RETT综合征患儿一例MECP2基因突变分析

目的通过对甲基化CpG结合蛋白2(mehty1-Cp G binding protein 2,MECP2)基因的突变分析,对1例典型的Rett综合征患儿进行基因诊断,并为该家庭提供遗传咨询。方法采用聚合酶链反应和DNA直接测序对先证者及其父母MECP2基因的4个外显子进行序列分析,同时对患儿进行染色体核型分析以排除染色体异常。结果患儿核型正常。针对MECP2基因进行突变分析发现患者存在c.473CT(T158M)杂合突变,其父母未检测到该突变。结论错义突变T158M是导致该RETT家系患者临床表型的主要原因,通过对RETT家系个体Mecp2基因分析可对REET家系进行有效的遗传咨询。     

围术期心理干预的护理研究

目的 探讨心理干预在缓解围术期应激反应的作用及对术后恢复的影响。方法 术前应用松弛训练、矫正认知、信息疗法等心理学方法对病人进行心理干预。结果 通过心理干预手段位病人围术期的血压、心率较对照组明显稳定；术后疼痛心理评分，6h以后明显低于对照组；术后在肠蠕动恢复及下地时间方面较对照组明显提前。结论 心理干预能减轻患者围术期的应激反应，并能促进患者术后的恢复。     

抗vWF-A3区特异性单抗SZ-123单链抗体的构建和表达

目的 以能与vWF-A3区特异结合的单克隆抗体SZ-123为模板,构建其单链抗体,为深入研究vWF-A3结构与功能的关系提供有力的工具.方法 通过逆转录及多聚酶链反应,扩增并克隆单抗SZ-123的可变区基因VH、VL,经测序后加入连接肽,构建成SZ-123单链抗体(SZ-123 ScFV),并在大肠杆菌中进行表达.采用酶联免疫吸附试验( ELISA)和Western blot方法验证SZ-123 ScFV的免疫原性.结果 VH、VL可变区基因符合小鼠抗体可变区特征,SZ-123 ScFV基因拼接正确.表达产物除少量为分泌型外,主要以包涵体形式存在,表达量占菌体蛋白的20％.经过变性和复性后,获得具有生物活性的高纯度单链抗体片段.结论 成功表达了SZ-123单链抗体,该小分子抗体具有特异结合vWF-A3区的能力.     

大鼠抗小鼠重组膜联蛋白A2单克隆抗体的制备与鉴定

目的建立能够分泌特异性抗小鼠膜联蛋白A2（AnxA2）的大鼠-小鼠融合单克隆抗体杂交瘤细胞系。方法利用重组小鼠AnxA2蛋白作为抗原免疫Wistar大鼠,通过细胞融合和杂交瘤筛选技术建立特异分泌抗小鼠AnxA2蛋白的大鼠-小鼠融合单抗杂交瘤细胞株。采用酶联免疫吸附方法确定单克隆抗体亚型,免疫印迹和流式细胞术验证所筛选的单克隆抗体的特异性。结果通过对30多个分泌抗体的融合细胞克隆的筛选,获得一株分泌特异性抗小鼠AnxA2的大鼠-小鼠融合单克隆抗体杂交瘤细胞系SZ-158,连续培养传代后染色体分析证实,Wistar大鼠脾细胞与小鼠SP2/0细胞稳定融合;免疫印迹和流式细胞术显示该单克隆抗体可以特异识别小鼠AnxA2重组蛋白。结论该研究所获得的大鼠-小鼠融合单克隆抗体特异性针对小鼠AnxA2蛋白,可用于AnxA2转基因小鼠中AnxA2表达的检测,为深入研究AnxA2的功能提供了物质保障。     

硼替佐米对内皮细胞迁移及血管新生的影响

Objective To investigate the effects of bortezomib on the migration of endothelial cells and the expression of angiogenesis-related molecules,and explore the mechanism of its antiproliferation of tumor cells.Methods Cell count kit CCK-8 was used to detect the relative proliferation activity of cells after treated by bortezomib at different concentrations for 12 h and 24 h,respectively.Transwell model was uesd to detect the migration rate of cells.Expression levels of VEGF and Annexin A2 genes were determined by realtime quantitative PCR.Annexin A2 protein was validated by Western blot.Results After treated with bertezomib at concentrations of 2.5、5.0 and 10 nmol/L for 12h,respectively,the HMEC-1 cell proliferation activity was 1.004±0.002,0.793±0.021 and 0.874 4-0.062,respectively,being no statistical difference from that of control group(1.000)P<0.05);while the migration rates of them were 0.697±0.060,0.597±0.090 and 0.874±0.062,respectively,being significantly lower than that of control group(1.000)(P<0.05 ) and so did for the expression of VEGF and Annexin A2 genes.After treated with 5 nmol/L bortezomib for 12 h.the Annexin A2 and VEGF gene relative expression level of HMEC-1 cells was 0.540±0.001 and 0.793±0.153,respectively,being of statistical difference from that of control group(1.000)P<0.05).The conspicuous downregulation of Annexin A2 protein was also confirmed by Western Blot.Conclusions Bortezomib can inhibit migration of endothelial cell HMEC-1 by downregulating the expression of VEGF and Annexin A2, displaying a new mechanism of bortezomib for inhibition of tumor proliferation.     

VEGF与COX-2在多发性骨髓瘤病人中的表达及意义

本研究探讨血管内皮生长因子（VEGF）和环氧化酶-2（Cox-2）在多发性骨髓瘤（MM）患者中的表达及临床意义。收集29例MM患者血清标本,用ELISA法测定血清中VEGF含量,Western blot法测定血清中Cox-2含量。结果表明：MM患者血清VEGF含量（365．34±65．63）Pg／ml明显高于正常对照组VEGF含量（122．52±39．29）pg／ml（P〈0．05）,在疾病进展期VEGF含量（395．07±54．90）pg／ml高于稳定期VEGF含量（300．33±44．22）pg／ml（P〈0．05）。MM患者血清Cox-2表达阳性率为31％,高于正常对照组阳性率0％（P〈0．01）,疾病进展期Cox-2表达阳性率为50％,高于稳定期阳性率21％（P〈0．01）。结论：VEGF和Cox-2水平在MM发病中有一定意义,可作为评估病情的重要指标。     

恶性血液病患者骨髓AnnexinⅡ基因的表达

本研究探讨恶性血液病患者骨髓细胞annexinⅡ基因的表达水平及其在恶性血液病发生发展中的作用。用实时定量PCR方法检测了81例初治急性白血病患者、6例多发性骨髓瘤患者（MM）及20例缺铁性贫血患者（对照）骨髓annexinⅡmRNA的表达水平。结果显示：annexinⅡ在初治AML—M3患者骨髓中的表达显著高于其它类型恶性血液病患者和对照者,此外在初治AML—M5、AML—M4、MM患者骨髓中也有较高表达,而在初治AML—M,以及AML—M,患者中表达量与对照者相比无统计学差异。结论：annexinⅡ除在AML—M,中高表达外,在临床观察具有高度侵袭浸润能力的类型M5、M4、MM中也有较高表达,推测annexinⅡ可能通过增强恶性血液病细胞降解细胞外基质等机制参与恶性血液病侵袭、浸润过程。     

产前诊断ins(10；13)(q11.2；q31q33)mat染色体异常1例

患者 女, 34岁, G3P2, 表型及智力未见异常, 于2017年8月5日因继发不孕症来郑州大学第一附属医院就诊。遗传咨询后夫妇双方进行外周血染色体检查, 患者核型为46, XX, ins(10;13)(q11.2;q31q33)(图1), 患者丈夫核型未见异常。患者要求通过植入前遗传学检测技术进行辅助生殖, 成功受孕后, 于孕16+4周时进行低深度全基因组拷贝数变异测序(copy number variation sequencing, CNV-seq)和染色体核型分析, 胎儿CNV-seq分析结果未见异常, 胎儿的核型分析结果为46, XX, ins(10;13)(q11.2;q31q33)mat(图2)。本研究通过了郑州大学第一附属医院伦理委员会的审查(KS-2018-KY-36), 夫妻双方均签署了临床研究知情同意书。     

FISH快速检测自然流产绒毛染色体非整倍体异常的临床价值

目的:探讨应用荧光原位杂交技术(FISH)对早期自然流产绒毛染色体非整倍体检测的临床价值。方法:对30例因自然流产行清宫术的绒毛组织行FISH分析,使用7种探针对13、16、18、21、22号和X、Y染色体进行了检测,并对这30例流产夫妇行外周血淋巴细胞染色体常规核型分析。结果:FISH分析的30例自然流产的绒毛组织中,有17例检测出了异常信号,检出率为57%,其中8例16-三体、2例22-三体、2例13-三体和5例三倍体。30例自然流产夫妇外周血淋巴细胞染色体核型未见异常。结论:FISH技术可以快速、简便地检测出流产物绒毛组织染色体非整倍体的异常,FISH技术的应用可以为自然流产夫妇遗传咨询提供重要的信息。     

Effects of thalidomide on Annexin Ⅱ gene regulation

Objective To investigate the effect of thalidomide on Annexin Ⅱ (AnxA2) gene regula-tion in multiple myeloma cell line RPMI8226 and human microvascular endothelial cell line HMEC-1 cells in vitro, and explore the potential mechanism of thrombosis induced by thalidomide. Methods RPMI8226 and HMEC-I cells were cultivated in vitro. Real time quantitative PCR (RQ-PCR) was used to detect the influ-ence of thalidomide at different concentration on the expression of AnxA2 mRNA, flow cytometry(FCM) and confocal microscopy were used to detect the cell surface protein level after the samples were stimulated with different concentrations of thalidomide. Results AnxA2 mRNA level in RPMI8226 cells treated with thalido-mide at 12.5 μg/ml, 25.0 μg/ml and 50.0 μg/ml was decreased compared with the control group (0.60 ±0. 15, 0.33 ± 0. 14, 0.42 ±0. 16, vs 1.07 ±0. 16, respectively, P <0.05)and did so in HMEC-1 cells (0.21 ±0.20, 0.08 ±0.08, 0.17 ±0. 16 vs 1.16 ±0.24, respectively, P <0.05). The AnxA2 protein lev-el in RPMI8226 cells treated with above mentioned concentrations of thalidomide was also decreased compared with the control (3.39 ± 0.32, 2.82 ± 0.28, 3.21 ± 0.23 vs 5.53 ± 0.32, repectively, P < 0. 05) and that did so in HMEC-1 cells (0.72±0. 11, 0.64 ±0.08, 0.67 ±0.08 vs 1.40 ±0. 15, respectively, P<0.05). Conclusions Thalidomide can inhibit the expression of AnxA2 mRNA and protein in RPMI8226 and HMEC-1 cells, which may be one of the mechanisms for the development of thrombosis induced by thalido-mide in multiple myeloma patients.