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BACKGROUND: Studies have shown that a certain dose of interleukin-4 intervention can yield the appropriate ratio of M1/M2 macrophage profile to generate a microenvironment conducive to bone healing and promoting bone regeneration. OBJECTIVE: To investigate the effect of interleukin-4 on NLRP3 inflammasome activation and osteoclast differentiation during bone tissue regeneration guided by bone replacement materials. METHODS: Forty-eight 6-8-week-old male SD rats were selected to establish a 5-mm diameter cranial bone defect model on the left and implanted with BioOss bone replacement material at the same time, and the periosteum was sutured. Rat models were randomly divided into control and experimental groups (n=24) at postoperative 3 days. Interleukin-4 and PBS were injected locally into the cranial bone defect area for the experimental and control groups, once a day, for 5 consecutive days. SD rats were executed at 1 and 2 weeks after surgery and skull samples were taken. Immunohistochemical staining was performed to detect the expression of cleaved caspase-1 and interleukin-1β protein. Immunofluorescence staining was performed to observe the expression of inducible nitric oxide synthase (M1 surface marker) and cleaved caspase-1 (NLRP3 indicator). Real-time fluorescence quantitative PCR was performed to detect the expression of related inflammatory factors caspase-1, interleukin-1β and osteoclast factor histone K gene. The differentiation and number of osteoclasts were observed by anti-tartrate acid phosphatase staining. At 6 and 12 weeks after surgery, micro-CT and hematoxylin-eosin staining were performed to observe the osteogenesis of the skull. RESULTS AND CONCLUSION: Immunofluorescence staining showed that the number of inducible nitric oxide synthase and cleaved caspase-1 doublestained cells in the experimental group was significantly lower than that in the control group at 1 and 2 weeks (P < 0.05). Immunohistochemical staining showed that the expression of cleaved caspase-1 and interleukin-1β in the experimental group was significantly lower than that in the control group at 1 week (P < 0.05). RT-qPCR suggested that the expression of caspase-1 mRNA in the experimental group was lower than that in the control group at 1 week (P < 0.05). The expression intensity of interleukin-1β and osteoclast factor histone K mRNA was significantly lower in the experimental group than that in the control group at postoperative 1 and 2 weeks (P < 0.05). Anti-tartrate acid phosphatase staining showed that the number of osteoclasts was significantly lower in the experimental group at 1 and 2 weeks than that in the control group (P < 0.05). Micro-CT results showed that the bone volume fraction and bone mineral density in the bone defect area were significantly higher in the experimental group than those in the control group at 6 and 12 weeks after injection (P < 0.05). Hematoxylin-eosin staining showed that 12 weeks after injection, multiple ossification centers were formed in the center of the defect in the experimental group, with scattered mature bone cells and lacunae, and a large number of osteoblasts around the material arranged into a single layer to participate in the formation of bone matrix. The results suggest that interleukin-4 may be involved in downregulating NLRP3 inflammasome expression, inhibiting the activation of caspase-1 and thus reducing the secretion of interleukin-1β, reducing the inflammatory state of the local microenvironment, as well as inhibiting osteoclast differentiation and playing a role in promoting new bone production during bone tissue regeneration guided by bone replacement materials. [ABSTRACT FROM AUTHOR]