促性腺激素释放激素拮抗剂对大鼠睾丸支持细胞雄激素受体和Fas配 …

本实验利用注射促性腺激素释放激素拮抗体剂（ＧｎＲＨ－Ａ）引起睾丸生精细胞程序化死亡的大鼠模型，用地高辛标记的大鼠雄激素受体和Ｆａｓ配体的ＲＮＡ探针对大鼠睾丸石蜡发片进行原位杂交，观察雄激素受体和Ｆａｓ配本基因在生精细胞凋亡中的变化。结果显示，ＧｎＲＨ－Ａ处理后曲细精管有退行性变，原位杂交表明此时Ｓｅｒｔｏｌｉ细胞的雄激素受体表达明显减低，而Ｆｓａ配体表达显著升高。提示Ｆａｓ配体的表达可能与生精上皮     

睾丸免疫豁免细胞的分离及其功能测定

目的 探讨睾丸产生免疫豁免的机理。方法 应用胶原酶、胰蛋白酶及脱氧核糖核酸酶消化制备睾丸Ｓｅｒｔｏｌｉ细胞，体外与活性淋巴细胞共同培养，了解其对淋巴细胞的杀伤作用。并用ＳＡＢＣ法标记睾丸组织Ｓｅｒｔｏｌｉ细胞Ｆａｓ配体的表达。结果 Ｓｅｒｔｏｌｉ细胞可杀伤与之共培养的活性淋巴细胞，睾丸组织Ｓｅｒｔｏｌｉ细胞有明显的Ｆａｓ配体表达。结论 Ｆａｓ配体阳性的Ｓｅｒｔｏｌｉ细胞对活性淋巴细胞的杀伤作用可能     

神经生长因子mRNA水平与睾丸生精细胞成熟和功能的关系

以地高辛素标记人β－神经生长因子ＤＮＡ为探针，采用原位杂交的方法观察了小鼠睾丸曲精小管中ＮＧＦｍＲＮＡ的分布情况，发现阳性细胞分布于大部分曲精小管基底至管腔的各级生精细胞中，随着生精细胞成熟度的增加，ＮＧＦｍＲＮＡ表达水平逐步增加，细胞核与间质呈阴性。结果提示：ＮＧＦ在睾丸中具有自分泌与旁分泌作用的可能，不断促进精子自身的成熟与活力。     

神经生长因子mRNA水平与睾丸生精细胞成熟和功能的关系

以地高辛素标记人β－神经生长因子ＤＮＡ为探针，采用原位杂交的方法观察了小鼠睾丸曲精小管中ＮＧＦｍＲＮＡ的分布情况，发现阳性细胞分布于大部分曲精小管基底至管腔的各级有细胞中，随着生精细胞成熟度的增加，ＮＧＦｍＲＮＡ表达水平逐步增加，细胞核与间质呈阴性。结果提示：ＮＧＦ在睾丸中具有自分泌与旁分泌作用的可能，不断促进精子自身的成熟和活力。     

前列腺癌细胞中表皮生长因子受体的表达和激活

目的：探讨前列腺癌（ＰＣａ）细胞中表皮生长因子受体（ＥＧＦＲ）表达和激活及其与雄激之间的相互作用。方法：采用免疫沉淀和Ｗｅｓｔｅｒｎ ｂｌｏｔ方法,检测雄激素依赖人ＰＣａ细胞株ＬＮＣａＰ及雄激素非依赖人ＰＣａ细胞株ＤＵ－１４５、ＰＣ－３中ＥＧＦＲ的表达和磷酸化激活,测定雄激素对ＬＮＣａＰ ＥＧＦＲ表达和激活的影响。结果：在ＤＵ－１４５和ＰＣ－３中,ＥＧＦＲ表达和激活的基础与表皮生长因子（ＥＧＦ）处     

Fas／FasL与肾小球疾病

Ｆａｓ是一种跨膜糖蛋白，属于ＴＮＦ／ＮＧＦ受体家族，广泛分布于各种组织细胞上，与其相应的Ｆａｓ配体（Ｆａｓ Ｌｉｇａｎｄ，ＦａｓＬ）结合可诱导该细胞凋亡。Ｆａｓ／ＦａｓＬ主要生物作用为参与机体免疫     

Fas/FasL与肾小球疾病

Ｆａｓ是一种跨膜糖蛋白，属于ＴＮＦ／ＮＧＦ受体家族，广泛分布于各种组织细胞上，与其相应的Ｆａｓ配体（ＦａｓＬｉｇａｎｄ，ＦａｓＬ）结合可诱导该细胞凋亡。Ｆａｓ／ＦａｓＬ主要生物作用为参与机体免疫系统细胞生成选择，调节机体免疫应答和参与ＴＣＬ的细胞毒性作用。Ｆａｓ／ＦａｓＬ的功能增强或缺陷均可导致肾小球疾病的发生和发展，这就为如何防治肾小球疾病提供了新的理论思路。     

Fas,FasL与泌尿系肿瘤

Ｆａｓ是分子量为 40 ,0 0 0～ 45 ,0 0 0的跨膜蛋白质 ,属于神经生长因子 (ＮＧＦ) /肿瘤坏死因子 (ＴＮＦ)受体超家族成员 ,与Ｆａｓ抗体或配体 (ＦａｓＬ)结合 ,可引起Ｆａｓ敏感细胞的凋亡 ,多种细胞表面表达Ｆａｓ。ＦａｓＬ是一种属于ＴＮＦ家族的细胞表面分子 ,主要是由活性Ｔ细胞表达。Ｆａｓ/ＦａｓＬ主要参与免疫反应下调及Ｔ细胞介导的细胞毒性。Ｆａｓ系统功能障碍引起免疫增生紊乱和加速自身免疫性疾病。近年来 ,国内外不断报告肿瘤细胞Ｆａｓ的表达及由Ｆａｓ、ＦａｓＬ及其细胞内信号传导分子共同构成的Ｆａｓ系统在诱导细胞…     

Fas／FasL表达和细胞凋亡在人类同种异体移植肾急性排斥中的 …

目的 探讨移植肾急性排斥（ＡＲ）时细胞凋亡与Ｆａｓ和Ｆａｓ配体表达的作用及其临床意义。方法 分别用原位末标记技术（ＴＵＮＥＬ法）和免疫组织化学方法检测２６例移植肾ＡＲ标本中细胞凋亡和Ｆａｓ／ＦａｓＬ表达情况。结果 细胞凋亡和Ｆａｓ／ＦａｓＬ表达主要在ＡＲ移植肾上管上皮发生且凋亡指数和Ｆａｓ／ＦａｓＬ表达与肾损害程度平行。与正常肾对照线和对照肾功能稳定组比较差异显著。结论肾小管上皮雕恨在ＡＲ所致的移     

Fas系统、细胞凋亡与移植免疫

Ｆａｓ系统介导的细胞凋亡在移植免疫中起着十分重要的作用 ,它参与淋巴细胞的发育、自我耐受的形成、Ｔ淋巴细胞克隆的限制、免疫应答的实现等诸多免疫过程。本文就Ｆａｓ系统与Ｔ淋巴细胞凋亡、移植排斥和移植耐受的研究进展予以综述。一、Ｆａｓ、ＦａｓＬ的结构和分布在第五届人白细胞分型国际会议上Ｆａｓ被正式命名为ＣＤ95。Ｆａｓ抗原的天然配体ＦａｓＬ则是Ｓｕｄａ于 1993年从大鼠细胞毒性Ｔ淋巴细胞 (ＣＴＬ)中纯化确定的[1] 。Ｆａｓ属于肿瘤坏死因子受体(ＴＮＦＲ)和神经生长因子受体 (ＮＧＦＲ)超家族 ,其分子结构由三部分组…     

The Fas system in the seminiferous epithelium and its possible extra-testicular role

The Fas system is involved in the control of immune system homeostasis and nonfunctional Fas system leads to autoimmune disease in mice and humans. The Fas system is a mechanism through which cells expressing Fas ligand (FasL) induce apoptosis of Fas expressing cells. In mouse and rat, the testis represents the main source of constitutive FasL in the body. The roles so far proposed for this molecule in the testis, such as maintenance of immunoprivilege and regulation of physiological germ cell apoptosis, need to be reconsidered as both hypotheses are based on an erroneous cellular location of FasL in the seminiferous epithelium. Recently, we demonstrated that in rodents FasL mRNA is present in germ cells and not in Sertoli cells, and that FasL protein is displayed on the surface of spermatozoa. Here we propose that, for the mouse spermatozoa, the FasL may represent a self-defence mechanism against lymphocytes present in the female genital tract. To verify this hypothesis, we performed crossings between males gld, with nonfunctional FasL, and syngenic or nonsyngenic females. We observed a significant decrease of litter size in outbred crossings with gld males compared with wild-type males, suggesting a possible role of FasL in immunoprotection of the sperm in the female genital tract. The possibility that in humans, by analogy with mouse, FasL plays a self-protective role for the spermatozoon cannot be excluded, and awaits experimental information on the expression of FasL on human sperm cells.     

Germ cell apoptosis induced by ureaplasma urealyticum infection

Aim: To study the effect of Ureaplasma urealyticum (UU) infection on germ cell apoptosis of male rats. Methods: Male rats were infected artificially with UU serotype 8 (T960). Morphological changes of germ cells in the seminiferous tubules and the lumen of the epididymides were observed under the light microscope. Fluorescence-conjugated polyclonal antibodies to Fas and Fas ligand (FasL) were used to localize Fas and FasL. TUNEL staining of germ cells and Sertoli cells was performed by the AKPase method. TUNEL-positive rate ( % positive cells) and TUNEL-positive area (area occupied by stained cells) were analysed by KS400 Image Analysis System. The DNA laddering analysis was performed by agarose gels electrophoresis. Results: In those rats infected with UU: (1) Exfoliated germ cells were dramatically increased. Many multinucleated giant cells were found in the seminiferous tubules and the lumen of the epididymides. (2) The number of TUNEL-positive cells and the TUNEL-positive area were significantly increased. (3) The expression of Fas and FasL in germ cells and Sertoli cells was up-regulated. (4) Discrete bands of fragmented DNA were found in the testicular cells. Conclusion: In male rats, germ cell apoptosis was increased in UU infection.     

Proapoptotic Fas ligand is expressed by normal kidney tubular epithelium and injured glomeruli

Fas ligand (FasL) is a cell membrane cytokine that can promote apoptosis through activation of Fas receptors. Fas receptor activation induces glomerular cell apoptosis in vivo and participates in tubular cell death during acute renal failure. However, there is little information on the expression of FasL in the kidney. This study reports that FasL mRNA and protein are present in normal mouse and rat kidney. In situ hybridization and immunohistochemistry showed that proximal tubular epithelium is the main site of FasL expression in the normal kidney. In addition, increased total kidney FasL mRNA and de novo FasL protein expression by glomerular cells were observed in two different models of glomerular injury : rat immune-complex proliferative glumerulonephritis and murine lupus nephritis. Both full-length and soluble FasL were increased in the kidneys of the mice with nephritis. Cultured murine proximal tubular epithelial MCT cells and primary cultures of murine tubular epithelial cells expressed FasL mRNA and protein. Tubular epithelium-derived FasL induced apoptosis in Fassensitive lymphoid cell lines but not in Fas-resistant lymphoid cell lines. By contrast, MCT cells grown in the presence of the survival factors of serum were resistant to FasL, and only became partially sensitive to apoptosis induced by high concentrations (100 ng/ml) of FasL upon serum deprivation. However, MCT cells stimulated with inflammatory mediators (tumor necrosis factor-alpha, interferon-gamma, and lipopolysaccharide) increased cell surface Fas expression and were sensitized to apoptosis induced by FasL (FasL 55 +/- 5% versus control 8.3 +/- 4.1% apoptotic cells at 24 h, P < 0.05). Cytokine-primed primary cultures of tubular epithelial cells also acquired sensitivity to FasL-induced apoptosis. These results suggest that FasL expression by intrinsic renal cells may play a role in cell homeostasis in the normal kidney and during renal injury.     

Sertoli cells induce xenolymphocyte apoptosis in vitro

BACKGROUND: Testicular Sertoli cells can protect pancreatic islet grafts from allo- and autoimmune destruction; however, the mechanisms underlying immune privilege of the testicle are not well understood, especially in xenotransplantation. The purpose of this study was to investigate whether rat Sertoli cells could induce mouse lymphocyte apoptosis in vitro. METHODS: Testis was isolated from 2- to 4-week-old Sprague Dawley (SD) rats. Sertoli cells were successfully prepared by digestion with collagenase type V, trypsin, and DNase I, and then identified by electron microscope. Viability and apoptosis of cultured cells were measured by flow cytometry. We examined the apoptosis rates of Balb/c mouse lymphocytes, which were cocultured with SD rat Sertoli cells by FACS. The expression of Fas ligand (Fasl), transforming growth factor (TGF)-beta1 and clusterin on Sertoli cells were detected by immunocytochemistry. RESULTS: In the cocultured system, Sertoli cells accounted for more than 93%. With our isolation method, the viability of Sertoli cells was more than 95% and the apoptosis rate was 10.87% +/- 3.87%. The lymphocyte apoptosis ratio was 15.52 +/- 0.17 (P < .01, compared with the control groups). SABC immunochemistry staining showed that the sertoli cells could express FasL, TGF-beta, and clusterin. CONCLUSIONS: In our coculture in vitro, rat Sertoli cells expressed FasL and TGF-beta1 as well as induced the apoptosis of mouse lymphocytes. These results indicated that the expression of FasL and TGF-beta1 on Sertoli cells might relate to immune privilege in xenotransplantation.     

凋亡相关蛋白及粘附分子在大鼠心脏急性排斥反应中的作用

目的 探讨凋亡及凋亡相关因子Fas／FasL、Bcl-2/Bax，粘附分子ICAM-1、P-selectin、E-selectin、L-Selectin、PECAM-1和VCAM-1在心脏移植中的表达及意义。方法 用免疫组织化学方法和原位杂交法，检测大鼠心脏移植后不同时间(1、3、5、7d)、Fas／FasL、ICAM-1、P-selectin、L-selectin、E-selectin蛋白、Bcl-2/Bax、VCAM-1、PECAM-1 mRNA的表达情况。结果 随着移植后天数的增加，细胞凋亡增多，Fas／FasL、Bax表达增加，粘附分子ICAM-1、VCAM-1、PECAM-1表达也增加，Selectin表达较少。正常大鼠肝脏未见细胞凋亡及粘附分子的表达。结论 细胞凋亡和粘附分子ICAM-1、VCAM-1、PECAM-1表达增加可能与心脏移植后急性排斥反应有关，而Fas／FasL、Bax可能促进了凋亡的发生。     

Fas和FasL 系统在非梗阻性无精子症睾丸中的表达

目的：为探讨睾丸生精功能障碍与细胞凋亡的关系，研究Fas和FasL系统在非梗阻性无精子症睾丸支持、间质和生精细胞中的表达。方法：对20例非梗阻性无精子症患者行睾丸开放性活检，常规病理检查，按Johnson评分法评价精子发生和发生障碍的程度；采用免疫组化SABC法对睾丸支持、间质和生精细胞进行Fas和FasL表达的检测。结果：睾丸活检生精功能评为8分有14例，3分有2例，6、5、4和2分各有1例。在20例非梗阻性无精子症睾丸的间质、支持和生精细胞均有Fas和FasI。的表达；而支持细胞Fas和FasL的阳性和强阳性表达率明显高于间质和生精细胞。结论：非梗阻性无精子症的睾丸支持、间质及生精细胞Fas和FasL的高表达与精子生成障碍是一致的，非梗阻性无精子症可能与生殖细胞过度凋亡密切相关。     

Fas/FasL表达和细胞凋亡在人类同种异体移植肾急性排斥中的作用

目的探讨移植肾急性排斥（ＡＲ）时细胞凋亡与Ｆａｓ和Ｆａｓ配体（ＦａｓＬ）表达的作用及其临床意义。方法分别用原位末端标记技术（ＴＵＮＥＬ法）和免疫组织化学方法检测２６例移植肾ＡＲ标本中细胞凋亡和Ｆａｓ／ＦａｓＬ表达情况。结果细胞凋亡和Ｆａｓ／ＦａｓＬ表达主要在ＡＲ移植肾小管上皮发生,且凋亡指数和Ｆａｓ／ＦａｓＬ表达与肾组织病理损伤程度平行,与正常肾对照组和移植肾功能稳定组比较差异显著（Ｐ＜０．０１）。结论肾小管上皮细胞凋亡在ＡＲ所致的移植肾损伤中起重要作用,Ｆａｓ／ＦａｓＬ系统可能参与移植肾ＡＲ,是造成肾小管上皮细胞凋亡的重要因素。ＴＵＮＥＬ法检测细胞凋亡可作为判断移植肾病理变化和预后的重要指标     

Analysis of the Fas system and Bcl-2 in rat liver allograft rejection.

BACKGROUND: Apoptosis is involved in the mechanism of cell death observed in liver allograft rejection. The liver cells are sensitive to Fas-mediated apoptosis; however, little is known about the involvement of the Fas system in liver allograft rejection. We used rat models to investigate the expression of Fas/Fas ligand and apoptosis-related proteins during liver allograft rejection. MATERIALS AND METHODS: DA rats to Lewis, and Lewis to Lewis orthotopic liver transplantation were performed; liver samples were collected on days 1, 3, 5, 7, and 9 postoperatively (each n = 3). Apoptosis was monitored by TUNEL and electron microscopy. The expression of Fas, FasL, bcl-2, and bax was examined at the mRNA level and by means of immunohistochemistry. RESULTS: The TUNEL index in the allografts and isografts on day 7 was 20.1 +/- 1.5 and 7.7 +/- 2.6/1000 cells, respectively. Fas and bax mRNA were constitutively expressed in both of the groups. The expression of Fas ligand mRNA in the allografts which rose on day 5 was 10 times stronger compared to that in the isografts. On the other hand, bcl-2 mRNA was generally expressed in the isografts while it decreased in the allografts. The immunohistochemical analysis also showed an increased reactivity of Fas ligand on day 5 in the allograft, which was observed both in parenchymal and nonparenchymal cells. CONCLUSIONS: These results strongly suggest that Fas/Fas ligand interaction mediates the liver injury during allograft rejection. In addition, other regulatory factors of apoptosis, such as bcl-2, might also be involved in this pathogenesis.