Abstract：
BACKGROUND: HuoXueTongFu Formula (HXTF) is a traditional Chinese herbal formula that has been used as a supplement and alternative therapy for intraperitoneal adhesion (IA). However, its specific mechanism of action has not been fully understood.
OBJECTIVE: In surgery, IA presents an inevitable challenge, significantly impacting patients\' physical and mental well-being and increasing the financial burden. Our previous research has confirmed the preventive effects of HXTF on IA formation. However, the precise mechanism of its action still needs to be understood.
METHODS: In this study, the IA model was successfully established by using the Ischemic buttons and treated with HXTF for one week with or without Mer Tyrosine Kinase (MerTK) inhibitor. We evaluated the pharmacodynamic effect of HXTF on IA mice. The MerTK/phosphoinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway-associated proteins were detected by Western blotting. Neutrophil extracellular traps (NETs) were detected by immunofluorescence. Macrophage phenotype was assessed by immunohistochemistry and flow cytometry. Inflammatory cytokines were detected by Real Time Quantitative PCR and Western blotting.
RESULTS: HXTF reduced inflammatory response and alleviated IA. HXTF significantly enhanced MerTK expression, increased the number of M2c macrophages, and decreased the formation of NETs. In addition, the MerTK/PI3K/AKT pathway was significantly activated by HXTF. However, after using MerTK inhibitors, the role of HXTF in inducing M2c macrophage through activation of the PI3K/AKT pathway was suppressed and there was no inhibitory effect on NETs formation and inflammatory responses, resulting in diminished inhibition of adhesion.
CONCLUSIONS: HXTF may improve IA by activating the MerTK/PI3K/AKT pathway to induce M2c polarization, which removes excess NETs and attenuates the inflammatory response.
OBJECTIVE: In surgery, IA presents an inevitable challenge, significantly impacting patients\' physical and mental well-being and increasing the financial burden. Our previous research has confirmed the preventive effects of HXTF on IA formation. However, the precise mechanism of its action still needs to be understood.
METHODS: In this study, the IA model was successfully established by using the Ischemic buttons and treated with HXTF for one week with or without Mer Tyrosine Kinase (MerTK) inhibitor. We evaluated the pharmacodynamic effect of HXTF on IA mice. The MerTK/phosphoinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway-associated proteins were detected by Western blotting. Neutrophil extracellular traps (NETs) were detected by immunofluorescence. Macrophage phenotype was assessed by immunohistochemistry and flow cytometry. Inflammatory cytokines were detected by Real Time Quantitative PCR and Western blotting.
RESULTS: HXTF reduced inflammatory response and alleviated IA. HXTF significantly enhanced MerTK expression, increased the number of M2c macrophages, and decreased the formation of NETs. In addition, the MerTK/PI3K/AKT pathway was significantly activated by HXTF. However, after using MerTK inhibitors, the role of HXTF in inducing M2c macrophage through activation of the PI3K/AKT pathway was suppressed and there was no inhibitory effect on NETs formation and inflammatory responses, resulting in diminished inhibition of adhesion.
CONCLUSIONS: HXTF may improve IA by activating the MerTK/PI3K/AKT pathway to induce M2c polarization, which removes excess NETs and attenuates the inflammatory response.
摘要：
背景：活血通复方（HXTF）是一种传统的中草药配方，已被用作腹膜内粘连（IA）的补充和替代疗法。然而,其具体作用机制尚未完全了解。
目标：在外科手术中，IA提出了不可避免的挑战，显着影响患者的身心健康，增加经济负担。我们先前的研究已经证实了HXTF对IA形成的预防作用。然而,其作用的确切机制仍需了解。
方法：在本研究中，通过使用缺血纽扣成功建立IA模型，并在有或没有Mer酪氨酸激酶（MerTK）抑制剂的情况下用HXTF治疗一周。我们评估了HXTF对IA小鼠的药效学作用。Western印迹法检测MerTK/磷酸肌醇3激酶(PI3K)/蛋白激酶B(AKT)通路相关蛋白。免疫荧光法检测中性粒细胞胞外诱捕网(NETs)。通过免疫组织化学和流式细胞术评估巨噬细胞表型。通过实时定量PCR和Western印迹检测炎性细胞因子。
结果：HXTF降低了炎症反应，缓解了IA。HXTF显著增强MerTK表达，增加M2c巨噬细胞的数量，并减少了NET的面积。此外,MerTK/PI3K/AKT通路被HXTF显著激活。然而,使用MerTK抑制剂后，HXTF通过激活PI3K/AKT途径诱导M2c巨噬细胞的作用被抑制，对NETs形成和炎症反应没有抑制作用，导致粘附抑制减弱。
结论：HXTF可能通过激活MerTK/PI3K/AKT通路诱导M2c极化来改善IA，去除过量的NETs并减弱炎症反应。
目标：在外科手术中，IA提出了不可避免的挑战，显着影响患者的身心健康，增加经济负担。我们先前的研究已经证实了HXTF对IA形成的预防作用。然而,其作用的确切机制仍需了解。
方法：在本研究中，通过使用缺血纽扣成功建立IA模型，并在有或没有Mer酪氨酸激酶（MerTK）抑制剂的情况下用HXTF治疗一周。我们评估了HXTF对IA小鼠的药效学作用。Western印迹法检测MerTK/磷酸肌醇3激酶(PI3K)/蛋白激酶B(AKT)通路相关蛋白。免疫荧光法检测中性粒细胞胞外诱捕网(NETs)。通过免疫组织化学和流式细胞术评估巨噬细胞表型。通过实时定量PCR和Western印迹检测炎性细胞因子。
结果：HXTF降低了炎症反应，缓解了IA。HXTF显著增强MerTK表达，增加M2c巨噬细胞的数量，并减少了NET的面积。此外,MerTK/PI3K/AKT通路被HXTF显著激活。然而,使用MerTK抑制剂后，HXTF通过激活PI3K/AKT途径诱导M2c巨噬细胞的作用被抑制，对NETs形成和炎症反应没有抑制作用，导致粘附抑制减弱。
结论：HXTF可能通过激活MerTK/PI3K/AKT通路诱导M2c极化来改善IA，去除过量的NETs并减弱炎症反应。
