Abstract

Mesenchymal stem cells (MSCs) can differentiate into multiple cell lineages, including osteoblasts and adipocytes. We reported previously that glucocorticoid-induced leucine zipper (GILZ) inhibits peroxisome proliferator-activated receptor gamma-2 (Ppargamma2) expression and blocks adipocyte differentiation. Here we show that overexpression of GILZ in mouse MSCs, but not MC3T3-E1 osteoblasts, increases alkaline phosphatase activity and enhances mineralized bone nodule formation, whereas knockdown of Gilz reduces MSC osteogenic differentiation capacity. Consistent with these observations, real-time reverse transcription-PCR analysis showed that both basal and differentiation-induced transcripts of the lineage commitment gene Runx2/Cbfa1, as well as osteoblast differentiation marker genes including alkaline phosphatase, type I collagen, and osteocalcin, were all increased in GILZ-expressing cells. In contrast, the mRNA levels of adipogenic Ppargamma2 and C/ebpalpha were significantly reduced in GILZ-expressing cells under both osteogenic and adipogenic conditions. Together, our results demonstrate that GILZ functions as a modulator of MSCs and that overexpression of GILZ shifts the balance between osteogenic and adipogenic differentiation of MSCs toward the osteogenic pathway. These data suggest that GILZ may have therapeutic value for stem cell-based therapies of metabolic bone diseases, such as fracture repair.

Highlights

  • Bone marrow-derived mesenchymal stem cells (MSCs)3 are pluripotent and can give rise to several distinct cell lineages, such as osteoblasts, adipocytes, chondrocytes, myocytes, and even neurons under appropriate conditions (1– 4)

  • Our results demonstrate that GC-induced leucine zipper (GILZ) functions as a modulator of MSCs and that overexpression of GILZ shifts the balance between osteogenic and adipogenic differentiation of MSCs toward the osteogenic pathway

  • In this report we address the question whether GILZ, while inhibiting adipocyte differentiation, can stimulate osteoblast differentiation of MSCs using GILZ gain-of-function and loss-of-function studies

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Summary

EXPERIMENTAL PROCEDURES

Isolation and Characterization of Mouse Bone Marrow MSCs— The bone marrow MSCs used in this study were isolated from 18-month-old male C57BL/6 mice (NIA, National Institutes of Health). All animal procedures were approved by the Institutional Committee for Animal Care and Use Committee (IACUC) at the Medical College of Georgia. The MSCs were isolated from long bones of 6 mice as a mixture population using a protocol modified from Gimble et al (17), Peister et al (18), and Tropel et al (19). These MSCs, which are negative for CD-11b, CD-11c, JOURNAL OF BIOLOGICAL CHEMISTRY 4723

GILZ Enhances MSC Osteogenic Differentiation
RESULTS
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Findings
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