芝麻素通过抑制AKT/GSK3β/NFATc1信号通路减轻2型糖尿病大鼠骨质疏松症

Sesamin Alleviates Type 2 Diabetic Osteoporosis in Rats through Inhibiting AKT/GSK3β/NFATc1 Signaling Pathway

目的:探究芝麻素对2型糖尿病性骨质疏松症(T2DOP)的治疗作用及其可能机制。方法:将健康雄性SD大鼠分为对照组、模型组、低剂量芝麻素组(L-Ses组)和高剂量芝麻素组(H-Ses组),每组10只。模型组、L-Ses组和H-Ses组大鼠给予高脂高糖饲料喂饲4周,并一次性腹腔注射链脲佐菌素(STZ,剂量35 mg/kg)溶液建立2型糖尿病性骨质疏松症大鼠模型。造模成功后,L-Ses组和H-Ses组大鼠分别按照10 mg/kg和40 mg/kg的剂量给予芝麻素灌胃处理,每日1次,持续药物处理8周。对照组和模型组大鼠同时间段给予等量的生理盐水灌胃处理。分别检测大鼠血清中空腹血糖值(Fasting plasma glucose,FPG)、空腹胰岛素(Fasting insulin,FINS)、糖化血红蛋白(Glycosylated hemoglobin,HbA1c)、血清总胆固醇(Serum total cholesterol,TC)、甘油三酯(Triglyceride,TG)、低密度脂蛋白胆固醇(Low-density lipoprotein cholesterol,LDL-C)、高密度脂蛋白胆固醇(High-density lipoprotein cholesterol,HDL-C)、超氧化物歧化酶(Superoxide dismutase,SOD)、过氧化氢酶(Catalase,CAT)、谷胱甘肽(Glutathione,GSH)和丙二醛(Malondialdehyde,MDA)的水平。采用苏木素-伊红(Hematoxylin-eosin staining,HE)染色检测大鼠股骨组织形态。μCT扫描仪分析大鼠股骨干骺端三维结构以及骨密度(Bone mineral density,BMD)、骨小梁数(Trabecular number,Tb.N)和骨小梁厚度(Trabecular thickness,Tb.Th)参数,Western blot检测骨形成指标骨碱性磷酸酶(Bone alkaline phosphate,BALP)、骨钙素(Osteocalcin,OCN)、成骨相关基因2(Runt-related transcription factor 2,Runx2),以及磷脂酰肌醇-3激酶(Phosphoinositide 3 kinase,PI3K)/蛋白激酶B(Protein kinase B,Akt)/糖原合成酶激酶3β(Glycogen synthase kinase 3β,GSK3β)/活化T细胞核因子1(Nuclear factor of activated T cells c1,NFATc1)信号通路相关蛋白的表达。结果:与对照组比较,模型组大鼠血清FPG、FINS、HBA1c、TC、TG、LDL-C和MDA水平均升高(P<0.05),HDL-C、SOD、CAT和GSH水平均降低(P<0.05);股骨组织中骨小梁断裂状态较多,排列紊乱,骨质严重流失,BMD、Tb.N和Tb.Th均降低(P<0.05),股骨组织中BALP、OCN和Runx2蛋白表达水平均降低(P<0.05),AKT和NFATc1蛋白磷酸化水平均升高(P<0.05),GSK3β蛋白磷酸化水平降低(P<0.05)。与模型组比较,L-Ses组和H-Ses组大鼠血清中FPG、FINS、HBA1c、TC、TG、LDL-C和MDA水平均降低(P<0.05),HDL-C、SOD、CAT和GSH水平均升高(P<0.05);股骨组织中骨小梁断裂状态较少,排列、形态结构和骨质流失均呈现较大改善,BMD、Tb.N和Tb.Th均升高(P<0.05),股骨组织中BALP、OCN和Runx2蛋白表达水平均升高(P<0.05),AKT和NFATc1蛋白磷酸化水平均降低(P<0.05),GSK3β蛋白磷酸化水平升高(P<0.05)。与L-Ses组比较,H-Ses组大鼠血清中FPG、FINS、HBA1c、TC、TG、LDL-C、SOD、CAT、GSH和MDA水平均降低(P<0.05),HDL-C水平升高(P<0.05);大鼠股骨组织和形态稍有缓解,BMD、Tb.N和Tb.Th均升高(P<0.05),股骨组织中BALP、OCN和Runx2蛋白表达水平均升高(P<0.05),AKT和NFATc1蛋白磷酸化水平均降低(P<0.05),GSK3β蛋白磷酸化水平升高(P<0.05)。结论:芝麻素可能通过抑制AKT/GSK3β/NFATc1信号通路,促进骨形成,进而缓解T2DOP大鼠的骨质疏松症进展。

Objective:To investigate the therapeutic effect and its mechanism of Sesamin(Ses)on type 2 diabetic osteoporosis(T2DOP)model rats.Methods:Healthy male SD rats were divided into control group,model group,low-dose of Ses group(L-Ses group)and high-dose of Ses group(H-Ses group),with 10 rats in each group.The rats in model group,L-Ses group and H-Ses group were fed with high-fat and high-sugar diet for 4 weeks,and a one-time intraperitoneal injection of streptozotocin(STZ,dose 35 mg/kg)solution was used to establish a type 2 diabetic osteoporosis rat model.After successful modeling,rats in L-Ses group and H-Ses group were given Ses intragastric treatment at the dose of 10 mg/kg and 40 mg/kg,once a day,for 8 weeks.Rats in control group and model group were given the same amount of normal saline intragastric treatment at the same time.The indexes in serum of fasting plasma glucose(FPG),fasting insulin(FINS),glycosylated hemoglobin(HbA1c),serum total cholesterol(TC),triglyceride(TG),low-density lipoprotein cholesterol(LDL-C),high-density lipoprotein cholesterol(HDL-C),superoxide dismutase(SOD),catalase(CAT),glutathione(GSH)and malondialdehyde(MDA)in rats were detected according to the methods described in the detection kit.The morphology of rat femur was detected by Hematoxylin-eosin(HE)staining.The three-dimensional structure and bone mineral density(BMD),trabecular number(Tb.N),trabecular thickness(Tb.Th)parameters of femoral shaft epiphysis in rats were analyzed byμCT scanner.The protein expression levels of bone formation indicators bone alkaline phosphate(BALP),osteocalcin(OCN),runt-related transcription factor 2(Runx2)and AKT/GSK3β/NFATc1 signaling pathway were detected byWestern blot.Results:Compared with control group,the levels of FPG,FINS,HBA1c,TC,TG,LDL-C and MDA in serum of model group rats were increased(P<0.05),while the levels of HDL-C,SOD,CAT and GSH were decreased(P<0.05),the trabecular bone fracture in the femur tissue was more,the arrangement was disordered,and the bone loss was serious,and the BMD,Tb.N and Tb.Th were all decreased(P<0.05),the protein expression levels of BALP,OCN and Runx2 in the femur tissue were all decreased(P<0.05),the phosphorylation levels of AKT and NFATc1 were increased(P<0.05),while the phosphorylation levels of GSK3βwere decreased(P<0.05).Compared with model group,the levels of FPG,FINS,HBA1c,TC,TG,LDL-C and MDA in serum of L-Ses group and H-Ses group rats were decreased(P<0.05),while levels of HDL-C,SOD,CAT and GSH were increased(P<0.05),fracture of trabeculae was less in femur tissue,the arrangement,morphological structure and bone loss were greatly improved,the BMD,Tb.N and Tb.Th were all increased(P<0.05),and the protein expression levels of BALP,OCN and Runx2 in femur tissue were all increased(P<0.05),the phosphorylation levels of AKT and NFATc1 were decreased(P<0.05),while the phosphorylation levels of GSK3βwere increased(P<0.05).Compared with L-Ses group,the levels of FPG,FINS,HBA1c,TC,TG,LDL-C,SOD,CAT,GSH and MDA in serum of H-Ses group rats were decreased(P<0.05),while levels of HDL-C was increased(P<0.05),the tissue and morphology of rat femur were slightly relieved,the BMD,Tb.N and Tb.Th were all increased(P<0.05),and the protein expression levels of BALP,OCN and Runx2 in femur tissue were all increased(P<0.05),the phosphorylation levels of AKT and NFATc1 were decreased(P<0.05),while the phosphorylation levels of GSK3βwere increased(P<0.05).Conclusion:Sesamin may promote bone formation by inhibiting AKT/GSK3β/NFATc1 signaling pathway,thereby alleviating osteoporosis progression in T2DOP rats.