苦杏仁苷对镉暴露大鼠骨损伤的防治作用及对miR-302b-NF-κB-mTOR信号通路的影响

Prevention and treatment of amygdalin on bone injury in cadmium-exposed rats and its effect onmiR-302b-NF-κB-mTOR signaling pathway

目的 苦杏仁苷对镉中毒大鼠骨损伤的防治作用及对微小RNA-302b(microRNA-302b,miR-302b)-核因子κB(nuclear factor kappa-B,NF-κB)-雷帕霉素靶蛋白(rapamycin target protein,mTOR)信号通路的影响。方法 清洁级SD大鼠100只分成对照组、镉暴露模型组(30 mg/kg二氯化镉)、苦杏仁苷低、中、高剂量组(50.0、100.0和200.0 mg/kg)。镉暴露模型组、苦杏仁苷低、中、高剂量组大鼠每日腹腔注射二氯化镉溶液,持续12周,期间,苦杏仁苷低、中、高剂量组大鼠每日予以相应剂量的苦杏仁苷灌胃,试验结束后获取双侧股骨组织测量股骨远端的骨密度(bone density,BMD)、骨小梁数量、骨小梁厚度、骨小梁分散度,股骨骨钙素(osteocalcin,OC)、骨桥蛋白(osteopontin,OPN)、骨保护素(osteoprotegerin,OPG)、股骨miR-302b、NF-κB、mTOR水平。结果 与对照组比较,镉暴露模型组股骨BMD、骨小梁数量、骨小梁厚度、股骨OC、OPN、OPG、miR-302b水平明显降低(均P<0.05),骨小梁分散度、股骨NF-κB、mTOR mRNA蛋白水平明显升高(均P<0.05)。与镉暴露模型组比较,苦杏仁苷各剂量组股骨BMD[(1.21±0.19)vs(1.38±0.23)(1.55±0.26)(1.61±0.27)g/cm^(3)]、骨小梁数量[(2.36±0.39) vs(3.98±0.67)(4.69±0.78)(5.87±0.94)个/mm^(2)]、骨小梁厚度[(0.04±0.01) vs(0.07±0.01)(0.09±0.02)(0.12±0.02)mm]、股骨OC[(328.91±105.85)vs(585.25±184.20)(895.25±298.52)(1 250.21±358.52)moL/mg]、OPN[(589.52±185.54)vs(854.29±258.22)(1 358.96±420.59)(1 852.20±595.20)moL/mg]、OPG [(485.63±121.65)vs (741.25±222.69)(1 259.95±489.52)(1752.63±652.85)moL/mg]、miR-302b(0.69±0.11 vs 0.95±0.16、1.29±0.21、1.65±0.26)水平明显升高(均P<0.05)。骨小梁分散度[(0.68±0.14)vs(0.52±0.11)(0.48±0.09)(0.31±0.07)mm]、股骨NF-κB(3.56±0.73 vs 1.37±0.21、2.90±0.41、2.02±0.23、0.95±0.48 vs 0.87±0.14、0.72±0.25、0.43±0.12)、mTOR mRNA蛋白(4.37±0.76 vs 3.52±0.25、2.57±0.65、1.55±0.21、1.02±0.36 vs 0.82±0.47、0.68±0.54、0.38±0.35)水平明显降低(均P<0.05)。伴随着苦杏仁苷给药剂量的增加,苦杏仁苷各剂量组BMD、骨小梁数量、骨小梁厚度、股骨OC、OPN、OPG、miR-302b水平逐渐增加,骨小梁分散度、股骨NF-κB、mTOR mRNA蛋白水平逐渐降低,均有明显剂量反应关系(均P<0.05)。结论 苦杏仁苷能明显改善骨小梁结构数量,增加骨密度,对Cd暴露骨损伤有明显治疗作用,其机制与苦杏仁苷可激活miR-302b进而抑制NF-κB、mTOR表达有关。

Objective To investigate the preventive effect of amygdalin on bone damage in cadmium poisoning rats and its effect on microRNA-302b(miR-302b)-nuclear factor kappa-B(NF-κB)-rapamycin target protein(mTOR)signaling pathway.Methods Totally 100 clean-grade SD rats were divided into the control group,the cadmium exposure model group(30 mg/kg cadmium dichloride),amygdalin low,medium and high dose groups(50.0,100.0,200.0 mg/kg).The rats in the cadmium exposure model group,and the low-,middle-and high-dose amygdalin groups were given daily intraperitoneal injection of cadmium dichloride solution for 12 weeks.During this period,the rats in the low-,medium-and high-dose amygdalin groups were given the corresponding doses of amygdalin by gavage every day.The bilateral femoral tissues were obtained after the experiment,to measure the bone density(BMD),trabecular number,trabecular thickness,trabecular dispersion,and femoral osteocalcin(OC)of the distal femur,as well as the levels of osteopontin(OPN),osteoprotegerin(OPG),femoral miR-302b,NF-κB,mTOR.Results Compared with the control group,the femoral BMD,trabecular number,trabecular thickness,femoral OC,OPN,OPG,and miR-302b levels in the cadmium exposure model group were significantly decreased(all P<0.05),and the trabecular bone dispersion,femoral NF-κB,mTOR mRNA protein levels were significantly increased(all P<0.05).Compared with the cadmium exposure model group,the femoral BMD[(1.21±0.19)vs(1.38±0.23)(1.55±0.26)(1.61±0.27)g/cm^(3)],trabecular number[(2.36±0.39)vs(3.98±0.67)(4.69±0.78)(5.87±0.94)/mm^(2)],trabecular thickness[(0.04±0.01)vs(0.07±0.01)(0.09±0.02)(0.12±0.02)mm],femoral OC[(328.91±105.85)vs(585.25±184.20)(895.25±298.52)(1250.21±358.52)moL/mg)],OPN[(589.52±185.54)vs(854.29±258.22)(1358.96±420.59)(1852.20±595.20)moL/mg],OPG[(485.63±121.65)vs(741.25±222.69)(1259.95±489.52)(1752.63±652.85)moL/mg],miR-302b(0.69±0.11 vs 0.95±0.16,1.29±0.21,1.65±0.26)were significantly increased(all P<0.05).And the trabecular bone dispersion[(0.68±0.14)vs(0.52±0.11)(0.48±0.09)(0.31±0.07)mm],femoral NF-κB(3.56±0.73 vs 1.37±0.21,2.90±0.41,2.02±0.23,0.95±0.48 vs 0.87±0.14,0.72±0.25,0.43±0.12),mTOR mRNA protein levels(4.37±0.76 vs 3.52±0.25,2.57±0.65,1.55±0.21,1.02±0.36 vs 0.82±0.47,0.68±0.54,0.38±0.35)were significantly decreased(all P<0.05).With an increase in the dosage of amygdalin administration,the BMD,trabecular number,trabecular thickness,femoral OC,OPN,OPG,and miR-302b levels in the amygdalin groups gradually increased,while the trabecular bone dispersion,femoral NF-κB,and mTOR mRNA protein levels gradually decreased,with a significant dose response relationship(all P<0.05).Conclusion The amygdalin can significantly improve the number of trabecular bone structures,increase bone mineral density,and have a significant therapeutic effect on cadmium-exposed bone injury,its mechanism is related to the activation of miR-302b by amygdalin and the inhibition of NF-κB and mTOR expression.