Generation of CD2＋CD8＋ NK cells from c-kit＋ BM cells 173 NK cells. As a results, in present study, we show that a fraction of porcine c-kit＋ BM cells can develop into CD2＋ CD8＋ NK cells under the influence of cytokines (IL-2, IL-7, IL-15, IL-21, SCF, and FLT3L), HDCs, and stromal cells. But, there are several other possibilities for the low fre- quency of CD2 and CD8 molecules, porcine NK cell markers. First, the levels of cytokines required for gen- eration of NK cells from BM cells may have narrow ranges. Second, only a small portion of porcine c-kit＋ BM cells de- spite the right levels of cytokines can acquire NK cell properties. Consistent with this idea, CD2＋CD8＋ NK cells differentiated from c-kit＋ BM cells were a mere about 35.5% (Fig. 2). But, these cytokines-induced CD2 ＋ CD8＋ NK cells expressed the perforin, granzyme B, and NKG2D mRNA when mRNA levels were analyzed. And, their pro- tein levels were increased more than those in c-kit＋ BM cells not pretreated with cytokines (Fig. 3 and Fig. 4), sug- gesting that the cytokines-induced CD2＋CD8＋ NK cells possess a mechanisms that express the factors related to NK properties. Therefore, we speculate that these cytokines are required in the stage of NK cells generation from por- cine c-kit＋ BM cells. Another unsolved question is whether these cytokines we identified play roles completely in differ- entiation from c-kit＋ BM cells to CD2＋CD8＋ NK cells. Because some cytokines may be released from NK cells dur- ing the differentiation, we can also assume that these dif- ferentiation are partly involved in the machinery regulated by the earlier differentiated NK cells. Therefore, this study provides the first in vitro evidence of a possible role for a various of cytokines in CD2＋CD8＋ NK cells development from porcine c-kit＋ BM cells. Additionally, in previous study, Sotiriadis et al. described the pig model, specifically germ-free pigs and specific pathogen-free (SPF) mini-pigs, had been used to study various aspects of the immune sys- tem, specifically innate immunity [22]. Specifically, it is re- ported that at least for the first 4 weeks of life, pigs are immunocompetent, owing to complete absence of foreign antigen exposure and subsequent lack of immunoglobulin production [45]. Furthermore, it is known that the porcine molecules are much more similar to those in humans [46]. This allows for study of therapeutic applications in an or- ganism without any xenoreactivity with ADCC or direct cell lysis as well as graft-vs-host disease. Also, this confers a much more immediate pertinence to the use of pigs without immune rejection in human disease. But, although xeno- graft rejection has been largely resolved, other problems may remain major hurdles. 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