Stress-Induced Disruption of Circadian and Neuroimmune Networks in Lacrimal Glands Drives Dry Eye Pathogenesis

Chronic psychological stress is an important but poorly defined risk factor for dry eye disease. This study investigated how sustained stress alters circadian regulation in the lacrimal gland (LG) and the role of neuroendocrine signaling in stress-induced dry eye disease.

Male C57BL/6J mice underwent two weeks of daily restraint stress with or without β-adrenergic blockade or glucocorticoid synthesis inhibition. Tear secretion was assessed by pilocarpine-stimulated phenol red thread test. LGs were collected every three hours across 24 hours for bulk RNA sequencing, and rhythmicity was analyzed with Jonckheere-Terpstra-Kendall cycle (JTK_CYCLE) algorithm. Single-cell RNA sequencing at zeitgeber time 6 mapped stress hormone receptor expression across LG cell types. Histology, immunohistochemistry, and plasma hormone assays characterized structural, immune, metabolic, and neural changes.

Stress reduced tear secretion, altered LG morphology, and activated sympathetic and hypothalamic-pituitary-adrenal axis. Transcriptomic profiling showed a marked loss of rhythmic gene expression in immune, metabolic, neural, and proliferative pathways, whereas core clock oscillators remained intact, defining circadian output decoupling. This was associated with disrupted diurnal immune trafficking, reduced neuronal activity, impaired metabolism, and diminished proliferation. Single-cell analysis revealed β-adrenergic and glucocorticoid receptor expression in epithelial, endothelial, and immune populations. Propranolol or metyrapone partially restored rhythmic transcription, glandular architecture, and tear secretion.

Chronic stress induces circadian output decoupling in the lacrimal gland, driving neuroimmune and metabolic dysfunction and dry eye disease pathogenesis. Partial rescue by β-adrenergic and glucocorticoid pathway inhibition highlights neuroendocrine signaling as a therapeutic target and establishes the LG as a peripheral model of stress-induced circadian disruption.