Transplantation of Osteogenically Differentiated Mouse iPS Cells for Bone Repair

Abstract

Induced pluripotent stem (iPS) cells are a type of undifferentiated cell that can be obtained from differentiated cells and have the pluripotent potential to differentiate into the musculoskeletal system, the myocardium, vascular endothelial cells, neurons, and hepatocytes. We therefore cultured mouse iPS cells in a DMEM containing 15% FBS, 10 −7 M dexamethasone, 10 mM β-glycerophosphate, and 50 µg/ml ascorbic acid for 3 weeks, in order to induce bone differentiation, and studied the expression of the bone differentiation markers Runx2 and osteocalcin using RT-PCR in a time-dependent manner. Osteocalcin, a bone differentiation marker in bone formation, exhibited the highest expression in the third week. In addition, the deposition of calcium nodules was observed using Alizarin red S staining. iPS cells cultured for bone differentiation were transplanted into severe combined immunodeficiency (SCID) mice, and the osteogenic potential exhibited after 4 weeks was studied. When bone differentiation-induced iPS cells were transplanted into SCID mice, bone formation was confirmed in soft X-ray images and tissue specimens. However, teratoma formation was confirmed in 20% of the transplanted models. When mouse iPS cells were treated with irradiation of 2 Gray (Gy) prior to transplantation, teratoma formation was inhibited. When mouse iPS cells treated in a likewise manner were xenotransplanted into rats, bone formation was confirmed but teratoma formation was not observed. It is believed that irradiation before transplantation is an effective way to inhibit teratoma formation.

Notch signaling partly regulates the osteogenic differentiation of retinoic acid-treated murine induced pluripotent stem cells

“…Although successful bone healing/ regeneration occurred when using these cells, teratoma formation was observed in 20% of the mice with transplanted osteogenically-induced iPSCs (30). These results suggest that differentiation of cells may be incomplete and that undifferentiated cells persist after osteogenic induction.…”

Bone tissue engineering via human induced pluripotent, umbilical cord and bone marrow mesenchymal stem cells in rat cranium

“…However, when transplanted back into patients for specific usage, the hiPSCs need to be induced into high-quality progenitor cells like MSCs, or fully-differentiated homogenous mature cells. Mixed undifferentiated cells are undesirable as they may undergo spontaneous differentiation and give rise to teratomas in vivo [55]. There are various protocols to derive MSCs or osteogenic cells from hiPSCs.…”

Comparative Analysis of Mouse-Induced Pluripotent Stem Cells and Mesenchymal Stem Cells During Osteogenic Differentiation In Vitro

“…To date, the differentiation mechanisms of MSCs have been well studied, and methods that guide MSC differentiation into mature osteoblasts using osteogenic induction factors, such as dexamethasone, ascorbic acid, and b-glycerophosphate, have been well established [9]. Using these classical osteogenic induction factors, several studies have examined the in vitro-directed mineralization capacity [10][11][12][13][14] and in vivo bone formation potential [15,16] of mouse iPSCs. In these studies, the in vitro calcification potential of iPSCs was evaluated using a combination of osteogenic molecular markers, gene expression, and histological staining [10][11][12][13][14][15][16], but no study to date has assessed the structural properties or elemental composition of the mineralized extracellular matrix (ECM).…”

“…Using these classical osteogenic induction factors, several studies have examined the in vitro-directed mineralization capacity [10][11][12][13][14] and in vivo bone formation potential [15,16] of mouse iPSCs. In these studies, the in vitro calcification potential of iPSCs was evaluated using a combination of osteogenic molecular markers, gene expression, and histological staining [10][11][12][13][14][15][16], but no study to date has assessed the structural properties or elemental composition of the mineralized extracellular matrix (ECM). It is often difficult to determine whether osteogenically induced cells, indeed, produce bonelike calcium phosphate mineral similar to native osteoblasts, especially when using osteogenic marker gene expression and histological staining techniques, such as the von Kossa and Alizarin Red methods [17].…”