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1.
位全芳  朱琼  常宇骁  杨劲 《海南医学》2014,25(6):785-787
目的siRNA靶向沉默p53蛋白后,观察其对PCNA泛素化修饰及细胞凋亡的影响。方法将p53siRNA及阴性对照siRNA通过脂质体分别转染入HeLa细胞中;转染后48h,顺铂处理不同细胞系,Westernblot法检测p53低表达对细胞PCNA泛素化修饰(ubi-PCNA)及凋亡诱导基因Caspase-3的影响。结果siRNA可显著抑制p53的表达;Westernblot结果提示,与对照组比较,顺铂损伤后p53低表达细胞系其PCNA泛素化修饰蛋白表达增加,同时Caspase-3蛋白表达下降,细胞存活率升高。结论靶向沉默p53的表达,可促进顺铂损伤诱导的PCNA泛素化修饰,降低细胞凋亡率。  相似文献   

2.
目的 探讨肌球蛋白重链9(MYH9)对非小细胞肺癌(NSCLC)细胞增殖、凋亡和对顺铂敏感性的影响并探讨其作用机制。方法 常规培养6株NSCLC细胞系(A549、H1299、H1975、SPCA1、H322、H460)和1株正常支气管上皮细胞系(16HBE)采用Western blot法检测MYH9的表达情况;采用免疫组织化学染色法检测NSCLC患者组织MYH9的表达,实验分为癌组织(49例)和癌旁组织(43例);常规培养H1299和H1975细胞系,使用CRISPR/Cas9技术构建MYH9敲除的NSCLC细胞系,实验分为sgNC组,sgMYH9组,采用细胞计数试剂盒-8(CCK-8)、克隆形成实验检测敲除MYH9对肺癌细胞增殖的影响;Western blot实验、细胞凋亡流式细胞术检测MYH9对细胞凋亡的影响;通过IC50法检测敲除MYH9的肺癌细胞对顺铂敏感性的影响,实验分为sgNC组、sgMYH9组;培育小鼠肿瘤异种移植物模型验证MYH9体内生物学功能,实验分为sgNC组、sgMYH9组。结果与癌旁组织相比,MYH9在癌组织中上调(P<0.001),高表达患者具有更短的生...  相似文献   

3.
目的:研究蟾毒灵对人非小细胞肺癌(NSCLC)A549细胞、H1975细胞的顺铂化疗增敏作用并探讨其可能的参与机制。方法以不同浓度蟾毒灵、顺铂及二者联合分别处理A549细胞、H1975细胞,采用CCK-8法、流式细胞术等方法检测药物对细胞增殖和凋亡的影响;Western blotting 技术检测药物作用后核因子-κB (NF-κB )、c-myc等的表达。结果蟾毒灵可显著抑制A549细胞、H1975细胞增殖并诱导细胞凋亡,且具有一定的时间、浓度依赖性;低剂量蟾毒灵可明显增加A549细胞、H1975细胞对顺铂的化疗敏感性,且此过程伴随NF-κB、c-myc的表达下调。结论蟾毒灵显著增加A549细胞、H1975细胞对顺铂的敏感性;抑制NF-κB 的活化及其下游分子c-myc的表达是其可能的参与机制之一。  相似文献   

4.
目的 研究脱嘌呤脱嘧啶核酸内切酶-1(APE1)对非小细胞肺癌细胞放射敏感性的影响。方法 采用免疫组化染色检测手术标本切片(非小细胞肺癌组织20例及癌旁组织5例)中APE1的表达,Western blot与实时荧光定量(qRT)-PCR检测非小细胞肺癌细胞系(A549,H460,H1299)及肺正常上皮细胞系中APE1的表达;Western blot检测辐照A549与H460细胞0~6 Gy后APE1的表达;构建沉默APE1的A549与H460稳定细胞株后,Western blot检测其辐照6 Gy后APE1的蛋白表达;细胞克隆形成实验检测细胞克隆形成率;CCK-8检测细胞增殖;细胞术流式检测细胞凋亡。结果 非小细胞肺癌组织及细胞系中APE1的表达明显上调;辐照能诱导非小细胞肺癌细胞APE1的表达,且表达随辐照剂量增高而上调;沉默APE1能有效地抑制辐照诱导A549与H460细胞中APE1的表达;成功构建沉默APE1的A459、H460稳定细胞株;沉默APE1联合辐照进一步抑制了非小细胞肺癌细胞克隆形成率、细胞增殖率,同时促进细胞凋亡( PAPE1可增强非小细胞肺癌细胞的放射敏感性。  相似文献   

5.
目的观察川楝素对顺铂抗肺癌活性的协同效应并研究其机制。方法人肺癌细胞系A549分为对照组、川楝素组、顺铂组、川楝素+顺铂组及川楝素+顺铂+X连锁凋亡抑制蛋白(XIAP)质粒组。MTT实验检测各组A549细胞活力抑制率;流式细胞实验检测各组A549细胞凋亡;免疫共沉淀及Western blot实验检测各组A549细胞XIAP表达水平及其与半胱天冬酶-9(Caspase-9)和半胱天冬酶-3(Caspase-3)的相互作用;Western blot实验检测各组A549细胞Caspase-9和Caspase-3的活化。结果川楝素+顺铂组A549细胞活力抑制率[(66.8±5.9)%]显著高于顺铂单治疗组[(15.2±1.2)%,P0.05]和川楝素+顺铂+XIAP质粒组[(20.4±1.8)%,P0.05];川楝素+顺铂组A549细胞凋亡率[(34.5±2.6)%]显著高于顺铂单治疗组[(9.7±0.8)%,P0.05]和川楝素+顺铂+XIAP质粒组[(13.3±1.2)%,P0.05];免疫共沉淀及Western blot实验显示川楝素能显著抑制A549细胞XIAP的表达及其与Caspase-9和Caspase-3的相互作用,川楝素促进顺铂对Caspase-9和Caspase-3的活化。结论川楝素通过下调XIAP表达增强顺铂抗肺癌活性。  相似文献   

6.
目的 探究miR-4286对非小细胞肺癌细胞的顺铂(DDP)敏感性的调节作用及初步机制。方法 采用qRT-PCR法检测A549细胞和A549顺铂耐药细胞(A549/DDP)中miR-4286的表达水平;采用miR-4286模拟物和抑制剂成功构建miR-4286的过表达和低表达细胞模型,分为4组:模拟物阴性对照组、miR-4286模拟物组、抑制剂阴性对照组和miR-4286抑制剂组;采用CCK-8法、平板克隆法和流式细胞术等方法检测改变miR-4286后非小细胞肺癌细胞对顺铂的敏感性的改变;采用TargetScan、qRT-PCR法和蛋白印迹法检测miR-4286和核输出蛋白1(XPO1)的靶向关系;采用蛋白印迹法检测在顺铂的作用下改变miR-4286后非小细胞肺癌细胞的凋亡蛋白表达量的改变。结果 miR-4286在A549细胞中的表达高于A549/DDP细胞(P<0.01)。在0~2g/ml顺铂的作用下,过表达miR-4286后A549细胞增殖率降低、凋亡率和对顺铂的敏感性增加(P<0.05或P<0.01);低表达miR-4286后结果与之相反(P<0.05或P<0.01)。过表达miR-4286后,A549细胞中XPO1的mRNA和蛋白量表达减少,低表达miR-4286后结果与之相反(P<0.05或P<0.01)。在顺铂作用下,下调miR-4286抑制A549细胞凋亡蛋白的表达,同时也下调XPO1能负调控这一作用(P<0.05或P<0.01)。结论 miR-4286能靶向XPO1调控非小细胞肺癌A549细胞对顺铂的敏感性,其机制可能与细胞的凋亡通路相关。  相似文献   

7.
目的 探讨小分子NEK2干扰RNA (siRNA-NEK2)逆转人肺癌耐药细胞(A549/DDP)耐药性的研究.方法 应用Real-time PCR法及Western blot法验证NEK2基因在人肺腺癌耐药细胞系(A549/DDP)细胞系呈现高表达;并采用脂质体为转染介质,将NEK2小片段RNA导入A549/DDP细胞沉默NEK2基因;应用MTT法观察顺铂、阿霉素对沉默NEK2基因后的A549/DDP细胞的细胞毒活性变化.结果 NEK2-mRNA、NEK2蛋白在A549/DDP细胞系表达水平均显著高于A549细胞系,差异有统计学意义(P<0.05);顺铂、阿霉素对A549/DDP细胞的半数抑制浓度(IC50)分别是(53.08±0.56)、(8.09±0.31) μg/mL,对siRNA-control A549/DDP细胞的IC50分别是(53.09±0.78)、(8.10±0.98) μg/mL,而对siRNA-NEK2 A549/DDP细胞的1C50有显著下降,其值分别是(18.59±0.10)、(2.30±0.14) μg/mL,两组比较,差异均有统计学意义(P<0.05).结论 小分子NEK2干扰RNA沉默NEK2可以提高耐药的肺癌细胞对化疗药物的敏感性.NEK2可能成为逆转肺癌耐药的一个新的靶点.  相似文献   

8.
目的探讨蛋白酶体抑制剂MG132联合顺铂对肺癌NCI—H1299细胞凋亡及半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)及半胱氨酸天冬氨酸蛋白酶-8(Caspase-8)蛋白表达的影响。方法将肺癌NCI—H1299细胞行体外原代及传代培养,将培养好的细胞分为4组:对照组(C组,加入磷酸盐缓冲液3 ml)、顺铂组(D组,加入顺铂20 mg/L)、MG132组(M组,加入MG-132 2.5μmol/L)、顺铂+MG132组(D+M组,加入顺铂20 mg/L+MG-132 2.5μmol/L)。孵育30 min,应用MTT比色检测法测定各组肺癌NCI-H1299细胞生长抑制情况,倒置显微镜观察肺癌细胞形态学的变化,流式细胞仪测定肺癌细胞凋亡情况,应用Western blot法对Caspase-3、Caspase-8蛋白表达进行定性及定量分析。结果 MTI结果显示,M组、D+M组肺癌NCI-H1299细胞增殖率显著高于C组及D组(P<0.05),而D+M组肺癌细胞增殖率高于M组(P>0.05);肺癌NCI-H1299细胞培养48 h后M组、D+M组细胞凋亡率高于C组及D组(P<0.05),且D+M组肺癌细胞凋亡率高于M组;Western blot法显示M组、D+M组Caspase-3、Caspase-8蛋白表达水平低于C组及D组(P均<0.05)。结论 MG132与顺铂联合应用可抑制TGF—β1诱导肺癌NCI-H1299细胞凋亡,可能作用机制是MG132能有效下调Caspase-3、Caspase-8蛋白表达。  相似文献   

9.
目的:分析人非小细胞肺癌(non-small cell carcinoma,NSCLC)组织中c-Myc基因的表达,探讨沉默c-Myc基因在非小细胞肺癌基因治疗中的前景?方法:应用实时荧光定量PCR法检测84例非小细胞肺癌组织及其相应癌旁组织中c-Myc基因mRNA的表达情况?通过脂质体LipofactAMINE2000将c-Myc基因siRNA瞬时转染至非小细胞肺癌H1299癌株细胞中,顺铂(Cisplatin,cis)药物毒性实验验证c-Myc基因沉默对肿瘤细胞的抑制效果?应用RT-PCR检测H1299细胞中c-Myc基因RNA的表达,CCK-8法检测抑制细胞的效果?结果:非小细胞肺癌组织中c-Myc基因mRNA的表达水平高于相应的癌旁组织,差异有统计学意义(P < 0.05)?c-Myc siRNA瞬时转染后的H1299细胞中,c-Myc基因mRNA的表达水平显著下降(P < 0.05)?将c-Myc基因沉默和顺铂联合使用,还可以显著提高顺铂对H1299细胞增殖的抑制效果(P < 0.05)?结论:c-Myc基因在非小细胞肺癌组织中的表达水平明显高于相应癌旁组织,沉默c-Myc基因能显著提高顺铂抑制非小细胞肺癌H1299 细胞增殖的作用,c-Myc基因有可能成为NSCLC基因治疗的新靶点?  相似文献   

10.
目的探究线粒体雌激素受体β(ERβ)抑制凋亡刺激诱导非小细胞肺癌细胞死亡的分子机制。方法免疫荧光及western
blotting分析细胞内源ERβ的线粒体定位;检测顺铂与十字孢碱(staurosporine,STS)处理过表达或沉默线粒体ERβ后细胞凋亡
的变化;免疫共沉淀和western blotting分析线粒体ERβ与促凋亡蛋白Bad的相互作用。结果ERβ存在于A549及201T细胞的
线粒体中;过表达或沉默ERβ可以显著减少或增加顺铂与STS诱导的Bax激活及细胞凋亡;线粒体ERβ与促凋亡蛋白Bad相互
作用可能阻抑Bax的活化及线粒体转位。结论线粒体雌激素受体β可以通过与Bad相互作用抑制顺铂或STS诱导非小细胞肺
癌细胞的凋亡,提示线粒体ERβ可能是治疗非小细胞肺癌的一个新靶点。
  相似文献   

11.
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.  相似文献   

12.
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.  相似文献   

13.
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.  相似文献   

14.
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.  相似文献   

15.
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.  相似文献   

16.
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.  相似文献   

17.
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.  相似文献   

18.
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.  相似文献   

19.
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.  相似文献   

20.
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.  相似文献   

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