Chinese Journal of Tissue Engineering Research ›› 2026, Vol. 30 ›› Issue (24): 6165-6173.doi: 10.12307/2026.184

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Bushen Tongshi Pills improves osteogenic disorders in alcoholic femoral head necrosis rats

Wang Weiwei1, Ding Qiang1, Rong Xiangbin2, Guo Liang1, Zhao Canbin1, Tao Hongcheng1, Niu Chicheng1, Liu Jinfu3, Zeng Ping3   

  1. 1Guangxi University of Chinese Medicine, Nanning 530029, Guangxi Zhuang Autonomous Region, China; 2Ruikang Hospital Affiliated to Guangxi University of Chinese Medicine, Nanning 530011, Guangxi Zhuang Autonomous Region, China; 3The First Affiliated Hospital of Guangxi University of Chinese Medicine, Nanning 530011, Guangxi Zhuang Autonomous Region, China
  • Received:2025-06-09 Revised:2025-09-30 Online:2026-08-28 Published:2026-01-27
  • Contact: Zeng Ping, PhD, Doctoral supervisor, The First Affiliated Hospital of Guangxi University of Chinese Medicine, Nanning 530011, Guangxi Zhuang Autonomous Region, China
  • About author:Wang Weiwei, MD candidate, Guangxi University of Chinese Medicine, Nanning 530029, Guangxi Zhuang Autonomous Region, China
  • Supported by:
    The National Natural Science Foundation of China, No. 82160913 (to ZP); Guangxi Natural Science Foundation, No. 2024GXNSFAA010228 (to ZP)

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Abstract: BACKGROUND:  Bushen Tongshi Pills has been proven to delay the progression of collapse in alcoholic femoral head necrosis, but the mechanism is still unclear.
OBJECTIVE: To explore the mechanism by which Bushen Tongshi Pills improve osteogenic disorders of alcoholic femoral head necrosis. 
METHODS: Fifty male Sprague-Dawley rats were randomly divided into a control group, a model group, and low, medium, and high dose groups of Bushen Tongshi Pills, with 10 rats in each group. Except for the control group, the other groups were fed with Lieber DeCarli liquid food containing ethanol for 8 weeks to establish an alcoholic femoral head necrosis model. At the same time, the low, medium, and high dose groups of Bushen Tongshi Pills were given 1.05, 2.1, and 4.2 g/kg of Bushen Tongshi Pills by gavage per day, respectively. After 8 weeks, Micro-CT was used to observe the overall morphology of the femoral head. Hematoxylin-eosin staining was used to observe the pathological morphology of the femoral head. Enzyme-linked immunosorbent assay was used to detect the levels of interleukin-1β and interleukin-18 in rat serum. Immunohistochemistry staining and western blot assay were used to detect nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing protein 3, Caspase-1, Gasdermin D, Runt related transcription factor 2, osteocalcin, and type I collagen, while the mRNA expression levels of the above genes were quantified by RT-qPCR. 
RESULTS AND CONCLUSION: (1) Micro-CT and hematoxylin-eosin staining results showed that the medium- and high-dose Bushen Tongshi Pills could significantly improve the bone loss and pathological morphology development of the femoral head in the model rat of alcoholic femoral head necrosis. (2) The enzyme-linked immunosorbent assay results showed that compared with the control group, the levels of interleukin-1β and interleukin-18 in the serum of the model group rats were significantly increased (P < 0.05); compared with the model group, the levels of interleukin-1β and interleukin-18 in the serum of rats were significantly decreased in each dose group of Bushen Tongshi Pills (P < 0.05), and decreased in a dose-dependent manner. Among them, the high-dose group of Bushen Tongshi Pills showed the most significant decrease (P < 0.05). (3) Immunohistochemistry, western blot assay, and RT-qPCR results showed that compared with the control group, the protein and mRNA expression levels of the nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing protein 3, Caspase-1, Gasdermin D in the model group were significantly upregulated (P < 0.05), while the protein and mRNA expression levels of Runt-related transcription factor 2, osteocalcin, and type I collagen were significantly downregulated (P < 0.05). Compared with the model group, the protein and mRNA expression levels of the nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing protein 3, Caspase-1, Gasdermin D in each dose group of Bushen Tongshi Pills were downregulated in a dose-dependent manner, while the protein and mRNA expression levels of Runt-related transcription factor 2, osteocalcin, and type I collagen were upregulated in a dose-dependent manner. Among them, the high-dose group of Bushen Tongshi Pills showed the most significant upregulation (P < 0.05). (4) The western blot assay results showed that compared with the control group, the expression levels of cleaned-Caspase-1, dermatin D-N protein, cleaned-Caspase-1/Caspase-1 ratio, and dermatin D-N/dermatin D ratio in the femoral head of the model group rats were significantly upregulated (P < 0.05). Compared with the model group, the expression levels of cleaned-Caspase-1, dermatin D-N protein, and dermatin D-N/dermatin D ratio were significantly downregulated in each dose group of Bushen Tongshi Pills (P < 0.05). Compared with the model group, the cleaned-Caspase-1/Caspase-1 ratio was significantly downregulated in the medium and high dose groups of Bushen Tongshi Pills (P < 0.05). To conclude, Bushen Tongshi Pills potentially attenuate inflammatory cytokine release through the nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing protein 3/Caspase-1/Gasdermin D pathway and enhance osteogenic differentiation, thereby promoting bone tissue regeneration in the rat model of alcoholic femoral head necrosis.


Key words: Bushen Tongshi Pills, alcoholic femoral head necrosis, pyroptosis, nucleotide-binding oligomerization domain,leucine-rich repeat and pyrin domain-containing protein 3 (NLRP3), Caspase-1, Gasdermin D (GSDMD), osteogenesis, pathway

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