红藻氨酸作用后海马神经元胞膜表面的超微结构的原子力显微镜观察

目的 观察原代培养神经元在红藻氨酸(KA)作用下细胞表面形态的三维构像变化。方法 利用原子力显微镜(AFM)对大鼠海马神经元经不同浓度的KA(25和250μmol／L，50和100nmol／L)分别作用10min和100min后的表面结构进行纳米级水平扫描和观测。结果 正常海马神经元表面光滑，起伏均匀、规律；KA作用后神经元呈退行性改变，表现为胞体肿胀，胞膜表面粗糙，出现隆起和“孔洞”样结构，并且其变化程度分别与作用时问和KA浓度呈量-效关系。结论 KA对海马神经元毒性作用后胞膜表面超微结构所产生的明显变化，可借助AFM清晰观察，并定量分析。     

Preparation and identification of monoclonal antibodies against Penicillium marneffei]

OBJECTIVE: To prepare and identify monoclonal antibodies (mAbs) against Penicillium marneffei. METHODS: Recombinant mannoprotein1 (MP1) of Penicillium marneffei was used to immunize BALB/c mice, and anti-MP1 mAbs were obtained by means of hybridoma. Screening and identification of the mAbs were subsequently performed with indirect enzyme-linked immunosorbent assay and Western blotting, respectively. RESULTS: Four hybridomas producing antibodies against Penicillium marneffei were obtained, and the IgG isotypes of the 4 mAbs were identified as IgG1 with affinity constants (K) of 8.2x10(-9), 4.7x10(-9), 6.5x10(-9) and 2.7x10(-9), respectively. Western blotting demonstrated specific recognition of Aspergillus fumigatus MP1 by the obtained mAbs. CONCLUSION: The 4 hybridomas producing anti-MP1 mAbs with high specificity and affinity can be of significant value in the diagnosis of Penicilliosis marneffei infections.     

川芎嗪及复方丹参注射液辅助治疗妊娠高血压综合征60例临床分析

目的 比较川芎嗪注射液及复方丹参注射液在辅助治疗妊娠高血压综合征(PHI)中的作用，探讨两药在改善微循环及胎儿宫内缺氧方面的差异，进一步指导临床用药。方法 选取2001年6月～2002年6月在我院住院治疗的60例PHI患者，平均分为2组，在有效镇静、解痉、降压、低流量吸氧等治疗的基础上，分别给予静脉滴注川芎嗪注射液及复方丹参注射液治疗1个疗程。结果 川芎嗪组抑制纤溶、改善微循环的效果显著；复方丹参组主要降低血液粘稠度、降低胆圊醇及血酯。结论 川芎嗪及复方丹参对PHI有肯定的治疗作用，由于两药的临床作用不同，需有针对性用药。     

TNP-470对裸鼠体内人结肠癌细胞增殖和凋亡的影响

目的 研究TNP-470对裸鼠体内人结肠癌细胞增殖和凋亡的影响。方法 将16只人结肠癌皮下移植瘤裸鼠随机分成实验组和对照组，实验组隔日予以TNP-47030mg/kg，皮下注射，对照组予以相同体积的生理盐水，4周后处死，应用免疫组织化学法（免疫组化）和图象分析技术检测肿瘤细胞中增殖细胞核抗原（PCNA）的表达，TdT介导的dUTP缺口末端标记（TUNEL）法检测肿瘤细胞的凋亡，透射电镜观察凋亡细胞的超微结构。结果 TNP-470明显抑制了肿瘤的生长，抑瘤率为45.53%。实验组肿瘤中PCNA的表达显低于对照组，凋亡指数上升，电镜下可见典型的细胞凋亡形态学变化。结论 TNP-470除了通过抗血管生成作用抑制肿瘤生长外，还可通过抑制LOVO细胞本身的增殖和诱导凋亡发挥抗肿瘤作用。     

经蓝碟(LapDisc)手助腹腔镜结直肠癌根治术

目的 探讨手助腹腔镜结直肠癌根治术的临床效果。方法 应用LapDisc手助腹腔镜技术完成27例结直肠癌根治术。结果 手术全部成功，无一例中转开腹。手术时间90～260min，平均140min。术中出血50～200ml，平均110ml。术后无死亡及吻合口漏等并发症。随访6～23个月，平均8．6月，未见切口种植复发。结论 手助腹腔镜结直肠癌根治具有安全、创伤小、术后恢复快及降低标准腹腔镜手术难度等优点，值得临床推广应用。     

子宫内膜异位症猕猴动物模型的建立

目的构建子宫内膜异位症(EM)猕猴动物模型。方法选择5只健康、月经规律的雌性猕猴(4只实验猴、1只对照猴),在月经第8~15天,雌激素高峰的第3~5天,手术将猕猴的子宫内膜种植到盆腔子宫腔以外的部位,对照猕猴用大网膜取代内膜种植。在术后第2、4个月动物月经周期的第8~15天开腹或腹腔镜探查。结果术后第2个月探查,4只实验猴中,3只异位内膜种植存活,其中2只形成异位囊肿。术后第4个月,对2只已形成EM的猕猴行腹腔镜再次探查,其盆腔粘连,异位内膜生长情况与术后第2个月基本相似;3只形成EM的猕猴精神状态、饮食、体质量增长情况均较对照猕猴差,2只发生肠套叠死亡。结论本研究根据种植学说原理成功地构建了子宫内膜异位症猕猴动物模型,但猕猴的个体差异是模型是否成功的一个关键因素,因而认为遗传因素是子宫内膜异位症发病中的另一重要机制,子宫内膜异位生长对机体的整体机能有较为明显的影响。     

肝吸虫病合并胆管癌29例报告

目的 探讨肝吸虫病与胆管癌之间的病理机制及其临床诊治经验。方法 回顾性分析肝吸虫病合并胆管癌病人29例。结果男性21例，女性8例，平均年龄62．5岁，高分化腺癌13例(44．8％)，低分化腺癌16例(55．2％)。根治性肿瘤切除术21例(72．4％)，姑息性内引流术5例，剖腹探查活检术3例；生存10年者2例，5年者3例，3年者6例，1年者13例，6个月者4例，3个月者1例。结论 (1)肝吸虫感染、肝吸虫卵的沉积是诱发胆管上皮细胞增生、癌变的重要因素；(2)本病诊断有赖于病理，术中病理活检的广泛应用是提高其诊断符合率的有效措施；(3)治疗仍是以手术为主的综合治疗，能根治性切除则预后佳，不能根治者解决胆道梗阻也可延长生命。     

人自然杀伤细胞抑制性受体P58.2基因克隆与鉴定

目的：P58．2基因克隆与鉴定。方法：采用RT—PCR技术，从正常人外周血单个核细胞中扩增自然杀伤细胞抑制性受体P58．2基因全长cDNA，经酶切后构建重组克隆载体，双酶切和测序鉴定。结果：RT—PCR获得了预期的扩增产物P58．2全长cDNA，成功构建了pSPORT1-P58．2重组克隆载体，酶切、酶谱分析与预期结果相符。DNA测序结果与GenBank登记的人P58．2cDNA全长碱基序列一致。结论：pSPORT1-P58．2重组克隆载体构建成功；P58．2基因序列与文献报道一致，为下一步构建表达载体和基因转染等工作打下良好的基础。     

血管内皮生长因子受体启动子驱动重组腺病毒双自杀基因系统选择性杀伤大肠癌细胞

OBJECTIVE: To observe the selective killing effect of adenovirus (Ad)-mediated double suicide gene driven by kinase domain-containing receptor(KDR) promoter on human colorectal cancer LoVo cells and human umbilical vein endothelial ECV304 cells. METHODS: The plasmid pAdEasy-KDR-CDglyTK was transfected into 293 packaging cells for amplification of the infectious Ad and used to infect the KDR-producing cells (ECV304 and LoVo) and the KDR-nonproducing cells (LS174T) respectively. The three cells were treated with the prodrugs 5-flurocytosine (5-FC) and ganciclovir (GCV) at different concentrations after infection. The killing effects of the fusion gene system on the cells were evaluated. The distribution of cell cycle was detected by flow cytometry. RESULTS: The infection rates of the recombinant Ad were similar among the 3 cells, gradually increasing with the increment of multiplicity of infection (MOI) and reaching 100% with the MOI of 200. The LoVo cells and ECV304 cells infected with Ad-KDR-CDglyTK were highly sensitive to both of the prodrugs (P>0.1), whereas the infected LS174T cells failed to exhibit similar sensitivity (P<0.001). The killing effect of CD/TK fusion gene on the target cells was much stronger than that of either suicide gene (P<0.001). The cell cycle of LoVo cells was arrested at G1 phase. CONCLUSION: The CD/TK fusion gene system driven by KDR promoter can selectively kill KDR-expressing human colorectal cancer LoVo cells and endothelial cells.     

双阴性选择法纯化培养人骨髓多能成体祖细胞

OBJECTIVE: To establish a method for purifying and culturing human multipotent adult progenitor cells (MAPCs) in vitro. METHODS: Mononuclear cells were separated from the bone marrow of healthy adult volunteers by density gradient centrifugation and cultivated in adherent culture. The plastic-adherent cultured bone marrow cells were isolated by magnetic-activated cell sorting with CD45 and GlyA magnetic microbeads, and the purity of CD45-/GlyA- cells evaluated by flow cytometry. Phase-contrast microscopy was used to detect morphological changes of the cells in different stages of culture. RESULTS: Approximately (5-10)x10(4)/ml MAPCs could be separated from every 1x10(6)/ml bone marrow mononuclear cells by magnetic- activated cell sorting. The viability of the cells before and after separation was (96.7+/-1.7)% and (96.0+/-2.4)%, respectively. The isolated MAPCs grew well in a self-prepared culture medium till the16th passage. The purity of CD45-/GlyA- separated from the bone marrow was more than 98% as examined by flow cytomety even till the 12th passage. CONCLUSIONS: MAPCs derived from adult human bone marrow can be purified by magnetic-activated cell sorting with CD45 and GlyA microbeads and retain the undifferentiated state for a long time. The self-prepared culture medium is appropriate for MAPCs cultures in vitro.