Abstract: Objective The objective of this study is to investigate the mechanism by which Runmu Ling Formula(RML)ameliorates ocular surface damage in rats with dry eye through activation of the LC3-ATG5 autophagy pathway and inhibition of inflammatory factor expression. Methods A dry eye model was established via subcutaneous injection of scopolamine hydrobromide. A total of 60 Sprague-Dawley(SD)rats were randomly assigned to six groups,with ten rats per group: a blank control group, a model group, an RML-treated group, an autophagy activator group, an autophagy inhibitor group, and an RML combined with autophagy inhibitor group. The ocular surface function of the subjects was assessed using a variety of methods, including corneal fluorescein staining(CFS)scores, tear break-up time(TBUT), and the phenol red thread test(PRTT). The corneas were subjected to a comprehensive evaluation that entailed the application of hematoxylin-eosin(HE)staining and transmission electron microscopy. RT-qPCR and Western blotting were employed to measure the expression of autophagy-related markers(LC3, ATG5, P62)and inflammatory factors(IL-1β, IL-18, TNF-α). Results In comparison with the blank control group, the model group demonstrated a significant reduction in TBUT(P<0.001), elevated CFS scores(P<0.001), diminished PRTT values(P<0.001), and augmented vacuolation of corneal epithelial cells. RML intervention led to a significant restoration of ocular surface homeostasis, as evidenced by substantial improvements in TBUT, CFS, and PRTT(all P<0.05). The transmission electronmicroscopy revealed an increase in the formation of autophagosomesin the RML group compared to the model group. Mechanistically, RML significantly activated the LC3-ATG5 autophagy pathway, as evidenced by increased LC3-II/LC3-I ratios(P<0.05)and elevated ATG5 expression(P<0.05). Concurrently, RML potently suppressed the expression of inflammatory factors TNF-α, IL-1β, and IL-18(all P<0.05). Conclusion The efficacy of Runmu Ling Formula in alleviating ocular surface inflammation and injury in rats with dry eye has been demonstrated by its ability to activate the LC3-ATG5 autophagy pathway and inhibit the expression of key inflammatory factors, including TNF-α, IL-1β, and IL-18.

Key words: Runmu Ling Formula, Dry eye, Animal model, Autophagy, Inflammatory factors