Heterotopic endochondral ossification in periarticular supporting tissues, including tendons and ligaments, often occurs after trauma or surgery and contributes to joint stiffness and pain. Postmenopausal women exhibit a higher incidence of such conditions, implicating estrogen in the maintenance of musculoskeletal homeostasis. However, the molecular mechanisms by which estrogen prevents tendon ossification remain poorly understood.
To investigate this, we used a murine model of estrogen deficiency combined with tendon injury. Adult female mice underwent bilateral ovariectomy (OVX), followed by Achilles tendon needle puncture (ATP model) to induce endochondral ossification. A separate group of OVX mice received subcutaneous estrogen supplementation. Histological analysis using Safranin-O staining revealed cartilage-like matrix formation in the tendon core of OVX-ATP mice, whereas estrogen-treated OVX-ATP mice exhibited reduced Safranin-O-positive regions. Micro-computed tomography confirmed the presence of ossification at 8 to 12 weeks post-injury in OVX-ATP mice, with only minimal ossification observed in estrogen-treated mice.
RNA sequencing analysis of the injured tendon tissues identified Sparc (secreted protein acidic and rich in cysteine) as significantly upregulated in OVX-ATP mice. SPARC is a matricellular protein known to promote osteogenesis. Intratendinous injection of recombinant human SPARC in non-OVX ATP model mice led to marked enhancement of tendon ossification, indicating a causal role. In vitro, estrogen treatment significantly suppressed Sparc mRNA expression in NIH3T3 fibroblasts, suggesting that SPARC is a downstream effector of estrogen signaling.
These findings suggest that estrogen inhibits tendon ossification at least in part by downregulating SPARC. This pathway may contribute to tendon homeostasis and presents SPARC as a potential therapeutic target for preventing heterotopic ossification in estrogen-deficient states such as postmenopause.