A crucial role of caspase-3 in osteogenic differentiation of bone marrow stromal stem cells
Masako Miura, … , Marian Young, Songtao Shi
Masako Miura, … , Marian Young, Songtao Shi
Published December 15, 2004
Citation Information: J Clin Invest. 2004;114(12):1704-1713. https://doi.org/10.1172/JCI20427.
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Article Bone biology

A crucial role of caspase-3 in osteogenic differentiation of bone marrow stromal stem cells

Abstract

Caspase-3 is a critical enzyme for apoptosis and cell survival. Here we report delayed ossification and decreased bone mineral density in caspase-3–deficient (Casp3–/– and Casp3+/–) mice due to an attenuated osteogenic differentiation of bone marrow stromal stem cells (BMSSCs). The mechanism involved in the impaired differentiation of BMSSCs is due, at least partially, to the overactivated TGF-β/Smad2 signaling pathway and the upregulated expressions of p53 and p21 along with the downregulated expressions of Cdk2 and Cdc2, and ultimately increased replicative senescence. In addition, the overactivated TGF-β/Smad2 signaling may result in the compromised Runx2/Cbfa1 expression in preosteoblasts. Furthermore, we demonstrate that caspase-3 inhibitor, a potential agent for clinical treatment of human diseases, caused accelerated bone loss in ovariectomized mice, which is also associated with the overactivated TGF-β/Smad2 signaling in BMSSCs. This study demonstrates that caspase-3 is crucial for the differentiation of BMSSCs by influencing TGF-β/Smad2 pathway and cell cycle progression.

Authors

Masako Miura, Xiao-Dong Chen, Matthew R. Allen, Yanming Bi, Stan Gronthos, Byoung-Moo Seo, Saquib Lakhani, Richard A. Flavell, Xin-Hua Feng, Pamela Gehron Robey, Marian Young, Songtao Shi

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Proliferation and osteogenic differentiation of BMSSCs in caspase-3–defi...
Proliferation and osteogenic differentiation of BMSSCs in caspase-3–deficient mice. (A) Appearance of CFU-F derived from WT, Casp3+/–, and Casp3–/– mice (left). There was a significant difference in the number of colonies between caspase-3–deficient mice (Casp3–/– and Casp3+/–) and WT mice (right). Error bars represent the mean ± SD (n = 10; *P < 0.001). (B) BrdU incorporation of BMSSCs. The proliferation rate of cultured BMSSCs was assessed by BrdU incorporation assay for 24 hours. Representative pictures are shown at left. Original magnification, ×400. The number of BrdU-positive cells is indicated as a percentage of the total number of counted BMSSCs and averaged from 5 replicated cultures. Error bars represent mean ± SD (n = 5; *P < 0.001). (C) Alizarin red staining of BMSSCs cultured under the osteogenic inductive condition. Casp3–/– and Casp3+/– BMSSCs showed a lower calcium accumulation than WT BMSSCs. (D) Bone formation by BMSSCs in vivo. BMSSCs were transplanted into immunocompromised mice with HA/TCP (HA). Bone formation assessed by H&E staining was decreased in Casp3–/– and Casp3+/– transplants compared with WT transplants. B, bone; C, connective tissue; green arrows, hematopoeitic marrow elements. Original magnification, ×200. The BFR was calculated as the percentage of newly formed bone area per total area of transplant at the representative cross-sections. The graph represents mean ± SD of the percentage of WT (WT, n = 3; Casp3+/–, n = 5; Casp3–/–, n = 4; **P < 0.0001).