Bone marrow (BM) cell harvesting is a crucial element in the isolation of mesenchymal stem cells (MSCs). A simple method for harvesting cat BM cells is described. The results show that a large number of BM cells can rapidly be harvested from the cat by this simple procedure. MSCs prepared by density-gradient method were spindle-shaped morphology with bipolar or polygonal cell bodies and strongly positive for CD9 and CD44 and negative for CD18 and CD45-like. They were capable of differentiation to adipocytic and osteocytic phenotypes when exposed to appropriate induction media. The advantages of this method are its rapidity, simplicity, low invasiveness, and low donor attrition and good outcome.

Bone marrow mesenchymal stem cells (BMMSCs) have the capacity for self-renewal and differentiation into a variety of cell types. They represent an attractive source of cells for gene and cell therapy. The purpose of this study is to direct the specific expression of the DsRed reporter gene in Sca-1+ BMMSCs differentiated into a cardiomyogenic lineage. We constructed the prMLC-2v-DsRed vector expressing DsRed under the control of the 309 tp fragment of the rat MLC-2v 5'-flanking region. The specific expression of the DsRed reporter gene under the transcriptional control of the 309 bp fragment of the rat MLC-2v promoter was tested in 5-azacytidine healed-Sca-1+ BMMSCs over 2 weeks after the prMLC-2v-DsRed transfection. The prMLC-2v-DsRed was specifically expressed in the Sca-1+ BMMSCs with cardiomyogenic lineage differentiation and it demonstrates that the 309 bp sequences of the rat MLC-2v 5'-flanking region is sufficient to confer cardiac specific expression on a DsRed reporter gene. The cardiac-specific promoter-driven reporter vector provides an important tool for the study of stem cell differentiation and cell replacement therapy in ischemic cardiomyopathy.

Mesenchymal stem cells (MSCs) are considered to be attractive approaching in gene or drug delivery for cancer therapeutic strategies. In this study, the ability and feasibility of human bone marrow derived MSCs expressing the cytosine deaminase (CD)/5-Fluorocytosin (5-FC) prodrug was evaluated to target human osteosarcoma cell line Cal-72. At first, the fibroblast-like cells were successfully obtained from human bone marrow and demonstrated that they contained full of stem characteristics by the ability of differentiation into adipocyte/osteocyte and expression of typical mesenchymal markers CD90, CD44, while negative for CD34 and CD133 markers. We established the stable CD-expressing MSCs cell line (CD-MSCs) by transfection of pEGFP-C3 containing cytosine deaminase::uracil phos-phoribosyltransferase (CD::UPRT) gene into MSCs, and confirmed that the manipulated MSCs still remained full characteristics of multipotent cells and shown migration toward human osteosarcoma cancer cells Cal-72 as high as origin MSCs. Based on bystander effect, the therapeutic CD-MSCs significantly augmented the cytotoxicity on cancer cell Cal72 in either direct co-culture or conditioned medium in the presence of 5-FC. Moreover, in osteosarcoma cancer- bearing mice, the therapeutic CD/5-FC MSCs showed the inhibition of tumor growth compared with control mice which was s.c injected with only Cal72. Our findings suggest that these therapeutic CD-MSCs may be suitable and viable cellular vehicles for targeting human osteosarcoma cancer.