蛋白依赖激酶6（cyclin-dependent ki-nase6,CDK6）和细胞周期蛋白D1（cyclin D1）的表达。 结果：模型组骨髓及肿瘤的G0/G1期细胞比例均低于空白组（P〈0.05）,PI和CDK4、CDK6、cyclin D1表达高于空白组（P〈0.05）。治疗组骨髓G0/G1期细胞比例低于模型组及空白组,PI及CDK4、CDK6、cyclinD1表达均高于模型组及空白组;肿瘤组织G0/G1期细胞比例高于模型组及空白组,PI及CDK4、CDK6、cyc-lin D1表达均低于模型组及空白组。 结论：双黄升白颗粒对骨髓抑制Lewis肺癌荷瘤鼠的细胞周期具有双重调控作用,其机制可能与双重调节cyclin D1、CDK4和CDK6表达有关。     

叶黄素对人食管鳞癌细胞诱导分化效应与机制探讨

目的 探讨叶黄素对食管癌EC9706 细胞诱导分化的影响及其机制.方法 食管癌EC9706细胞采用开放式单层贴壁培养.实验设溶剂对照及叶黄素3个剂量组,采用MTT法及流式细胞术,对EC9706 细胞的增殖和细胞周期进行分析,HE染色对细胞的形态学进行观察,酸解DNA甲绿-派洛宁技术了解增殖与分化型细胞比例,免疫组化检测cyclin D1的表达.结果 与溶剂对照组比较,叶黄素(100 μg/mL和150 μg/mL)可明显抑制EC9706 细胞的增殖,阻滞细胞周期于G0/G1期, 降低细胞增殖指数,细胞形态趋向良性分化,cyclin D1 表达水平降低.结论 叶黄素可通过抑制cyclin D1 表达而介导食管癌EC9706细胞增殖抑制和诱导成熟分化.     

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.     

APPLICATION OF LORNOXICAM TO PATIENT-CONTROLLED ANALGESIA IN PATIENTS UNDERGOING ABDOMINAL SURGERIES

FOR anesthesiologis s ,treatingpostoperativepainhas alwaysbeen a problem.Althoughopioidshave been provedtobe effective,theirsideeffectscouldnotbeignored.With thedevelopmentofscienceand pharmacology,many drugs with aspectsof satisfactoryanalgesicefficacyand couldbe welltoleratedby patientshave been developed.And lornoxicamisone of them, which isa non-steroidalanti-inflammatorydrug (NSAID ), with analgesic, anti-infl-ammatory,andantipyreticproperties.Itseliminationhalf-time(3 to 5 hours) isle…     

Acupuncture Combined with Spinal Tui Na for Treatment of Primary Dysmenorrhea in 30 Cases

Objective: To observe the therapeutic effects in acupunture treatment of primary dysmenorrhea combined with spinal Tui Na, and study its mechanism. Methods: Thirty cases of the treatment group were treated by acupuncture combined with spinal Tui Na, and thirty cases in the control group were treated by routine acupuncture. Results: The total effective rate was 93.3% in the treatment group, and 73.3% in the control group, with a significant difference between the two groups (P<0.05). Conclusions: Acupuncture combined with spinal Tui Na has good prospects for treatment of primary dysmenorrhea.     

Luo Lingjie's Experience in Treating Chronic Nephropathy

In treating chronic nephropathy,Luo Lingjie,a chief physician,pays attention to regulating the balance between yin and yang,treating infection if present,and removing pathogenic factors.He prescribes gentle drugs and uses carefully strongly warming-tonifying ones,emphasizes the importance of persuading the patient to persist in treatment with medication and nurse one's health for recuperation,and is good at combined use of TCM and western medicine therapy and brings the merits of various therapies into full play,with obvious theraoeutic effects.     

Dr.Zhang Ren's Experience in the Acupuncture Treatment of Different Diseases with the Same Therapeutic Principle

Dr.Zhang Ren,the chief physician,is the chairman of Shanghai Acupuncture and Moxibustion Association.Having been engaged in medicine for about 40 years,he is experienced in treating various intractable diseases.In his long years of clinical practice,he advocates taking the TCM differentiation as the basis to seek for the acupuncture method for treatment of modern intractable diseases.The author of this essay had the fortune to follow Dr.Zhang in study.The following is a summary of Dr.Zhang's experience in the acupuncture treatment for different intractable diseases with the same therapeutic principle.     

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.     

猪肺磷脂注射液联合经鼻持续气道正压通气对呼吸衰竭早产儿的临床疗效及肌酸激酶同工酶活性的影响

目的 探讨猪肺磷脂注射液联合经鼻持续气道正压通气(NCPAP)对呼吸衰竭早产儿的临床疗效及肌酸激酶同工酶活性(CK-MB)的影响.方法 选取呼吸衰竭早产儿80例,分为观察组和对照组各40例.对照组采用NCPAP给氧治疗,观察组给予NCPAP给氧联合猪肺磷脂气管内给药.观察两组患儿治疗前及治疗12h、24 h后PaO2、PaCO2、血氧饱和度(SaO2)、pH的变化情况,检测治疗前及治疗5d后血清CK-MB水平;评估两组患儿的临床治疗效果.结果 两组患儿PaO2、PaCO2、SaO2、pH比较,差异均有统计学意义(P＜0.05),其中观察组治疗后的PaO2、SaO2、pH均高于对照组,PaCO2则低于对照组.两组的PaO2、SaO2、pH均随观察时间延长而升高(P＜0.05),PaCO2均随观察时间的延长而降低(P＜0.05).观察组治疗有效率为87.5％,显著高于对照组的70.0％ (P ＜0.05).治疗5d后两组患儿血清CK-MB水平均较前降低(P＜0.05),且观察组明显低于对照组(P＜0.05).结论 猪肺磷脂注射液气管内给药联合NCPAP可以显著降低呼吸衰竭早产儿CK-MB的含量,提高治疗有效率,起到很好的呼吸循环支持作用.