摘要
目的探讨温肾通络法对糖尿病肾病相关骨质疏松(nephropathy-related osteoporosis,DNOP)大鼠的作用及潜在机制。方法9周龄雄性Sprague-Dawley(SD)大鼠通过链脲佐菌素处理以诱导DNOP大鼠模型。实验分组为:对照组(n=10)、DNOP组(n=10)、温肾通络方低剂量组(n=10)、温肾通络方中剂量组(n=10)、温肾通络方高剂量组(n=10)和阿仑膦酸钠(alendronate,ALN)组(阳性对照组,n=10)。酶联免疫吸附试验试剂盒检测肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-6(interleukin-6,IL-6)、白细胞介素-8(interleukin-6,IL-8)、白细胞介素-1β(interleukin-6,IL-1β)、碱性磷酸酶(Alkaline phosphatase,ALP)和酒石酸盐抗性酸性磷酸酶(tartrate resistant acid phosphatase,TRAP)的水平。蛋白质免疫印迹分析测定Wnt/YAP/TAZ通路相关蛋白、上皮细胞-间充质转化(epithelial-mesenchymal transition,EMT)相关标志物以及成骨细胞分化相关蛋白的表达。结果在对一般特征的影响中,与DNOP组大鼠相比,中剂量温肾通络方组大鼠的体重显著增加(P<0.05),空腹血糖(P<0.01),血清肌酐(P<0.05)和尿素氮(BUN)水平(P<0.05)降低,尿肌酐水平(P<0.01)升高;高剂量温肾通络方组大鼠的体重显著增加(P<0.01),空腹血糖(P<0.01),血清肌酐(P<0.05)、BUN水平(P<0.01)、尿量(P<0.01)、尿白蛋白(ALB)水平(P<0.05)、肾脏重量(P<0.01)和肾脏指数水平降低(P<0.01);ALN治疗组的肾脏重量和肾脏指数水平降低(P<0.01)。在对EMT相关标志物的影响中,与DNOP组大鼠相比,中剂量、高剂量温肾通络方组和ALN治疗组α-SMA和Vimentin的表达降低,E-cadherin的表达升高。在炎症水平中,高剂量和中剂量温肾通络方组的TNF-α、IL-6、IL-8和IL-1β水平显著降低(P<0.05)。在对骨代谢的影响中,与DNOP组相比,高剂量温肾通络方组和ALN组的尿钙和磷减少(P<0.05)。在对骨密度(BMD)和显微结构、对股骨生物力学特性以及对骨骼Wnt/YAP/TAZ信号通路和分化的影响中,与DNOP组相比,温肾通络方和ALN组的BMD和骨体积(BV)显著增加(P<0.05),损失弹性模量、极限载荷和抗弯强度均显著降低(P<0.05)、成骨细胞分化相关蛋白BMP-4、BMP-7、Runx2、OCN和Col1a1的表达显著增加(P<0.01)。结论该研究表明,温肾通络方对DNOP具有治疗作用,可能是通过下调Wnt/YAP/TAZ通路所实现的。
Objective To investigate the effects and potential mechanisms of Wenshen Tongluo Formula in rats with diabetic nephropathy-induced osteoporosis(DNOP).Methods Male Sprague-Dawley(SD)rats,aged 9 weeks,were induced with DNOP using streptozotocin(STZ).The rats were divided into the following groups:Control Group(n=10),DNOP Group(n=10),Low-dose Wenshen Tongluo Formula Group(n=10),Medium-dose Wenshen Tongluo Formula Group(n=10),High-dose Wenshen Tongluo Formula Group(n=10),and Alendronate(ALN)Group(positive control,n=10).Enzyme-linked immunosorbent assay(ELISA)kits were used to detect tumor necrosis factor-alpha(TNF-α),interleukin-6(IL-6),interleukin-8(IL-8),interleukin-1β(IL-1β),alkaline phosphatase(ALP),and tartrate-resistant acid phosphatase(TRAP)levels.Western blotting was used to analyze the expression of proteins related to the Wnt/YAP/TAZ signaling pathway,epithelial-mesenchymal transition(EMT)markers,and osteoblast diferentiation proteins.Results Treatment with Wenshen Tongluo Formula significantly improved the overall health,kidney function,and pathological changes in the kidneys of DNOP rats.Wenshen Tongluo Formula reduced the levels of TNF-α,IL-6,IL-8,IL-1β,ALP,and TRAP,increased serum calcium and phosphorus levels,and decreased urinary calcium and phosphorus levels.Additionally,the formula significantly improved bone mineral density(BMD),bone volume(BV),and bone microstructure in DNOP rats.The elastic modulus,ultimate load,and bending strength of the femur were also significantly enhanced.After treatment,the expression ofα-SMA,vimentin,Wnt Family Member 3A(Wnt3a),phosphorylated Yes-associated protein(p-YAP),and TAZ was significantly downregulated,while the expression of E-cadherin,Bone Morphogenetic Protein 4(BMP-4),Bone Morphogenetic Protein 7(BMP-7),Runt-related transcription factor 2(Runx2),osteocalcin,and collagen type I alpha 1 chain(Collal)was significantly upregulated.Conclusion This study suggests that Wenshen Tongluo Formula has therapeutic effects on DNOP,which may be achieved through the downregulation of the Wnt/YAP/TAZ signaling pathway.
作者
徐英凯
蒋剑锋
王磊
XU Ying-kai;JIANG Jian-feng;WANG Iei(Department of Orthopedics and Traumatology,Hai'an Hospital of Traditional Chi-nese Medicine Afiliated to Nanjing University of Traditional Chinese Medicine,Hai'an 226600,Jiangsu,China;不详)
出处
《广东医学》
2025年第5期672-680,共9页
Guangdong Medical Journal
基金
江苏省科学技术厅项目(BE2022801)
南通市市级科技计划(指导性)项目(JCZ19078)。
