Nankai University School of Medicine, Tianjin, PR China; Department of Plastic and Reconstructive Surgery, PLA General Hospital, Beijing 100853, PR China.
Department of Plastic and Reconstructive Surgery, PLA General Hospital, Beijing 100853, PR China; Medical School of Chinese PLA, Beijing 100853, PR China.
Biochem Biophys Res Commun. 2018 Jun 2;500(2):310-317. doi: 10.1016/j.bbrc.2018.04.065. Epub 2018 Apr 14.
Mesenchymal stem cell (MSC)-derived exosomes have been recognized as new candidates for the treatment of ischemic disease or injury and may be an alternative treatment for cell therapy. This aim of the study was to evaluate whether exosomes derived from adipose mesenchymal stem cell (ADSC) can protect the skin flap during ischemia-reperfusion (I/R) injury and induce neovascularization.
To investigate the effects of exosomes in the I/R injury of flap transplantation in vivo, flaps were subjected to 6 h of ischemia by ligating the left superficial inferior epigastric vessels (SIEA) followed by blood perfusion. Exosomes derived from normal ADSC (ADSC-exos) and exosomes derived from ADSC preconditioned with HO (HO-ADSC-exos) were injected into the flaps. Then, the blood perfusion unit (BPU) of the flaps was measured by Laser Doppler Perfusion Imaging (LDPI) and microvessel density was determined by the endothelial with cell marker CD31 with Immunohistochemistry (IHC) staining. Inflammatory cell infiltration of the skin flap and apoptosis were detected by hematoxylin & eosin staining (H&E) and the TdT-mediated biotinylated dUTP nick end-labeling (TUNEL) technique.
In vivo, exosomes significantly increased flap survival and capillary density compared to I/R on postoperative day 5, and decreased the inflammatory reaction and apoptosis in the skin flap (P < 0.05). Furthermore, HO-ADSC-exos had better outcomes compared to normal exosomes (P < 0.05). ADSC-exos could significantly increase human umbilical vein endothelial cell (HUVEC) proliferation (P < 0.05), but no statistic difference was found in exosomes derived from different microenvironments (P > 0.05). HUVEC co-cultured with HO-ADSC-exos increased the migration ratio and generated more cord-like structures compared to ADSC-exos and the control group (P < 0.05).
ADSC-exos can enhance skin flap survival, promote neovascularization and alleviate the inflammation reaction and apoptosis in the skin flap after I/R injury. The use of a specific microenvironment for in vitro stem cell culture, such as one containing a low concentration of HO, will facilitate the development of customized exosomes for cell-free therapeutic applications in skin flap transplantation.
间充质干细胞（MSC）衍生的外泌体已被认为是治疗缺血性疾病或损伤的新候选药物，可能是细胞治疗的替代方法。本研究旨在评估脂肪间充质干细胞（ADSC）衍生的外泌体是否可以在缺血再灌注（I/R）损伤期间保护皮瓣，并诱导血管生成。
为了研究外泌体在皮瓣移植 I/R 损伤中的体内作用，通过结扎左侧腹壁下动脉（SIEA）使皮瓣缺血 6 小时，然后再进行血液灌注。将来自正常 ADSC 的外泌体（ADSC-exos）和经过 HO 预处理的 ADSC 衍生的外泌体（HO-ADSC-exos）注射到皮瓣中。然后，通过激光多普勒灌注成像（LDPI）测量皮瓣的血流灌注单位（BPU），并用内皮细胞标志物 CD31 通过免疫组织化学（IHC）染色来确定微血管密度。通过苏木精和伊红染色（H&E）和末端转移酶介导的生物素化 dUTP 缺口末端标记（TUNEL）技术检测皮瓣的炎性细胞浸润和细胞凋亡。
在体内，与术后第 5 天的 I/R 相比，外泌体显著增加了皮瓣的存活率和毛细血管密度，并减少了皮瓣中的炎症反应和细胞凋亡（P<0.05）。此外，与正常外泌体相比，HO-ADSC-exos 具有更好的效果（P<0.05）。ADSC-exos 可显著增加人脐静脉内皮细胞（HUVEC）的增殖（P<0.05），但来自不同微环境的外泌体之间没有统计学差异（P>0.05）。与 ADSC-exos 和对照组相比，与 HO-ADSC-exos 共培养的 HUVEC 迁移率更高，形成的管状结构更多（P<0.05）。
ADSC-exos 可提高皮瓣存活率，促进血管生成，并减轻 I/R 损伤后皮瓣的炎症反应和细胞凋亡。在体外干细胞培养中使用特定的微环境，例如含有低浓度 HO 的微环境，将有助于开发用于皮瓣移植的无细胞治疗应用的定制外泌体。
