Mechanism of Azaline B-induced Apoptosis in Rat Testis / 대한비뇨기과학회지

PURPOSE: To identify the mechanism of azaline B-dependent apoptosis, the regulation of Fas and FasL genes has been investigated. MATERIALS AND METHODS: Azaline B was subcutaneously injected into Sprague-Dawley rats. The levels of Fas receptor (Fas) and Fas ligand (FasL) were detected by reverse transcription-polymerase chain reaction (RT- PCR). Azaline B-dependent apoptosis was determined by terminal deoxynucleotidyl transferase-mediated dUTP nick and labeling (TUNEL) and DNA fragmentation assay. Transacting factor of FasL promoter was identified by DNase I footprinting and DNA mobility shift assay. RESULTS: The azaline B-treated testis (250microgram/kg body wt/day) had decreased to 70+/-2.5% and 38+/-1.8% of the normal testis weight at 3 and 5 days after the injection, respectively, but the weights of the testis were not changed after pretreatment of follicle-stimulating hormone (FSH) and testosterone. Apoptosis of the testis was detected by DNA fragmentation assay and TUNEL assay after the azaline B treatment. The levels of Fas and FasL mRNA were increased by the treatment of azaline B in both time- and dose-dependent manners. In DNase I footprinting assay with FasL promoter, the nuclear factor prepared from control was bound with at least four sites: SP-1 binding site at 283, NF-kappa B binding site at 219, TATA at 132 and the gamma-interferon response element (gamma-IRE) at 78. gamma-IRE was completely protected by the nuclear extract prepared from azaline B-treated rat testis. In DNA mobility shift assay, the binding activity of gamma-IRE binding protein was increased after azaline B treatment. CONCLUSIONS: These results suggest that Fas-FasL system may be important to azaline B-dependent apoptosis in rat testis and that gamma-IRE binding protein is related to the azaline B-dependent regulation of FasL gene.

The Study on Apoptosis and Expression of Fas, Fas-ligand, Bax, and Bcl-2 in Human Fragmented Embryos / 대한불임학회지

OBJECTIVE: The present study was performed to investigate whether apoptosis occur in human embryos by annexin staining and detect the expression of Fas, Fas-ligand (FasL), Bax, and Bcl-2 in human fragmented embryos derived from IVF-ET by immunofluorescence and Western blot analysis. MATERIALS AND METHODS: Using annexin staining, immunofluorescence and Western blot analysis on normal and fragmented embryos, we were able to detect apoptotsis and apoptotic gene products in fragmented embryos. RESULTS: Phosphatidylserine (PS) translocation, the marker for apoptosis, were detected frequently in fragmented embryos. Bcl-2 and Bax protein were detected in both fragmented and non-fragmented embryos. When fragmented embryos compared to normal embryos, immunofluorescent intensity of Bcl-2 tended to be lower in fragmented embryos. Bax gene expression increased in the fragmented embryos compared to the normal embryos. This result supports a model in which the molar ratio of Bcl-2 to Bax determines whether apoptosis induced or inhibited in human embryo. Fas was highly expressed in human preimplantation embryos but not FasL. It suggests that embryo may undergo apoptosis by binding with FasL produced by follicular or immune cells. CONCLUSION: The over expression of Bax and Fas will trigger apoptosis to lead embryo fragmentation and change embryo to be nonviable.

The Clinical Significance of Fas and Fas Ligand Expression in Thyroid Neoplasms

PURPOSE: Recent studies indicate that Fas and Fas Ligand (Fas-L) are implicated in autoimmune endocrine diseases and tumors of the thyroid. In this study we tried to elucidate the expression stati of Fas and Fas-L in some kinds of thyroid neoplasms, and their relationships with 4 prognostic factors in papillary thyroid cancer (i.e., size, lymph node metastasis, capsule invasion, age). METHODS: 66 cases of thyroid neoplasm including 45 cases of papillary cancer (PTC), 3 of a follicular cancer (FTC), 1 of a poorly differentiated cancer (PDC), 1 of a undifferentiated cancer (UC), 7 of follicular adenoma (FA), and 9 of nodular hyperplasia (NH) were examined, and estimated as negative, weak positive and strong positive about the Fas and Fas-L expression by the immunohistochemical staining intensities. We then collected and compared the differrences between benign and malignant tumors. The expressions of Fas and Fas-L in papillary thyroid cancers were evaluated relating to the differences in the prognostic factors (i.e., the size, lymph node status, capsule invasion, and age of the patients). RESULTS: Malignant thyroid tumors revealed stronger staining intensity than benign neoplasms. In papillary thyroid cancers, Fas-L staining intensities were significantly stronger in the cases with perithyroidal lymph node metastasis, or in those of 45 years old or over than in those with no lymph node metastasis, or younger than 45 years. CONCLUSION: Both Fas and Fas-L are implicated in thyroid tumorigenesis and revealed stronger staining intensities in malignant than benign tumors, and the Fas-L staining intensities may have some prognostic implications at least in papillary thyroid cancers.