Amtil-Ouahdi Ilias, Vergara Fabián, Rio Carlos, González-Martínez Coral, Jahn Andreas, Forteza-Genestra María Antonia, Gayá Antonio, Calvo Javier, Sala-Llinas Ernest, Navarrete Bernardino Alcázar, Romero-Ortiz Ana Dolores, Monjo Marta, Ramis Johana M, Ortega Francisco G
Health Research Institute of the Balearic Islands (IdISBa), Palma, Spain.
Cell Therapy and Tissue Engineering Group, Research Institute on Health Sciences (IUNICS), Palma, Spain.
Biofactors. 2025 May-Jun;51(3):e70021. doi: 10.1002/biof.70021.
Pulmonary fibrosis (PF) is a progressive, life-threatening disease marked by excessive scarring of lung tissue. Recently, extracellular vesicles (EVs) have emerged as a promising antifibrotic therapy due to their regenerative and anti-inflammatory properties. However, the success of EV-based therapies depends on the route of administration, which can significantly influence their biodistribution and therapeutic effects. Furthermore, in PF, aging is a significant risk factor for the disease and, until today, EV treatment efficacy has not been studied in aged tissues. Specifically, we studied EVs derived from human umbilical cord mesenchymal stem cells and compared the biodistribution of these vesicles delivered via three routes: intravenous (IV), intrapleural (IP), and intratracheal (IT). A protocol was developed to set EV staining and concentration, minimizing animal use while maximizing the accuracy of results. To evaluate therapeutic effects, we conducted three experimental setups: (i) to assess their ability to reverse established fibrosis; (ii) to evaluate their effect on fibrosis progression; and (iii) to study early inflammation and macrophage polarization. Lung fibrosis and inflammation were assessed by analyzing fibrotic markers, inflammatory cytokines, collagen deposition, and bronchoalveolar lavage (BAL) fluid cell analysis, providing insights into EVs therapeutic potential in aged, fibrotic lung tissue. In the biodistribution study, IV administration was identified as the most effective route, successfully delivering EVs to both normal and fibrotic lung tissues. In the therapeutic study, antifibrotic effects were observed only when EVs were administered prophylactically, before the establishment of fibrosis. Under this protocol, IV-administered EVs reduced fibrotic mRNA biomarkers, collagen deposition, inflammatory cell infiltration, and macrophage polarization in BAL, as well as altering cytokine. Our findings emphasize the critical importance of selecting the appropriate route of administration for EV-based therapies. Notably, our work with an aging model reveals that EV treatments primarily exhibit prophylactic effects, with a marked reduction in their regenerative potential compared to previous studies conducted in younger models.
肺纤维化（PF）是一种进行性、危及生命的疾病，其特征是肺组织过度瘢痕化。近来，细胞外囊泡（EVs）因其再生和抗炎特性而成为一种有前景的抗纤维化治疗手段。然而，基于EVs的治疗成功与否取决于给药途径，这会显著影响其生物分布和治疗效果。此外，在PF中，衰老也是该疾病的一个重要风险因素，而迄今为止，尚未在老年组织中研究过EV治疗的疗效。具体而言，我们研究了源自人脐带间充质干细胞的EVs，并比较了通过三种途径递送这些囊泡后的生物分布：静脉内（IV）、胸膜内（IP）和气管内（IT）。我们制定了一个方案来确定EV染色和浓度，在尽量减少动物使用的同时最大化结果的准确性。为了评估治疗效果，我们进行了三个实验设置：（i）评估它们逆转已形成纤维化的能力；（ii）评估它们对纤维化进展的影响；（iii）研究早期炎症和巨噬细胞极化。通过分析纤维化标志物、炎性细胞因子、胶原沉积以及支气管肺泡灌洗（BAL）液细胞分析来评估肺纤维化和炎症，从而深入了解EVs在老年纤维化肺组织中的治疗潜力。在生物分布研究中，静脉内给药被确定为最有效的途径，成功地将EVs递送至正常和纤维化肺组织。在治疗研究中，仅在纤维化形成之前预防性给予EVs时才观察到抗纤维化作用。在此方案下，静脉内给予的EVs减少了BAL中纤维化mRNA生物标志物、胶原沉积、炎性细胞浸润和巨噬细胞极化，同时改变了细胞因子。我们的研究结果强调了为基于EVs的治疗选择合适给药途径的至关重要性。值得注意的是，我们在衰老模型中的研究表明，与之前在年轻模型中进行的研究相比，EV治疗主要表现出预防作用，其再生潜力显著降低。
