Abstract
Introduction: Evidence exists concerning the anti-oxidant properties of oestrogen in protecting neuronal cells from oxidative stress. The withdrawal of oestrogen after menopause is the major factor determining age related bone loss and apoptotic death of osteocytes. While oestrogen replacement demonstrates clear oestrogen receptor mediated benefits to bone cells little is known regarding oestrogens’ anti-oxidant effects in bone.
Methods: Here we have used MLO-Y4 osteocyte-like cell line to determine whether oestrogen saving effects on osteocytes involves its activities as an anti-oxidant.
MLO-Y4 cells were treated with physiological doses (10−8)M of either 17-beta E2 or the oestrogen receptor inactive stereoisomer 17-alpha E2 with or without the specific oestrogen receptor antagonist ICI 182,780 prior to the addition of 0.4milliM 30% (v/v) H2O2. Cellular apoptosis was determined using morphological and biochemical criteria.
Results: H2O2 induced an increase in apoptosis of MLO-Y4 (14.3 ± 3 SD vs control 1.4 ± 0.9). Pre-treatment of the cells with 17-beta E2 significantly reduced H2O2 induced apoptosis (2.4 ± 0.96). Pre-treatment of cells with 17-alpha E2 or ICI 182,780 also reduced oxidant induced apoptosis to 3.4 ± 1.5 SD and 7.0 ± 2.3 respectively.
The cellular production of reactive oxygen species was determined using the free radical indicator 2′7′- dichlorodihydrofluorescein diacetate. H2O2 induced increases in the number of ROS positive cells (34.6 ± 9.07 SD vs control 0.22 ± 0.39 SD). In contrast pre-treatment with both 17-beta E2 and 17-alpha E2 reduced the number of ROS positive cells associated with H2O2 treatment (Fig 1).
Conclusion: These data suggest that oestrogens ability to save osteocytes from oxidant induced death is independent of the oestrogen receptor and may be related to oestrogens known activity as an anti-oxidant. This raises the possibility that loss of osteocytes during oestrogen insufficiency may occur through a failure to suppress the activity of naturally occurring or disease associated production of oxidant molecules.
Correspondence should be addressed to Mr Carlos Wigderowitz, Honorary Secretary BORS, University Dept of Orthopaedic & Trauma Surgery, Ninewells Hospital & Medical School, Dundee DD1 9SY.
