康复新凝胶剂对小鼠创面组织修复的作用及相关机制探讨

Study on the Effect and Related Mechanism of Kangfuxin Gel on the Repair of Mice Wound Tissue

目的:观察康复新凝胶剂对小鼠皮肤创面损伤的作用,并探讨其可能的作用机制。方法:60只C57BL/6J小鼠随机分成模型组、康复新凝胶剂组(凝胶剂组)、林可霉素利多卡因凝胶剂组(林可霉素组),每组20只,采用全层皮肤切除法在C57BL/6J小鼠背部制备伤口,伤口造模后开始按组对应给药10d;于治疗第3、7、10天拍照观察伤口图像并计算创面愈合率;酶联免疫吸附法(ELISA)检测血液中白细胞介素-6(IL-6)与转化生长因子-β(TGF-β)的含量;于治疗后第10天处死各组小鼠,苏木精-伊红染色法(HE)观察创面组织变化;免疫组化染色检测创面组织血管内皮生长因子(VEGF)、M1型巨噬细胞标志物(iNOS)与M2型巨噬细胞标志物(CD206)蛋白表达;荧光定量PCR(qRT-PCR)检测创面组织中IL-6、TGF-β、肿瘤坏死因子-α(TNF-α)与白细胞介素-10(IL-10)mRNA表达水平。结果:与模型组比较,康复新凝胶剂组小鼠于治疗后第3、7、10天伤口愈合较为明显,创面愈合率显著增加,差异有统计学意义(P<0.05);血清中IL-6含量显著下降且TGF-β含量显著升高(P<0.05)。治疗10d后,与模型组比较,康复新凝胶剂组小鼠创面炎性细胞浸润明显,新生致密肉芽组织,有大量新生毛细血管形成,同时VEGF阳性表达率显著增加(P<0.05),IL-6、TNF-αmRNA表达水平显著下降(P<0.05),IL-10、TGF-βmRNA表达水平显著升高(P<0.05),iNOS阳性细胞数显著减少而CD206阳性细胞数显著增加(P<0.05),且与林可霉素利多卡因凝胶剂作用等效。结论:康复新凝胶剂能够促进小鼠伤口愈合,减轻伤口部位炎症反应,其机制可能与创面巨噬细胞表型转化有关。

Objective To observe the effect of Kangfuxin Gel on skin wounds in mice and explore its possible mechanism.Methods 60 C57BL/6J mice were randomly divided into the model group,the Kangfuxin gel group(gel group)and the lincomycin lindocaine gel group(lincomycin group),20 mice in each group.Used full-thickness skin excision method to prepare wounds on the back of C57BL/6J mice.After wound modeling,start to administer drugs according to the group for 10 days.Taked pictures and observe wound images on the 3,7 and 10 days of treatment and calculate the wound healing rate.Detectted the levels of interleukin-6(IL-6)and transforming growth factor-β(TGF-β)in the blood by enzyme-linked immunosorbent assay(ELISA).The mice of each group were sacrificed on the 10th day after treatment.Hematoxylin-Eosin staining method(HE)observes the changes of wound tissue.Immunohistochemical staining was used to detect the protein expression of vascular endothelial growth factor(VEGF),M1 macrophage marker(iNOS)and M2 macrophage marker(CD206)in wound tissue.Fluorescence quantitative PCR(qRT-PCR)was used to detect the mRNA expression levels of IL-6,TGF-β,tumor necrosis factor-α(TNF-α)and interleukin-10(IL-10)in wound tissues.Results Compared with the model group,mice in the Kangfuxin gel group had more obvious wound healing on the 3,7 and 10 days after treatment,the wound healing rate increased significantly,the differences were statistically significant(P<0.05).The serum IL-6 content decreased significantly and the content of TGF-βincreased significantly(P<0.05).After 10 days of treatment,compared with the model group,the inflammatory cell infiltration in the wound surface of the Kangfuxin gel group was obvious,new dense granulation tissue,a large number of new capillaries were formed,and the VEGF positive expression rate was significantly increased(P<0.05),IL-6 and TNF-αmRNA expression levels decreased significantly(P<0.05),IL-10 and TGF-βmRNA expression levels increased significantly(P<0.05),the number of iNOS positive cells decreased significantly while the number of CD206 positive cells increased significantly(P<0.05),and was equivalent to lincomycin and lindocaine gel.Conclusion Kangfuxin gel can promote wound healing in mice and reduce the inflammatory response at the wound site.The mechanism may be related to the phenotypic transformation of macrophages on the wound surface.