Molecular and Cellular Biochemistry

, Volume 342, Issue 1–2, pp 191–199 | Cite as

Osteogenic role of endosomal chloride channels in MC3T3-E1 cells

Article
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Abstract

ClC-3, ClC-4, and ClC-5 belong to the voltage gated chloride channels (ClCs) and facilitate the endosomal acidification. The mutations of these endosomal chloride channel genes cause different genetic diseases with various bone disorders. We hypothesized that these endosomal ClCs might be involved in the bone development or osteoblast differentiation. Here we used MC3T3-E1 osteoprogenitor cell line and primarily cultured mouse osteoblasts and detected the expression of Clcn3, Clcn4, and Clcn5 in these cells. We analyzed the relationships between three endosomal ClCs and the osteogenic phenotype using osteoinductive treatment, overexpressing of ClCs and RNAi of ClCs. We found the increased mRNA levels of osteogenic markers [alkaline phosphatase (Alp), osteocalcin (Oc), bone sialoprotein (Bsp), and runt-related transcription factor 2 (Runx2)] were in parallel to that of Clcn3, Clcn4 and Clcn5 with osteoinductive treatment and overexpressed ClCs. Overexpressed ClCs were localized in intracellular periphery and also promoted the mineralization of cells in vitro. While RNAi mediated gene silencing of ClC-3, ClC-4, and ClC-5 down regulated the expression of the four osteogenic markers. The positive relationship between endosomal ClCs and the osteogenic markers suggested a new function of endosomal ClCs in osteogenic differentiation.

Keywords

Chloride channel Endosome Osteoblast Differentiation Osteogenic gene 

Abbreviations

ClC

Voltage gated chloride channel

CLCN3, Clcn3

Chloride channel 3 gene

CLCN4, Clcn4

Chloride channel 4 gene

CLCN5, Clcn5

Chloride channel 5 gene

TGF-β1

Transforming growth factor, beta 1

ALP, Alp

Alkaline phosphatase

BSP, Bsp

Bone sialoprotein

OC, Oc

Osteocalcin

RUNX2, Runx2

Runt-related transcription factor 2

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Notes

Acknowledgments

We would like to thank Dr. Sandra E. Guggino of Johns Hopkins University for providing ClCs expression vectors and Dr. Zhang Yan of the Department of Biochemistry and Molecular Biology of Fourth Military Medical University for providing MC3T3-E1 cells. We are also thankful for support from the Foundation of Key of Projects of Science and Technology of Xi’an City [SF09027(4)], the National Natural Science Foundation of China (NSFC) (30371540) and the Foundation of Key of Projects of Science and Technology of Shaanxi Province (2009K17-06). Thanks for the technique support from Equipment Center, PLA Institute of Stomatological Research.

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Copyright information

© Springer Science+Business Media, LLC. 2010

Authors and Affiliations

  • Huan Wang
    • 1
    • 2
  • Na Huo
    • 3
  • Feifei Li
    • 1
  • Shanmin Fu
    • 1
  • Yang Xue
    • 2
    • 4
  • Ting Yang
    • 2
  • Xuan Wen
    • 2
  • Yin Ding
    • 1
  • Xiaohong Duan
    • 2
  1. 1.Department of Orthodontics, School of StomatologyFourth Military Medical UniversityXi’anPeople’s Republic of China
  2. 2.Department of Oral Biology, School of StomatologyFourth Military Medical UniversityXi’anPeople’s Republic of China
  3. 3.Institute of StomatologyChinese General Hospital of PLABeijingPeople’s Republic of China
  4. 4.Department of Oral and Maxillofacial Surgery, School of StomatologyFourth Military Medical UniversityXi’anPeople’s Republic of China

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