Rapid and efficient generation of cartilage pellets from mouse induced pluripotent stem cells by transcriptional activation of BMP-4 with shaking culture

Autor: Maolin Zhang, Kunimichi Niibe, Takeru Kondo, Phoonsuk Limraksasin, Hiroko Okawa, Xinchao Miao, Yuya Kamano, Masahiro Yamada, Xinquan Jiang, Hiroshi Egusa
Jazyk: angličtina
Rok vydání: 2022
Předmět:
Zdroj: Journal of Tissue Engineering, Vol 13 (2022)
Druh dokumentu: article
ISSN: 2041-7314
20417314
DOI: 10.1177/20417314221114616
Popis: Induced pluripotent stem cells (iPSCs) offer an unlimited source for cartilage regeneration as they can generate a wide spectrum of cell types. Here, we established a tetracycline (tet) controlled bone morphogenetic protein-4 ( BMP-4 ) expressing iPSC (iPSC- Tet/BMP-4 ) line in which transcriptional activation of BMP-4 was associated with enhanced chondrogenesis. Moreover, we developed an efficient and simple approach for directly guiding iPSC- Tet/BMP-4 differentiation into chondrocytes in scaffold-free cartilaginous pellets using a combination of transcriptional activation of BMP-4 and a 3D shaking suspension culture system. In chondrogenic induction medium, shaking culture alone significantly upregulated the chondrogenic markers Sox9, Col2a1 , and Aggrecan in iPSCs- Tet/BMP-4 by day 21. Of note, transcriptional activation of BMP-4 by addition of tet (doxycycline) greatly enhanced the expression of these genes. The cartilaginous pellets derived from iPSCs- Tet/BMP-4 showed an oval morphology and white smooth appearance by day 21. After day 21, the cells presented a typical round morphology and the extracellular matrix was stained intensively with Safranin O, alcian blue, and type II collagen. In addition, the homogenous cartilaginous pellets derived from iPSCs- Tet/BMP-4 with 28 days of induction repaired joint osteochondral defects in immunosuppressed rats and integrated well with the adjacent host cartilage. The regenerated cartilage expressed the neomycin resistance gene, indicating that the newly formed cartilage was generated by the transplanted iPSCs- Tet/BMP-4 . Thus, our culture system could be a useful tool for further investigation of the mechanism of BMP-4 in regulating iPSC differentiation toward the chondrogenic lineage, and should facilitate research in cartilage development, repair, and osteoarthritis.
Databáze: Directory of Open Access Journals
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