河虾原肌球蛋白诱导的肥大细胞脱颗粒对巨噬细胞极化的体外研究OA

[Objective]Macrophages and mast cells,as vital components of the immune system,jointly form a key cellular network within the immune microenvironment.However,the effects and mechanisms of mast cell degranulation on macrophages in the context of food allergies remain poorly understood.This study aims to investigate the impact of allergen-induced mast cell degranulation on macrophage polarization.[Method]Highly purified macrophages and mast cells were first obtained through directed differentiation of mouse bone marrow stem cells.An in vitro degranulation model was established using mast cells as the primary cell type and the Macrobrachium nipponense allergen tropomyosin(TM).Optimal sensitization conditions were determined by optimizing mouse serum dilution ratio,TM concentration,and stimulation duration.Building upon this,direct contact co-culture and non-contact Transwell co-culture systems were established between mast cells and macrophages.Transcriptome sequencing technology was employed to systematically evaluate the effects of mast cell degranulation on macrophage proliferation,morphology,expression of polarization markers,and cytokine secretion.[Result](1)High-purity macrophages and mast cells were successfully induced from mouse bone marrow stem cells.The optimal stimulation conditions for the in vitro degranulation model are as follows:mouse serum dilution ratio 1∶30,TM stimulation concentration 100 µg/mL,and stimulation time 30 min.The results of co-culture showed that the degranulation of mast cells had no significant effect on macrophage proliferation but induced elongated morphology and increased pseudopod formation.In both direct co-culture and Transwell co-culture systems,mast cell degranulation significantly upregulated macrophage iNOS and Arg1 gene expression while promoting IFN-γ and IL-10 secretion.(2)Transcriptome sequencing results indicate that compared with the M0 group,genes associated with M1 polarization(Nos2,IL1β)and genes associated with M2 polarization(Arg1,CHIL3)showed an upward trend in the M0+MC group.(3)GO and KEGG enrichment analyses revealed that differentially expressed genes were significantly enriched in biological processes such as immune response and outer plasma membrane,as well as in the cytokine-cytokine receptor interaction pathway.[Conclusion]Under allergen TM stimulation,mast cell degranulation can induce macrophages to shift toward a mixed polarization state exhibiting both M1 and M2 characteristics through paracrine pathways,thereby reshaping their immune responses,which provides experimental evidence for understanding the synergistic regulation of immune cells in allergic reactions.