黄体生成素和卵泡刺激素在大鼠垂体前叶的细胞共定位研究

为探讨卵泡刺激与黄体生成生素分泌及基因表达与调控用抗人ＬＨ－β亚单位单克隆抗体和扩ＦＳＨ－β亚单位单克隆抗体进行免疫细胞化学定位的研究。通过连续切片免疫酶技术，免疫细胞化学双标记及电镜免疫金双标记技术，在大鼠垂体前叶进行ＦＳＨ和ＬＨ的免疫细胞化学共定位研究。本文对垂体前叶ＧＴＨ细胞存在着３种激素基因表达形式的可能原因进行了探讨。     

基础卵泡刺激素水平在体外受精周期中的应用

目的探讨基础卵泡刺激素（bFSH）对体外受精（IVF-ET）助孕效果的影响。方法 970个接受IVF-ET周期前1-6个周期,于月经第3天抽血测定bFSH水平,按bFSH水平不同分为A组bFSH     

卵泡刺激素及其受体在大鼠胰腺的共定位观察

目的观察卵泡刺激素(FSH)及其受体(FSHR)在大鼠胰腺中的共存关系,以了解FSH对胰腺的功能意义。方法应用免疫荧光组织化学双标记染色技术,在激光共聚焦扫描显微镜下观察染色结果。结果部分外分泌腺细胞、导管上皮细胞及部分胰岛细胞呈FSH反应阳性,呈强绿色荧光。小血管内皮细胞也呈FSH反应弱阳性,呈弱的绿色荧光。部分胰岛细胞呈FSHR反应阳性,呈强红色荧光。一些外分泌腺上皮细胞呈FSHR反应弱阳性,呈淡红色荧光。有一些胰岛细胞呈FSH和FSHR双阳性反应,呈黄色荧光或红、绿相间荧光。结论FSH及其受体均存在于大鼠胰腺中,胰岛中的部分细胞共同存在FSH及其受体,FSH可能通过旁分泌或自分泌的途径对胰腺的功能起调节作用。     

转化生长因子β信号通路对卵泡刺激素调节卵巢颗粒细胞功能的影响

目的 探讨大鼠卵巢颗粒细胞中卵泡刺激素(FSH)和转化生长因子β(TGF-β)信号通路之间的相互作用.方法 取21 dSD大鼠卵泡分离的颗粒细胞原代培养,实验分为对照组、FSH处理组和转化生长因子β Ⅱ型受体(TGF-β R Ⅱ)中和组.通过免疫细胞化学和Western blotting定位和检测TGF-β R Ⅱ以及...     

补肾活血方对小鼠卵泡刺激素及其受体mRNA表达的影响

目的:探讨补肾活血复方中药对免疫性卵巢早衰小鼠卵泡刺激素(Follicle stimulating hormone,FSH)及其受体(Follicle stimulating hormone receptor,FSHR)mRNA表达水平的影响。方法:以小鼠透明带3(Zona pellucida3)为抗原,皮下多点注射免疫雌性小鼠建立免疫性卵巢早衰模型,以生理盐水注射免疫的小鼠作为空白对照组。设补肾活血中药低、中、高不同剂量的3组治疗组,以泼尼松、倍美力为西药治疗对照组,用酶联免疫竞争法检测小鼠治疗后血清中抗透明带抗体(ZPAb)、FSH和雌二醇(E2)浓度,用实时荧光定量逆转录-多聚酶链反应(RT-PCR)检测小鼠卵巢组织FSHR mRNA表达水平。结果:模型组小鼠血清ZPAb的含量明显比空白对照组高,但血清E2的含量比空白对照组低,差异都有显著性意义(P0.01);补肾活血中药治疗的3组小鼠血清E2的含量都比模型组有显著性升高,差异有非常显著性意义(P0.001)。模型组小鼠血清FSH的含量与空白对照组比较,差异没有统计学意义(P0.05),补肾活血中药治疗的3组小鼠血清FSH的含量与模型组比较,含量都有显著性升高,差异有显著性意义(P0.05)。模型组小鼠卵巢组织FSHR表达的mRNA量比空白对照组低,但差异没有统计学意义(P0.05);补肾活血中药治疗的3组小鼠卵巢组织FSHR表达的mRNA量比模型组高,差异有显著性意义(P0.05)。小鼠血清E2的含量与卵巢组织FSHR mRNA表达量呈线性正相关关系(R2=0.359,F=64.888,P0.001)。结论:补肾活血复方中药通过促进垂体分泌FSH的增加和卵巢颗粒细胞表面FSHR表达量的增加,提高卵巢组织对FSH的反应性,促使卵巢颗粒细胞分泌雌激素量增加,并且卵巢颗粒细胞分泌E2的量与卵巢颗粒细胞表面FSHR表达量呈正相关关系。     

雄激素受体和卵泡刺激素受体在成年大鼠睾丸中的期依赖性表达

目的　确定雄激素受体 (AR)和卵泡刺激素受体 (FSHR)在大鼠睾丸中的细胞定位和表达变化。　方法　利用地高辛标记的cRNA探针 ,在成年大鼠睾丸冰冻切片上进行原位杂交 ;同时利用透光显微切割技术将成年大鼠曲细精管分为Ⅱ～Ⅵ、Ⅶ～Ⅷ、Ⅸ～Ⅻ、ⅩⅢ～Ⅰ 4个阶段 ,提取总RNA ,用α 32 P标记的cDNA探针进行斑点杂交 ,观察AR和FSHRmRNA表达的细胞定位和期依赖性变化。利用图像分析系统 ,对阳性杂交信号单位面积平均辉度进行定量分析。　结果　ARmRNA阳性杂交信号位于支持细胞和间质细胞 ,于Ⅶ～Ⅷ期最强 ,Ⅸ～Ⅰ期最弱 ;FSHRmRNA阳性杂交信号位于支持细胞 ,于ⅩⅢ～Ⅰ期最强 ,Ⅶ～Ⅷ期最弱。各阶段之间具有非常显著性差异 (P<0 0 1)。　结论　AR和FSHR期依赖性表达的不同规律提示T(睾酮 )和FSH(卵泡刺激素 )作用于精子发生的不同阶段 ,说明睾酮和卵泡刺激素协同作用调节成年大鼠的精子发生。这一结论将为人类生育调控和不孕症的治疗提供新思路     

大鼠胰腺卵泡刺激素(β)及其受体的免疫荧光定位及原位杂交

目的探讨大鼠胰腺组织中是否表达卵泡刺激素(FSH)及其受体(FSHR),为进一步研究FSH对胰腺的功能调节提供理论依据。方法采用单标记免疫荧光技术和原位杂交方法进行研究。结果大鼠胰岛大部分细胞及部分胰腺外分泌部细胞呈FSH蛋白和mRNA,FSHR蛋白和mRNA免疫荧光反应阳性,阳性物质分布于细胞质,细胞核呈阴性反应。结论大鼠胰腺细胞具有自身合成和分泌FSH及其受体的功能,说明FSH对胰腺细胞的作用可能通过旁分泌或自分泌的作用实现的。     

雄激素受体和卵泡刺激素受体在成年大鼠睾丸中期的依赖性表达

目的 确定雄激素受体（AR）和卵泡刺激素受体（FSHR）在大鼠睾丸中的细胞定位和表达变化。方法 利用地高辛标记的cRNA探针,在成年大鼠睾丸冰冻切片上进行原位杂交;同时利用透光显微切割技术将成年大鼠曲细精管分为Ⅱ-Ⅵ、Ⅶ-Ⅷ、Ⅸ-Ⅻ、XIII－Ⅰ4个阶段,撮总RNA,用α－^32P标记的cDNA探外进行斑点杂交,观察AR和FSHR mRNA表达的细胞定位和期依赖性变化。利用图像分析系统,对阳性杂交信号单位面积平均辉度进行定量分析。结果 AR mRNA阳性杂交信号位于支持细胞和间质细胞,于Ⅶ-Ⅷ期最强,Ⅸ-Ⅰ期最弱;FSHR mRNA阳性杂交信号位于支持细胞,于XIII－Ⅰ期最强,Ⅶ-Ⅷ期最弱。各阶段之间具有非常显著性差异（P＜0.01）。结论 AR和FSHR期依赖性表达的不同规律提示T（睾酮）和FSH（卵泡刺激素）作用于精子发生的不同阶段,说明睾酮和卵泡刺激素协同作用调节成年大鼠的精子发生。这一结论将为人类生育调控和不孕症的治疗提供新思路。     

睾酮对小鼠卵泡体外发育的影响

目的探讨高浓度睾酮对体外培养的小鼠卵泡生长发育的影响。方法以出生后12日龄的ICR雌鼠体外培养卵泡为研究模型,将卵泡随机分为以下4个组,每组30个:正常对照组(control组)、10-4mol/L睾酮处理组、10-5mol/L睾酮处理组和10-6mol/L睾酮处理组。将卵泡随机分为以下3个组,每组30个:正常对照组(control组)、睾酮处理组(T组,T=10-5mol/L)和雄激素受体拮抗剂氟他胺预处理后再加睾酮处理组(F+T组,F=10-5mol/L,T=10-5mol/L)。在60mm无菌细胞培养皿上进行微滴培养,每天在倒置显微镜下观察卵泡轮廓,颗粒细胞增生情况,卵母细胞形态以及有无卵母细胞逸出等,并记录卵泡存活和退化的情况。结果 10-4mol/L睾酮能抑制卵泡的生长,而10-5mol/L睾酮能够促进颗粒细胞的增殖和卵泡的生长;同时,在卵泡生长前期,睾酮能刺激卵泡的发育,在后期则呈抑制作用,即卵泡发育停滞、退化或死亡。结论在初级和次级卵泡阶段,10-5mol/L睾酮能促进卵泡的发育,而在窦状卵泡阶段则抑制卵泡的发育。     

大鼠胃卵泡刺激素和促性腺激素释放激素受体的共定位

我们先前的研究已经研究,大鼠消化系统广泛分布有促性腺激素释放激素（GnRH）免疫反应性上皮细胞,而且证实这些细胞同样存在GnRH mRNA的杂交信号,说明胃、肠、胰腺能自身合成GnRH。随后我们又证实大鼠下颌下腺、胰腺和胃含有GnRH的受体,说明它们又是GnRH的靶器官。     

小鼠睾丸支持细胞的体外分离和纯化

目的寻求简便经济的方法分离纯化小鼠睾丸支持细胞,提高其获取量。方法用0.1%的胶原酶和0.25%的胰蛋白酶次第消化准备7-8天小鼠生殖细胞悬液,置于37℃,5%CO2孵箱内培养,4小时后换液,24小时后向培养板内加入20 mmol Tris-HCl低渗处理3分钟,去除精原细胞,即得到高纯度的支持细胞。结果在小鼠睾丸支持细胞的分离培养中,支持细胞占培养细胞90%以上。结论应用本实验方法分离小鼠睾丸支持细胞获得成功并简化了国内现行的分离方法。     

Glycosaminoglycans and PDGF Signaling in Mesenchymal Cells

Platelet derived growth factor (PDGF) is involved in the autocrine growth stimulation of normal and malignant cells, the stimulation of angiogenesis, and the recruitment and regulation of tumor fibroblasts. PDGF has been shown to physically interact with glycosaminoglycans which are abundant in the extracellular microenvironment. The present review discusses the effects of glycosaminoglycans on the functions mediated by the PDGF on cells of mesenchymal origin. Recent studies have demonstrated that both soluble and surface bound glycosaminoglycan chains can modulate PDGF-BB isoform signaling depending on the cell type. These data demonstrated that the microenvironment rich in GAGs/PGs is able to significantly modify the cellular response to PDGF-BB signaling in a critical way for cell growth and differentiation.     

大鼠睾丸支持细胞的分离纯化与鉴定

目的　简化大鼠睾丸支持细胞的分离纯化方法 ,通过不同方法对Sertoli细胞进行形态学观察鉴定。方法　取鼠龄 16～ 2 2天的雄性Wistar大鼠睾丸 ,采用酶次第消化 ,培养过程中纯化Sertoli细胞 ,并用多种方法对Sertoli细胞进行鉴定。结果　大鼠睾丸支持细胞占培养细胞总数的 90 %以上。HE染色 ,Sertoli细胞突起很多 ,核仁清晰 ,在胞质中可见吞噬物和大小不等的空泡 ;甲基绿 派洛宁染色 ,Sertoli细胞富含RNA ;Feulgen染色和透射电镜 ,核仁周围可见卫星核小体。结论　本实验培养方法可获得更多、纯度高的Sertoli细胞 ,HE染色、甲基绿 派洛宁染色、Feulgen染色和透射电镜是鉴定Sertoli细胞的有效方法     

Human Autoantibody to Sertoli Cells Detected in Healthy Individuals

Human autoantibody to Sertoli cells was detected in normal human sera. This IgM type autoantibody was undetectable during the neonatal period and was found in 11.5% of 365 serum samples taken from adult healthy persons of both sexes. This human autoantibody to Sertoli cells exhibited quite the same target-organ specificity and multi organ reactivity (salivary gland ductules, pancreatic intercalated duct, renal lower nephron, pituitary acidophilic cells) as those of the murine monoclonal autoantibody (IgM class) to Sertoli cells (TM-1: WHO registered code T43). The TM-1 monoclonal antibody could recognize testicular antigens with molecular weights of 67,000 and 23,000 in Sertoli cells, and had already been demonstrated capable of inducing murine experimental spermatogenic disturbance when administrated together with murine monoclonal autoantibody to seminiferous tubular basement membrane (TNI 2: WHO registered code T44). These observations may suggest that human spermatogenic disturbance could be easily induced by the multi-organ reactive autoantibody to Sertoli cells even in healthy individuals under particular conditions where this autoantibody can be allowed to reach the target Sertoli cells across the barrier of seminiferous tubular wall by either autoantibody to seminiferous tubular basement membrane or other toxic damage. Acta Pathol Jpn 41: 879-888, 1991.     

Degeneration of Sertoli and Spermatogenic Cells in Homozygous and Heterozygous Weaver Mice

In the neurological mutant mouse weaver (wv/wv), the majority of males are infertile due to hypospermatogenesis. Heterozygous weaver mice (wv/+) cease mating successfully when males reach an average age of 3.5 months. The contents of epididymal fluid were scored for the number of sperm and sperm motility in wv/wv, wv/+ and controls. Testes were examined in mice of the three genotypes at various ages using light and electron microscopy. In wv/+ males, sperm counts were significantly lower than in controls and were significantly higher than in wv/wv. The seminiferous epithelium in weaver mice appears depleted soon after puberty and a wide range of degenerative changes was identified in both germ cells and Sertoli cells. Analogous cellular aberrations were detected in heterozygous males, but they appeared at an older age and were not as severe as in wv/wv. We hypothesize that in weaver homo- and heterozygosity the damage of Sertoli cells may induce degeneration of germinal cells and particularly affect the most advanced spermatogenic cells.     

Signaling Pathways in Cancer and Embryonic Stem Cells

Cancer cells have the ability to divide indefinitely and spread to different parts of the body during metastasis. Embryonic stem cells can self-renew and, through differentiation to somatic cells, provide the building blocks of the human body. Embryonic stem cells offer tremendous opportunities for regenerative medicine and serve as an excellent model system to study early human development. Many of the molecular mechanism underlying tumorigenesis in cancer and self-renewal in stem cells have been elucidated in the past decade. Here we present a systematic analysis of seven major signaling pathways implicated in both cancer and stem cells. We present on overview of the JAK/STAT, Notch, MAPK/ERK, PI3K/AKT, NF-kB, Wnt and TGF-β pathways and analyze their activation status in the context of cancer and stem cells. We focus on their role in stem cell self-renewal and development and identify key molecules, whose aberrant expression has been associated with malignant phenotypes. We conclude by presenting a map of the signaling networks involved in cancer and embryonic stem cells.