School of Health and Life Sciences, University of the West of Scotland, Hamilton International Technology Park, Lanarkshire Campus, Stephenson Place, South Lanarkshire, G72 0LH, Scotland, UK.
School of Infection and Immunity, University of Glasgow, Glasgow, G12 8TA, Scotland, UK.
Arthritis Res Ther. 2024 Sep 28;26(1):167. doi: 10.1186/s13075-024-03398-3.
Small extracellular vesicles (sEV) derived from synovial fibroblasts (SF) represent a novel molecular mechanism regulating cartilage erosion in osteoarthritis (OA). However, a comprehensive evaluation using disease relevant cells has not been undertaken. The aim of this study was to isolate and characterise sEV from OA SF and to look at their ability to regulate OA chondrocyte effector responses relevant to disease. Profiling of micro (mi) RNA signatures in sEV and parental OA SF cells was performed.
SF and chondrocytes were isolated from OA synovial membrane and cartilage respectively (n = 9). sEV were isolated from OA SF (± IL-1β) conditioned media by ultracentrifugation and characterised using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Particle size was confirmed by nanoparticle tracking analysis (NTA). sEV regulation of OA chondrocyte and cartilage effector response was evaluated using qPCR, ELISA and sulphated glycosaminoglycan assay (sGAG). RNA-sequencing was used to establish miRNA signatures in isolated sEV from OA SF.
OA SF derived sEV were readily taken up by OA chondrocytes, with increased expression of the catabolic gene MMP 13 (p < 0.01) and decreased expression of the anabolic genes aggrecan and COL2A1 (p < 0.01) observed. Treatment with sEV derived from IL-1β stimulated OA SF significantly decreased expression of aggrecan and COL2A1 (p < 0.001) and increased SOX 9 gene expression (p < 0.05). OA chondrocytes cultured with sEV from either non-stimulated or IL-1β treated OA SF, resulted in a significant increase in the secretion of IL-6, IL-8 and MMP-3 (p < 0.01). Cartilage explants cultured with sEV from SF (± IL-1β) had a significant increase in the release of sGAG (p < 0.01). miRNA signatures differed between parental SF cells and isolated sEV. The recently identified osteoclastogenic regulator miR182, along with miR4472-2, miR1302-3, miR6720, miR6087 and miR4532 were enriched in sEV compared to parental cells, p < 0.01. Signatures were similar in sEVs derived from non-stimulated or IL-1β stimulated SF.
OA SF sEV regulate chondrocyte inflammatory and remodelling responses. OA SF sEV have unique signatures compared to parental cells which do not alter with IL-1β stimulation. This study provides insight into a novel regulatory mechanism within the OA joint which could inform future targeted therapy.
滑膜成纤维细胞（SF）来源的小细胞外囊泡（sEV）代表了一种调节骨关节炎（OA）软骨侵蚀的新型分子机制。然而，尚未使用相关疾病的细胞进行全面评估。本研究的目的是分离和鉴定 OA SF 来源的 sEV，并观察其调节与疾病相关的 OA 软骨细胞效应反应的能力。对 sEV 和 OA SF 细胞中的微小（mi）RNA 特征进行了分析。
从 OA 滑膜膜和软骨中分别分离 SF 和软骨细胞（n=9）。通过超速离心从 OA SF（±IL-1β）条件培养基中分离 sEV，并通过扫描电子显微镜（SEM）和透射电子显微镜（TEM）进行特征分析。纳米颗粒跟踪分析（NTA）确认粒径。通过 qPCR、ELISA 和硫酸化糖胺聚糖测定（sGAG）评估 sEV 对 OA 软骨细胞和软骨效应物反应的调节作用。使用 RNA 测序确定 OA SF 来源 sEV 中的 miRNA 特征。
OA SF 来源的 sEV 可被 OA 软骨细胞轻易摄取，观察到 catabolic 基因 MMP13 的表达增加（p<0.01），而 anabolic 基因 aggrecan 和 COL2A1 的表达减少（p<0.01）。用 IL-1β 刺激的 OA SF 衍生的 sEV 处理显著降低了 aggrecan 和 COL2A1 的表达（p<0.001），并增加了 SOX9 基因的表达（p<0.05）。用来自非刺激或 IL-1β 处理的 OA SF 的 sEV 培养的 OA 软骨细胞，导致 IL-6、IL-8 和 MMP-3 的分泌显著增加（p<0.01）。用 SF（±IL-1β）衍生的 sEV 培养的软骨外植体显著增加了 sGAG 的释放（p<0.01）。与亲本细胞相比，sEV 中存在的 miRNA 特征不同。最近发现的破骨细胞生成调节剂 miR182 以及 miR4472-2、miR1302-3、miR6720、miR6087 和 miR4532 在 sEV 中丰富，而在亲本细胞中则没有，p<0.01。非刺激或 IL-1β 刺激的 SF 衍生的 sEV 中的特征相似。
OA SF sEV 调节软骨细胞炎症和重塑反应。OA SF sEV 与亲本细胞相比具有独特的特征，而这些特征不会因 IL-1β 刺激而改变。本研究提供了 OA 关节内一种新型调节机制的见解，可为未来的靶向治疗提供信息。
