Two piglets and one juvenile pig were used to investigate closely what types of cells express green fluorescent protein (GFP) and if any, whether the GFP-tagged cells could be used for stem cell transplantation research as a middle-sized animal model in bone marrow cells of recloned GFP pigs. Bone marrow cells were recovered from the tibia, and further analyzed with various cell lineage markers to determine which cell lineage is concurrently expressing visible GFP in each individual animal. In the three animals, visible GFP were observed only in proportions of the plated cells immediately after collection, showing 41, 2 and 91% of bone marrow cells in clones #1, 2 and 3, respectively. The intensity of the visible GFP expression was variable even in an individual clone depending on cell sizes and types. The overall intensities of GFP expression were also different among the individual clones from very weak, weak to strong. Upon culture for 14 days in vitro (14DIV), some cell types showed intensive GFP expression throughout the cells; in particular, in cytoskeletons and the nucleus, on the other hand. Others are shown to be diffused GFP expression patterns only in the cytoplasm. Finally, characterization of stem cell lineage markers was carried out only in the clone #3 who showed intensive GFP expression. SSEA-1, SSEA-3, CD34, nestin and GFAP were expressed in proportions of the GFP expressing cells, but not all of them, suggesting that GFP expression occur in various cell lineages. These results indicate that targeted insertion of GFP gene should be pursued as in mouse approach to be useful for stem cell research. Furthermore, cell- or tissue-specific promoter should also be used if GFP pig is going to be meaningful for a model for stem cell transplantation.

As an aquaculture effluent is recognized as a source of water pollution, many methods are being researched to solve this problem. It has been reported that the aquaculture effluent contains many organic compounds containing nitrogen, phosphorus, potassium, and calcium. The purpose of this study was to investigate the possibility of using the effluent from bio-floc technology (BFT) inland aquaculture for fertilizing blueberry ‘Duke’ to promote the growth. The experiment was conducted in a commercial blueberry farm in Jinju, Gyeongsangnam-do, where blueberries are actually cultivated in order to find out the effects of fertilizers in situ. The experiment was carried out with five treatments: control with only irrigation, conventional nutrient solution fertigation, and the fertigation with three concentrations (×1, ×0.5, or ×0.25) of effluent from BFT (Bio-Floc Technology) inland aquaculture. The treatment period was from the beginning of April, when a new leaf of blueberry began to develop, to the middle of June, when the blueberry was harvested, for a total of 10 weeks. As a result of the experiment, the treatments with effluent from BFT inland aquaculture showed better growth qualities than those of control and conventional cultivation. However, there was no statistically significant difference among all treatments regarding the total fruit production. Based on these results, it was concluded that the effluent from BFT inland aquaculture can be used for the cultivation of blueberry because it is equivalent to conventional cultivation method. This study provided a new viewpoint of recycling the effluent from BFT for agricultural purpose to reduce water pollution problems due to aquaculture wastewater.

Use of nature-derived matrices of a part of body tissues has been used to repair damaged tissues in practical terms. Recently, the same idea has also been applied to regenerate whole organs including the heart, liver, lung, and pancreas etc. Thus, so-called bio-artificial organ technology becomes a promising way of overcoming the lack of donor organs and immune rejections in organ transplantation if we can obtain recipient stem cells. Although the regenerated heart in vitro so far may demonstrate some typical organ's responses in vitro and vivo, it is still far from a fully functional organ for transplantation. We initiated a study to look at changes occurring during the generation of bio-artificial organ using the mouse model. Adult hearts were dissected out and perfused for acellularization with SDS-containing buffer and washed several times. Enzymatic treatment also evaluated the acellular purity by isolating genomic DNA and total RNA before and after DNase and RNase treatments. For recellularization, differentiating H9C2 cell or cells derived from P19 EC cells along with mesenchymal stem cells were seeded on the finally obtained heart matrix several times before submerging culture for generating the heart. Histological analyses revealed that complete removal of cellular components. The intensive staining of alcian blue (pH 1.5 and 2.5) suggests that acid mucopolysaccharides, glycocomponents and sulfate-containing saccharides are widely spread within the heart matrix. There was little DNA and RNA in the heart matrix after the enzymatic treatments as judging by the DAPI or PI staining. Cell seeding and subsequent submerging culture showed substantial heart tissue development as evidenced by immunocytochemistry and RT-PCR in the recellularized and grown heart. From these results, we suggest that each procedure for bio-artificial organ has to be carefully examined to improve the entire process.