Osteocytes up-regulate the terminal differentiation of pre-osteoblasts via gap junctions

• Published in: Biochemical and biophysical research communications 2015-01, Vol.456 (1), p.1-6
• Main Authors: Nishikawa, Yoichi, Akiyama, Yuko, Yamamoto, Kiyofumi, Kobayashi, Masayuki, Watanabe, Eri, Watanabe, Nobukazu, Shimizu, Noriyoshi, Mikami, Yoshikazu, Komiyama, Kazuo
• Format: Article
• Language: English
• Subjects: 3T3 Cells; Alkaline phosphatase; Alkaline Phosphatase - metabolism; Animals; Bone sialoprotein; Carbenoxolone - chemistry; Carrier Proteins - metabolism; Cell Communication; Cell Cycle; Cell Differentiation; Coculture Techniques; Gap junction; Gap Junctions - physiology; Gene Expression Regulation; Green Fluorescent Proteins - metabolism; Humans; Integrin-Binding Sialoprotein - metabolism; Mice; Osteoblasts; Osteoblasts - cytology; Osteocytes; Osteocytes - cytology; Osteocytes - metabolism; Patch-Clamp Techniques; RNA, Messenger - metabolism
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2014.10.128

•We examined cell-to-cell interaction between pre-osteoblasts and osteocytes.•ALP and BSP expression in pre-osteoblasts co-cultured with osteocytes was increased.•Gap junctions were detected between pre-osteoblasts and osteocytes.•The gap junction inhibitor suppressed the increased expression of ALP and BSP.•Osteocytes up-regulate the terminal differentiation of pre-osteoblasts via gap junctions. We examined cell-to-cell interaction between pre-osteoblasts and osteocytes using MC3T3-E1 and MLO-Y4, respectively. First, GFP expressing MC3T3-E1 (E1-GFP) cells were generated to isolate the cells from co-culture with MLO-Y4. No changes were observed in the expression of osteogenic transcription factors Runx2, Osterix, Dlx5 and Msx2, but expression of alkaline phosphatase (ALP) and bone sialoprotein (BSP) in E1-GFP co-cultured with MLO-Y4 was 300–400-fold greater than that in mono-cultured E1-GFP. In addition, mineralized nodule formation was drastically increased in co-cultured E1-GFP cells compared to mono-cultured cells. Patch clamp assay showed the presence of gap junctions between E1-GFP and MLO-Y4. Furthermore, when the gap junction inhibitor carbenoxolone (CBX) was added to the culture, increased expression of ALP and BSP in E1-GFP co-cultured with MLO-Y4 was suppressed. These results suggest that gap junction detected between pre-osteoblasts and osteocytes plays an important role on the terminal differentiation of pre-osteoblasts.