著者

Kentaro ENDO Naoki FUJITA Takayuki NAKAGAWA Ryohei NISHIMURA

出版者

JAPANESE SOCIETY OF VETERINARY SCIENCE

雑誌

Journal of Veterinary Medical Science (ISSN:09167250)

巻号頁・発行日

pp.18-0551, (Released:2019-06-04)

被引用文献数

13

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Mesenchymal stem cells (MSCs) are proposed to be useful in cartilage regenerative medicine, however, canine MSCs have been reported to show poor chondrogenic capacity. Therefore, optimal conditions for chondrogenic differentiation should be determined by mimicking the developmental process. We have previously established novel and superior canine MSCs named bone marrow peri-adipocyte cells (BM-PACs) and the objective of this study was to evaluate the effects of growth factors required for in vivo chondrogenesis using canine BM-PACs. Spheroids of BM-PACs were cultured in chondrogenic medium containing 10 ng/ml transforming growth factor-β1 (TGF-β1) with or without 100 ng/ml bone morphogenetic protein-2 (BMP-2), 100 ng/ml growth differentiation factor-5 (GDF-5) or 100 ng/ml insulin-like growth factor-1 (IGF-1). Chondrogenic differentiation was evaluated by the quantification of glycosaminoglycan and Safranin O staining for proteoglycan production. The expression of cartilage matrix or hypertrophic gene/protein was also evaluated by qPCR and immunohistochemistry. Spheroids in all groups were strongly stained with Safranin O. Although BMP-2 significantly increased glycosaminoglycan production, Safranin O-negative outer layer was formed and the mRNA expression of COL10 relating to cartilage hypertrophy was also significantly upregulated (P<0.05). GDF-5 promoted the production of glycosaminoglycan and type II collagen without increasing COL10 mRNA expression. The supplementation of IGF-1 did not significantly affect cartilaginous and hypertrophic differentiation. Our results indicate that GDF-5 is a useful growth factor for the generation of articular cartilage from canine MSCs.

著者

Cheng-Shu CHUNG Naoki FUJITA Naoya KAWAHARA Sho YUI Eunryel NAM Ryohei NISHIMURA

出版者

公益社団法人 日本獣医学会

雑誌

Journal of Veterinary Medical Science (ISSN:09167250)

巻号頁・発行日

pp.12-0470, (Released:2013-02-19)

被引用文献数

12 39

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Stem cell transplantation is one of the most promising yet enigmatic treatments for spinal cord injury (SCI), a common problem in dogs. As pre-differentiated mesenchymal stem cells (MSCs) can be expanded and differentiated into neurospheres in vitro, before being transplanted back, they may prove to be more beneficial for treating SCI. Therefore, we compared the endogenous differentiation potential, including the neuronal cell differentiation, of neurospheres from canine bone marrow MSCs (cBMMSCs) with that of the adipose tissue-derived MSCs (cADMSCs). Nestin-positive neurospheres were generated from MSCs derived from the bone marrow and adipose tissue. Neuronal cells were differentiated from the neurospheres derived from both these tissues. Gene expression analysis revealed that Nestin, βIII-tubulin, NCAM, OCT4 and SOX2 were expressed in MSCs and the corresponding neurospheres. Notably, cBMMSC-derived neuronal cells expressed higher levels of βIII-tubulin. The mRNA expressions of NANOG, Nestin, OCT4 and SOX2 were upregulated in neurospheres derived from both. Immunofluorescence analysis detected the expression of neuronal markers, namely, βIII-tubulin, GFAP, S100, NF200 and MAP2, in differentiated neuron-like cells. Our findings highlight that both cBMMSCs and cADMSCs could be differentiated into neurospheres and neuron-like cells, and therefore, these cells are suitable candidates for cell transplantation. Further, cADMSCs form a more suitable cell source, as larger number of cells could be harvested from cADMSC-derived neurospheres. Future studies employing in vivo transplantation models to investigate the effectiveness of MSCs for treating SCI are warranted.