炎症激活间充质干细胞源微泡对糖尿病慢性创面愈合的促进作用观察

Lipopolysaccharides priming mesenchymal stem cells accelerate diabetic wound healing via exosomes

目的探讨炎症激活条件下间充质干细胞分泌的微泡(LPS-exosomes)对糖尿病慢性创面的治疗作用。方法用终浓度为100ng/ml的脂多糖预处理间充质干细胞2d,收集上清液,利用超速梯度离心法分离提取微泡,并对其进行鉴定。建立糖尿病大鼠皮肤缺损模型,将实验大鼠随机分为3组:未治疗组、间充质干细胞源微泡治疗组、炎症激活间充质干细胞源微泡治疗组,分次多点注射微泡60μg于伤口周边,1次/d,连续10d。于治疗后第3、7、14天进行创面大体观察,q RT-PCR法检测创面炎症相关因子表达水平及巨噬细胞亚型表面标记物的表达改变。结果与未治疗组相比,炎症激活间充质干细胞源微泡治疗组创面愈合速度明显加快,治疗后第7、14天促炎因子IL-1、IL-12及M1型巨噬细胞表面标记物i NOS表达水平明显降低,同时抗炎因子IL-10、TGF-β及M2型巨噬细胞表面标记物CD163表达明显增加,差异均有统计学意义(P<0.05)。结论炎症激活间充质干细胞源微泡可能通过调节巨噬细胞表型改变,抑制创面局部慢性炎症反应,进而加速糖尿病皮肤创面的愈合。

Objective To study the therapeutic effect of exosome derived from lipopolysaccharides（LPS） priming mesenchymal stem cells（MSCs） for diabetic wound healing.Methods Human umbilical cord MSCs were treated with LPS（100ng/ml） for 2 days,the supernatant were then collected,and exosomes were harvested by density gradient centrifugation and identified.Diabetic cutaneous wounds were prepared and the animals were divided into the following three groups：control group,untreated MSCs derived exosome（un-exosome） treatment group and LPS primed MSCs derived exosome（LPS-exosome） treatment group.Exosomes（60μg） were injected dispersively into the wound edge daily for 10 days.After treatment,the therapeutic results were evaluated by gross obser vation of the wounds,the expression levels of inflammation related factors and macrophage subtype markers in the injured sites were detected by q RT-PCR at day 3,7 and 14 after treatment.Results Compared with control group,the diabetic wound healing was obviously improved in LPS-exosome treatment group after treatment for 7 and 14 days,with faster wound close,depressed expression of pro-inflammatory factors IL-1,IL-12 and M1 macrophage surface marker i NOS,and upregulation of anti-inflammatory factors IL-10,TGF-β and M2 macrophage surface marker CD163,the differences were significant（P〈0.05）.Conclusions LPS-exosome may balance macrophage plasticity,restrain chronic inflammation and accelerate diabetic cutaneous wound healing.