乙醛脱氢酶2调控巨噬细胞M2型极化促进创面愈合OACSTPCD
Aldehyde dehydrogenase 2 promotes wound healing by regulating M2 macrophage polarization
目的 验证乙醛脱氢酶 2(aldehyde dehydrogenase 2,ALDH2)在创面愈合中的作用并探讨其分子机制.方法 构建小鼠全层皮肤切除模型,随机分为对照组和Alda-1(ALDH2 激动剂)组,观察创面愈合情况并检测创面组织ALDH2 活性.采用Masson染色观察创面胶原纤维含量,免疫组化染色检测ALDH2 和α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)水平,免疫荧光染色检测Ⅰ型胶原(Col-Ⅰ)、Ⅲ型胶原(Col-Ⅲ)表达和F4/80+、iNOS+、CD206+细胞数.体外实验采用白介素 4(interleukin-4,IL-4)诱导RAW264.7 细胞M2 型极化,分别用Alda-1 和ALDH2抑制剂CVT-10216处理细胞.流式细胞仪检测F4/80+CD206+细胞比例,酶联免疫分析测定细胞上清中促炎因子和抗炎因子水平,Western印迹法检测AKT/mTOR信号通路相关蛋白表达.结果 与对照组相比,Alda-1 组小鼠创面愈合率显著提高,创面的Col-Ⅰ、Col-Ⅲ及α-SMA表达增多;F4/80+细胞差异无统计学意义,iNOS+细胞显著减少,CD206+细胞显著增多.体外实验结果显示,与IL-4组相比,IL-4+Alda-1组F4/80+CD206+细胞比例、抗炎因子和p-AKT、p-mTOR蛋白表达显著升高,促炎因子表达降低;IL-4+CVT-10216 组F4/80+CD206+细胞比例、抗炎因子和p-AKT、p-mTOR蛋白表达显著降低,促炎因子表达升高.结论 ALDH2通过AKT/mTOR通路诱导巨噬细胞M2型极化,促进小鼠创面愈合.
Objective To verify the role of aldehyde dehydrogenase 2(ALDH2)in wound healing and to explore the underlying mechanisms.Methods A murine excisional wound model was developed and mice were randomly assigned to control group and Alda-1(ALDH2 agonist)group.Wound healing rate and activity of ALDH2 were measured.Masson staining was used to observe the collagen fiber content,and immunohistochemistry was used to detect the expression of ALDH2 and α-smooth muscle actin(α-SMA).The expression of collagen typeⅠandⅢ(Col-Ⅰ,Col-Ⅲ),and the number of F4/80,inducible nitric oxide synthase(iNOS)and CD206 positive(F4/80+,iNOS+,CD206+)cells were analyzed using immunofluorescences.Furthermore,interleukin(IL)-4-stimulated RAW264.7 cells were treated with Alda-1 or CVT-10216(an ALDH2 inhibitor)in vitro.The proportion of F4/80+CD206+cells were analyzed using flow cytometry.Anti-inflammatory and pro-inflammatory cytokines were examined using ELISA.The expression of protein associated with the AKT/mTOR pathway were detected via western blotting.Results Compared with control group,the wound healing rate of mice in the Alda-1 group was significantly improved,and the expression of Col-Ⅰ,Col-Ⅲ,and α-SMA in the wound surface increased.Although the number of F4/80+cells in wounds did not significantly differ,there was a decrease in iNOS+cells and an increase in CD206+cells.In vitro,compared to IL-4 group,IL-4+Alda-1 group exhibited an increased proportion of F4/80+CD206+macrophages and higher levels of anti-inflammatory factors,alongside a reduction in levels of pro-inflammatory factors.Conversely,IL-4+CVT-10216 group demonstrated a decreased proportion of F4/80+CD206+macrophages,lower levels of anti-inflammatory factors with a concomitant increase in pro-inflammatory factors.Additionally,the protein expressions of p-AKT and p-mTOR were significantly elevated in IL-4+Alda-1 group and diminished in IL-4+CVT-10216 group.Conclusion ALDH2 induces M2 macrophage polarization via AKT/mTOR pathway to promote wound healing in mice.
张思敏;亓发芝
上海市老年医学中心整形外科,上海 201104上海市老年医学中心整形外科,上海 201104||复旦大学附属中山医院整形外科,上海 200032
临床医学
创面愈合巨噬细胞极化乙醛脱氢酶2Alda-1
wound healingmacrophage polarizationaldehyde dehydrogenase 2Alda-1
《中国临床医学》 2024 (005)
724-733 / 10
国家自然科学基金(82172212).Supported by National Natural Science Foundation of China(82172212).
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