背景:促进受损的前交叉韧带(ACL)组织的愈合过程对于患者安全恢复运动至关重要。干细胞来源的外泌体已显示出增强受损肌腱/韧带再生的积极作用。然而,外泌体在储存和预组装方面的临床应用具有挑战性.我们假设来自人脐带干细胞(hUSC-EX)的冻干外泌体可以增强慢性损伤的ACL细胞的细胞活性。
方法:我们在IACUC下从兔中收获了8周损伤的ACL细胞(编号:110232)批准。从人脐带干细胞的培养基中纯化研究的外泌体(IRB批准号A202205014),冻干储存,和水合使用。我们比较了来自相同兔的外泌体处理的细胞与非外泌体处理的细胞(对照组)。我们检查了细胞活力,扩散,I型和III型胶原蛋白的迁移能力和基因表达,TGFβ,VEGF,hUSC-EX治疗后8周损伤的ACL细胞的肌腱发生。
结果:水合后,HUSC-EX的平均尺寸为84.5±70.6nm,细胞的Alix检测呈阳性,TSG101、CD9、CD63和CD81蛋白,但α-微管蛋白阴性。治疗24小时后,hUSC-EX显著提高了细胞活力,与无外泌体治疗组相比,8周损伤ACL细胞的增殖和迁移能力。此外,胶原蛋白合成的表达,TGFβ,VEGF,在24hhUSC-EX递送后8周损伤的ACL细胞中,肌腱发生基因均显着增加。
结论:冻干外泌体易于储存,水合后易于使用,从而保持它们的特性。用冻干的hUSC-EX处理改善了8周损伤的ACL细胞的活性和基因表达。
结论:冻干hUSC-EX保留了外泌体的特性,并能改善慢性损伤(8周)的ACL细胞。冻干的hUSC-EX可以作为有效和安全的生物材料,可以在室温下使用,以增强部分ACL撕裂患者和残余保留ACL重建后的细胞活性。
BACKGROUND: Facilitating the healing process of injured anterior cruciate ligament (ACL) tissue is crucial for patients to safely return to sports. Stem cell derived exosomes have shown positive effects on enhancing the regeneration of injured tendons/ligaments. However, clinical application of exosomes in terms of storage and pre-assembly is challenging. We hypothesized that lyophilized exosomes derived from human umbilical cord stem cells (hUSC-EX) could enhance the cell activity of chronically injured ACL cells.
METHODS: We harvested the 8 weeks injured ACL cells from rabbit under IACUC (No. 110232) approval. The studied exosomes were purified from the culture medium of human umbilical cord stem cells (IRB approval No. A202205014), lyophilized to store, and hydrated for use. We compared exosome treated cells with non-exosome treated cells (control group) from the same rabbits. We examined the cell viability, proliferation, migration capability and gene expression of type I and III collagen, TGFβ, VEGF, and tenogenesis in the 8 weeks injured ACL cells after hUSC-EX treatment.
RESULTS: After hydration, the average size of hUSC-EX was 84.5 ± 70.6 nm, and the cells tested positive for the Alix, TSG101, CD9, CD63, and CD81 proteins but negative for the α-Tubulin protein. After 24 h of treatment, hUSC-EX significantly improved the cell viability, proliferation and migration capability of 8 weeks injured ACL cells compared to that of no exosome treatment group. In addition, the expression of collagen synthesis, TGFβ, VEGF, and tenogenesis gene were all significantly increased in the 8 weeks injured ACL cells after 24 h hUSC-EX delivery.
CONCLUSIONS: Lyophilized exosomes are easily stored and readily usable after hydration, thereby preserving their characteristic properties. Treatment with lyophilized hUSC-EX improved the activity and gene expression of 8 weeks injured ACL cells.
CONCLUSIONS: Lyophilized hUSC-EX preserve the characteristics of exosomes and can improve chronically injured (8 weeks) ACL cells. Lyophilized hUSC-EX could serve as effective and safe biomaterials that are ready to use at room temperature to enhance cell activity in patients with partial ACL tears and after remnant preservation ACL reconstruction.