Shikonin Ameliorates Glucocorticoid Induced Osteoporosis by Regulating RANKL/RANK/TRAF6 and Its Mediated NF-κB/MAPKs Signaling Pathway and Oxidative Stress
10.11842/wst.20221126001
- VernacularTitle:紫草素通过调节RANKL/RANK/TRAF6及其介导的NF-κB/MAPKs信号通路与氧化应激来改善激素性骨质疏松
- Author:
Long SANG
1
;
Kedi WU
;
Jiazheng JIANG
Author Information
1. 海南西部中心医院 儋州 571700
- Keywords:
Shikonin;
Osteoporosis;
NF-κB/MAPKs;
RANKL/RANK/TRAF6
- From:
World Science and Technology-Modernization of Traditional Chinese Medicine
2023;25(11):3758-3768
- CountryChina
- Language:Chinese
-
Abstract:
Objective To evaluate the effect and potential mechanism of Shikonin on hormone-induced osteoporosis in rats.Methods RAW264.7 cell lines were treated with indicated concentrations of Shikonin,and the effects of Shikonin on cell viability were evaluated by CCK8 assay.Then the cells were divided into five groups:control group,dexamethasone(osteoporosis agent)group,dexamethasone + recombinant human parathyroid(anti-osteoporosis agent)group,dexamethasone + Shikonin 0.3 μmol·L-1 group,and dexamethasone + Shikonin 1.2 μmol·L-1 group.The effect of Shikonin on osteoclast function was evaluated by TRAP staining and bone resorption area measurement.The rat model of osteoporosis induced by dexamethasone was established,and divided into five groups(n=8):dexamethasone group,dexamethasone + PTH group,dexamethasone + Shikonin 0.3 μmol·L-1 group and dexamethasone + Shikonin 1.2 μmol·L-1 group,while control group was set up,then each group given separately 0.9%NaCl 6 mL·kg-1,PTH 20 μg·kg-1 3 days,Shikonin 0.5 mg·kg-1 day(0.3 μmol·L-1),Shikonin 2 mg·kg-1 day(1.2 μmol·L-1),and 0.9%NaCl 6 mL·kg-1for 3 weeks.ELISA was used to detect Serum type I collagen(CTX),Cathepsin K(Cathepsin K)and oxidative stress related indicators.HE staining was used to detect the changes of bone structure,and dual-energy X-ray scanning was used to detect the changes of bone density(BMD).Western blot and immunofluorescence were used to detect the effect of Shikonin on NF-κB/MAPKs signaling pathway,Western blot and qRT-PCR were used to detect the expression of RANKL-related proteins and mRNA,and Western blot was used to detect the blocking effect of Shikonin on RANKL/TRAF6.Results 1.2 μmol·L-1 Shikonin had the least damage to cells.Shikonin decreased the number of TRAP positive cells(P<0.05),decreased bone absorption area(P<0.05).and decreased the expression of CTX and Cathepsin K(P<0.05),and in 1.2 μmol·L-1 group decreased more significantly(P<0.05).Shikonin increased rat femur BMD(P<0.05),and increased more significantly in 1.2 μmol·L-1 group(P<0.01).Shikonin reversed the reduction and thinning of bone trabeculae induced by dexamethasone.The expression of SOD and GSH in rats increased(P<0.05),while the expression of MDA decreased(P<0.05).Shikonin inhibited the phosphorylation of NF-κB pathway related proteins in vitro.Shikonin inhibited the expression of RANK,RANKL,TRAF6,c-fos and NFATc1 protein and mRNA.Moreover,Shikonin blocked the expression of TRAF6.Conclusion Shikonin can ameliorate glucocorticoid induced osteoporosis by inhibiting RANKL/RANK/TRAF6 and its mediated NF-κB/MAPKs signaling pathway and oxidative stress.
