Fibroblast specialization across microanatomy in a single-cell atlas of human Achilles tendon.

Tendons are transitional tissues linking muscle to bone, enabling locomotion and fine motor control. The cellular biology across the Achilles tendon unit is poorly understood, yet is critical for interpreting normal function and pathological changes across its microanatomically defined functional zones. We generated a spatially resolved transcriptomic atlas of adult (age 45-76) nontendinopathic human Achilles tendon, sampling the tendon-bone junction (enthesis), midbody, myotendinous junction, and adjoining muscle. Six fibroblast subtypes were identified, with distinct transcriptional profiles and spatial distributions, suggesting specialized functional roles across the tendon. Two dominant fibroblast types were specifically positioned in the tendon midsubstance and paratenon (vessel-rich region surrounding the tendon fibrils); other populations included perineural, myotendinous junction-specific, muscle-specific, and lining-layer fibroblasts. These findings demonstrate how cellular diversity across a transitional tissue may underlie microanatomical-specific roles. This atlas provides a foundation for understanding cellular functions across the tendon and adjoining muscle and will be essential for comparisons with diseased tissue, identifying pathogenic mediators and treatment targets for autoimmune and degenerative pathologies of the Achilles tendon.NEW & NOTEWORTHY We present the first spatially resolved single-cell atlas of the human Achilles tendon. By sampling across the microanatomy of the tendon from enthesis to muscle, we demonstrate changes in fibroblast composition across this transitional tissue. Distinct fibroblast subsets were discovered with specific transcriptomic signatures, and several were found in distinct spatial locations corresponding to putative functional roles in tendon and adjoining muscle. These findings demonstrate how cellular diversity across a transitional tissue may underlie microanatomical-specific roles.