Ultrastructure of Amelanotic Melanocytes from Human Hair Follicles

Objective: To investigate the ultra structure of amelanotic melanocytes (AMMC). Methods: The hair follicles obtained from normal human scalp by 0.50% collagenase type V treatment were washed with 0.1mol/L phosphate buffer salt (PBS). Hair-follicle cell suspensions were prepared by trypsin treatment and cultured in melanocyte medium. Remaining keratinocytes were removed by differential trypsinization. 100μg/ml geneticin was used to eliminate the contaminating fibroblasts. At third passage, the cells were trypsinized, and then washed in phosphate-buffered saline and processed for transmission electron microscopy. Results: Under transmission electron microscope, the cultured cells showed round or oval shape, with single large nuclear and the karyotheca were double deck. There were obvious euchromosome within the nucleus, and sparse heterochromosome. There were various organelles in the cytoplasm, including plentiful melanosomes with nearly similar size, mitochondria, rough endoplasmic reticule (RER) and ribosome. The electron density granules in most of the melanosomes disposed along concentric circularities. Golgi apparatus in the cells was inconspicuous. Conclusion: The ultra structure of AMMC from human hair follicles is different from that of epidermal melanocytes, and these characteristics determine the functional immature of AMMC.     

Identification of testis development and spermatogenesis-related genes in human and mouse testes using cDNA arrays

We have constructed cDNA microarrays from the human testis large insert cDNA library, containing 9216 genes, together with several housekeeping genes. The cDNA microarrays were used to identify gene expression differences between human fetal and adult testes. Of >8700 hybridized clones, 731 exhibited significant differential expression characteristics. About 7500 genes were identified when the same cDNA microarrays were used for hybridization with cDNA probes from mouse testis, with 256 genes having significant differential expression between the age of 1-4 weeks. Among these genes, 101 were identified as critically related to testis development and possibly to spermatogenesis since they were found in both human and mouse testes, and expressed differentially at different stages of testis development. Of the 101 development-related genes, 59 full-length cDNAs have been sequenced previously, while the full-length cDNAs of the other 42 genes have not been published. We have obtained 11 full-length sequences of the 42 genes and deposited them in the GenBank. The conserved testis development-related genes found in both human and mouse testes may include genes that are likely to be involved in testicular functions, especially spermatogenesis, thus providing a basis for further functional characterization of the genes in mouse models.     

机械牵张对大鼠心室肌蛋白激酶B磷酸化及心钠素分泌的影响

目的:研究机械牵张对大鼠心肌蛋白激酶B(Akt)活化和心钠素(ANF)分泌的影响。方法:采用Langendorff方法灌流大鼠心脏,膨胀球囊持续牵张左心室,从左心室游离心肌,提取胞浆蛋白,用Western blot检测磷酸化Akt、总Akt水平;收集冠脉流出液,用放射免疫分析法检测冠脉流出液中ANF含量。结果:持续牵张不影响灌流心脏的心率和冠脉流出量。但经过20min持续牵张,牵张组心脏灌流液中ANF含量(209.89±65.45pg/ml)较对照组(108.84±25.18pg/ml)明显增高(P<0.01);牵张组大鼠左心室心肌组织磷酸化Akt水平(0.76±0.03)明显高于对照组(0.32±0.02),而总Akt水平与对照组相比无显著性差异。结论:机械牵张可引起心脏心钠素的分泌增加,其机制可能与胞内Akt信号通路的激活有关。     

阿尔茨海默病、阿尔茨海默病混合型及血管性痴呆患者心理及行为症状的比较

目的 比较阿尔茨海默病(Alzheimer Disease，AD)、AD混合型痴呆(Mixed dementia，MD)、血管性痴呆(Vascular dementia，VD)心理和行为症状(Psychological and behavioral symptoms of dementia，PBSD)的特征。方法 AD、MD及VD患者各30名参加本研究。采用Alzheimer病行为症状评定量表(The Begavioral Pathlolgy in Alzheiner Disease Rating Scale，BEHAVE—AD)、Cohen—Masfield激惹性问卷(The Cohen Mansfield Agitation Inventory，CMAI)评定痴呆患者BPSD。结果 AD患者激惹、焦虑与恐惧发生率较高，VD患者无目的游荡发生率、严重程度较低，MD患者BPSD症状无特异性。结论 AD、VD患者BPSD症状有特异性，MD患者BPSD表现无特异性。     

Molecular cloning and expression of a novel alternative splice variant of BRDT gene

In this study, a human adult testis cDNA microarray was constructed and hybridized with (33)P-labeled human adult testis, embryo testis and sperm cDNA probes, respectively. A novel alternative splice variant of BRDT gene, named BRDT-NY, presumably involved in testicular function was cloned. It was expressed 3.96-fold more in human adult than embryo testis and also expressed in human spermatozoa. Similarly, RT-PCR revealed a differential expression pattern of this gene in human adult testes and fetal testes. The full length of BRDT-NY was 3438 bp and contained a 2883 bp open reading frame, encoding a 960-amino-acid protein. Sequence analysis showed that it has two bromodomains in N-terminal of the protein. Multiple tissue RT-PCR results showed that BRDT-NY was exclusively expressed in testis. mRNA expression of BRDT-NY gene was deleted in some azoospermic patients' testes. These experiments suggested that BRDT-NY gene may have an important role in the process of spermatogenesis and may be correlated with male infertility.     

抗胸腺细胞血清性肾炎模型大鼠肾小球中C5b-9的沉积及NO、TNFα含量分析

目的: 探讨抗胸腺细胞血清性肾炎(ATSN)大鼠肾小球内C5b-9复合物的沉积与某些炎症介质和细胞因子如:一氧化氮(NO)和肿瘤坏死因子α(TNFα)的含量情况。 方法: 大鼠一次性静脉注射抗胸腺细胞抗血清(ATS)建立ATSN模型,定期对ATSN大鼠肾小球中的补体C5b-9复合物进行免疫组化染色定位、显微图像扫描半定量分析;并对有C5b-9包绕的肾小球系膜细胞(MC)进行计数。测定ATSN大鼠肾中诱生型NO合酶(iNOS) mRNA的表达、尿液中NO的代谢产物(NO^-₂／NO^-₃)及TNFα的排泄量。 结果: ATSN模型大鼠肾小球MC先溶解坏死后继发增生,病变早期(溶解时相)补体C5b-9复合物主要定位于肾小球系膜区及MC表面;随着病程的进展,被C5b-9包绕的MC逐渐减少, 病程初期ATSN大鼠肾小球MC有明显的 iNOS mRNA表达, 尿液中NO^-₂／NO^-₃ 和TNFα的排泄量也明显增加。在ATSN病变的增生阶段(一般7 d后),上述指标的变化逐渐趋缓。 结论: ATSN模型大鼠肾小球中MC逐渐溶解与补体C5b-9沉积及NO和TNFα的合成与释放有一定关系。     

人外周血CD4+CD25+调节性T细胞的分离、鉴定和功能特征

目的： 分离人外周血CD4＋ CD25＋ Treg细胞, 并检测其功能.方法： RT-PCR技术检测CD4＋ CD25＋ Treg细胞中Foxp3的mRNA表达;与CD8＋ T细胞和CD4＋ CD25- T细胞共同培养, 或加入外源性IL-2及IL- 4, 检测其抑制功能;流式细胞术检测IFN-γ、 IL- 4和IL-10.结果： CD4＋ CD25＋ Treg细胞高表达Foxp3, 主要分泌IL-10, 能够抑制CD8＋ T细胞和CD4＋ CD25- T细胞的增殖, 高浓度IL-2能够阻断CD4＋ CD25＋ Treg细胞的抑制功能.结论： CD4＋ CD25＋ Treg细胞是一群具有免疫抑制功能的调节性T细胞, 这种抑制作用能够被高浓度IL-2阻断.     

编码N端截短的IκBα突变体重组腺病毒的构建与鉴定

目的：通过去除N端丝氨酸32/36磷酸化位点，获得人胎盘组织IκBα突变体（IκBαM）基因，构建其复制缺陷型重组腺病毒（AdIκBαM），并进行体外表达和活性检测。方法:PCR定点克隆IκBαM基因（203-1 003 bp），亚克隆至pShuttle和pGEM-T，进行PCR、双酶切、DNA测序和同源性分析。将重组质粒pShuttle-IκBαM中含CMV启动子、IκBαM cDNA和PolyA信号的表达单元定向插入Ad5腺病毒载体，构建成重组腺病毒AdIκBαM，再经脂质体介导共转染293细胞进行包装。Western blotting检测AdIκBαM在293细胞中蛋白表达情况，电泳迁移率实验观察AdIκBαM抑制佛波酯诱导的ECV304细胞核因子κB（NF-κB）激活的作用。结果:成功克隆长801 bp的新型IκBαM基因，与GenBank中登陆的IκBα基因（接受号M69043）相应核苷酸序列一致。所制备的AdIκBαM滴度为4.0×1012 pfu/L。AdIκBαM介导IκBαM基因在293细胞中表达，并以剂量依赖性方式显著抑制佛波酯诱导的ECV304细胞NF-κB活化 。结论：AdIκBαM有效介导IκBαM基因表达并特异性抑制NF-κB活性，有望应用于哮喘的基因治疗。     

国产重组人促卵泡激素在辅助生殖技术控制性超促排卵的临床应用

目的评价国产重组人促卵泡激素(recombinant human follicle-stimulating hormone,rhFSH)用于辅助生殖技术(assisted reproductive technology,ART)控制性超促排卵(controlled ovarian hyperstimulation,COH)的有效性及安全性。方法本试验采用多中心、随机、双盲、阳性平行对照、非劣效研究方法,于2017年7月至2019年6月间选取6家生殖医学中心纳入卵巢储备正常的不孕女性进行ART的COH治疗。受试者随机分为试验组(国产rhFSH,n=134)和对照组(进口rhFSH,n=133),研究过程中因各种因素排除受试者共8例,试验组7例,对照组1例,最终依照研究方案完成试验的受试者试验组127例,对照组132例。比较两组受试者COH周期中获得的卵母细胞总数、rhFSH用药情况、卵母细胞受精率、优质胚胎数、临床妊娠率、活产率、新生儿情况及不良反应发生率等指标。结果试验组和对照组在COH周期中获得的卵母细胞总数分别为(13.0±5.8)枚和(12.9±5.7)枚,差异无统计学意义(P>0.05);在82例卵胞质内单精子显微注射(intracytoplasmic sperm injection,ICSI)受试者中,试验组(39例)获得MII卵母细胞数[(9.9±3.9)枚]显著高于对照组(43例)[(7.5±3.0)枚,P=0.003];卵母细胞受精率试验组[63.82%(1048/1642)]显著高于对照组[56.19%(958/1705),P<0.001]。rhFSH用药时间和总量、优质胚胎数、临床妊娠率、早产率、活产率、新生儿异常发生率、新生儿体质量、Apgar评分等两组间差异无统计学意义(P均>0.05);治疗期间卵巢过度刺激综合征(ovarian hyperstimulation syndrome,OHSS)和其他不良反应发生率差异无统计学意义(P均>0.05),且均为进口rhFSH已知的不良反应。结论在卵巢储备正常的不孕女性中使用相同卵巢刺激治疗方案,国产rhFSH有效性及安全性与进口rhFSH相当。