学会発表Characteristics of bovine fibroblast cells after freeze drying
In general, mammalian somatic cells are preserved in the ultra low temperature freezer or the liquid nitrogen. However, the maintenance and transportation of frozen cells are costly, and there are problems with a safety aspect on liquid nitrogen. Freeze drying, which removes moisture through sublimation, has been used for a long-term storage of food, drug, yeast. To our knowledge, little work has been devoted to freeze dry for mammalian somatic cells. The objective of this study was to assess the physical properties on freeze-dried (FD) products of bovine fibroblast cells, and evaluate the characteristics. Bovine fibroblast cells were obtained from ear skin tissue of a Japanese Brown Cattle-Kochi. The cells were suspended in a freeze-drying solution, and lyophilized for 11.5 h. The eutectic point of freeze-drying solution was about -28℃, therefore, we compared the freezing method between rapid freezing and slow freezing (-0.3℃/min) at -30℃. The DNA damage of the FD cells was evaluated with comet assay. The FD cells were also observed by Transmission Electron Microscopy (TEM). The moisture content and glass transition temperature (Tg) were measured by Karl Fischer titration and Differential Scanning Calorimetry, respectively. The rate of DNA-damaged cells was significantly lower in slow freezing (14%; 68/500) than in rapid freezing (24%; 122/500) (P<0.05). After freeze-drying, the plasma membrane was of all cells was completely damaged, however, the nuclear membrane was intact. In the FD products, the moisture content was 27% and Tg was -28℃. These results suggest that bovine FD fibroblast cells would be useful as a donor cell for nuclear transfer, because they preserve their nuclear structure. However, FD conditions should be improved for a long-term storage at room temperature.
学会発表Effects of feeding yuzu peel in Japanese Brown Cattle-Kochi under heat stress
In general, heat stress reduces feed intake in beef cattle, and it causes a decrease of meat production in summer. Yuzu (Citrus junos), which is a citrus fruit native to northeast Asia, contains various flavonoids and antioxidants. However, after the juice is extracted from fruit, most of the peel is treated as waste in Japan. In this study, we expected the function of anti-heat stress in yuzu, and investigated the effects of feeding yuzu peel in Japanese Brown Cattle-Kochi at the late stage of the fattening period in summer. This experiment was conducted from June to July in 2015. A total of 6 cattle (yuzu feeding group: 4, non-feeding group: 2) was examined. Yuzu peel paste of 2.5% was added to feed for one month before slaughter. The consumption of concentrate, roughage, and water was recorded every day. The body weight was measured before and after experiment. After slaughter, skeletal muscle, a liver, and fat were collected. Moreover, the skeletal muscle and the fat were vacuum-packed and stored at 0°C for 20 days to analyze fatty acids, amino acids, and metabolites. No significant differences in water intake were observed between yuzu feeding group and non-feeding group. The consumption of concentrate was higher in non-feeding group, and roughage was higher in yuzu feeding group. Yuzu feeding group gained weight more than non-feeding group during the experiment, therefore, dairy gain and feed efficiency in yuzu feeding group were higher than in non-feeding group. Yuzu feeding also altered the metabolism in the liver and the blood plasma. In the skeletal muscle, toxic substances, e.g. diethanolamine, N-glycolylneuraminic acid and histamine, were reduced in yuzu feeding group. These results suggest that feeding of yuzu peel maintains homeostasis in Japanese Brown Cattle-Kochi against heat stress, and improves the meat production in summer.