Enhanced mitochondrial biogenesis contributes to Wnt induced osteoblastic differentiation of C3H10T1/2 cells.
Mitochondria play a key role in cell physiology including cell differentiation and proliferation. We investigated the changes of mitochondrial biogenesis during Wnt-induced osteoblastic differentiation of murine mesenchymal C3H10T1/2 cells. Scanning electron microscopy demonstrated that activation of Wnt signaling by Wnt-3A conditioned medicum (CM) resulted in significant increase in the number of mitochondria in C3H10T1/2 cells. In addition, the induction of alkaline phosphatase (ALP) activities by Wnt-3A CM was accompanied by significant increase in mitochondrial mass (p<0.05), mitochondrial membrane potential (p<0.05), intracellular reactive oxygen species production (p<0.05), resting oxygen consumption rate (p<0.05), cellular ATP content (p< or =0.05) and mtDNA copy number (p<0.05) compared to the cells with control CM (L292-CM) treatment. Moreover, co-treatment with Dkk-1 or WIF-1, both of which are Wnt inhibitors, abrogated the Wnt-3A-induced ALP activities as well as mitochondrial biogenesis markers. Upregulation of mitochondrial biogenesis by overexpression of mitochondrial transcription factor A (Tfam) significantly enhanced Wnt-induced osteogenesis as measured by ALP activities. In contrast, inhibition of mitochondrial biogenesis by treatment with Zidovudine (AZT) resulted in significant inhibition of ALP activities. Finally, ALP activities in human osteosarcoma cell line devoid of mitochondrial DNA (rho(0) cells) was significantly suppressed both in basal and Wnt-3A stimulated state compared to those from mitochondria-intact cells (rho+ cells). As a mechanism for Wnt-mediated mitochondrial biogenesis, we found that Wnt increased the expression of PGC-1alpha, a critical molecules in mitochondrial biogenesis, through Erk and p38 MAPK pathway independent of beta-catenin signaling. We also found that increased mitochondrial biogenesis is in turn positively regulating TOPflash reporter activity as well as beta-catenin levels. To summarize, mitochodrial biogenesis is upregulated by Wnt signaling and this upregulation contributes to the osteoblastic differentiation of mouse mesenchymal C3H10T1/2 cells.
An
Jee Hyun
JH
Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Korea.
Yang
Jae-Yeon
JY
Ahn
Byung Yong
BY
Cho
Sun Wook
SW
Jung
Ju Yeon
JY
Cho
Hwa Young
HY
Cho
Young Min
YM
Kim
Sang Wan
SW
Park
Kyong Soo
KS
Kim
Seong Yeon
SY
Lee
Hong Kyu
HK
Shin
Chan Soo
CS
D050156:Adipogenesis / Q000187:drug effects; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D017077:Culture Media, Conditioned / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D051379:Mice; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology / Q000648:ultrastructure; D010012:Osteogenesis / Q000187:drug effects; D000071248:Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors; D014158:Transcription, Genetic / Q000187:drug effects; D051153:Wnt Proteins / Q000494:pharmacology*; D060508:Wnt3 Protein; D060509:Wnt3A Protein; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism
D017077:Culture Media, Conditioned; D000071248:Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; C485290:Ppargc1a protein, mouse; D015534:Trans-Activators; D014157:Transcription Factors; C555510:WNT3A protein, human; D051153:Wnt Proteins; D060508:Wnt3 Protein; D060509:Wnt3A Protein; C555511:Wnt3a protein, mouse; D051176:beta Catenin; D020928:Mitogen-Activated Protein Kinases