LacZ and Interleukin-3 ExpressionIn Vivoafter Retroviral Transduction of Marrow-Derived Human Osteogenic Mesenchymal Progenitors

Abstract

Human marrow-derived mesenchymal progenitor cells (hMPCs), which have the capacity for osteogenic and marrow stromal differentiation, were transduced with the myeloproliferative sarcoma virus (MPSV)-based retrovirus, vM5LacZ, that contains the LacZ and neo genes. Stable transduction and gene expression occurred in 18% of cells. After culture expansion and selection in G418, approximately 70% of neo(r) hMPCs co-expressed LacZ. G418-selected hMPC retain their osteogenic potential and form bone in vivo when seeded into porous calcium phosphate ceramic cubes implanted subcutaneously into SCID mice. LacZ expression was evident within osteoblasts and osteocytes in bone developing within the ceramics 6 and 9 weeks after implantation. Likewise, hMPCs transduced with human interleukin-3 (hIL-3) cDNA, adhered to ceramic cubes and implanted into SCID mice, formed bone and secreted detectable levels of hIL-3 into the systemic circulation for at least 12 weeks. These data indicate that genetically transduced, culture-expanded bone marrow-derived hMPCs retain a precursor phenotype and maintain similar levels of transgene expression during osteogenic lineage commitment and differentiation in vivo. Because MPCs have been shown to differentiate into bone, cartilage, and tendon, these cells may be a useful target for gene therapy.

Engineering mesenchymal stem cells for immunotherapy

“…In an experimental SCID/hu model, human MSCs engineered to produce IL-3 were seeded into osteo-inductive ceramic cubes that were then subcutaneously implanted into mice. 32 In this study, MSC differentiation occurred inside the cubes and resulted in bone formation and IL-3 secretion into the systemic circulation for at least 12 weeks. MSCs are easily transduced by retroviral vectors, with a gene transfer efficacy between 50 and 85%.…”

“…This tolerance induction may be useful in allogeneic transplantations, where low incidence of GVHD was observed when the hematopoietic graft was coinjected with MSCs. 32 The tolerigenic effect of MSC delivery was equivalent whatever was the route of cell injection, intravenously or loaded onto an osteo-inductive matrix.…”

“…Although the origin of these stem cells in the joint is unknown, their presence in the normal synovium has also been demonstrated in humans and in rabbits, where TGFb stimulation was shown to induce cartilage tissue formation. 32,35 Engineering MSCs MSCs can be used in different therapeutic strategies either as immunosuppressive agents or genetically engineered to express molecules acting against the autoimmune process. As MSCs are not rejected and traffic to injuried mesenchymal tissues, in particular bone, they represent the opportunity to deliver therapeutic proteins.…”

Antitumor effect of genetically engineered mesenchymal stem cells in a rat glioma model

“…Since MSCs possess high affinity for glioma and exert organ-protective effects, utilizing MSCs as a vehicle for gene therapy in the treatment of brain cancer represents a rational clinical approach. However, the efficiency of exogenous gene transfer is relatively low using physiochemical methods, 42 and even employing viral vectors, such as retroviral 43,44 or adenoviral vectors, 15,42 the method is not efficient. Refractoriness to viral vector infection may be mainly due to a deficit of native viral receptors.…”

Enhanced engraftment of EPO-transduced human bone marrow stromal cells transplanted in a 3D matrix in non-conditioned NOD/SCID mice

“…4,8,9 However, in most, if not all cases, the engraftment was detected by PCR, and an evaluation of the percentage of survival was never provided. Furthermore, results obtained by systemic infusion of mouse BMSCs may be misleading, because it is a common experience that these cells are contaminated by a large number of macrophages that are easily transplanted in this way.…”