Early choroidal vascular remodeling predicts atropine treatment response in myopia: clinical cohort and experimental evidence

Low-dose atropine is widely used for myopia control in children, yet treatment response varies between individuals, highlighting the clinical need for early predictors of long-term efficacy. Although choroidal thickness has been investigated as a potential biomarker, its association with atropine response remains inconsistent. Therefore, we aimed to identify more reliable choroidal biomarkers, develop models for predicting treatment response, and investigate biological processes potentially involved in atropine-induced choroidal remodeling. This secondary analysis of the AMIXT randomized controlled trial included 290 myopic children (0.01% atropine: 195; placebo: 95) with 2-year OCTA follow‑up. We developed one-dimensional (1D) /2D/3D choroidal parameters partitioned into luminal and stromal compartments. Random-intercept cross-lagged panel models (RI-CLPM) assessed compartmental directional interplay. Prediction models were developed and assessed for predicting continuous change from baseline in 24-month cycloplegic spherical equivalent (SE) and treatment response (defined as SE change ≤ 0.5 diopter [D]/year) using baseline factors and early changes in choroidal parameters. Mechanistically, single-cell RNA sequencing was performed on retinal pigment epithelium/choroid complexes from form-deprivation myopia mice treated with atropine or vehicle, followed by in vitro and in vivo validation using flow cytometry and immunofluorescence. Among choroidal metrics, 3D parameters exhibited superior predictive value, particularly 4-month changes in luminal volume within the 3 mm perifoveal zone (△LV_3), which performed best in linear and logistic prediction models (R 2 : 0.41; Area under the curve: 0.89). RI-CLPM identified a significant cross-lagged association from luminal volume at 4 months to stromal volume at 2 years. Single cell transcriptomic analyses revealed atropine enhanced nitric oxide (NO) response signatures and attenuated reactive oxygen species (ROS) related programs in arterial choroidal endothelial cells (ECs). These findings were supported in murine choroidal tissue and primary human choroidal ECs and further suggested that atropine may improve NO bioavailability supporting the potential involvement of reducing endothelial ROS. Early choroidal vascular expansion serves as a promising biomarker for atropine efficacy. Mechanistically, atropine may influence choroidal vascular remodeling through modulation of endothelial oxidative stress and NO signaling. These findings provide biological plausibility for the involvement of the choroidal ROS-NO axis, although its direct contribution to clinical choroidal remodeling requires further investigation. Chinese Clinical Trial Registry Registration number (ChiCTR2000039827||http//www.chictr.org.cn/), Registration Date 2020-11-11.