中国畜牧兽医2025，Vol.52Issue(12)：5529-5538,10.DOI:10.16431/j.cnki.1671-7236.2025.12.001
混合谱系激酶结构域样蛋白对小鼠巨噬细胞和成纤维细胞间相互作用的影响
Effect of MLKL on the Interaction Between Macrophages and Fibroblasts in Mice
摘要
Abstract
[Objective]This experiment aimed to explore the effects of mixed lineage kinase domain-like protein(MLKL)on macrophages and fibroblasts during the process of skin wound healing in mice and its regulatory mechanism.[Method]The back trauma models of C57BL/6J mice and MLKL gene knockout(MLKL-/-)mice were established.Immunohistochemistry was used to detect the dynamic expression of MLKL,as well as the recruitment of macrophages and fibroblasts after skin injury in mice.Through in vitro experiments,a co-culture system of fibroblasts and M1/M2 macrophages conditioned medium(M1/M2ø CM)was established.Western blotting was performed to evaluate the protein expression of transforming growth factor-β(TGF-β),fibroblast growth factor-2(FGF-2),and matrix metalloproteinase-9(MMP-9)in fibroblasts.M1/M2 macrophages were co-cultured with fibroblast-conditioned medium(FBCM).The secretion of interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)in M1 macrophages was measured by ELISA.The secretion of IL-10 and the expression of Arginase protein in M2 macrophages were determined by ELISA and Western blotting,respectively.[Result]The immunohistochemical staining results showed that positive MLKL signals were detected on the 3rd,5th and 7th days of skin wound healing in C57BL/6J mice.The expression level of MLKL increased with the prolongation of wound healing time.Moreover,the number of CD86-positive M1 macrophages,CD163-positive M2 macrophages and Vimentin-positive fibroblasts around the wound in MLKL-/-mice was less than that in C57BL/6J mice.In co-culture experiments,compared with the untreated group,the treatment of M1ø CM and M2ø CM derived from C57BL/6J significantly increased the protein expression levels of TGF-β,FGF-2 and MMP-9 in fibroblasts(P＜0.05),whereas after treatment with M1ø CM and M2ø CM derived from MLKL-/-,there was no significant difference in the protein expression of FGF-2 and MMP-9 in fibroblasts(P＞0.05),while the expression level of TGF-β protein was significantly decreased(P＜0.05).After macrophages were treated with C57BL/6J FBCM,the contents of IL-6 and TNF-α in M1 macrophages,the content of IL-10 and the expression level of Arginase protein in M2 macrophages were significantly increased(P＜0.05).After macrophages were treated with MLKL-/-FBCM,the contents of related cytokines and protein expression levels were both lower than those in C57BL/6J FBCM treatment group(P＜0.05).[Conclusion]MLKL could promote skin wound healing by regulating the interaction between macrophages and fibroblasts,influencing the recruitment of macrophages,thereby promoting the healing of skin wounds.关键词
成纤维细胞/巨噬细胞/MLKL/伤口愈合Key words
fibroblast/macrophages/MLKL/wound healing分类
农业科技引用本文复制引用
董妍沁,刘博,赵佳敏,包文慧,巩志国,任佩佩,王奕心,代佳山,赵非凡,张双翼..混合谱系激酶结构域样蛋白对小鼠巨噬细胞和成纤维细胞间相互作用的影响[J].中国畜牧兽医,2025,52(12):5529-5538,10.基金项目
国家自然科学基金项目(32202879) （32202879）
内蒙古自然科学基金项目(2024MS03064) （2024MS03064）
内蒙古农业大学一流学科科研专项(YLXKZX-NND-061) （YLXKZX-NND-061）
