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目的:探讨动脉粥样硬化中钾通道Kv1.3阻断剂抗hKv1.3E314抗体对巨噬细胞分化的影响。方法:培养THP-1单核细胞,给予佛波酯100μg/L诱导72h使其分化为巨噬细胞。将巨噬细胞洗涤后重新接种在6孔板中,分为A组、B组和C组:A组加入LPS 10ng/L和IFN-γ20μg/L诱导24h使其分化;B组给予E314抗体(300nmol/L)在37℃孵育2h后加入LPS 10ng/L和IFN-γ20μg/L诱导24h使其分化;C组细胞不做任何处理。倒置相差显微镜下观察细胞形态学变化;流式细胞术鉴定巨噬细胞分型;ELLISA检测细胞上清液中IL-10、IL-12表达水平。结果:经流式细胞术鉴定:A组细胞iNOS抗体强阳性,为M1型;B组细胞CD206抗体强阳性,为M2型;C组流式结果为阴性。ELLISA结果显示A组IL-12表达水平高于B组(P<0.05),A组IL-10表达水平低于B组(P<0.05)。结论:体外诱导巨噬细胞分化,炎性因子促使其向M1型分化,具有促进炎症反应的作用;加入Kv1.3通道阻断剂后,炎性因子同样诱导分化的巨噬细胞会向M2型抗炎方向发展。通过阻断巨噬细胞上的Kv1.3通道可以促进巨噬细胞向着具有抗炎作用的M2型巨噬细胞分化。
Objective: To investigate the effect of anti-hKv1.3E314 antibody against potassium channel Kv1.3 blocker on macrophage differentiation in atherosclerosis. Methods: THP-1 monocytes were cultured and induced by phorbol ester 100μg / L for 72h to differentiate into macrophages. The macrophages were washed and re-seeded into 6-well plates and divided into groups A, B and C: Group A was treated with LPS 10ng / L and IFN-γ20μg / L for 24 hours to induce differentiation; Group B was given E314 antibody 300nmol / L) were incubated at 37 ℃ for 2h and then induced by differentiation of LPS 10ng / L and IFN-γ 20μg / L for 24 h. Cells in group C were treated without any treatment. Morphological changes were observed under inverted phase contrast microscope. Flow cytometry was used to identify macrophages. ELLISA was used to detect the expression of IL-10 and IL-12 in supernatant. Results: The results of flow cytometry showed that the iNOS antibody in group A was strongly positive for type M1; the antibody in group B was strongly positive for type M2 and the type C was negative in flow cytometry. ELLISA results showed that the expression of IL-12 in group A was higher than that in group B (P <0.05). The expression of IL-10 in group A was lower than that in group B (P <0.05). CONCLUSION: Macrophage differentiation induces in vitro and induces differentiation into M1 type and promotes the inflammatory response. After Kv1.3 channel blocker is added, macrophages differentiated from inflammatory cytokines will also differentiate into M2 type Anti-inflammatory direction. By blocking Kv1.3 channels on macrophages, macrophages can be promoted to differentiate toward anti-inflammatory M2 macrophages.