Saraf Namita, Ramachandran Rajalakshmy Ayilam, Cao Mou, Lemoff Andrew, Baniasadi Hamid, Robertson Danielle M
Department of Ophthalmology, UT Southwestern Medical Center, Dallas, TX, USA.
Department of Ophthalmology, UT Southwestern Medical Center, Dallas, TX, USA.
Ocul Surf. 2024 Oct;34:317-325. doi: 10.1016/j.jtos.2024.08.009. Epub 2024 Aug 17.
Autologous serum is widely used for the treatment of severe ocular surface disease with mixed efficacy. Extracellular vesicles (EVs) are small membrane bound structures present in all body fluids, including serum. This study compared the proteomic, metabolomic, and inflammatory cytokine composition of serum-derived EVs (SDEVs) to that of the soluble free protein fraction and the subsequent capacity of SDEVs to induce corneal epithelial cell migration and inflammation.
SDEVs were isolated from human serum using size exclusion chromatography. SDEVs were analyzed using nanoparticle tracking analysis, transmission electron microscopy, and western blotting. The effects of SDEVs on corneal epithelial cell migration were tested using a standard scratch assay. Inflammatory cytokines in SDEVs and the free protein fraction were quantified using a microarray. A mutli-omics approach was further used to define SDEV cargo. The ability of SDEVs to modulate inflammation in corneal epithelial cells was quantified using ELISAs.
Western blot and TEM confirmed the presence of SDEVs. Proinflammatory cytokines, along with complement proteins and TGF-β, were decreased in SDEVs compared to serum. Metabolites present in SDEVs were mostly involved in amino acid biosynthesis, the TCA cycle and oxidative phosphorylation. SDEVs exhibited pro-migratory effects similar to serum however, SDEVs did not induce secretion of IL-6 or IL-8.
SDEVs exhibit reduced levels of pro-inflammatory cytokines while retaining the beneficial wound healing properties of serum. Unlike serum, SDEVs do not induce inflammation. SDEVs may represent an alternative option for patients with severe ocular surface disease where traditional autologous serum has failed.
自体血清被广泛用于治疗严重眼表疾病，但疗效不一。细胞外囊泡（EVs）是存在于包括血清在内的所有体液中的小膜结合结构。本研究比较了血清来源的细胞外囊泡（SDEVs）与可溶性游离蛋白组分的蛋白质组学、代谢组学和炎性细胞因子组成，以及SDEVs随后诱导角膜上皮细胞迁移和炎症的能力。
使用尺寸排阻色谱法从人血清中分离SDEVs。使用纳米颗粒跟踪分析、透射电子显微镜和蛋白质印迹法对SDEVs进行分析。使用标准划痕试验测试SDEVs对角膜上皮细胞迁移的影响。使用微阵列对SDEVs和游离蛋白组分中的炎性细胞因子进行定量。进一步采用多组学方法来确定SDEVs的货物。使用酶联免疫吸附测定法对SDEVs调节角膜上皮细胞炎症的能力进行定量。
蛋白质印迹和透射电子显微镜证实了SDEVs的存在。与血清相比，SDEVs中的促炎细胞因子以及补体蛋白和转化生长因子-β减少。SDEVs中存在的代谢物大多参与氨基酸生物合成、三羧酸循环和氧化磷酸化。SDEVs表现出与血清相似的促迁移作用，然而，SDEVs不会诱导白细胞介素-6或白细胞介素-8的分泌。
SDEVs中促炎细胞因子水平降低，同时保留了血清有益的伤口愈合特性。与血清不同，SDEVs不会诱导炎症。对于传统自体血清治疗失败的严重眼表疾病患者，SDEVs可能是一种替代选择。
