Identification of the occurrence and potential mechanisms of heterotopic ossification associated with 17-beta-estradiol targeting MKX by bioinformatics analysis and cellular experiments

BACKGROUND: Tendon heterotopic ossification (HO) is a common condition occurring secondary to tendon injury or surgical trauma that significantly affects the patient’s quality of life. The treatment of tendon HO remains challenging due to a lack of clarity regarding the pathological mechanism. Mohawk (MKX) is a key factor in preventing tendon HO; however, its upstream regulatory mechanism remains to be understood. This study aimed to identify potential compounds that target and regulate MKX and explore their functional mechanisms. METHODS: Bioinformatics analysis of MKX-related compounds and proteins was performed based on data from the STITCH and OncoBinder databases. Subsequently, the SymMap database was used to study MKX-related traditional Chinese medicine drugs and symptoms. Next, the OncoBinder genomic and proteomic discovery model was applied to identify potential regulators of MKX. The analytical tool Expert Protein Analysis System for proteomics was used to predict the three-dimensional structure of MKX, and the AutoDockTools software was used to identify pockets of activity at potential sites for molecular docking. Furthermore, we evaluated the effect of different doses of 17-beta-estradiol on bone marrow-derived mesenchymal stem cells (BM-MSCs). RESULTS: By predicting the three-dimensional structure of MKX and simulating molecular docking, Pro-Tyr and 17-beta-Estradiol were found to target and bind to MKX. Analysis of the STITCH and OncoBinder databases showed that MKX had a significant regulatory correlation with suppressor interacting 3 A/histone deacetylase 1 (SIN3A/HDAC1). The GO and KEGG pathway enrichment analysis revealed that the functions of MKX and its associated proteins were mainly enriched in osteogenic-related pathways. Assessment of the proliferation of BM-MSCs revealed that 17-beta-estradiol possibly upregulated the mRNA expression of the HDAC1-SIN3A/BMP pathway-related RUNX2, thereby promoting the proliferation of BM-MSCs. CONCLUSIONS: The compounds Pro-Tyr and 17-beta-Estradiol may bind to MKX and thus affect the interaction of MKX with SIN3A/HDAC1.