The Regulation Of Sclerostin On Biological Function Of Cementoblasts And The Potential Related Mechanism Under Mechanical Stress

During orthodontic treatment, absorption and restoration of dental root may appear, even under light force; and when the balance of absorption and restoration is broken, there often occur the negative effect Orthodontically Induced Inflammatory Root Resorption( OIIRR); however, effective methods for prevention and restoration have not yet been found.Cementoblasts are located on the surface of cementum. Sclerostin is a natural inhibitor of wnt/β-catenin signaling pathway, which can inhibit the bone formation mediated by osteoblast. Research believes that BMP2 in osteoblasts inhibit Wnt signals and reduce the formation of bone mass by up-regulating the expressions of sclerostin(SOST) and DKK1. But sclerostin is also regulated by mechanical stimulation. Sclerostin is highly expressed in mature osteoblasts, and mechanical load can reduce the protein and gene expressions of sclerostin. This experiment aims to study the regulation of sclerositin on biological function of cementoblasts and the potential related mechanism under mechanical stress, and this experiment was divided into three parts:1. Time-based protein expressions of sclerostin in OCCM-30 under mechanical compressive stressObjective: To explore the time-based protein expressions of osclerostin in OCCM-30 under mechanical compressive stress.Methods: OCCM-30 cells were loaded with uniaxial compressive stress(size of 2000 μstrain and frequency of 0.5Hz); loading duration was 3h, 6h,12 h and 24 h, and loaded group and non-loaded group were set at each time-point. WB was used to detect the expression of sclerostin, and RT-PCR was used to detect SOST m RNA level.Results: Osteosclerosis was detected in each group. And compared with non-loaded group, the protein expressions of osteosclerosis was up-regulated in 3h loaded group and 6h loaded group. And there were no significant differences between 12 h and 24 h groups.Conclusion: There is endogenous osteosclerosis in cementoblasts, and mechanical compression stress can enhance the protein expressions of osteosclerosis.2. Regulation of sclerostin on wnt/β-catenin signaling pathway and BMP/samd pathway under mechanical compressive stress.Objective: To study the regulation of sclerostin on cementification-related wnt/β-catenin signaling pathway and BMP/smad pathway under mechanical compressive stress.Methods: Cells were cultivated with medium containing different concentrations of sclerostin(0ng/ml, 25ng/ml, 50ng/ml, and 100ng/ml), and loaded with uniaxial compressive stress(size of 2000 μstrain and frequency of 0.5Hz) for 6h. With cells without compressive stress and sclerostin as the control group, WB was used to detect β-catenin, smad1/5/8, and p-smad1/5/8.Results: The expression of p-smad1/5/8 in the loaded group was higher than that of the non-loaded group when concentration of sclerostin was0ng/ml, and the expression of p-smad1/5/8 was significantlydown-regulated as the increase of concentration of sclerostin. There were no obvious differences between β-catenin and smad1/5/8.Conclusion: Under the mechanical comprehensive stress, sclerostin can both regulate the expression of the BMP signaling pathway.3. The influences of sclerostin on OCCM-30-related factors under mechanical compressive stressObjective: To study the influences of sclerostin on OCCM-30-related factors under mechanical compressive stress.Methods: Cells were cultivated with medium containing different concentrations of sclerostin(0ng/ml, 25ng/ml, 50ng/ml, and 100ng/ml), and loaded with uniaxial compressive stress(size of 2000 μstrain and frequency of 0.5Hz) for 6h. With cells without compressive stress and sclerostin as the control group, WB was used to detect the expressions of Runx-2, OCN, BSP,RANKL and OPG. And activity of ALP was also detected.Results: Compared with non-loaded group, expression of Runx-2 was down-regulated after loading; expressions of Runx-2 was also down-regulated with the increase of sclerostin concentration under loading condition, and expressions of BSP and OCN were similar as that of Runx-2.Expressions of RANKL and OPG: compared with non-loaded group,expression of RANKL was up-regulated while expression of OPG was down-regulated in loaded group; and with the increase of sclerostin concentration, expression of RANKL was gradually increased while expression of OPG was down-regulated. ALP activity: The activity of ALP was increased under mechanical compressive stress; and with the increase of concentration of sclerostin, ALP activity showed a decreasing trend.Conclusion: Osteosclerosis inhibits the mineralization of cementoblasts under mechanical compressive stress, that is to say, this effect is achieved by inhibiting the expressions of osteogenesis factors(Runx2, OCN and BSP,etc.) and promoting the ratio of cementoclast related factors RANKL/OPG through BMP signaling pathway...