Effects Of Benefit-bone-capsule On Wnt/β-catenin Signal Pathway Inbone Tissue Of Ovariectomized Rats

Objective:To explore the mechanism of Benefit-Bone-Capsule(BBC) on OVX-induced osteoporosis, we investigate the effect of BBC on Wnt/β-Catenin signaling which emerged as a novel key pathway for promoting bone formation. Methods:1. After acclimated feeding for 1 week, 84 female SD rats of 3-month old were randomly assigned into ovariectomized group(70 rats) and sham-operation group(14 rats).The ostoporotie model was established by ovariectomy.2. After 12 weeks of the model established, the bone density of these rats was detected by dual energy X-ray absorptionmetry to assure the successful rate of the OVX model.3. Ovariectomized group were body-weight-matched and randomly diveded into the following five groups: model control group(14 rats), L-BBC group(14 rats), M-BBC group(14 rats), H-BBC group(14 rats) and positive control group(14 rats). The rats in BBC group were respectively treated with gastric perfusion of BBC solution that amounted to 1, 3and 9 times of the dose adult patients using in clinical. Those in positive control group were treated with fosamax.4. After 12 weeks of the drug intervention, the BMD detection of lumbar spine and femoral was carried on.5. The expression of wnt3 a and wnt10 b was measured by immunohistochemical technique.6. FQ-PCR was used to detect β-Catenin, GSK-3β, and Runx2 m RNA expression of femur.7. The protein of β-Catenin was measured by Western-blotting. Results: 1. Compared with the Sham group, the BMD of the OVX rats was significantly reduced P<0.05).The BMD of OVX rats treated with BBC were significantly higher than thatwithout treatment(P<0.05). 2. Immunohistochemical technique vadilated that there is an improvement of the expression of nt3a and Wnt10 b after BBC intervention(P<0.05). 3. Results of FQ-PCR showed that β-Catenin, Runx2 m RNA expression decreased significantly n OVX group compared with the Sham group(P<0.05), while GSK-3β m RNA increased ignificantly(P<0.05). Compared with OVX group, β-Catenin, Runx2 m RNA expression ignificantly increased in the BBC-H group(P<0.05), while GSK-3β m RNA was ignificantly decrease(P<0.05). 4. Results of Western-blotting showed that β-Catenin protein was significantly decreased in VX group(P<0.05), while the same protein was significantly increased in BBC-H group fter the drug intervention(P< 0.05). Conclusion:Results showed that BBC can decreased bone mineral elements loss, improved biomechanics parameters in OVX rats.Wnt3a, Wnt10 b activation can promote the dissociation of β-catenin complexes, release of β-catenin monomer and inhibition of β-catenin monomer degradation. BBC decreased GSK-3β m RNA, and increased β-catenin and Runx2 proteins expression in femur of OVX rats.These data suggest that OVX suppresses the canonical Wnt signal in OVX rats, partially through the enhancement of the GSK-3β. BBC act as activator of the canonical Wnt signal by subsequently upregulating β-catenin gene.