[通过不同途径应用人脐带间充质干细胞外泌体治疗小鼠全厚皮肤缺损伤口的效果]。

H. Y. Wang, T. Ba, B Zhou, Z. Q. Yan, R. J. Wang, L. Y. Liu
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According to the random number table, the injured mice were divided into control group (without drug administration), local wound application group, wound margin subcutaneous injection group, and tail vein injection group (with 30 mice in each group). Mice in the latter three groups were given 0.2 mL phosphate buffer solution containing 200 μg hUCMSC exosomes by local wound application, subcutaneous injection at the wound margin, and tail vein injection, respectively. On post injury day (PID) 7, 14, and 21, the general condition of the wound was observed, and the wound healing rate was calculated; the wound tissue was collected, the pathological changes and collagen fibers were observed respectively by hematoxylin-eosin staining and Masson staining, the number of new microvessels was observed by CD31 immunohistochemical staining, and the content of tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) was detected by enzyme-linked immunosorbent assay. The sample number was 10 in each group at each time point. Results: On PID 7, 14, and 21, the wounds of mice in the 4 groups all healed gradually, and the wound healing of the mice in wound margin subcutaneous injection group was the best; the wound healing rates of mice in the three administration groups were significantly higher than those in control group (P<0.05), the wound healing rates of mice in wound margin subcutaneous injection group and tail vein injection group were significantly higher than those in local wound application group (P<0.05), and the wound healing rates of mice in wound margin subcutaneous injection group were significantly higher than those in tail vein injection group (P<0.05). On PID 7, 14, and 21, the growth and epithelialization speed of the wound tissue of mice in the three administration groups were significantly accelerated, and the collagen fibers in the wounds of mice in the three administration groups were larger in number and more neatly arranged in comparison with the control group. On PID 7, 14, and 21, under every 200-fold visual field, the number of new microvessels in the wound tissue of mice in local wound application group was 24.1±2.5, 50.7±4.1, and 44.2±2.3, respectively, the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group was 32.2±2.9, 67.5±4.9, and 53.6±3.7, respectively, and the number of new microvessels in the wound tissue of mice in tail vein injection group was 27.8±2.4, 59.1±3.7, and 49.6±2.6, respectively, which was significantly more than 20.6±1.7, 46.7±3.4, and 40.9±2.8 in control group (P<0.05); the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group and tail vein injection group was significantly more than that in local wound application group (P<0.05); the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group was significantly more than that in tail vein injection group (P<0.05). On PID 7, 14, and 21, the content of TNF-α and IL-6 in the wound tissue of mice in the three administration groups was significantly less than that in control group (P<0.05), the content of TNF-α and IL-6 in the wound tissue of mice in wound margin subcutaneous injection group and tail vein injection group was significantly less than that in local wound application group (P<0.05), and the content of TNF-α and IL-6 in the wound tissue of mice in wound margin subcutaneous injection group was significantly less than that in tail vein injection group (P<0.05). Conclusions: Local wound application, subcutaneous injection at the wound margin, and tail vein injection of hUCMSC exosomes can all promote the wound healing of full-thickness skin defects in mice through alleviating excessive inflammatory response and promoting angiogenesis. 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According to the random number table, the injured mice were divided into control group (without drug administration), local wound application group, wound margin subcutaneous injection group, and tail vein injection group (with 30 mice in each group). Mice in the latter three groups were given 0.2 mL phosphate buffer solution containing 200 μg hUCMSC exosomes by local wound application, subcutaneous injection at the wound margin, and tail vein injection, respectively. On post injury day (PID) 7, 14, and 21, the general condition of the wound was observed, and the wound healing rate was calculated; the wound tissue was collected, the pathological changes and collagen fibers were observed respectively by hematoxylin-eosin staining and Masson staining, the number of new microvessels was observed by CD31 immunohistochemical staining, and the content of tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) was detected by enzyme-linked immunosorbent assay. 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On PID 7, 14, and 21, the growth and epithelialization speed of the wound tissue of mice in the three administration groups were significantly accelerated, and the collagen fibers in the wounds of mice in the three administration groups were larger in number and more neatly arranged in comparison with the control group. On PID 7, 14, and 21, under every 200-fold visual field, the number of new microvessels in the wound tissue of mice in local wound application group was 24.1±2.5, 50.7±4.1, and 44.2±2.3, respectively, the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group was 32.2±2.9, 67.5±4.9, and 53.6±3.7, respectively, and the number of new microvessels in the wound tissue of mice in tail vein injection group was 27.8±2.4, 59.1±3.7, and 49.6±2.6, respectively, which was significantly more than 20.6±1.7, 46.7±3.4, and 40.9±2.8 in control group (P<0.05); the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group and tail vein injection group was significantly more than that in local wound application group (P<0.05); the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group was significantly more than that in tail vein injection group (P<0.05). On PID 7, 14, and 21, the content of TNF-α and IL-6 in the wound tissue of mice in the three administration groups was significantly less than that in control group (P<0.05), the content of TNF-α and IL-6 in the wound tissue of mice in wound margin subcutaneous injection group and tail vein injection group was significantly less than that in local wound application group (P<0.05), and the content of TNF-α and IL-6 in the wound tissue of mice in wound margin subcutaneous injection group was significantly less than that in tail vein injection group (P<0.05). Conclusions: Local wound application, subcutaneous injection at the wound margin, and tail vein injection of hUCMSC exosomes can all promote the wound healing of full-thickness skin defects in mice through alleviating excessive inflammatory response and promoting angiogenesis. 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引用次数: 0

摘要

目的研究人脐带间充质干细胞(hUCMSC)外泌体通过伤口局部涂抹、伤口边缘皮下注射和尾静脉注射治疗小鼠全厚皮肤缺损伤口的效果,并探索 hUCMSC 外泌体治疗伤口的最佳给药途径。研究方法从内蒙古包钢医院妇产科 3 名 25-35 岁正常分娩妇女的废弃脐带组织中提取 hUCMSC 外泌体并成功鉴定。选取 120 只 6-8 周龄的雄性 BALB/c 小鼠,在其背部制备全厚皮肤缺损创面。根据随机数字表将受伤小鼠分为对照组(不给药)、伤口局部涂药组、伤口边缘皮下注射组和尾静脉注射组(每组 30 只)。后三组小鼠分别接受 0.2 mL 含有 200 μg hUCMSC 外泌体的磷酸盐缓冲液局部伤口涂抹、伤口边缘皮下注射和尾静脉注射。在损伤后第 7、14 和 21 天,观察伤口的总体情况,并计算伤口愈合率;收集伤口组织,分别用苏木精-伊红染色法和马森染色法观察病理变化和胶原纤维,用CD31免疫组化染色法观察新生微血管的数量,用酶联免疫吸附法检测肿瘤坏死因子α(TNF-α)和白细胞介素-6(IL-6)的含量。每个时间点每组 10 个样本。结果PID7、14、21时,4组小鼠伤口均逐渐愈合,以创缘皮下注射组小鼠伤口愈合最好;3个给药组小鼠伤口愈合率均显著高于对照组(P<0.05),创缘皮下注射组和尾静脉注射组小鼠的伤口愈合率明显高于局部伤口应用组(P<0.05),创缘皮下注射组小鼠的伤口愈合率明显高于尾静脉注射组(P<0.05)。在 PID 7、14 和 21 日,三个给药组小鼠伤口组织的生长和上皮化速度明显加快,且与对照组相比,三个给药组小鼠伤口中的胶原纤维数量更多,排列更整齐。PID7、14和21日,在每200倍视野下,局部伤口给药组小鼠伤口组织中新生微血管的数量分别为(24.1±2.5)、(50.7±4.1)和(44.2±2.3,创缘皮下注射组小鼠创面组织新生微血管数分别为(32.2±2.9)、(67.5±4.9)、(53.6±3.7),尾静脉注射组小鼠创面组织新生微血管数分别为(27.8±2.4)、(59.1±3.7)、(49.6±2.6,分别明显多于对照组的20.6±1.7、46.7±3.4和40.9±2.8(P<0.05);创缘皮下注射组和尾静脉注射组小鼠创面组织新生微血管数明显多于创面局部应用组(P<0.05);创缘皮下注射组小鼠创面组织新生微血管数量明显多于尾静脉注射组(P<0.05)。PID7、14和21日,三组小鼠创面组织中TNF-α和IL-6的含量均显著低于对照组(P<0.05),创缘皮下注射组和尾静脉注射组小鼠创面组织中TNF-α和IL-6的含量显著低于创面局部涂药组(P<0.05),创缘皮下注射组小鼠创面组织中 TNF-α 和 IL-6 的含量明显低于尾静脉注射组(P<0.05)。结论局部伤口涂抹、伤口边缘皮下注射和尾静脉注射 hUCMSC 外泌体都能通过缓解过度炎症反应和促进血管生成来促进小鼠全厚皮肤缺损的伤口愈合。其中,伤口边缘皮下注射的治疗效果更好,表明伤口边缘皮下注射是 hUCMSC 外泌体治疗伤口的最佳给药途径。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
[Effects of applying human umbilical cord mesenchymal stem cell exosomes through different pathways to treat full-thickness skin defect wounds in mice].
Objective: To investigate the effects of human umbilical cord mesenchymal stem cell (hUCMSC) exosomes in the treatment of full-thickness skin defect wounds in mice through local wound application, subcutaneous injection at the wound margin, and tail vein injection, and to explore the optimal administration route of hUCMSC exosomes for wound treatment. Methods: This study was an experimental study. hUCMSC exosomes were extracted from the discarded umbilical cord tissue of three normal delivery women aged 25-35 years in the Department of Obstetrics and Gynecology of Baogang Hospital of Inner Mongolia and successfully identified. Totally 120 male BALB/c mice aged 6-8 weeks were selected, and full-thickness skin defect wounds were prepared on the back of them. According to the random number table, the injured mice were divided into control group (without drug administration), local wound application group, wound margin subcutaneous injection group, and tail vein injection group (with 30 mice in each group). Mice in the latter three groups were given 0.2 mL phosphate buffer solution containing 200 μg hUCMSC exosomes by local wound application, subcutaneous injection at the wound margin, and tail vein injection, respectively. On post injury day (PID) 7, 14, and 21, the general condition of the wound was observed, and the wound healing rate was calculated; the wound tissue was collected, the pathological changes and collagen fibers were observed respectively by hematoxylin-eosin staining and Masson staining, the number of new microvessels was observed by CD31 immunohistochemical staining, and the content of tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) was detected by enzyme-linked immunosorbent assay. The sample number was 10 in each group at each time point. Results: On PID 7, 14, and 21, the wounds of mice in the 4 groups all healed gradually, and the wound healing of the mice in wound margin subcutaneous injection group was the best; the wound healing rates of mice in the three administration groups were significantly higher than those in control group (P<0.05), the wound healing rates of mice in wound margin subcutaneous injection group and tail vein injection group were significantly higher than those in local wound application group (P<0.05), and the wound healing rates of mice in wound margin subcutaneous injection group were significantly higher than those in tail vein injection group (P<0.05). On PID 7, 14, and 21, the growth and epithelialization speed of the wound tissue of mice in the three administration groups were significantly accelerated, and the collagen fibers in the wounds of mice in the three administration groups were larger in number and more neatly arranged in comparison with the control group. On PID 7, 14, and 21, under every 200-fold visual field, the number of new microvessels in the wound tissue of mice in local wound application group was 24.1±2.5, 50.7±4.1, and 44.2±2.3, respectively, the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group was 32.2±2.9, 67.5±4.9, and 53.6±3.7, respectively, and the number of new microvessels in the wound tissue of mice in tail vein injection group was 27.8±2.4, 59.1±3.7, and 49.6±2.6, respectively, which was significantly more than 20.6±1.7, 46.7±3.4, and 40.9±2.8 in control group (P<0.05); the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group and tail vein injection group was significantly more than that in local wound application group (P<0.05); the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group was significantly more than that in tail vein injection group (P<0.05). On PID 7, 14, and 21, the content of TNF-α and IL-6 in the wound tissue of mice in the three administration groups was significantly less than that in control group (P<0.05), the content of TNF-α and IL-6 in the wound tissue of mice in wound margin subcutaneous injection group and tail vein injection group was significantly less than that in local wound application group (P<0.05), and the content of TNF-α and IL-6 in the wound tissue of mice in wound margin subcutaneous injection group was significantly less than that in tail vein injection group (P<0.05). Conclusions: Local wound application, subcutaneous injection at the wound margin, and tail vein injection of hUCMSC exosomes can all promote the wound healing of full-thickness skin defects in mice through alleviating excessive inflammatory response and promoting angiogenesis. Among them, subcutaneous injection at the wound margin has a better therapeutic effect, indicating subcutaneous injection at the wound margin is the optimal administration route for hUCMSC exosomes in wound treatment.
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