Abstract
<title>Abstract</title> <p>Stem cell-based therapies have gained recently increasing attention as potential treatment options for retinal diseases, including glaucoma. Among these approaches, the secretome or the conditioned medium (CM) derived from mesenchymal stromal cells (MSCs) represents a promising cell-free therapeutic alternative due to its regenerative and immunomodulatory properties. However, prior to exploring therapeutic efficacy in animal disease models, its ocular and systemic safety must be established. The present study aimed to evaluate the tolerance of human umbilical cord (hUC) MSCs-derived CM following subconjunctival (SbC) and intravitreal (ITV) administration in healthy Wistar Hannover rats. Twenty-four adult male rats were randomly assigned to four experimental groups based on administration route (SbC or ITV) and euthanasia timepoint (5 or 28 days). CM was administered unilaterally to the right eye, with the contralateral eye serving as an internal control. Ocular tolerance was assessed through comprehensive ophthalmic examination, intraocular pressure (IOP) monitoring, electroretinography (ERG), and histological evaluation of ocular tissues. Systemic safety was evaluated through histopathological analysis of liver, kidney, and spleen samples. No relevant persistent ocular abnormalities or sustained IOP changes were observed. Retinal function remained largely preserved, although ITV administration induced mild alterations in selected electrophysiological parameters. Histological analysis revealed overall preservation of the retinal architecture, despite a reduction in outer nuclear layer thickness in ITV groups at the later timepoint. Histopathological examination of systemic organs showed preserved tissue architecture with only mild incidental findings. Overall, these results support the ocular and systemic tolerability of MSCs CM in healthy rats, with SbC delivery showing a more stable functional and structural profile.</p>