Screening Mouse Genes Related to Blastocyst Implantation by Using PCR Subtraction

Objective To identify genes that may be related to embryo implantation Materials & Methods The PCR subtraction technique was applied at implantation and inter-plantation sites on day 4. 5 of pregnancy in mice. Two novel Expressed Sequence Tags (ESTs ), EST8 and EST81 were identified; their expression in tissues was analyzed by Northern blotting, and their full-length cDNAs were synthesized by PCR.Results We found that these two novel ESTs (EST8and EST81) were noticeably expressed in implantation site in the mouse on day 4. 5 of pregnancy. EST8 was expressed at high level in livers and implantation sites of the mice, while at low level in ovaries and inter-plantation sites. EST81 was predominantly expressed in implantation site and ovary, and at low level in all other tissues. Their complete cDNAs, 1 665bp and 1 264 bp respectively, were synthesized by using PCR.Conclusion The two full-length cDNAs were responsible for embryo implantation,and their functions need to be further studied.     

小鼠胚泡着床相关基因的差减筛选

目的:筛选新的小鼠胚泡着床相关基因。方法:对孕4.5 d小鼠着床和非着床位点的子宫内膜组织进行PCR差减。获得了2个新的在小鼠胚泡着床位点高表达的EST(EST8和EST81)。结果:EST8在孕4.5 d小鼠着床位点、肝脏中表达较高,在非着床位点和卵巢中亦有微量表达。EST81主要在孕4.5 d小鼠子宫着床组织和卵巢中表达,其它各种组织中也有微量表达。用PCR方法获得了相应的全长。cDNA,长度分别为1665bp和1364bp。结论:用差减筛选获得的两种cDNA是孕小鼠着床相关基因,其功能有待进一步研究。     

卡巴胆碱对烧伤延迟复苏大鼠免疫功能的影响

Objective To explore the changing trend of Ia on monocyte, lymphocyte apoptosis rate, TNF-α and IL-6 in abdominal aorta of burned rats with delayed resuscitation and the influence of application of carbachol on them. Methods Adult male Wistar rats were randomly divided into normal control group(n =8), scald group(n =48) and scald with carbachol treatment group(n =48). In latter two groups, rats were inflicted with 30% total body surface area (TBSA) full-thickness scald and delayed fluid resuscitation. All scald rats were sacrificed at the 6th hours or 1st, 2nd, 3rd, 7th, 14th day after scald, with 8 rats at each time point. Expression of Ia antigen on monocyte and lymphocyte apoptosis rate were determined by direct immunofluorescence on a flow cytometer, and TNF-α and IL-6 was measured by ELISA. Results Expression of la on monocyte was obviously lower than that of controls. The lowest levels were recorded on the 6th hours and 1st day after scald. Subsequently, Ia was elevated gradually, but still lower than that of normal rats(P <0. 01). After administration of carbachol, Ia expression was obviously promoted, compared with the simple scald group (P <0. 01). Lymphocyte apoptosis rate, TNF-α and IL-6 was higher than that of controls(P <0. 01). After administration of cavachol, , lymphocyte apoptosis rate and TNF-α and IL-6 was obviously down-regulated on the 6th hours, 1st day, 2nd day and 3rd day after scald injury, compared with the simple scald group (P < 0. 01 or 0. 05). Conclusion After severe burn with delayed fluid resuscitation, there is a low la expression, high lymphocyte apoptosis rate and increased releasing of proinflammatory cytokine. Immune function was suppressed. Carbacho] could improve the immune function of scald rats with delayed fluid resuscitation.     

Establishment of Germ-line Competent C57BL/6J Embryonic Stem Cell Lines

Objective To establish C57BL/6J embryonic stem （ES） cell lines with potential germ- line contribution Methods ES cells were isolated from blastocyst inner cell mass of C5 7BL/6J mice, and cultured for 15 passages, and then injected into blastococels of ICR mice blastocysts to establish chimeric mice. Results Three ES cell lines （mC57ES1,mC57ES3, mC57ES7） derived from the inner cell mass of C57BL/6J mice blastocysts were established. They were characteristic of undifferentiated state, including normal XY karyotype, expression of a specific cell surface marker “stage-specific embryonic antigen-I” and alkaline phosphatase in continuous passage. When injected into immunodeficient mice, mC57ES1 cells consistently differentiated into derivatives of all three embryonic germ layers. When mC57ES1 cells were transferred into ICR mice blastocysts, 4 chimeric mice have been obtained. One male of them revealed successful germ-line transmission. Conclussion We have obtained C57BL/6J ES cell lines with a potential germ-line contribution, which can be used to generate transgenic and gene knock-out mice.     

地震救援中实施远程医疗服务:把最好的医学专家送到灾区人民身边

目的和方法 总结解放军总医院远程医疗会诊系统在2013年“4·20”四川芦山地震救援中发挥其远程医疗服务的经验,探讨远程医疗会诊系统在灾难救援中的作用.结果和结论 在2013年4月20日四川芦山地震后,解放军总医院专家医疗队及时将无线便携式远程医疗会诊系统发送至地震灾区伤员集合点医院,第一时间组织开通面向灾区服务的远程医疗会诊系统,与位于地震灾区的多家医院及地震救援队救护车远程医疗系统联通,提供24h包括急诊和危重症救治、骨科、脑外科、肝胆外科、妇产科等相关专业的远程救援服务,对地震伤员的抢救提出了诊断和救治意见,形成了救灾现场专家与后方专家的良性互动,使灾区专家医疗队的救治能力倍增.解放军总医院在芦山地震中共进行远程医疗会诊110余例,在对灾难现场的受伤人员进行有效的紧急救治中实现了实时高清会诊和现场手术指导,并在业务层面实现了多媒体通信系统与医院医疗信息系统的对接,在功能层面实现了各联盟医院的视频、音频和数据以及医疗业务的互联互通,为我国在灾难救援中实施远程医疗抢救提供了可借鉴的经验.     

微小RNA调节骨骼肌发育与损伤修复的作用及其机制

microRNAs是一类非编码小RNAs分子,新近发现其具有重要的调节基因表达的功能,它能通过抑制翻译和降解靶mRNA来负性调控转录后水平的基因表达,miRNAs已经被证实在肌肉发育和肌细胞增殖和分化的调节中具有重要作用。最近研究发现,肌肉特异性转录因子控制一些microRNAs的表达,通过多种机制调节肌肉发育和功能。结合信息学、生物化学和遗传基因学方法,不仅将阐明骨骼肌microRNAs调控网络,更好地理解肌肉组织的调节,还将通过鉴定候选microRNAs的潜在临床应用靶点,增加肌肉营养不良的治疗干预的新机会。     

Plumbagin attenuates cancer cell growth and osteoclast formation in the bone microenvironment of mice

Aim： To investigate the effects of plumbagin, a naphthoquinone derived from the medicinal plant Plumbago zeylanica, on human breast cancer cell growth and the cancer cell-induced osteolysis in the bone microenvironment of mice.
Methods： Human breast cancer cell subline MDA-MB-231SA with the ability to spread and grow in the bone was tested. The cell proliferation was determined using the CCK-8 assay. Apoptosis was detected with Annexin V/PI double-labeled flow cytometry. Red fluorescent protein-labeled MDA-MB-231SArfp cells were injected into the right tibia of female BALB/c-nu/nu mice. Three days after the inoculation, the mice were injected with plumbagin （2, 4, or 6 mg/kg, ip） 5 times per week for 7 weeks. The growth of the tumor cells was monitored using an in vivo imaging system. After the mice were sacrificed, the hind limbs were removed for radiographic and histological analyses.

Results： Plumbagin （2.5–20 μmol/L） concentration-dependently inhibited the cell viability and induced apoptosis of MDA-MB-231SA cells in vitro （the IC50 value of inhibition of cell viability was 14.7 μmol/L）. Administration of plumbagin to breast cancer bearing mice delayed the tumor growth by 2–3 weeks and reduced the tumor volume by 44%–74%. The in vivo imaging study showed that plumbagin dose-dependently inhibited MDA-MB-231SArfp cell growth in bone microenvironment. Furthermore, X-ray images and micro-CT study demonstrated that plumbagin reduced bone erosion area and prevented a decrease in bone tissue volume. Histological studies showed that plumbagin dose-dependently inhibited the breast cancer cell growth, enhanced the cell apoptosis and reduced the number of TRAcP-positive osteoclasts.

Conclusion： Plumbagin inhibits the cell growth and induces apoptosis in human breast cancer cells in mice bone microenvironment, leading to significant reduction in osteolytic lesions caused by the tumor cells.     

高渗晶胶复合液静脉补液在犬烧伤早期防治休克疗效观察

目的研究应用高渗晶胶复合液快速静脉补液对35%TBSA烧伤犬伤后早期休克的防治作用及对血流动力学的影响。方法 Beagle犬12只,采用凝固汽油燃烧法制作Ⅲ度烧伤面积35%TBSA模型,随机分为常规补液组(A组)和高渗补液组(B组)。A组按照4ml/(kg.1%TBSA)输入乳酸钠林格氏液,在前8h输入总量的一半,B组采用高渗氯化钠羟乙基淀粉40注射液,按照8ml/kg快速补液后终止补液。分别于伤前、伤后1h、2h、4h、6h、8h进行血流动力学检测,测定各组犬的胸腔内血容量指数(ITBI)、血管外肺水量(EVLW)、血容量(PV)及血浆渗透压。结果烧伤后两组ITBI均呈逐步降低趋势,B组经高渗补液后,伤后1h其ITBI降低与A组相比差异无统计学意义(P0.05),至伤后2h开始明显降低,4h后显著低于A组,两组相比差异有统计学意义(P0.05);A组EVLW显示伤后渗出增多,而B组经高渗补液,EVLW显示伤后2们h内血管外渗出较伤前差异无统计学意义(P0.05),渗出量显著低于A组,两组相比差异有统计学意义(P0.05);伤后A组PV明显降低,B组经高渗补液,PV在伤后1、2、4h降低缓慢,且PV明显高于A组,两组相比差异有统计学意义(P0.05),伤后4h后PV迅速减少,并显著低于A组,两组相比差异有统计学意义(P0.05);伤后A组血浆渗透压有所降低,B组经高渗补液,血浆渗透压在伤后1、2h较伤前增高,且显著高于A组(P0.05),两组相比差异有统计学意义,伤后4h后逐步降低,至伤后8hB组显著低于A组,两组相比差异有统计学意义(P0.05)。结论烧伤早期快速静脉补充高渗晶胶复合液,在4h内能够有效维持血流动力学指标稳定,并减少组织渗出,从而起到大大减少补液量、节约医疗资源、降低并发症、提高复苏疗效的作用,适于危重烧伤早期院外现场急救。     

四川省分离的基因1型乙型脑炎病毒分子特征分析

目的 从四川省巴中市采集的蚊虫标本中分离乙型脑炎(简称乙脑)病毒(JEV),确定其基因型别,并分析相关的基因1型乙脑病毒PrM和E基因区段氨基酸序列特征.方法 对2004年采集蚊虫标本进行病毒分离,对新分离的乙脑病毒进行生物学、血清学及分子生物学鉴定.逆转录聚合酶链反应(RT-PCR)扩增新分离JEV的PrM、E区段核苷酸序列,测序后应用Clustal X软件做碱基配对分析,MEGA4软件完成病毒进化分析,GENEDOC(3.2)软件完成氨基酸位点分析,根据蜱传脑炎病毒可溶性蛋白晶体结构为模板进行乙脑病毒E蛋白三维结构模拟预测分析.结果 共采集4668只蚊虫标本,主要是骚扰阿蚊和库蚊,分离到6株病毒,经鉴定均属于基因1型的乙脑病毒.将四川省分离的6个毒株结合我国新分离的基因1型乙脑病毒与减毒活疫苗株SA14-14-2株的PrM区段和E区段氨基酸比较,发现PrM区段在PrM2、64和65位存在基因1型乙脑病毒独有的氨基酸位点差异,E区段存在14处共同的氨基酸位点差异,其中在E129、222、327和366位点为中国目前分离到的基因1型乙脑病毒所特有的位点特征.结论 从四川省巴中市首次分离到基因1型的乙脑病毒,并发现基因1型乙脑病毒与减毒活疫苗株之间PrM、E基因区段存在氨基酸差异,但现行疫苗株理论上可以保护新分离的基因1型乙脑病毒.     

异种去细胞真皮基质在体内转归的初步研究

目的 初步研究异种（猪）去细胞真皮基质在体内的转归。方法 经曲通（Triton）X-100、胰蛋白酶处理得到异种（猪）去细胞真皮基质后，将其包埋于SD大鼠皮下，于术后1、2、4、6、8、12、16、20和30周观察异种（猪）去细胞真皮基质的大体形态及组织学变化。结果 制备的异种（猪）去细胞真皮基质去细胞成分，胶原纤维排列有序，基底膜结构完整。大体观察异种（猪）去细胞真皮基质原始形态随时间延长而逐渐模糊．组织学观察异种（猪）去细胞真皮基质1周以炎性细胞浸润为主，以后成纤维细胞、毛细血管逐渐增多，真皮胶原排列逐渐规则、致密。结论 异种（猪）去细胞真皮基质可在体内作为支架长期存在，并可诱导自身成纤维细胞及毛细血管有序长入。