Use of human tissue and cells is the central theme in bio-medical research. Under the strong pressure and severe competition in international society, Japanese government has just started to settle the guidelines in the area. In year 2000, we will have at least 7 independent guidelines from the government. Since in practice a research project will be related to pleural guidelines, the use of human material in the bio-medial area will be more complicated than ever. Therefore, we will be cooperatively work with Institutional Review Board and find the rational pathway to realize public acceptance for our research. On the way of our planning we had the reports of Ministry of Health and Welfare Committee (Chair, Dr. K. Kurokawa,1) and Ethical Committee in Japanese Tissue Culture Association (Chair, Dr. Matumura,2). An author of this report (T. M. ) had been given a chance to participate in the discussion of the Committee of Ethical Issues in JTCA. In this report we will describe the process of our effort to establish the acceptable status in use of human material as public research resources.

View full abstract

This paper introduces the comparative quality evaluations of bovine embryos developed from in vitro-matured (IVM) and -fertilized (IVF) oocytes cultured in different culture systems using serum-free or serum-containing media. Recently, we developed serum-free culture systems for in vitro development of bovine embryos with or without somatic cell (bovine cumulus/granulosa cells: BCGC) cocultures. The improved serum-free media (IVMD101 in the presence of BCGC and IVD101 in the absence of BCGC cocultures) can offer several advantages over the culture in serum-containing medium, including increased rates of blastocyst formation and cell numbers. Conspicuous differences in the morphological features between the embryos cultured in serum-free media and serum-supplemented medium were observed at the morula and blastocyst stages. Morulae and blastocysts cultured in serum-supplemented medium had cells containing numerous lipid droplets (LD), especially immature mitochondria were frequently observed in the blastomeres at the morula stage. On the other hand, the blastomeres of morulae and blastocysts cultured in serum-free media contained many elongated mitochondria and lysosome-like vesicles, but LD were not abundant. The histochemical examination demonstrated that large sudanophilic LD increased dramatically in the embryos cultured in serum-supplemented medium from the morula to the blastocyst stages. These findings showed that bovine embryos cultured in serum-containing medium abnormally accumulated cytoplasmic lipids into their cytoplasm. Furthermore, the bovine plasma lipoproteins may directly affect on the accumulation of cytoplasmic lipids. The survival and hatching rates of embryos (morulae, early blastocysts, blastocysts) produced in serum-free media after postthaw culture were superior to those of embryos produced in serum-containing medium. The abnormal accumulation of cytoplasmic LD in embryos may affect the sensitivity of embryos to chillig and freezing. These serum-free culture systems are beneficial, not only for the production of good quality embryos from IVM-IVF bovine oocytes, but also for the study of the influences of serum factors on the embryo quality.

View full abstract

[in Japanese]

View full abstract