术前经动脉灌注化疗对晚期乳腺癌细胞凋亡的影响

目的 :探讨术前动脉灌注化疗诱导乳腺癌细胞凋亡发生情况及其对患者预后的影响。方法 :分别采用TUNEL法检测及电镜观察术前动脉灌注化疗 40例和术前未化疗的 42例晚期乳腺癌细胞凋亡发生情况 ,计算凋亡发生率及凋亡指数 ,并进行对比分析。结果 :术前动脉灌注化疗组与术前未化疗组凋亡发生率分别为 92 5 %和 78 5 % ,凋亡指数分别为 19 37± 6 49和 9 2 6± 5 0 4(P <0 0 1)。凋亡指数高低与局部晚期乳腺癌的无病生存率相关 (P <0 0 1)。结论 :局部晚期乳腺癌术前动脉灌注化疗可诱导乳腺癌细胞的凋亡 ,提高患者无病生存率。     

丙型肝炎病毒非结构蛋白NS3对端粒酶活性的影响

目的研究丙型肝炎病毒非结构区3(HCV NS3)蛋白对端粒酶活性的影响,以探讨HCV NS3蛋白在HCV致癌中的作用,并观察端粒酶活性原位检测法的应用价值.方法利用HCV NS3真核细胞表达质粒pRcHCNS3-5′(表达HCV NS3 N端多肽),pRcHCNS3-3′(表达HCV NS3C端多肽)和空白质粒pRcCMV转染NIH3T3细胞,分别获得11、11和8个阳性克隆;采用链霉素抗生物素-过氧化物酶法(SP)免疫组织化学方法检测转染的NIH3T3细胞中HCV NS3蛋白表达,并通过端粒酶活性原位检测法和端粒酶聚合酶链反应(PCR)酶联免疫吸附反应(ELISA)技术分别检测转染前后NIH3T3细胞端粒酶活性的定位和定量变化.结果 HCV NS3表达质粒pRcHCNS3-5′或pRcHCNS3-3′转染的NIH3T3细胞均表达HCV NS3蛋白,HCV NS3蛋白阳性信号均位于细胞质中,并以前者表达的阳性信号为强(χ2=6.667,P<0.05),各组细胞端粒酶活性存在显著差异(F=143.083,P<0.01),其中质粒pRcHCNS3-5′转染的NIH3T3细胞端粒酶活性最强,11个克隆均呈阳性,质粒pRcHCNS3-3′转染的细胞次之(P<0.05),空白质粒pRcCMV转染细胞和未转染NIH3T3细胞最弱;HCV NS3蛋白的表达水平和端粒酶活性强度之间具有显著相关性(rs=0.808 4,P<0.01);采用端粒酶活性原位检测方法和端粒酶PCR ELISA技术检测结果具有较好的一致性(rs=0.501 96,P<0.01).结论 (1) HCV NS3蛋白可能是通过内源性机制激活细胞端粒酶导致宿主细胞恶性转化;(2) HCV NS3蛋白 N端多肽对宿主细胞端粒酶的激活作用强于C端多肽;(3) 进一步证实端粒酶活性原位检测法是一种适合于病理形态与功能研究的技术.     

Construction and expression of nucleic acid vaccine pVAXl-h-alpha S1-140 coding human alpha-synuclein

BACKGROUND： The deposition of α -synuclein （ α -syn） aggregates is a neuropathological feature of Parkinson＇s disease. It remains impossible to involve α-syn aggregation in the treatment of Parkinson＇s disease. A nucleic acid vaccine will provide a new pathway to immunotherapy for Parkinson＇s disease. OBJECTIVE： To construct a recombinant eukaryotic expression vector pVAX1 coding human α -syn and to observe its expression level in COS-7 cells. DESIGN AND SETTING： The present bioengineering and molecular biology experiment was performed at Department of Neurology, First Affiliated Hospital of Chongqing Medical University ＆ Chongqing Key Laboratory of Neurology. MATERIALS： The eukaryotic expression plasmid pVAXI, human embryonic brain tissue, healthy human blood cells, and COS-7 cells were purchased from Promega Company, USA. METHODS： The full-length CDS sequence of the human a -syn gene was amplified by RT-PCR, which contained restriction sites for the enzymes Kpn Ⅰ, Xba Ⅰ and Kozak consensus sequence. Then the PCR products and eukaryotic expression vector pVAX1 were digested with Kpn Ⅰ and Xba Ⅰ simultaneously, and were extracted and ligated by T4 ligase. The recombinant constructs pVAX1-h α -S1-140 were transformed into competent E. coli TOP 1 0 cells and the positive clones were screened and selected using PCR analysis, restriction digestion analysis, and DNA sequencing. The constructs were then tested for protein expression in COS-7 cells by RT-PCR and Western blotting. MAIN OUTCOME MEASURES： Identification of an eukaryotic expression vector containing the human α -syn gene, pVAX1-h α-S1-140, and detection of the expression in mammalian cell COS-7. RESULTS： The pVAX1 vector was successfully cloned with human α -syn in the correct orientation and in-frame. The DNA vaccine constructs pVAX 1-h α-S1-140 with the human α-syn gene were shown to be expressed in COS-7 cells. Human α-syn was successfully expressed in the mammalian cell line and was detected by RT-PCR     

Clinical and Experimental Studies on Composite Divitriol Infusion in Treating Tinea Manum

CompositeDivitriolinfusion(CDI),aChineseherbalpreparationderivedfromErFanDecoction(:6Ktk),isadrugforexternaluseintreatingtineamanum.Inrecentyears,clinicalandexperimentalstudieshavebeencarriedoutonitbytheauthorsandtheresultwasreportedasfollows.METHODSClinicalMaterialsThe216casesoftineamanum,withtheirdiagnosisconfirmedbytypicalsymptomandmicroscopicexaminationoffungus,weredividedrandomlyintotwogroups.Theaverageageofthe139patientsintheCDIgroup(75malesand64females)was35years,rangingfrom16--…     

Significance of antineutrophil cytoplasmic antibody in adult patients with Henoch-Sch(o)nlein purpura presenting mainly with gastrointestinal symptoms

AIM: To test the clinical significance of antineutrophil cytoplasmic antibody (ANCA) in evaluation of adult Henoch-Schonlein purpura (HSP) patients presenting mainly with abdominal symptoms.METHODS: Twenty-eight consecutive HSP patients who presented predominantly with abdominal symptoms were enrolled in this study. Control subjects included 27 age-and sex-matched patients with peptic ulcer disease, colon cancer, acute gastroenteritis, irritable bowel syndrome and colonic polyps. ANCA was measured by indirect immunofluorescence (IIF) in all patients, and follow-up ELJSA was performed in patients with positive IIF tests.RESULTS: ANCA was detected in 9 HSP patients by IIF (2 were positive for c-ANCA and 7 were positive for p-ANCA). No ANCA was found in the control group. The sensitivity and specificity of a positive ANCA test (either c- or p-ANCA) were 32.1% and 100% respectively. Only one out of the 9 patients with positive ANCA by IIF had positive ANCA by ELISA and the antigen was myeloperoxidase (MPO). The patients positive for ANCA had higher HSP clinical scores, and were more likely to have renal function impairment. Patients with late purpura development were also associated with more severe clinical manifestations.CONCLUSION: A positive ANCA test is associated with more severe symptoms in HSP. After inflammatory bowel disease is excluded, a positive ANCA test provides a clue to the diagnosis of HSP presenting predominantly with abdominal symptoms.     

The carboxyl-terminal domain of inducible Hsp70 protects from ischemic injury in vivo and in vitro.

Heat shock protein (Hsp)70 can suppress both necrosis and apoptosis induced by various injuries in vivo and in vitro. However, the relative importance of different functions and binding partners of Hsp70 in ischemic protection is unknown. To explore this question, we tested the ability of Hsp70-K71E, an adenosine triphosphate (ATP)ase-deficient point mutant, and Hsp70-381-640, a deletion mutant lacking the ATPase domain and encoding the carboxyl-terminal portion, to protect against ischemia-like injury in vivo and in vitro. Heat shock protein 70-wild type (-WT), -K71E, -381-640, and control vector plasmid LXSN were expressed in primary murine astrocyte cultures. Astrocytes overexpressing Hsp70-WT, -K71E, or -381-640 were all significantly protected from 4 h combined oxygen-glucose deprivation and 24 h reperfusion when assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay or propidium iodide staining and cell counting (P < 0.05). Brains of rats were transfected with plasmids encoding Hsp70-WT, -K71E, -381-640, or LXSN 24 h before 2 h middle cerebral artery occlusion followed by 24 h reperfusion. Animals that overexpressed either of the mutant proteins or Hsp70-WT had significantly better neurological scores and smaller infarcts than control animals. Protection by both mutants was associated with reduced protein aggregation, as assessed by ubiquitin immunohistochemistry and reduced nuclear translocation of apoptosis-inducing factor. The results show that the carboxyl-terminal portion of Hsp70 is sufficient for neuroprotection. This indicates that neither the ability to fold denatured proteins nor interactions with cochaperones or other proteins that bind the amino-terminal half of Hsp70 are essential to ischemic protection.     

FRMD7基因新突变位点导致先天性眼球震颤一家系的遗传学研究

目的探讨4.1埃兹蛋白－根蛋白－膜突蛋白结构域包含的7(FRMD7)基因新突变位点导致先天性眼球震颤(CN)一家系的遗传学特征。 方法2021年10月,收集于北京大学人民医院眼视光中心确诊CN家系四代19例的临床资料。采集家系中3例CN患者和9例眼正常者的外周血样本,检查受试者的最佳矫正视力(BCVA)、眼前节及眼底;评估眼球震颤的类型、头位、是否有中间带及眼位等。应用GenCap液相仪抓取目标基因技术,获取与眼部疾病相关的811个基因的外显子区域及其侧翼区域,对先证者进行高通量测序,筛选出致病基因和突变位点,并在家系中进行Sanger测序和共分离验证。使用Mutation Taster软件分析突变位点的基因突变类型;预测突变型蛋白质的三维结构和功能改变。 结果该家系的先证者(Ⅲ6)男性,11岁。此患者右眼BCVA 0.7,左眼BCVA 0.8。先证者表弟(Ⅲ9)男性,2岁,双眼视力检查配合度不佳。先证者祖父(Ⅰ1)男性,73岁。此患者右眼BCVA 0.3,左眼BCVA 0.4,双眼晶状体混浊。3患者均双眼正位,眼球震颤呈水平钟摆型,无中间带及代偿头位,眼底检查未见明显异常。此家系患者均为男性,呈现隔代遗传特征,符合X连锁隐性遗传。家系中12例受试者外周血基因检测结果显示,3例患者FRMD7基因第9外显子编码区发生半合子变异,Ⅱ6、Ⅱ8及Ⅲ8等3例眼正常女性者在该区域发生杂合变异,核苷酸变异c.822C>A,氨基酸变异p.Y274X,使终止密码子提前出现,造成蛋白质编码提前终止,大量氨基酸丢失。根据美国医学遗传学与基因组学学会遗传变异分类标准与指南,该变异被评定为疑似致病性变异。经Mutation Taster软件分析,结果显示突变性质为无义突变;突变型蛋白质三维结构和功能的预测显示该突变影响FRMD7蛋白结构的稳定性,可能使其功能受损。 结论FRMD7基因核苷酸变异c.822C>A(p.Y274X)为无义突变,属于新突变位点,此变异是该家系CN的可能致病原因,扩大了FRMD7基因突变频谱。     

Experimental and Numerical Investigation of Intact,Defective and Repaired Countersunk Composite Joints under Tensile Loading

Bolted joints are commonly used for assembling carbon fiber/resin composite structures. Since drilling may generate defects at hole edges which affect mechanical properties, it is of great engineering significance to develop proper repair methods to restore the mechanical properties of the defective parts. However, there are few studies on hole edge defects and their repair methods. Therefore, a novelty short fiber filling repair method was proposed to repair defective holes in this study. The mechanical properties of intact, defective and repaired countersunk composite joints were compared and investigated. Experimental tensile tests showed that defective joints had lower initial stiffness and failure loads compared to intact joints, while the mechanical properties were effectively restored after repair. Three-dimensional finite element models were also established to analyze the damage process of the joints. Results of numerical modelling were consistent with the experimental results. The simulations showed that changes in contact behaviors and local deformations caused by hole edge defects led to the low initial stiffness and stiffness transition point of the joint, while this phenomenon was reduced after repair. Additionally, despite different joint types, laminate failure mainly occurred around the hole and countersink.     

利培酮对首发精神分裂症患者的疗效观察及与血浆催乳素水平的相关性研究

目的探讨非典型抗精神病药利培酮对首发精神分裂症患者血浆催乳素(PRL)水平的影响及其与疗效的关系。方法对30例符合CCMD-Ⅲ诊断标准的首发精神分裂症患者(治疗组) 在治疗前后及30例健康自愿者(对照组)的血浆PRL水平用放射免疫法进行检测。用利培酮治疗12 周,采用阳性和阴性症状量表(PAN SS)在治疗前后分别进行评定。结果患者组治疗前血浆PRL水平低于对照组,差异有统计学意义(P<0．05);治疗组治疗后血浆PRL水平明显高于治疗前(P< 0．01);治疗前、后在PAN SS总分及其因子分上差异均有显著性意义(P<0．05)。治疗后PRL水平较高、治疗前后差异较大或治疗前PRL水平较高者,疗效较好,尤其是阳性症状改善明显。结论利培酮治疗明显增加血浆PRL水平;且治疗前后血浆PRL水平与疗效相关,主要与阳性症状的疗效有关。