紫朱软膏对金黄色葡萄球菌感染糖尿病足溃疡炎症反应及NF-κB/JNK/PI3K信号表达的影响

Effect of Zizhu Ointment on inflammatory response and NF-kB/JNK/PI3K signal expression of diabetic foot ulcer infected by staphylococcus aureus

关键词:  紫朱软膏  糖尿病足溃疡  金黄色葡萄球菌  炎症  汞  NF-κB

Key words:  Zizhu Ointment  diabetic foot ulcer  staphylococcus aureus  inflammation  mercury  NF-κB

摘要:

[目的] 糖尿病足溃疡是糖尿病的严重的下肢并发症，感染对溃疡的愈合具有明显的延迟作用。金黄色葡萄球菌是糖尿病足溃疡常见的致病菌。感染导致的持续的炎症是造成糖尿病足溃疡迁延难愈的重要因素。本研究旨在从感染性炎症反应的角度探讨中药紫朱软膏在金黄色葡萄球菌感染糖尿病足溃疡模型愈合中的作用机制。[方法] 采用高脂饲料配合链脲佐菌素（STZ）造成糖尿病，恒温恒压烫伤仪建立糖尿病足溃疡实验模型，通过皮下标准菌株注射造成感染，制成实验模型，以普通溃疡模型与实验模型作为对照研究感染性糖尿病足溃疡炎症反应的特点，通过对实验模型的干预对照包括生理盐水、西药贝复济、不同浓度梯度的紫朱软膏干预14 d，采用ELISA法检测血清一氧化氮（NO），C-反应蛋白（CRP）和白细胞介素-6（IL-6），蛋白质印迹法（Western blot）检测核因子-κB p65（NF-κB p65）、c-Jun N-末端激酶（JNK）和磷脂酰肌醇3-激酶（PI3K）信号通路信号分子，免疫组化法检测创面组织的血管内皮生长因子（VEGF）和血小板内皮细胞黏附分子-34（CD34），冷原子吸收法检测血汞（Hg）含量，研究紫朱软膏对金黄色葡萄球菌感染糖尿病足溃疡炎症的作用及药物安全性。[结果] 金黄色葡萄球菌感染的大鼠糖尿病足溃疡模型较普通溃疡模型炎症因子表达升高，紫朱软膏能够降低感染性糖尿病足溃疡大鼠炎症因子（NO，CRP和IL-6）表达。感染性糖尿病足溃疡模型中NF-κB、JNK、PI3Ks炎性信号通路激活，紫朱软膏能明显抑制JNK和NF-κB P65、JNK、PI3K的磷酸化，降低创面炎症反应，同时增加创面VEGF的表达，促进创面血管新生，同时对紫朱软膏用药大鼠血液汞含量的检测说明药物具有较好的安全性。[结论] 金黄色葡萄球菌感染的大鼠糖尿病足溃疡模型较普通溃疡模型炎症水平更高；紫朱软膏能够通过降低金葡菌感染的糖尿病足溃疡炎症水平、抑制炎症NF-κB、JNK、PI3K等炎症相关信号通路激活、增加血管生成因子CD34和VEGF刺激创面血管新生等途径促进组织修复，改善创面愈合，显示出紫朱软膏是一种安全有效的治疗糖尿病足溃疡的药物，具有良好的应用前景。

Abstract:

[Objective] Diabetic foot ulcer is a serious lower extremity complication of diabetes. The long-term treatment of diabetic foot ulcer is prone to secondary bacterial infection. Infection has a significant delay effect on the healing of ulcer. Staphylococcus aureus is a common infection pathogen of diabetic foot ulcers. Persistent inflammation caused by infection is an important factor that leads to the continuation of diabetic foot ulcers,which is also a clinical problem in the treatment of diabetic foot ulcers. Zizhu Ointment is an effective prescription for the treatment of diabetic foot ulcers in our hospital. This study aims to explore the mechanism of the effect of traditional Chinese medicine Zizhu ointment on the healing of diabetic foot ulcer model infected with Staphylococcus aureus from the perspective of infectious inflammation.[Methods] Using high fat feed composition chain urea with cephalosporins (STZ) caused by diabetes,constant temperature constant pressure burns instrument experimental diabetic foot ulcer model was established,through the subcutaneous standard strains caused by infection,made from experimental models,ordinary ulcer model in comparison with the experimental model with infective diabetic foot ulcer inflammatory response characteristics,through the contrast of interference from the experimental models including normal saline,Western medicine Bei Fuji,different concentration gradient of Zizhu Ointment intervention for 14 days,using ELISA method to detect serum nitric oxide (NO),C-reactive protein (CRP) and interleukin 6 (IL-6),Western blot (WB) to detect nuclear factor-kappa Bp65 predominate (p65 nf-kappa B),c-Jun-N-terminal kinase (JNK) and phosphatidyl inositol 3 kinase (PI3K) path way signal molecules,immunohistochemical method to detect wound tissue of vascular endothelial growth factor (VEGF) and platelet endothelial cell adhesion molecule,34 (CD34),cold atomic absorption method to detect the contents of blood mercury (Hg),study the Zizhu Ointment to staphylococcus aureus infection of diabetic foot ulcer inflammation effect and drug safety.[Results] The expression of inflammatory factors in the diabetic foot ulcer model with Staphylococcus aureus infection was higher than that in the normal ulcer model,and Zizhu Ointment could reduce the expression of inflammatory factors (NO,CRP and IL-6) in the diabetic foot ulcer model. Infective diabetic foot ulcer model in the nf-kappa B,JNK,PI3Ks inflammatory signaling pathways activated,Zizhu Ointment can obviously inhibit the JNK and nf-kappa B P65,JNK,phosphorylation of PI3K,reduce wound inflammation,at the same time increase the wound the expression of VEGF,promote angiogenesis,as well as the Zizhu Ointment medication rats blood mercury levels of testing instructions medication has good security.[Conclusion] The diabetic foot ulcer model of rats infected with Staphylococcus aureus has a higher level of inflammation than ordinary ulcer models;Zizhu ointment can reduce the inflammation level of diabetic foot ulcers infected by Staphylococcus aureus and inhibit the inflammation inflammation-related signaling pathway activation,increased angiogenesis factors CD34 and VEGF stimulate wound angiogenesis to promote tissue repair and improve wound healing,showing that Zizhu ointment is a safe and effective drug for treating diabetic foot ulcers and has good application prospects.