ABSTRACT. Diploid and triploid specimens of Japanese and Korean Fasciola sp. showed abnormality in their spermatogenesis. Live germ cells obtained from the testes were observed under a differential interference contrast microscope. In the stages from spermatogonium to spermatid, the cells combined together at the central cytoplasmic bridge during a series of divisions. One spermatogonium becomes a cell group of 8 primary spermatocytes through 3 mitoses. Until the primary spermatocyte stage, cells are divided in a uniform manner. In most of the diploid specimens, the primary spermatocytes are irregularly divided into non-uniform secondary spermatocytes, however, some specimens perform a regular division. In the majority of triploid flukes, the primary spermatocytes are divided in a regular pattern, but some of the specimens perform an irregular division. The non-uniform spermatids do not perform a spermiogenesis. In the diploid specimens, no spermatozoa were found that were produced by spermiogenesis. Whereas in the triploid specimens, some spermatids distributed uniformly on the surface went through a spermiogenesis. We observed some moving spermatozoa in one triploid specimen. The spermatozoa possibly retain their normal reproductive function. -KEY WORDS: chromosome, Fasciola, parthenogenesis, spermatogenesis, triploid.
Objective To examine the relationships between the form of cell death (apoptosis or necrosis), reactive oxygen species (ROS) generation, superoxide dismutase (SOD) activity and the level of heat-shock protein 70 (hsp 70) expression after thermotherapy of PC-3 prostate cancer cells; also assessed were the tumoricidal effects of combined treatment with both heat and the antioxidant inhibitor diethyldithiocarbamate (DDC). Materials and methods PC-3 cells were treated with thermotherapy at 42, 43 or 44uC for 30, 60, 90 or 120 min. Cell proliferation, ROS generation, SOD activity and cellular hsp 70 level were determined using tetrazolium-based cytotoxicity, fluorescent dichlorofluorescein (DCF) and nitroblue tetrazolium assays, Western blot analysis and flow cytometry, respectively. The apoptotic and necrotic cells were determined by staining with propidium iodide and fluorescein isothiocyanate-labelled annexin V. These variable were also measured after combined treatment of PC-3 cells with 1 mmol/L DDC and thermotherapy at 43 or 44uC for 60 min. Results Cell survival was significantly lower after heating cells at 43uC for 60, 90 and 120 min and at 44uC for all periods tested (P<0.05). At 43uC apoptosis increased with the duration of heating and was similarly enhanced after heating at 44uC for 30 min. Necrosis was not increased by heating at 42 or 43uC, but was markedly enhanced after heating at 44uC with both the duration of heating and with time after heating. Significant increases in DCF production were induced by heating at 43uC for 60, 90 and 120 min (P<0.05) and at 44uC at all times (P<0.010-0.005).There was a significant correlation between the level of ROS generation and necrosis (P<0.001) but no correlation between the ROS level and apoptosis. SOD activity increased in cells after heating at 43uC, with significant differences among cells heated for 60, 90 and 120 min (P<0.05). After heating at 44uC, SOD activity was maximal in cells heated for 30 min (P<0.005), by 30 min and then decreased with time after heating. There were significant increases in hsp 70 level in cells heated at 43uC for 90 and 120 min (P<0.05) and at 44uC for 30 and 60 min (P<0.05 and <0.025, respectively). Hsp 70 levels decreased after heating at 44uC for 90 and 120 min. The combination of DDC and heating significantly increased ROS generation and the percentage of cell death, and decreased SOD activity (P<0.05). Conclusions These findings show a qualitative change in the form of cell death induced by thermotherapy of PC-3 cells, which changed from apoptosis to necrosis according to the degree and duration of heating. Mild thermotherapy induced marginally low occurrence of apoptosis of PC-3 cells and DDC may represent a useful future strategy for the treatment of prostate carcinoma.
OBJECTIVE To determine whether oestrogen enhances platinum sensitivity, and if promoter CpG methylation of the oestrogen receptor‐α (ER‐α) gene determines the potential of cisplatin‐induced apoptosis in prostate cancer, as the high‐mobility group 1 (HMG1) preferentially binds to cisplatin‐modified DNA and is up‐regulated after oestrogen treatment in breast cancer cell line MCF‐7. MATERIALS AND METHODS The study comprised prostate cancer cell lines (LNCaP and PC‐3), 156 pathologically confirmed 156 radical prostatectomy samples and eight hormone‐refractory prostate cancer (HRPC) samples (from needle biopsy). Expression of HMG1 in cell lines was analysed by Western blotting or differential reverse‐transcription‐polymerase chain reaction (PCR). The methylation status of ER‐α was analysed by methylation‐specific PCR using bisulphite DNA as a template or bisulphite DNA sequencing. RESULTS In LNCaP cells, treatment with oestrogen increased HMG1 expression and co‐treatment with cisplatin and oestrogen reduced cell viability by accelerating apoptosis, compared with cisplatin alone. However, in PC‐3, oestrogen did not up‐regulate HMG1 or accelerate the cisplatin‐induced apoptosis. Although ER‐β was expressed in both LNCaP and PC‐3, ER‐α was expressed only in LNCaP. Bisulphite DNA sequencing of the ER‐α promoter showed partial methylation in LNCaP but complete methylation in PC‐3. ER‐α AS transfection diminished the cisplatin‐induced apoptosis in oestrogen‐treated LNCaP cells. In clinical samples there was ER‐α hypermethylation in 40% of prostate cancers this correlated with Gleason score (GS, 31% for GS < 7, 50% for GS = 7 and 56% for GS > 7). In addition, five of eight HRPC samples showed ER‐α hypermethylation. CONCLUSION These findings suggest that HMG1 induction as an enhancer of platinum sensitivity is mediated through interaction between oestrogen and ER‐α. As CpG hypermethylation of the ER‐α promoter is a frequent event in aggressive prostate cancer, negative conversion of ER‐α methylation is essential to achieve the most beneficial effect when combined chemotherapy of cisplatin with oestrogen is used in patients with prostate cancer.
ABSTRACT. Reproductive organs of stained and mounted whole specimens of different types of Fasciola (F. hepatica, F. gigantica, and parthenogenetic diploid and triploid flukes) were observed to clarify the structure of their reproductive organs. The results are as follows; 1. Basic structure differences could not be identified. 2. The flukes without sperm, or those with an extremely small quantity in the seminal vesicle, are parthenogenetic Fasciola sp. 3. It was newly discovered that the surface of the cirrus is surrounded by many shallow gutters, and that spines form a line in the gutters. 4. The structure of the reproductive organ on the genus Fasciola are shown in detail in the figures.
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