An important but unsolved question in this study is whether

An important but unsolved question in this study is whether ES cell-derived hematopoietic Gefitinib manufacturer and iPS cell-derived hematopoietic cells transduced with Ad-hHoxB4 have long-term hematopoietic reconstitution potential in vivo. Recent studies have demonstrated that some surface antigen expressions were different between bone marrow-derived HSPCs and ES cell-derived HSPCs, and that CD41+ cells had long-term repopulation ability in ES cell-derived HSPCs (McKinney-Freeman et al., 2009; Matsumoto et al., 2009). Our flow cytometric analysis revealed an increase of CD41+ cells in hHoxB4-transduced cells compared with non-transduced cells and LacZ-transduced cells (Fig. 3b). We also showed that Ad-hHoxB4-transduced cells could proliferate on OP9 stromal cells more efficiently than control cells (Fig. 2). Thus, these results suggest that immature hematopoietic cells were generated by transient hHoxB4 transduction, and that hHoxB4-transduced cells might have reconstitution potential in vivo. This in vivo transplantation analysis is now on-going in our laboratory. In the present study, we succeeded in the promotion of hematopoietic differentiation from mouse ES and iPS cells by Ad vector-mediated hHoxB4 transduction. Ad vector transduction can avoid the integration of transgene into host genomes, and multiple genes can be transduced by Ad vectors in an appropriate differentiation period. Thus, an even more efficient protocol for hematopoietic differentiation from ES and iPS cells could likely be established by co-transduction of HoxB4 and other genes involved in the hematopoiesis, such as Cdx4 (Wang et al., 2005) and Scl/Tal1 (Kurita et al., 2006), using Ad vectors. Taken together, our results show that Ad vector-mediated transient gene expression is valuable tool to induce hematopoietic cell from ES and iPS cells, and this strategy would be applicable to safe therapeutic applications, such as HSPC transplantation.
Materials and methods
Conflict of interest
Acknowledgments We thank Dr. S. Yamanaka for kindly providing the mouse iPS cell line 38C2 and 20D17. We would also like to thank Dr. J. Miyazaki and Dr. T. Imai for providing the CA promoter and anti-mouse CAR monoclonal antibody, respectively. We also thank Dr. K. Nishikawa (National Institute of Biomedical Innovation) for helpful comments. This work was supported by grants from the Ministry of Education, Culture, Sports, Science, and Technology (MEXT) of Japan and the Ministry of Health, Labour, and Welfare of Japan.
Introduction Stromal cells, typically identified as tissue-resident fibroblasts, form a supportive scaffold for both healthy and pathological tissues. Stromal cells implicated in disease can be divided into three broad types: mesenchymal stromal cells (MSCs), monocyte-derived stromal cells and stromal cells arising through epithelial-mesenchymal transition (EMT). Our unpublished data (Fig. 1) show the appearance of cells derived from these three alternative lineages in ex vivo culture. These cell populations are important players in development and tissue remodeling, regeneration of damaged organs, and fibrosis because they secrete growth/immunomodulatory factors and extracellular matrix (ECM) components. There are three key questions about stromal cells. First, due to the lack of specific markers, we do not know the relative contributions of MSCs, fibrocytes and EMT-derived cells to stroma in healthy and pathological organs. Second, much remains to be understood about whether these fibroblastic populations execute synergistic or antagonistic functions in disease. Third, it is unclear to what extant systemic mobilization and recruitment of progenitors from the bone marrow as opposed to their migration from extramedullary organs or resident tissues contributes to the formation of stroma.
Conclusion A number of questions regarding the etiology of stromal fibroblasts remain unanswered. The relative contribution of bone marrow, as opposed to other organs and resident tissue, to the pools of pathological stromal cells is to be defined in different pathological settings (Fig. 2). The relative content of mesenchymal progenitors, monocytes, and EMT-derived cells in pathological stroma is also yet to be characterized in disease-specific contexts. An important question is whether EMT-, MSC- and monocyte-derived stromal cells execute complementary or opposing functions during fibrotic pathogenesis (Fig. 3). It is unclear to which extent trans-differentiation of plastic fibroblast progenitors recruited into the lesion complicates the issue. Progress has been limited by the lack of reliable markers that could distinguish the distinct populations of stromal cells. In the past, various approaches have been undertaken to characterize molecular differences between stromal populations, however success in this endeavor has been limited. Recent identification of a new marker of adipose MSCs (Daquinag et al., 2011a) sets the stage for efforts toward systematic characterization of stromal progenitor markers. In the future, identification of differentially expressed cell surface molecules may enable targeting of stromal cell populations. Understanding the properties and roles of EMT-derived cells, MSCs, and monocytic stroma from bone marrow and other organs may outline strategies to differentially control the functions of the respective cell populations in order to suppress disease progression. We envision therapies aimed at specific stromal cell pools becoming a complementary treatment of fibrosis and cancer. New stromal cell markers will make it possible not only to quantify distinct populations of pathological stromal cells in disease, but also to ablate a desired subpopulation. Conversely, the approach of using MSCs as vehicles (Hall et al., 2007) might be expanded to fibrocytes and EMT-derived cells for directing treatment to desired sites for therapeutic purposes.