453. Tumorigenic Potential in Clones Derived from Rat Bone Marrow Cells

Abstract

Therapy using different stem cell populations has recently emerged as a complementary treatment for several diseases, and a wide variety of stem cell lineages have been explored. Mesenchymal stem cells (MSCs) represent an attractive option for use in cellular therapy because of their accessible nature and abundance in various tissues, such as bone marrow (BM) and adipose tissue. While MSCs are routinely isolated from BM, it is now evident that MSCs can also be isolated from many other tissues. These cells can be divided, based on their source of origin, into two broad categories: marrow-derived and non-marrow-derived. Although MSCs isolated from different sources share similar surface antigens and exhibit similar classical differentiation potential (bone, fat, and cartilage), these cells still exhibit heterogeneity in their phenotype and biological properties, which apparently depends on their tissue of origin and microenvironmental niche. Self-renewal is a property shared among cancer cells and stem cells. Stem cells can certainly be genetically susceptible and acquire mutations through the cell-culture process; even so, the potential tumorigenic risk of using MSCs from a heterogeneous cellular mixture has not yet been properly evaluated. Some cells derived from BM can present some markers associated with cancer (e.g., CD117 and CD34, which are always found in a gastrointestinal stromal tumor (GIST) diagnosis, the most common mesenchymal tumor of the gastrointestinal tract. Nonetheless, the presence of both markers on BM with tumorigenic potential has not been sufficiently explored yet. Interestingly, some subpopulations isolated from BM supernatant express CD34 and CD117 (c-Kit). CD117 is a tyrosine-kinase receptor activated by stem cell factor. Upon stimulation, CD117 activates multiple signaling pathways, including ERK and mTOR, which ultimately lead the activation of NFk-B; this nuclear factor is coincidently upregulated in various tumors. The goal of this work was to identify a direct correlation between immunophenotype and possible tumorigenic potential in a specific clone of adherent c-kit+/ CD34+ cells isolated by cell sorting from the supernatant of unfractionated rat bone marrow (SNBMC). We evaluated SNBMC tumorigenic potential in vivo and in vitro. We subcutaneously injected SNBMC CD117/CD34-positive cells and heterogeneous SNBMC (unsorted) into the bilateral flank of 55 male nude mice. We measured tumor size weekly for a month, and we found that tumors were present in 20% of the mice treated with unsorted cells and in 100% of the mice that received SNBMC CD117/CD34-positive cells. All of the tumors were solid neoplasias with small, undifferentiated cells and frequent mitotic figures; intratumoral necrotic areas were abundant, suggesting a rapid proliferation rate. Immunohistochemistry staining revealed intense immunoreactivity for cytokeratin, and we suggested a diagnosis of anaplastic carcinoma. Flow cytometry analysis from tumor-isolated cells revealed a loss of MSC surface markers such as CD90 and CD73; however CD117 and CD34 were preserved. Finally the participation of NFk-B in cell proliferation was evaluated by transfection with an IkB-α adenovirus and a significantly reduced proliferation rate was found. Nonetheless, given the increasing interest in using MSCs for regenerative medicine applications, the safety of administering MSCs to humans remains to be carefully evaluated in light of the potential role of MSCs in driving tumorigenicity, particularly cells expressing CD117/CD34. Understanding the tumorigenic process in a future will be important to determining the role of these markers on the molecular level.