Establishment of a TGF-β-Induced Rodent Myopia Model Using Subconjunctival Injections

Myopia is a major global public health concern characterized by excessive axial elongation and scleral extracellular matrix (ECM) remodeling. While experimental models are essential for investigating these mechanisms, traditional optical defocus methods in small rodents present technical limitations. Here, a standardizable pharmacological myopia model was established and evaluated via repeated subconjunctival injections of transforming growth factor-beta (TGF-β). To achieve targeted delivery, a dual-handed surgical stabilization technique was developed to evert the upper eyelid, allowing a 33-gauge needle to target the palpebral conjunctiva at a shallow 20°-30° angle. Through in vivo screening in rats, TGF-β2 (500 pg/eye) was identified as a prominent driver of posterior eye growth. Subsequent dose-response titration in mice revealed an inverted U-shaped profile. In the primary validation cohort, the lowest dose (250 pg/eye) induced pronounced axial elongation (0.197 ± 0.012 mm) and myopic refractive shifts (-1.58 ± 0.63 D). At the molecular level, repetitive TGF-β2 administration activated the retinal NOD-like receptor family pyrin domain-containing 3 (NLRP3) / interleukin-1β (IL-1β) inflammasome cascade and upregulated matrix metalloproteinase 2 (MMP2) expression in both species. Coupled with a two-day split biometric recording regimen that minimizes anesthesia-associated artifacts, this protocol provides a reliable, scalable platform for evaluating anti-myopia interventions.