Ovariectomy-induced bone loss in TNFα and IL6 gene knockout mice is regulated by different mechanisms

Abstract

We examined the effects of tumor necrosis factor-α (TNFα) and interleukin-6 (IL6) gene knockout in preserving the bone loss induced by ovariectomy (OVX) and the mechanisms involved in bone metabolism. Twenty female wild-type (WT), <i>TNFα</i>-knockout (TNFα^-/-) or <i>IL6-</i>knockout (IL6^-/-) mice aged 12 weeks were sham-operated (SHAM) or subjected to OVX and killed after 4 weeks. Bone mass and skeletal microarchitecture were determined using micro-CT. Bone marrow stromal cells (BMSCs) from all three groups (WT, TNFα^-/- and IL6^-/-) were induced to differentiate into osteoblasts or osteoclasts and treated with 17-β-estradiol. Bone metabolism was assessed by histological analysis, serum analyses and qRT-PCR. OVX successfully induced a high turnover in all mice, but a repair effect was observed in TNFα^-/- and IL6^-/- mice. The ratio of femoral trabecular bone volume to tissue volume, trabecular number and trabecular thickness were significantly decreased in WT mice subjected to OVX, but increased in TNFα^-/- mice (1.62, 1.34, 0.27-fold respectively; <i>P</i> < 0.01) and IL6^-/- mice (1.34, 0.80, 0.22-fold respectively; <i>P</i> < 0.01). Furthermore, we observed a 29.6% increase in the trabecular number in TNFα^-/- mice when compared to the IL6^-/- mice. Both, TNFα^-/- and IL6^-/- BMSCs exhibited decreased numbers of TRAP-positive cells and an increase in ALP-positive cells, with or without E2 treatment (<i>P</i> < 0.05). While the knockout of <i>TNFα</i> or <i>IL6</i> significantly upregulated mRNA expressions of osteoblast-related genes (<i>Runx2</i> and <i>Col1a1</i>) and downregulated osteoclast-related mRNA for <i>TRAP</i>, <i>MMP9</i> and <i>CTSK in vivo</i> and <i>in vitro</i>, <i>TNFα</i> knockout appeared to have roles beyond <i>IL6</i> knockout in upregulating <i>Col1a1</i> mRNA expression and downregulating mRNA expressions of WNT-related genes (<i>DKK1</i> and <i>Sost</i>) and TNF-related activation-induced genes (<i>TRAF6</i>). TNFα seemed to be more potentially invasive in inhibiting bone formation and enhancing TRAF6-mediated osteoclastogenesis than IL6, implying that the regulatory mechanisms of TNFα and IL6 in bone metabolism may be different.