支持细胞与其周围细胞之间的相互作用

支持细胞为生精细胞提供结构支架和营养,支持细胞分泌的细胞因子(如IL-1α和IL-6)可能涉及精子发生的局部调控。生精细胞分泌蛋白抑制因子或刺激因子来调节支持细胞的功能。生精细胞的分化,抑制支持细胞分泌睾丸素。支持细胞对间质细胞有营养作用,支持细胞释放的类黄体生成素释放激素(LH-RH)作用于间质细胞上的LH-RH受体,进而调节间质细胞的功能。间质细胞产生的β-内啡肽和黑素细胞刺激素可分别抑制和促进支持细胞的分泌。肌样细胞旁分泌因子(PModS)促进支持细胞合成雄激素结合蛋白(ABP)和转铁蛋白,而支持细胞可抑制肌样细胞增殖。     

Effect of the sampling sequence on the quality of Papanicolaou smear

The aim of the study was to determine whether the order of cell collection (ie, obtaining either endocervical first or ectocervical cells first) has an effect on the quality of the Papanicolaou smear. 1129 smears were obtained using an Ayre spatula and an endocervical brush. In 564 cases, the endocervical brush was used first, and in 565 cases, the spatula was used first. The number of smears obscured by blood, the smears without endocervical component, and the smears with poor fixation were compared between the two groups. More smears were partially obscured by blood when brush was used first (78, 13.8% compared with 48, 8.5%, P = 0.004). No endocervical component was found in seven (1.2%) smears from the brush-first group compared with five (0.9%) of the spatula-first group, which is an insignificant difference. There were no significant differences in the number of poor-fixated smears, too-thick smears, and satisfactory smears but limited by inflammation between the two methods. The quality of the Papanicolaou smear can be improved by using the Ayre spatula first followed by the endocervical brush. Fewer smears will be contaminated by blood which may result in more squamous intraepithelial lesions being detected.     

滋养层细胞培养方法研究进展

滋养层细胞在胚胎的植入过程中扮演重要角色 ,其增殖、功能调节失常与多种妊娠相关疾病的发生发展有密切关系。滋养层细胞的培养是进行这些相关实验的基础。然而 ,目前正被应用的滋养层细胞分离、培养、纯化方法均存在一定程度的缺陷。原代培养的滋养层细胞存在产率低、杂细胞污染等方面的缺憾 ,而滋养层细胞又在细胞特性上发生了变化。此外 ,目前在鉴定分离培养的细胞是否就是滋养层细胞这一关键问题上尚无统一标准     

子宫内膜腺上皮及基质细胞分离、培养作为子宫内膜异位症体外细胞模型的探索

目的 :探索在位子宫内膜腺上皮及基质细胞的分离、培养作为子宫内膜异位症 (EMs)的体外细胞模型。方法 :通过酶解、系列过滤、沉降及贴壁纯化等技术 ,分离、纯化及培养 15份EMs患者的在位子宫内膜腺上皮细胞及其基质细胞 ,并拟传代。结果 :11份标本获得成功 ;每 1g新鲜子宫内膜组织可得到 (8～ 12 )× 10 6个原代基质细胞及 (4～ 8)× 10 6个原代腺上皮细胞 ;基质细胞纯度率可达 95 %以上 ,腺上皮细胞的纯度率约为90 %。基质细胞可以有限的传代 ,腺上皮细胞不能传代。通过改良步骤 ,可提高基质细胞的产量。结论 :在位子宫内膜腺上皮及其基质细胞的分离、培养可作为EMs的体外细胞模型之一。     

Bisphenol A-induced morphological alterations in Sertoli and spermatogenic cells of immature Shiba goats <Emphasis Type="Italic">in vitro</Emphasis>: An ultrastructural study

Background and aims: There is no information currently available regarding the effects of bisphenol A (BPA) on testes in ruminants. Therefore, to establish and clarify the effects of BPA in ruminants, testicular tissue cultures were obtained from immature Shiba goats.
Methods: The testes of 2-month-old Shiba goats were cut into smaller pieces and seeded in medium. At 1, 3, 6 and 9 h after administration of various concentrations of BPA, the specimens underwent light and transmission electron microscopic observations
Results: At 1 h after BPA treatment, vacuolization and nuclear membrane rupture appeared within the nucleoplasm and cytoplasm of Sertoli cells. Such alterations tended to gradually increase in number in time- and dose-dependent manners. Thus, because of BPA treatment, apoptotic spermatogenic cells, necrotic spermatogenic cells, apoptotic Sertoli cells and necrotic Sertoli cells could be identified. Particularly in the Sertoli cell, ruptured vesicles could be found within the multivesicular nuclear body.
Conclusion: The treatment with BPA at a low concentration tends to lead spermatogenic and Sertoli cells to apoptosis, whereas a higher concentration tends to lead spermatogenic and Sertoli cells to necrosis. Therefore, this study showed that testicular tissue culture is an advantageous avenue for screening the testicular toxicity of chemicals in ruminants. (Reprod Med Biol 2004; 3 : 205–210)     

Sertoli cell and Sertoli—Leydig cell tumors of the ovary. A report of three cases with ultrastructural findings

Three tumors of the ovary containing Sertoli cells were studied by light and electron microscopy. Two of these tumors were well-differentiated neoplasms with epithelial cells often forming tubules. These cells were cylindrically shaped, contained round to oval nuclei and stood on a thin basement membrane. The cytoplasm was fibrillary and showed rough and smooth endoplasmic reticulum, lipid droplets and secretory granules. At the luminal borders the cells were often irregular and displayed apocrine-like activity. Having compared our data with results of studies from the literature of normal Sertoli cells, Sertoli cell adenomas of the testis and cells from other parts of the male reproductive system and those of normal ovarian stroma, we conclude that the Sertoli cell is most probably the cell of origin of these tumors. The third tumor was undifferentiated with a sarcomatoid appearance and contained islands of cartilage, which we consider to be metaplastic.     

Coculture of human spermatozoa with reproductive tract cell monolayers can enhance sperm functions better than coculture with vero cell monolayers

Purpose: In order to develop a better system for support of human sperm function in vitro, we conducted studies to evaluate whether reproductive tract cells are better than non-reproductive tract cells as an adjunt in that regard. Methods: Human spermatozoa were cocultured with Vero cells, with human oviduct cells and endometrial cells, and without cells (control) for either 1, 4, or 24 hr. Sperm motility was then analyzed with a computer-aided sperm analyzer (CASA-Hamiliton Thron, HTM IVOS Motility Analyzer). Aliquots of spermatozoa incubated for 24 hr were also stained with Hoechst 33258 and FITC-PNA to evaluate the status of acrosome in live cells. Results: Significant differences (P<0.05) between the oviduct cell and the control groups after 24 hr were evident in the curvilinear velocity (VCL) (81.4±13.4 vs 60.0±14.1 µm/sec) and amplitude of lateral head displacement (ALH) (5.2±0.6 vs 4.1±0.5 µm). The incidence of acrosome reaction of live sperm was significantly higher in the endometrial cell group than in the controls (25.4±9.9 vs 6.6±2.4%;P<0.001). Conclusions: Coculture with human reproductive tract cells seems to improve some functional parameters of human spermatozoa. Coincubation with such cell lines, especially oviduct cells, might be a feasible approach to optimization of human spermatozoa for assisted fertilization using subfertile or frozen-thawed samples. We think coincubating human spermatozoa with a human reproductive tract cell line, especially oviduct cells, might be a feasible approach in preparing human spermatozoa for assisted fertilization in subfertile and frozen-thawed semen samples.     

人子宫内膜腺癌细胞原代培养方法及细胞系鉴定

目的优化人子宫内膜癌细胞原代培养方法,建立体外培养细胞系。方法将辽宁医学院附属第一医院2009年10月至11月收治的3例子宫内膜癌患者手术切除的子宫内膜腺癌组织采用酶消化法制成单细胞悬液,用含血清DMEM/F-12培养基进行原代和传代培养,通过自然纯化、冻存及低血清培养法纯化腺上皮癌细胞。在倒置显微镜下观察细胞形态,行HE染色,免疫荧光染色对细胞进行鉴定,绘制生长曲线。结果 2例子宫内膜腺癌体外成功建系,传代数扩展至50～70代。细胞形态符合上皮癌细胞系特征,细胞角蛋白表达阳性,细胞生长曲线为S形。结论应用酶消化法原代培养子宫内膜腺癌细胞,并采用自然纯化、冻存及低血清培养法纯化腺上皮癌细胞简单、易行、高效。此2株细胞系可望作为子宫内膜腺癌研究的实验模型。     

精原干细胞在支持细胞饲养层上的长期增殖特征

目的:建立体外研究精原干细胞与支持细胞共培养的细胞模型,探讨体外精原干细胞在支持细胞层上的增殖特点。方法:应用两步酶消化法,分别从12-15d及6d龄昆明小鼠睾丸中分离支持细胞及精原干细胞,并用免疫细胞化学方法鉴定。将精原干细胞接种在支持细胞层上,观察不同时间点精原干细胞形成的细胞集落形态及数目。结果:精原干细胞与支持细胞层共培养24h,开始增殖分裂,呈双细胞形态。随着培养时间延长,双细胞集落数逐渐减少,而链状细胞集落逐渐增加。培养120h后,链状细胞局部融汇堆积,且细胞集落维持在相对稳定数量。在每4-5d更换培养液的条件下,细胞集落维持稳定状态的平均时间为51.2±5.8d。结论:精原干细胞在体外支持细胞层上呈集落样生长,并能长时间维持细胞集落形态及数目,这为体外研究药物或毒物干扰精原干细胞功能提供了细胞模型。     

两种人卵巢颗粒细胞分离提纯方法的比较研究

目的:比较2种人卵巢颗粒细胞分离提纯方法的效果。方法:收集体外受精-胚胎移植(IVF-ET)穿刺取卵后的卵泡液,分别用2种方法分离提取颗粒细胞。A法用红细胞裂解液去除混杂红细胞、酶消化卵泡液中黏液团、Percoll分离液与细胞悬液等体积叠加后离心;B法省略红细胞裂解步骤、舍弃黏液团、增加Percoll分离液用量。比较2种方法获取的颗粒细胞数量、活性、纯度以及实验耗时。结果:2种方法获取的颗粒细胞存活率均>90%,差异无统计学意义(P>0.05)。A法获取的颗粒细胞数量多于B法(P<0.05),但混杂较多细胞碎片,体外培养细胞生长状态欠佳;B法获取的颗粒细胞纯度高,体外培养生长状态良好,且实验耗时较A法短(P<0.05)。结论:2种方法对颗粒细胞存活率无明显影响。B法更为省时高效,有助于建立稳定的颗粒细胞体外培养体系。     

Prognostic determinants in patients with uterine and ovarian clear cell carcinoma: A SEER analysis

Introduction

The purpose of this study is to analyze and compare the demographics, treatment, and survival rates in patients with uterine clear cell carcinoma (UCCC) and ovarian clear cell carcinoma (OCCC).

Methods

The Surveillance, Epidemiology and End Results (SEER) program data for all 18 registries from 1988 to 2010 was reviewed to identify women with OCCC and UCCC. Demographic and clinical data were compared, and the impact of tumor site on survival was analyzed using the Kaplan–Meier method. Factors predictive of outcome were compared using the Cox proportional hazards model.

Results

The final study group consisted of 5421 women with clear cell histopathology. 3631 (67%) had OCCC and 1790 (33%) had UCCC. The mean age at diagnosis was 56 (± 12) years for women with OCCC and 67.7 (± 12.0) years for UCCC (P < 0.001). Patients with OCCC had a higher rate of late stage disease (38.9% vs. 21.2%; P < 0.001). Over the entire study period, after adjusting for known variables, there was no significant difference in cancer specific mortality between UCCC and OCCC, HR 1.05 (0.92–1.19). In the subset analysis by staging, in women with localized disease there was an improved survival in UCCC compared to OCCC. In contrast, in women with distant disease there was an increased mortality in women with UCCC.

Conclusion

In the entire population, there was no significant difference in cancer related mortality between the groups. However, in women with localized disease, UCCC had improved survival, but increased mortality in distant disease compared to OCCC.     

小儿严重肝病并棘刺红细胞症7例临床分析

目的探讨严重肝病并棘刺红细胞症的临床特点。 方法对2001年1月至2005年5月间桂林医学院附属医院儿科收治的7例严重肝病并棘刺红细胞症患儿的症状、体征，血细胞光镜检查，肝功能，血脂检查，预后，并结合文献资料进行总结分析。 结果7例中男4例，女3例，年龄5个月至9岁，主要临床表现有黄疸(6例)、出血(3例)、贫血(7例)、肝肿大(6例)或肝硬化(1例)、脾肿大(3例)，外周血中棘刺红细胞占所有红细胞的85%~944%，肝功能损害明显，血脂均有不同程度的改变。 结论严重肝病并棘刺红细胞症临床罕见，这7例患儿的共同特点是：(1)原发病各异；(2)病程长；(3)有程度不等的贫血，红细胞平均容积(MCV)以增大为主；(4)外周血中均有多少不等的棘刺红细胞；(5)肝功能损害明显；(6)血脂均有多项改变；(7)预后不良。     

卵巢无性细胞瘤伴绒毛膜促性腺激素显著升高1例

卵巢无性细胞瘤是一种中度恶性的卵巢生殖细胞肿瘤,好发于年轻女性,临床发病少见,手术+铂类为主的联合化疗使该病有良好的生存预后及妊娠结局.单纯卵巢无性细胞瘤无内分泌功能,临床检测绒毛膜促性腺激素(HCG)一般为阴性,仅6％～8％含有小簇合体滋养细胞而使HCG呈低反应.本文通过分析1例单纯卵巢无性细胞瘤伴HCG值高达549...     

青春期大鼠受己烯雌酚持续作用对成年后睾丸波形蛋白表达的影响

目的:探讨青春期大鼠受己烯雌酚(DES)持续作用,成年后睾丸曲细精管和间质区波形蛋白表达的影响。方法:将SD大鼠,随机分成对照组与3个DES实验组,对照组注射0.3ml纯玉米油,实验组分别注射含DES2μg·kg-1·d-1(低剂量组)、10μg·kg-1·d-1(中剂量组)、50μg·kg-1·d-1(高剂量组)的玉米油0.3ml,每组6只动物。各组隔日腹腔注射,28d后采集标本。采用免疫组化法观察睾丸波形蛋白的表达和变化。结果:在睾丸曲细精管内仅见波形蛋白表达于支持细胞胞浆,生精细胞中未见表达。在支持细胞中,波形蛋白由基膜基部绕核成环状,并随胞质呈辐射状向曲细精管管腔延伸。随着DES染毒剂量的增加,其表达变弱,向管腔的辐射状部分变短、消失。与对照组相比,DES实验组波形蛋白的阳性支持细胞表达率及表达量均显著下降(P<0.01),各实验组间比较均有显著性差异(P<0.01)。波形蛋白也见表达于睾丸间质区的间质细胞胞浆与结缔组织,3个实验组中的表达量分别与对照组相比均有显著性下降(P<0.01),低剂量组波形蛋白表达量与高剂量组相比也有显著减少(P<0.05)。各组波形蛋白在间质区的表达量均显著高于支持细胞胞浆的表达(P<0.01)。结论:青春期大鼠受DES持续作用后,成年大鼠睾丸支持细胞和间质区的波形蛋白表达量显著下降,且损伤呈剂量效应。     

醋酸棉酚与米索前列醇对离体培养的大鼠黄体细胞和人蜕膜细胞的影响

醋酸棉酚和米索前列醇对体外培养大鼠黄体细胞和人蜕膜细胞都有明显的损伤作用，呈现量-效相关性。棉酚单用对黄体、蜕膜细胞的LD50分别为1．27±0．09μg／ml和3．06±0．23μg／mg；但与来索5μg／ml（在黄体细胞），10μg／ml（在人蜕膜细胞）合用时，LD50分别降至0．89±0．25μg／ml和1．88±026μg／ml；与单用棉酚相比，有显著性差异。单用米索对黄体、蜕膜细胞的LD50各为14．29±1．29μg／ml和24．37±4．49μg／ml；但与棉酚0．5μg／ml（在黄体细胞），1．0μg／ml（在蜕膜细胞）合用时，分别下降为8．79±2．18μg／ml和17.29±1．56μg／ml，与单用米索相比，均有统计学差异，显示两药合用的协同效应。     

银杏黄酮对人卵巢恶性畸胎瘤细胞株PA-1及人卵巢颗粒细胞的影响

目的 探讨银杏黄酮对人卵巢恶性畸胎瘤细胞株PA-1及人卵巢颗粒细胞体外增殖的影响。方法 采用噻唑蓝[3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di- phenytetrazoliumromide,MTT]法观察质量浓度分别为0、175、200、225 mg/L的银杏黄酮作用于PA-1细胞24、48、72 h后对细胞生长的影响;流式细胞仪检测作用48 h后对PA-1细胞凋亡率及细胞生长周期的影响;选取2011年3月至2012年9月河北医科大学第二医院生殖中心行体外受精-胚胎移植（in vitro fertilization and embryo transfer ,IVF-ET）患者的人卵巢颗粒细胞,将同等剂量的银杏黄酮作用于人卵巢颗粒细胞,经MTT法观察其对颗粒细胞的影响、经化学发光法检测颗粒细胞所分泌的雌二醇（E2）值的变化。结果 随着银杏黄酮浓度增高,对PA-1细胞的增殖有明显的抑制作用,呈明显时间、剂量依赖性;凋亡率随剂量增加而增加,细胞周期被阻滞于G1期;而同等剂量的银杏黄酮作用于人卵巢颗粒细胞后对其生长没有抑制作用,且各组的E2值差异没有统计学意义（P<0 .05）。结论 银杏黄酮明显抑制人卵巢恶性畸胎瘤PA-1细胞生长,而对卵巢的功能细胞-颗粒细胞没有损伤。     

6B11抗独特型微抗体负载树突状细胞对卵巢癌细胞系细胞杀伤作用的体外研究

目的　用 6B1 1抗独特型微抗体体外负载树突状细胞 (DC) ,以期诱导出抗原特异性的抗肿瘤免疫反应。方法　分离培养HLA -A2 健康人外周血树突状细胞 ,培养过程中用 6B1 1微抗体负载 ,无关抗原F (ab)’2负载及未负载组为对照。光镜、电镜下观察树突的形态 ;流式细胞仪检测DC表面分子表达 ;3 H -TdR掺入法测DC刺激自体T细胞增殖 ;51 Cr 6h释放试验测激活的T细胞对卵巢癌细胞系的杀伤。结果　培养的树突细胞有典型形态 ,CD80、CD86、HLA -DR等表面分子呈高表达 (分别为 6 5 %、 86 2 %、 93 7% ) ;6B1 1微抗体负载的DC不仅能刺激自体T细胞的增殖 ,而且其诱导的CTL细胞对HLA -A2 、OC1 6 6 - 9 卵巢癌细胞系HOC1A有特异性的杀伤 ,结论　 6B1 1抗独特型微抗体模拟卵巢癌抗原OC1 6 6 - 9负载树突状细胞在体外可以诱导出抗原特异性的细胞毒T细胞 ,为进一步研究 6B1 1微抗体对卵巢癌免疫治疗作用提供了依据     

改良筛网法分离培养人子宫内膜细胞

目的:探索一种能获得较高纯度和较多数量子宫内膜细胞的分离培养方法。方法:取12例患者各1mg新鲜子宫内膜组织,经0.25%Ⅰ型胶原酶(含有0.005%DNA酶)消化后,将所得细胞悬液一分为二,分别采用传统筛网法和改良筛网法(二次筛网过滤和低速离心相结合)分离并记录所得细胞数;体外培养后采用光学显微镜观察,用免疫细胞化学和免疫荧光化学方法鉴定纯度,比较两种方法分离子宫内膜细胞的纯度和数量。结果:两种方法均有11例组织分离培养成功。传统筛网法分离的子宫内膜腺上皮细胞和间质细胞的纯度分别为(89.54±3.75)%和(92.82±1.60)%,细胞数分别为(0.47±0.09)×105个和(23.73±5.37)×105个。改良筛网法分离的腺上皮细胞和间质细胞纯度分别为(91.95±0.47)%和(99.28±0.21)%,细胞数分别为(8.09±1.24)×105个和(27.23±0.50)×105个。改良筛网法分离的腺上皮细胞数量和间质细胞纯度均明显高于传统筛网法,有统计学差异(P均=0.000)。结论:改良筛网法操作简单,能获得较高纯度的子宫内膜间质细胞及较多的子宫内膜腺上皮细胞。