Hydrogen sulfide generated by cystathionine gamma lyase inhibits lysyl oxidase and protects against calcific tendinopathy

Increased lysyl oxidase (LOX) activity favors pathologic cartilage and vessel calcification. LOX promotes disease through enhanced collagen cross-linking, inflammation, reactive oxygen species (ROS) production, cell trans-differentiation, and fibrosis. This study investigates the therapeutic potential of cystathionine gamma lyase (CSE)-generated hydrogen sulfide (H 2 S) to inhibit tendon calcification by targeting LOX in human samples and murine models of calcific tendinopathy (CT). Human shoulder supraspinatus tendons with varying degrees of CT were analyzed using Alizarin Red staining and LOX and CSE immunohistochemistry to evaluate the correlation between CSE and LOX/calcification. Mechanistic studies were performed using wild-type (WT) and CSE knockout murine tenocytes cultured in calcification-inducing medium with or without H 2 S donors or the LOX inhibitor β-aminopropionitrile (BAPN). Achilles tendon CT was induced in WT and CSE knockout mice via surgical intervention or aging. Tendon calcification, LOX expression, biomechanical integrity, and transcriptomic changes were assessed. Persulfidation of total proteins and recombinant human LOX (rhLOX) was measured using the dimedone-switch method. An inverse correlation between CSE levels and LOX/calcification was observed in human tendons and in the surgery-induced CT murine model. In murine tenocytes and in the aging murine model, CSE deficiency led to increased LOX expression, enhanced calcification, and reduced tendon biomechanical integrity.Transcriptomic analysis confirmed the negative association between CSE and LOX in murine CT. Mechanistically, H 2 S increased total cellular protein persulfidation, including rhLOX, resulting in inhibition of its enzymatic activity. Dysregulated LOX activity is a key driver of calcific tendinopathy. CSE-generated H 2 S effectively suppresses LOX activity, highlighting its potential as a therapeutic strategy for CT and other calcification-related disorders. This study identifies LOX as a therapeutic target in CT and supports H 2 S as a promising treatment strategy for this condition. Proposed mechanistic model In healthy tendon, CSE generates enough H 2 S to inhibit LOX-induced calcification, depicted in red. Contrarily, in tendon with trauma or in aged tendons, CSE-generated H 2 S is diminished ultimately leading to increased LOX activity and calcification. Overall, H 2 S could be of therapeutic relevance in calcific tendinopathy.