Effect of Niuxi Pills (牛膝丸) on Elderly Osteoporosis Model Mice through PI3K/Akt Pathway

Author：ZHOU Danwei1, JIN Lu1, WU Tong1, XIE Rixi1, GU Xiaoqiong1, ZHANG Xinjie2

Unit：1.Suzhou Integrated TCM & Western Medicine Hospital, Suzhou Jiangsu 215101, China; 2.Puzhuang Health Center, Linhu Town, Wuzhong District, Suzhou City, Suzhou Jiangsu 215101, China

Quote：引用：周丹微，金露，吴彤，谢日禧，顾小琼，张新杰.牛膝丸通过PI3K/AKT通路对老年性骨质疏松症模型小鼠的干预作用[J].中医药导报,2026,32(4):18-22,54.

DOI：10.13862/j.cn43-1446/r.2026.04.004

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Abstract：

Objective: To study the effect and mechanism of Niuxi pills on senile osteoporosis (SOP) in mice. Methods: Totally 6 four-month-old senescence-accelerated mouse resistant 1 (SAMR1) were used as control group, and 18 four-month-old senescence accelerated mouse prone 6(SAMP6) were randomly divided into model group, alendronate group and traditional Chinese medicine (TCM) group, with 6 mice in each group. After 2 weeks of adaptive feeding, the mice in TCM group were intragastrically administered with Niuxi pills at a dose of 0.23 g/(kg·d), and the mice in alendronate group was intragastrically administered with alendronate tablets at a dose of 1.53 mg/(kg·w). The mice in model group and control group were intragastrically administered with normal saline. After 12 weeks of treatment, the contents of procollagen I N-terminal propeptide (PINP), bone morphogenetic protein type 2 (BMP2), and bone alkaline phosphatase (BALP) in the serum of mice were measured by enzyme linked immunosorbent assay (ELISA). The femoral tissues of mice were stained using HE to observe the pathological changes of the femur under a microscope. The femurs of mice were scanned and analyzed using micro-CT (micro-CT). The parameters of bone mineral density (BMD), trabecular number (Tb.N), trabecular separation (Tb.Sp) and bone volume fraction (BV/TV) of the femur of mice were recorded. Biomechanical testing was performed on the femur of mice to determine the maximum load, maximum displacement and stiffness levels of the femur of mice. Western blotting technique was used to detect the protein expression levels of runt-related transcription factor 2 (RUNX2), BMP2, phosphorylated phosphatidylinyl-3-kinase (p-PI3K), PI3K, phosphorylated protein kinase B (p-Akt), and Akt in the femoral bone marrow of mice. Results: The serum levels of PINP, BMP2 and BALP in the model group were lower than those in control group (P<0.01). The serum levels of PINP, BMP2 and BALP in the alendronate group and TCM group were higher than those in the model group (P<0.01). HE staining showed that the model group mice had a significantly reduced number of trabecular bone, with widened and discontinuous intertrabecular spaces and numerous fracture sites. The control group mice had a significantly greater number of trabeculae than the model group. The TCM group and the alendronate group showed markedly improved trabecular bone quality, with widened trabeculae and increased trabecular number. The parameters of BMD, Tb.N, and BV/TV in the model group were significantly lower than those in the control group (P<0.01), while TB.Sp resolution was higher than that in the control group (P<0.01). The parameters of BMD, Tb.N, and BV/TV in the TCM group and alendronate group were significantly higher than those in the model group (P<0.05 or P<0.01), while TB.Sp resolution was lower than that in the model group (P<0.01). The femoral stiffness and maximum load level of mice in the model group were lower than those in the control group (P<0.01), while the maximum displacement was longer than that in the control group (P<0.01). The femoral stiffness and maximum load level of mice in the TCM group and alendronate group were higher than those in the model group (P<0.05 or P<0.01), while the maximum displacement was shorter than that in the model group (P<0.05 or P<0.01). The protein expression levels of RUNX2 and BMP2 in the model group were lower than those in the control group (P<0.01). The protein expression levels of RUNX2 and BMP2 and the ratios of p-PI3K/PI3K and p-Akt/Akt in the femurs of mice in the TCM group were significantly higher than those in the model group (P<0.05 or P<0.01). Conclusion: Niuxi pills can improve bone microarchitecture in SOP mice, and the mechanism may involve regulation of the PI3K/Akt signaling pathway to promote bone formation.

Key words：senile osteoporosis; Niuxi pills; PI3K/Akt signaling pathway; mouse

摘要：目的：研究牛膝丸对老年性骨质疏松症（SOP）小鼠的作用和机制。方法：以6只4个月龄的快速衰老对照小鼠（SAMR1）作为对照组，将18只4个月龄的快速衰老小鼠P6（SAMP6）随机分为模型组、阿仑膦酸钠组、中药组，每组6只。适应性喂养2周后，中药组小鼠灌胃牛膝丸[0.23 g/（kg·d）]，阿仑膦酸钠组小鼠灌胃阿仑膦酸钠片[1.53 mg/（kg·w）]，对照组、模型组小鼠灌胃生理盐水。12周后，酶联免疫吸附试验（ELISA）法检测小鼠血清Ⅰ型原胶原N端前肽（PINP）、骨形态发生蛋白2（BMP2）、骨碱性磷酸酶（BALP）含量；小鼠股骨组织HE染色，在显微镜下观察股骨病理学变化；使用微型CT（Micro-CT）扫描小鼠股骨，记录小鼠股骨骨密度（BMD）、骨小梁数量（Tb.N）、骨小梁间隙（Tb.Sp）及骨体积分数（BV/TV）；检测小鼠股骨生物力学，测定小鼠股骨最大载荷、最大位移及刚度水平；应用蛋白质印迹（Western blotting）法检测小鼠股骨组织Runt相关转录因子2（RUNX2）、BMP2、磷酸化磷脂酰基醇-3-激酶（p-PI3K）、磷脂酰基醇-3-激酶（PI3K）、磷酸化蛋白激酶B（p-Akt)、蛋白激酶B（Akt）蛋白表达水平。结果：模型组小鼠血清PINP、BMP2、BALP含量低于对照组（P<0.01）；中药组及阿仑膦酸钠组小鼠血清PINP、BMP2、BALP含量高于模型组（P<0.01）。HE染色显示模型组小鼠骨小梁数量明显减少，间隙增宽，且不连续，断裂部位较多；对照组小鼠骨小梁数量明显多于模型组；中药组及阿仑膦酸钠组小鼠骨小梁质量明显改善，骨小梁变宽，数量增加。模型组小鼠BMD、Tb.N、BV/TV低于对照组（P＜0.01），Tb.Sp高于对照组（P＜0.01）；中药组及阿仑磷酸钠组小鼠BMD、Tb.N、BV/TV高于模型组（P＜0.05或P＜0.01），Tb.Sp低于模型组（P＜0.01）。模型组小鼠股骨刚度、最大载荷低于对照组（P<0.01），最大位移明显长于对照组（P<0.01）；中药组及阿仑磷酸钠组小鼠股骨刚度及最大载荷均明显高于模型组（P<0.05或P<0.01），最大位移短于模型组（P<0.05或P<0.01）。模型组小鼠股骨组织RUNX2、BMP2蛋白相对表达量低于对照组（P<0.01）；中药组小鼠股骨组织RUNX2、BMP2蛋白相对表达量及p-PI3K/PI3K、p-Akt/Akt高于模型组（P<0.05或P<0.01）。结论：牛膝丸能改善SOP小鼠的骨微结构，机制可能为调节PI3K/Akt信号通路促进骨形成。

关键词：老年性骨质疏松症；牛膝丸；PI3K/Akt信号通路；小鼠

Release time：2026-04-26

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