Significance: Osteoblasts produce bone matrix and contribute to bone remodeling. We have established a procedure to directly convert human fibroblasts into osteoblasts by transducing some defined factors and culturing in osteogenic medium. Osteoblast-specific transcription factors, Runx2 and Osterix, in combination with Oct4 and L-Myc, drastically induced fibroblasts to produce calcified bone matrix and express osteoblast-specific markers. The directly converted osteoblasts (dOBs) showed similar gene expression profiles as normal osteoblasts and contributed to bone repair after transplantation into mice with bone defects. Furthermore, dOBs did not require continuous expression of the exogenous genes to maintain their phenotype. These findings strongly suggest successful direct reprogramming of fibroblasts into osteoblasts, which may be applicable to bone regeneration therapy.

Brown adipocytes (BAs) play important roles in body temperature regulation, energy balance, and carbohydrate and lipid metabolism. Activities of BAs are remarkably diminished in obese and diabetic patients, providing possibilities of transplanting functional BAs resulting in therapeutic benefit. Here, we show generation of functional BAs by cellular reprogramming procedures. Transduction of the PRDM16 gene into iPSC-derived embryoid bodies induced BA phenotypes (iBAs). Moreover, normal human fibroblasts were directly converted into BAs (dBAs) by C/EBP-β and C-MYC gene transduction. Approximately 90% of the fibroblasts were successfully converted within 12 days. The dBAs were highly active in mitochondrial biogenesis and oxidative metabolism. Mouse dBAs were induced by Prdm16, C/ebp-β, and L-myc genes, and after transplantation, they significantly reduced diet-induced obesity and insulin resistance in an UCP1-dependent manner. Thus, highly functional BAs can be generated by cellular reprogramming, suggesting a promising tailor-made cell therapy against metabolic disorders including type 2 diabetes mellitus.

Schwann cells (SCs) play pivotal roles in the maintenance and regeneration of the peripheral nervous system. Although transplantation of SCs enhances repair of experimentally damaged peripheral and central nerve tissues, it is difficult to prepare a sufficient number of functional SCs for transplantation therapy without causing adverse events for the donor. Here, we generated functional SCs by somatic cell reprogramming procedures and demonstrated their capability to promote peripheral nerve regeneration. Normal human fibroblasts were phenotypically converted into SCs by transducing SOX10 and Krox20 genes followed by culturing for 10 days resulting in approximately 43% directly converted Schwann cells (dSCs). The dSCs expressed SC-specific proteins, secreted neurotrophic factors, and induced neuronal cells to extend neurites. The dSCs also displayed myelin-forming capability both in vitro and in vivo. Moreover, transplantation of the dSCs into the transected sciatic nerve in mice resulted in significantly accelerated regeneration of the nerve and in improved motor function at a level comparable to that with transplantation of the SCs obtained from a peripheral nerve. The dSCs induced by our procedure may be applicable for novel regeneration therapy for not only peripheral nerve injury but also for central nerve damage and for neurodegenerative disorders related to SC dysfunction. Stem Cells Translational Medicine 2017;6:1207–1216

Cell fusion‐mediated formation of multinuclear osteoclasts (OCs) plays a key role in bone resorption. It is reported that 2 unique OC‐specific fusogens [ i.e. , OC‐stimulatory transmembrane protein (OC‐STAMP) and dendritic cell–specific transmembrane protein (DC‐STAMP)], and permissive fusogen CD9, are involved in OC fusion. In contrast to DC‐STAMP‐knockout (KO) mice, which show the osteopetrotic phenotype, OC‐STAMP‐KO mice show no difference in systemic bone mineral density. Nonetheless, according to the ligature‐induced periodontitis model, significantly lower level of bone resorption was found in OC‐STAMP‐KO mice compared to WT mice. Anti‐OC‐STAMP‐neutralizing mAb down‐modulated in vitro: 1 ) the emergence of large multinuclear tartrate‐ resistant acid phosphatase‐positive cells, 2) pit formation, and 3) mRNA and protein expression of CD9, but not DC‐STAMP, in receptor activator of NF‐κB ligand (RANKL)‐stimulated OC precursor cells (OCps). While anti–DC‐STAMP‐mAb also down‐regulated RANKL‐induced osteoclastogenesis in vitro , it had no effect on CD9 expression. In our mouse model, systemic administration of anti–OC‐STAMP‐mAb suppressed the expression of CD9 mRNA, but not DC‐STAMP mRNA, in periodontal tissue, along with diminished alveolar bone loss and reduced emergence of CD9 ⁺ OCps and tartrate‐resistant acid phosphatase‐positive multinuclear OCs. The present study demonstrated that OC‐STAMP partners CD9 to promote periodontal bone destruction by up‐regulation of fusion during osteoclastogenesis, suggesting that anti–OC‐STAMP‐mAb may lead to the development of a novel therapeutic regimen for periodontitis.—Ishii, T., Ruiz‐Torruella, M., Ikeda, A., Shindo, S., Movila, A., Mawardi, H., Albassam, A., Kayal, R. A., Al‐Dharrab, A. A., Egashira, K., Wisitrasameewong, W., Yamamoto, K., Mira, A. I., Sueishi, K., Han, X., Taubman, M. A., Miyamoto, T., Kawai, T. OC‐STAMP promotes osteoclast fusion for pathogenic bone resorption in periodontitis via up‐regulation of permissive fusogen CD9. FASEB J. 32, 4016–4030 (2018). www.fasebj.org

Transplantation of engineered three-dimensional (3D) bone tissue may provide therapeutic benefits to patients with various bone diseases. To achieve this goal, appropriate 3D scaffolds and cells are required. In the present study, we devised a novel nanogel tectonic material for artificial 3D scaffold, namely the nanogel-cross-linked porous (NanoCliP)-freeze-dried (FD) gel, and estimated its potential as a 3D scaffold for bone tissue engineering. As the osteoblasts, directly converted osteoblasts (dOBs) were used, because a large number of highly functional osteoblasts could be induced from fibroblasts that can be collected from patients with a minimally invasive procedure. The NanoCliP-FD gel was highly porous, and fibronectin coating of the gel allowed efficient adhesion of the dOBs, so that the cells occupied the almost entire surface of the walls of the pores after culturing for 7 days. The dOBs massively produced calcified bone matrix, and the culture could be continued for at least 28 days. The NanoCliP-FD gel with dOBs remarkably promoted bone regeneration in vivo after having been grafted to bone defect lesions that were artificially created in mice. The present findings suggest that the combination of the NanoCliP-FD gel and dOBs may provide a feasible therapeutic modality for bone diseases.

A procedure to generate functional osteoblasts from human somatic cells may pave the way to a novel and effective transplantation therapy in bone disorders. Here, we report that human fibroblasts were induced to show osteoblast phenotypes by culturing with ALK5 i II, which is a specific inhibitor for activin-like kinase 5 (ALK5) (tumor growth factor-β receptor 1 (TGF-β R1)). Cells cultured with ALK5 i II expressed osteoblast-specific genes and massively produced calcified bone matrix, similar to the osteoblasts induced from mesenchymal stem cells (MSC-OBs). Treatment with vitamin D3 in addition to ALK5 i II induced more osteoblast-like characters, and the efficiency of the conversion reached approximately 90%. The chemical compound-mediated directly converted osteoblasts (cOBs) were similar to human primary osteoblasts in terms of expression profiles of osteoblast-related genes. The cOBs abundantly produced bone matrix in vivo and facilitated bone healing after they were transplanted into immunodeficient mice at an artificially induced defect lesion in femoral bone. The present procedure realizes a highly efficient direct conversion of human fibroblasts into transgene-free and highly functional osteoblasts, which might be applied in a novel strategy of bone regeneration therapy in bone diseases.