[Frontiers in Bioscience 16, 2747-2755 , June 1, 2011]

The effect of dexamethasone and hypoxic stress on MC3T3-E1 cells

Zhen-Hong Zhu1, You-Shui Gao1, Bing-Fang Zeng1, Chang-Qing Zhang1

1Department of Orthopedic Surgery, Shanghai Sixth People's Hospital, Shanghai Jiao Tong University, 600 Yishan Road, Shanghai 200233, China

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Materials and methods 3.1. Cell culture and materials
3.2. Apoptosis assay
3.3. Cytochrome c release
3.4. Western blot analysis
3.5. Measurement of mitochondrial transmembrane potential
3.6. Quantitative real-time PCR
3.7. Statistical analysis
4. Results
4.1. Effects of Dex and hypoxic stress on murine osteoblastic MC3T3-E1 cells
4.2. The mitochondria-mediated apoptotic pathway is induced by Dex and hypoxic stress in MC3T3-E1 cells
4.3. The mitochondria-mediated apoptotic pathway is essential for Dex-induced apoptosis under hypoxic conditions
4.4. The effects of different apoptotic inhibitors on MC3T3-E1 cells 5. Discussion
6. Acknowledgements
7. References

1. ABSTRACT

Osteonecrosis of the femoral head (ONFH) can be caused by a decrease in the activity or numbers of osteoblasts, a process in which apoptosis may play an essential role. We investigated the effect of dexamethasone (Dex) combined with hypoxic stress on murine osteoblastic MC3T3-E1 cells. Flow cytometry, western blot and real-time quantitative PCR analyses revealed that hypoxia significantly enhanced Dex-induced apoptosis. Further data demonstrated that both the death receptor and the mitochondria-mediated pathway were involved in Dex-induced apoptosis under hypoxic conditions. However, the death receptor pathway had only a minor effect on this process. The expression levels of Bcl-2 and Bax, which regulate the mitochondria-initiated apoptotic cascade signaling pathway, were significantly different in response to Dex and hypoxia. The mitochondrial membrane potential collapsed, and the inhibitor brain- derived neurotrophic factor (BDNF) conferred effective protection against apoptosis. In summary, the mitochondria-mediated apoptotic pathway functions in osteoblast apoptosis that is induced by Dex in a hypoxic environment, and the present study may help us to gain further insight into the molecular mechanisms of steroid-induced ONFH.