背景:由糖尿病引起的缺血性疾病继续对健康构成重大挑战,并且对有效治疗的需求很高。间充质干细胞(MSCs)来源的外泌体作为一种无细胞治疗缺血性疾病的方法引起了广泛的关注。然而,脂肪间充质干细胞外泌体(ADSC-Exos)治疗糖尿病下肢缺血性损伤的疗效尚不清楚.
方法:通过差异超速离心从ADSCs培养上清液中分离外泌体,并通过EdU评估其对C2C12细胞和HUVECs的影响,Transwell,和体外试管形成测定分开。通过激光多普勒灌注成像评估ADSC-Exos治疗后肢体功能的恢复,肢体功能评分,和组织学分析。随后,进行miRNA测序和挽救实验以找出ADSC-Exos对糖尿病后肢缺血性损伤的保护作用的相关miRNA。最后,通过生物信息学分析和双荧光素酶报告基因测定证实了miRNA在C2C12细胞中的直接靶标。
结果:ADSC-Exos具有促进C2C12细胞增殖和迁移以及促进HUVEC血管生成的潜力。体内实验表明ADSC-Exos可以保护缺血骨骼肌,促进肌肉损伤的修复,加速血管再生.结合生物信息学分析,miR-125b-5p可能是该过程中的关键分子。miR-125b-5p转移到C2C12细胞中能够通过抑制ACER2过表达来促进细胞增殖和迁移。
结论:研究结果表明,源自ADSC-Exos的miR-125b-5p可能通过靶向ACER2在缺血性肌肉修复中发挥关键作用。总之,我们的研究可能为ADSC-Exos作为糖尿病下肢缺血的治疗选择提供新的见解.
BACKGROUND: Ischemic diseases caused by diabetes continue to pose a major health challenge and effective treatments are in high demand. Mesenchymal stem cells (MSCs) derived exosomes have aroused broad attention as a cell-free treatment for ischemic diseases. However, the efficacy of exosomes from adipose-derived mesenchymal stem cells (ADSC-Exos) in treating diabetic lower limb ischemic injury remains unclear.
METHODS: Exosomes were isolated from ADSCs culture supernatants by differential ultracentrifugation and their effect on C2C12 cells and HUVECs was assessed by EdU, Transwell, and in vitro tube formation assays separately. The recovery of limb function after ADSC-Exos treatment was evaluated by Laser-Doppler perfusion imaging, limb function score, and histological analysis. Subsequently, miRNA sequencing and rescue experiments were performed to figure out the responsible miRNA for the protective role of ADSC-Exos on diabetic hindlimb ischemic injury. Finally, the direct target of miRNA in C2C12 cells was confirmed by bioinformatic analysis and dual-luciferase report gene assay.
RESULTS: ADSC-Exos have the potential to promote proliferation and migration of C2C12 cells and to promote HUVECs angiogenesis. In vivo experiments have shown that ADSC-Exos can protect ischemic skeletal muscle, promote the repair of muscle injury, and accelerate vascular regeneration. Combined with bioinformatics analysis, miR-125b-5p may be a key molecule in this process. Transfer of miR-125b-5p into C2C12 cells was able to promote cell proliferation and migration by suppressing ACER2 overexpression.
CONCLUSIONS: The findings revealed that miR-125b-5p derived from ADSC-Exos may play a critical role in ischemic muscle reparation by targeting ACER2. In conclusion, our study may provide new insights into the potential of ADSC-Exos as a treatment option for diabetic lower limb ischemia.