Expression of functional Fas ligand in choriocarcinoma

PROBLEM: In the course of pregnancy, fetal trophoblast cells and in that of choriocarcinoma-etiology, trophoblast derived tumor cells, invade the uterine mucosa without causing rejection by decidual leukocytes. Fas ligand (FasL, CD95L, APO-IL), a central regulator of the immune system, has been implicated in the maintenance of immune privileged sites, such as the eye, the testis and the pregnant uterus by inducing apoptosis in activated infiltrating leukocytes. In normal pregnancy FasL, which is expressed by trophoblast cells, appears to contribute to the immune privilege of the pregnant uterus. As choriocarcinoma derives from trophoblast we wanted to assess the expression of FasL in this tissue. METHOD OF STUDY: Immunohistochemistry, immunofluorescence, TUNEL-assay, Western blotting, coculture experiments and flourescence-associated cell sorter-analysis were the techniques used. RESULTS: Expression of FasL was found on cells of choriocarcinoma in paraffin sections in situ and on three choriocarcinoma cell lines such as JEG-3, JAR and BeWo. These results were confirmed by Western blotting. In coculture experiments choriocarcinoma cells induced apoptosis in a Jurkat cell line - sensitive to FasL mediated killing. However, when the Jurkat cells were pre-incubated with a Fas-blocking monoclonal antibody, apoptosis was abolished to a great extent. CONCLUSION: Our findings show that choriocarcinoma cells express FasL and this aforementioned molecule is biologically active. We assume that FasL expression on choriocarcinoma cells may contribute to control of anti-tumor responses by inducing apoptosis in activated Fas bearing leukocytes.     

Expression and function of the Fas receptor on human blood and tissue eosinophils

The interaction between activated T cells and eosinophils has been proposed to play an important role in the pathogenesis of allergic diseases. T cell-derived cytokines such as interleukin-5 and granulocyte/macrophage colony-stimulating factor inhibit eosinophil apoptosis and may therefore contribute to the development of tissue and blood eosinophilia in these disorders. Withdrawal of these cytokines leads to eosinophil apoptosis in vitro. In contrast, the mechanisms which actively induce apoptosis in eosinophils are at present not completely understood. In this study, we demonstrate that freshly isolated human eosinophils express mRNA and protein for the Fas receptor. Using anti-Fas monoclonal antibody (mAb), we show that Fas activation accelerates apoptotic eosinophil death in vitro. Moreover, treatment of nasal polyps ex vivo with anti-Fas mAb decreased eosinophilic tissue inflammation. However, we observed that blood as well as tissue eosinophils derived from some eosinophilic donors do not express functional Fas receptors, although Fas protein is normally expressed in these cells. This implies that the susceptibility of the Fas receptor is a matter of regulation in eosinophils as previously observed in other systems. These data suggest that Fas ligand/Fas interactions are involved in the regulation of eosinophil apoptosis and that defects in this system could contribute to the accumulation of these cells in allergic and asthmatic diseases.     

Expression of the cystic fibrosis gene in human foetal tissues

In order to examine the onset of the cystic fibrosis (CF) diseaseprocess, the expression of the cystic fibrosis gene (CFTR) hasbeen examined in mid-trimester human foetal tissues by in situhybridization. CFTR mRNA was detected in the epithelia of pancreaticducts, small intestine, colon, genital ducts, lung and trachea.The majority of these sites of CFTR expression in the foetusare similar to those seen in adult tissues. However, epitheliaof the lung, that contain very little CFTR mRNA in the adult,express high levels of CFTR mRNA in the foetus. Since the lungis the major site of pathology and morbidity in CF these findingshave implications for treatment.     

Inflammatory neutrophils retain susceptibility to apoptosis mediated via the Fas death receptor

Apoptosis and clearance of neutrophils is essential for successful resolution of inflammation. Altered signaling via the Fas receptor could explain the observed prolongation of neutrophil lifespan and associated tissue injury at inflammatory sites. We therefore compared inflammatory neutrophils extracted from joints of rheumatoid arthritis patients, with peripheral blood neutrophils. Inflammatory neutrophils underwent constitutive apoptosis in culture more rapidly than peripheral blood neutrophils; this was not explained by changes in surface expression of Fas or by induction of Fas ligand. Inflammatory neutrophils remained sensitive to Fas-induced death, at levels comparable to those seen in peripheral blood neutrophils. Similarly, granulocyte-macrophage colony-stimulating factor reduced apoptosis but did not abolish signaling via Fas. These data provide evidence for the rate of apoptosis in inflammatory neutrophils being continually modulated by death and survival signals in the inflammatory milieu. This allows for rapid resolution of inflammation as levels of survival factors fall, and suggests new strategies for inducing resolution of inflammation.     

Expression of a receptor for IgM by human T cells in vitro

The capacity of human peripheral blood lymphocytes (PBL) to bind antigen-IgG antibody (EA(IgG)) and antigen-IgM antibody (EA(IgM)) complexes was investigated using a rosette technique with ox erythrocytes coated with rabbit IgG or IgM antibody. It was found that while EA(IgG)-rosette-forming cells (RFC) were detected on PBL freshly drawn from normal individuals, EA(IgM)-RFC were present only in suspensions kept in culture for 24 h in mediá supplemented with sera containing very low or no amounts of IgM. Experiments of simultaneous detection of EA(IgG)-RFC or EA(IgM)-RFC and other membrane markers for human T or B cells together with experiments on purified T or B cell populations indicated that EA(IgG)-RFC were formed by both T and B cells, while T cells only were capable of EA(IgM) rosette formation. The specificity of the receptors for IgG and IgM was determined by studying the inhibitory capacity of purified human IgM and IgG in the rosette assay. The receptor for IgG was inhibited by IgG and not by IgM, while the reverse was true for the receptor for IgM.     

Expression of low-affinity Fc gamma receptor by a human metastatic melanoma line

The class IIa of low-affinity receptors for the Fc region of IgG, Fc gamma RIIa, are expressed on immune cells. The cross-linking of Fc gamma RIIa by complexed IgG triggers activation of protein tyrosine kinase and internalization of immune complexes. In this report, we demonstrate the expression of Fc gamma RIIa by a human melanoma cell line (VIO) derived from a metastasis of a patient with regressive melanoma. The analysis of Fc gamma RIIa functions was performed in VIO cells and Fc gamma RlIa- or Fc gamma RIlb-transfected human melanoma cells (A375). The Fc gamma RIIa cross-linking induced protein tyrosine phosphorylation, including Fc gamma RIIa phosphorylation, and led to its internalization in a clathrin-independent way in human melanoma cells. Moreover, we showed that a part of internalized Fc gamma RIIa migrates in late endosomes, lysosomes and class II-containing compartments. These results suggest that melanoma cells can express functional Fc gamma RII, which might play a role in tumor-host relationships.     

可溶性Fas表达水平与人黑色素瘤相关性的初步研究

目的:分析可溶性Fas(soluble Fas,sFas)表达水平与人黑色素细胞瘤恶性程度的关系,探讨Fas、Fas配体与可溶性Fas在调控细胞凋亡信号转导途径中的作用.方法:对35位确诊黑色素细胞癌患者的皮肤组织标本和病例进行回顾性分析和归类,用免疫荧光的方法检测组织中Fas/FasL的表达水平.通过酶联免疫吸附反应(ELISA)方法对人血清中sFas的浓度进行分析.结果:35例黑色素细胞癌组织中,病理分型浅表型2例、结节型9例、肢端型18例、雀斑痣型6例.Fas在正常皮肤组织中高表达,恶性黑色素癌组织中低表达;而相反FasL在恶性黑色素癌组织中高表达,在正常皮肤组织中表达非常低.可溶性Fas在不同组织中的表达强度差异显著,sFas在恶性黑色素细胞癌患者、良性黑色素瘤和正常人血清中的浓度分别为(36.32±11.57) ng/ml、(7.33±3.78)ng/ml和(4.18±1.88)ng/ml.结论:可溶性Fas在恶性黑色素癌患者血清中含量较高,与黑色素细胞癌恶性程度呈正相关,sFas对细胞凋亡的竞争性抑制可能与黑色素细胞癌的进展密切相关.     

Expression of GABA-immunoreactivity by spinal motoneurons of some vertebrates

Electrophysiological and biochemical investigations have shown that gamma-aminobutyric acid (GABA) is the main inhibitory neurotransmitter in the vertebrate central nervous system. However, the present study shows that some motoneurons located in the spinal cord of young chickens and adult monkeys display a GABA-like immunoreactivity. The expression of GABA immunoreactivity in vertebrate motoneurons suggests that this inhibitory amino acid is colocalized with acetylcholine and could play a role in the neuromuscular transmission.     

Expression of a complete and functional complement system by human neuronal cells in vitro

We demonstrate in vitro expression of complement components, i.e. C3, factor H (FH), factor B (FB), C4, C1-inhibitor (C1-inh), C1q, C5, C6, C7 and C9, by four human neuroblastoma cell lines IMR32, SKNSH, SH-SY5Y and KELLY. Activating proteins C4, C9 and C1q, and regulatory proteins FH and C1-inh were produced constitutively by the four cell lines. C3, C6 and FB were mainly produced by SKNSH and SH-SY5Y. Western blot experiments showed that secreted proteins were structurally similar to their serum counterparts. An additional polypeptide of 43 kDa with FH immunoreactivity was detected, which could correspond to the N-terminal truncated form found in plasma. Regulation of complement expression by inflammatory cytokines, lipopolysaccharide and dexamethasone was tested in vitro. These factors had no significant effects on activating synthesis of components C3, FB and C4, but expression of regulating components C1-inh and FH was strongly increased particularly by IFN-gamma and tumor necrosis factor-alpha. The rate of synthesis of complement components was dependent on the differentiation of neuroblastoma cells. This effect of differentiation was also observed on normal rat neurons. Rat cerebellar granule cells constitutively expressed mRNA for C4 and C1q, but expression of C3 mRNA was induced by differentiation. This study shows that neurons could be another local source of complement in the brain, besides astrocytes and microglia. Human neuroblastoma cell lines can constitute an interesting model to analyze complement biosynthesis by human neurons. Local complement expression by neurons in vivo may be implicated in some physio-pathological processes.     

Expression of Fas and FasL in human serous ovarian epithelial tumors

The expression of Fas and FasL was studied in 86 patients with benign, borderline, and malignant serous ovarian lesions. Four normal ovaries, and monolayer epithelial cultures from a human fetal ovary, a borderline, and a serous adenocarcinoma were used for comparison. Expression of Fas and FasL was studied immunohistochemically and flowcytometrically. Fas was expressed in all 90 lesions; FasL in 57 lesions, including 2 normal ovaries. Fas expression was significantly increased in borderline tumors compared with benign (P = 0.005, t = -2.94) or malignant serous tumors (P = 0.0001, t = 4.15). FasL expression was significantly increased in malignant tumors compared with benign (P = 0.039, t = -2.10) and borderline tumors (P = 0.0016, t = -3.33). Flow cytometry showed a range of Fas expression in short-term cultures isolated from normal, borderline, and malignant ovarian serous tissue; in the few samples studied, FasL was not expressed. Expression in three serous ovarian cell lines was similar. Fas and FasL expression differed throughout the spectrum of ovarian lesions. FasL expression was increased in malignant tumors, and Fas expression was increased in borderline tumors. Changes in Fas/FasL expression in ovarian surface epithelium might play a functional role in the biology of ovarian tumors.     

Expression of epidermal growth factor receptor and transferrin receptor by human trophoblast populations

Epidermal growth factor (EGF) has several roles, including stimulation of cell division and differentiation. EGF receptor (EGFR) has been localized to villous syncytiotrophoblast, but expression by other human trophoblast populations has not been reported. EGFR expression was examined in normal and pathological placental tissues using a streptavidin-biotin-peroxidase technique; results were compared with expression of transferrin receptor (Tf-R) in similar tissues. EGFR was detected on villous syncytiotrophoblast in early and term pregnancy with labelling of the apical membrane, focal cytoplasmic reactivity, and patchy labelling of the trophoblast basement membrane. In contrast with other reports, EGFR was also consistently localized to villous cytotrophoblast, chorion laeve, and extravillous trophoblast populations in maternal uterine tissues. Maternal decidua showed diffuse labelling of stromal cells, particularly in the superficial zones. The reaction pattern in ectopic tubal pregnancy was similar to that in early intrauterine pregnancy. In molar pregnancy, EGFR was detected on villous syncytiotrophoblast and cytotrophoblast. In contrast, in normal, ectopic, and molar pregnancies labelling for Tf-R was confined to syncytiotrophoblast and to the proximal portions of the cytotrophoblast columns. Expression of EGFR by all trophoblast cells may represent a mechanism of placental growth and proliferation control. EGFR may also be involved with establishment of differentiated trophoblast functions including hormone secretion.     

Fas和β—雌二醇受体融合基因在L929细胞中的表达及活性鉴定

利用ＰＣＲ方法将人Ｆａｓ基因的信号肽、跨膜区、胞内区与人β－雌二醇受体的激素结合结构域基因融合，并插入高效真核表达载体ｐｃＤＮＡ３。用脂质体法转染Ｌ９２９细胞，加入Ｇ４１８４周后，筛选出的转化细胞经Ｗｅｓｔ－ｅｒｎｂｌｏｔ检测证明，该融合基因在Ｌ９２９细胞中得到较高表达。ＭＴＴ法检测进一步表明，在培养基中加入β－雌二醇可有效地诱导转化细胞发生凋亡，ＩＣ５０为１０－１０ｍｏｌ／Ｌ。     

Premature contraction of the ductus arteriosus: a cause of foetal death

Necropsy findings are presented of two cases of foetal heart failure secondary to premature, in-utero contraction should lead to a redirection of the right heart output either through the pulmonary vascular bed or to the systemic circulation via the foramen ovale. The muscular pulmonary arteries, however, were not dilated, indicating that no excessive pulmonary flow had occurred. The increased volume load on the right heart may have rendered the foramen ovale restrictive, with right-sided heart failures as a consequence. Premature closure of the ductus arteriosus should be considered a cause of foetal or early neonatal death.     

死亡受体5在小鼠神经增殖细胞中的表达

目的 探讨死亡受体5(DR5)对神经细胞增殖的影响.方法 采用5-溴脱氧尿嘧啶核苷(BrdU)、DR5、Doublecortin(DCX)等抗体免疫荧光标记法,检测各发育阶段(从胚胎期至生后成年,共100只小鼠)脑组织内DR5阳性细胞的表达变化,以及DR5阳性细胞与神经增殖细胞的关系.]结果在胚胎期和新生鼠中,DR5阳...     

Recruitment order of motoneurons during functional tasks

The recruitment order of motoneurons of first dorsal interosseous (1DI) muscle has been shown to be the same during either isometric abduction or flexion of the index finger. However, it is not known whether this stereotyped recruitment is the same for all functional tasks. This question was addressed by using the 1DI muscle, which contributes to many functional tasks involving the hand and fingers. Single motor units and surface electromyographic activity were recorded from 1DI in three human subjects. Each subject performed tasks of abduction of the index finger, rotation to unscrew a threaded knob and pincer to close a spring-loaded clip. The same motor units were activated for each of the three tasks, providing no evidence for the existence of task groups. The order of recruitment of pairs of selected motor units was the same for each of the three tasks.     

Recruitment order of motoneurons during functional tasks

The recruitment order of motoneurons of first dorsal interosseous (1DI) muscle has been shown to be the same during either isometric abduction or flexion of the index finger. However, it is not known whether this stereotyped recruitment is the same for all functional tasks. This question was addressed by using the 1DI muscle, which contributes to many functional tasks involving the hand and fingers. Single motor units and surface electromyographic activity were recorded from 1DI in three human subjects. Each subject performed tasks of abduction of the index finger, rotation to unscrew a threaded knob and pincer to close a springloaded clip. The same motor units were activated for each of the three tasks, providing no evidence for the existence of task groups. The order of recruitment of pairs of selected motor units was the same for each of the three tasks.