Nocth信号通路对糖尿病足溃疡炎症因子及转化生长因子表达的影响

Effects of Nocth signaling pathway on the expression of inflammatory factors and transforming growth factors in diabetic foot ulcers

目的:以糖尿病足溃疡大鼠为模型,通过对Notch信号干预,探索其对炎症因子、趋化因子及转化生长因子的作用。方法:实验模型采用高脂饲料配合链脲佐菌素(STZ)造成糖尿病,用恒温恒压烫伤仪建立糖尿病足溃疡实验模型,并以无糖尿病足溃疡模型作为普通溃疡对照组。随后,对糖尿病足溃疡实验模型干预7 d,包括以生理盐水、西药生长因子、Notch信号激动剂Jagged1、Notch信号抑制剂LY‑411575、中药紫朱软膏进行干预。采用ELISA法检测血清白介素‑1β(IL‑1β)、白介素‑6(IL‑6)、肿瘤坏死因子‑α(TNF‑α)、白介素‑17A(IL‑17A);采用Real‑TimePCR检测创面组织Notch信号相关的炎性因子(TNF‑α、IL‑1β、IL‑6、IL‑17)、趋化因子[白介素‑8(IL‑8)、IP‑10、单核细胞趋化蛋白1(MCP‑1)]及转化生长因子(TGF‑α、TGF‑β)。结果:ELISA检测结果显示,糖尿病足溃疡大鼠模型的血清IL‑1β、IL‑6、TNF‑α、IL‑17A水平明显高于普通溃疡模型(P<0.01)。与糖尿病足溃疡对照组相比,Notch信号抑制剂LY‑411575组与中药紫朱软膏组IL‑6、TNF‑α、IL‑17A含量明显减低(P<0.01)。创面组织Real‑timePCR结果显示,糖尿病足溃疡大鼠模型创面组织的炎症因子IL‑1β、IL‑6、TNF‑α、IL‑17和趋化因子IP‑10、IL‑8、MCP‑1水平明显高于普通溃疡模型(P<0.01),转化生长因子TGF‑β水平低于普通溃疡模型(P<0.01)。与糖尿病足溃疡对照组相比,Notch信号抑制剂LY‑411575明显降低糖尿病足溃疡大鼠炎症因子IL‑1β、IL‑6、TNF‑α、IL‑17水平和趋化因子IP‑10、IL‑8、MCP‑1含量(P<0.01),增加转化生长因子TGF‑α、TGF‑β表达(P<0.05),而Notch信号激动剂Jagged1则降低转化生长因子TGF‑α、TGF‑β表达(P<0.05),这表明了抑制Notch信号通路能够降低炎症因子IL‑1β、IL‑6、TNF‑α、IL‑17水平和趋化因子IL‑8含量,增加转化生长因子TGF‑α、TGF‑β表达。与糖尿病足溃疡对照组相比,中药紫朱软膏能够降低糖尿病足溃疡大鼠创面组织的炎症因子IL‑1β、IL‑6、TNF‑α、IL‑17和趋化因子IP‑10、IL‑8、MCP‑1水平(P<0.01),提高转化生长因子TGF‑α、TGF‑β表达(P<0.01)。Notch信号抑制剂LY‑411575则增强了紫朱软膏对转化生长因子TGF‑α、TGF‑β促表达作用。结论:糖尿病足溃疡大鼠模型较普通足溃疡血清及创面炎症因子、趋化因子表达偏高而转化生长因子表达不足。抑制Notch信号通路能够降低糖尿病足溃疡实验模型大鼠的炎症因子和趋化因子,增加转化生长因子表达,Notch信号通路具有促炎和部分促细胞增殖作用。紫朱软膏能够降低糖尿病足溃疡大鼠炎症因子和炎症趋化水平,提高转化生长因子表达,减轻创面炎症反应,其作用可能与抑制Nocth信号表达有关。

Objective:To explore the effects of Notch signaling pathway on inflammatory factors,chemokines,and trans-forming growth factors in a diabetic foot ulcer rat model.Methods:The experimental model used high-fat feed combined with streptozotocin to cause diabetes.The experimental model of diabetic foot ulcer was established with a constant temperature and pressure scald instrument,while non-diabetic foot ulcer model served as a normal control.Subsequently,intervention was conduct-ed on the experimental model of diabetic foot ulcer,by using physiological saline,Western medicine growth factor,Notch signal agonist Jagged1,Notch signal inhibitor LY-411575,and Chinese medicine Zizhu ointment for 7 days.ELISA was adopted to de-tect the serum IL-1β,IL-6,TNF-α,and IL-17A;Real-time PCR was used to test inflammatory factors(TNF-α,IL-1β,Il-6,IL-17),chemokines(IL 8,IP-10,MCP-1)and transforming growth factors(TGF-α,TGF-β),which all were related to Notch signal in wound tissues.Results:ELISA results showed the levels of serum IL-1β,IL-6,TNF-αand IL-17A in the diabetic foot ulcer rat model were significantly higher than those in the normal ulcer model(P<0.01).Compared to the diabetic foot ulcer con-trol group,the levels of IL-6,TNF-αand IL-17A in both the Notch signal inhibitor LY-411575 group and the Chinese medicine Zizhu ointment group significantly decreased(P<0.01).Real-time PCR results showed the levels of inflammatory factors IL-1β,IL-6,TNF-α,IL-17 and chemokines IP-10,IL-8,MCP-1 in wound tissue of diabetic foot ulcer rat model were significantly high-er than those in the normal ulcer model(P<0.01),while transforming growth factor TGF-βlevel was lower than that in the nor-mal ulcer model(P<0.01).Compared to the diabetic foot ulcer control group,the Notch signal inhibitor LY-411575 significantly reduced the level of inflammatory factors IL-1β,IL-6,TNF-α,IL-17 and chemokines IP-10,IL-8,MCP-1(P<0.01),and in-creased transforming growth factors TGF-αand TGF-βexpression in the diabetic foot ulcer rats(P<0.05),while the Notch sig-nal agonist Jagged1 significantly decreased the expression of TGF-αand TGF-β(P<0.05),which indicated inhibition of Notch signaling pathway could reduce the levels of the inflammatory factors IL-1β,IL-6,TNF-α,IL-17,IL-8 and increase transforming growth factors TGF-α,TGF-βexpression.Compared to the diabetic foot ulcer control group,Chinese medicine Zizhu ointment significantly reduced the levels of inflammatory factors IL-1β,IL-6,TNF-α,IL-17 and chemokines IP-10,IL-8,MCP-1 in the diabetic foot ulcer rats(P<0.01)and increased transforming growth factors TGF-α,TGF-βexpression(P<0.01).The Notch signal inhibitor LY-411575 enhanced the effect of Zizhu ointment on the expression of TGF-αand TGF-β.Conclusion:The dia-betic foot ulcer rat model has a higher expression of inflammatory factors and chemokines than the normal ulcer model,but the ex-pression of transforming growth factors is insufficient.Inhibition of the Notch signaling pathway can reduce the expression of in-flammatory factors and chemokines,and increase transforming growth factors in the experimental diabetic foot ulcer rats.The Notch signaling pathway has pro-inflammatory and partial cell proliferation effects.Zizhu ointment can reduce the levels of inflam-matory factors and chemokine chemotaxis in the diabetic foot ulcer rats,increase the expression of transforming growth factors,and reduce the inflammatory response on the wound.The role of Zizhu ointment may be related to the inhibition of Nocth signaling expression.