Function and Molecular Mechanism of DEFB1 Gene Polymorphism in Regulating Tendon Regeneration after Rotator Cuff Injury

Abstract

Purpose: The incidence of rotator cuff injury is as high as about 20% of shoulder joint diseases, causing functional limitations. Single-nucleotide polymorphism (SNP) of beta-defensin 1 gene (DEFB1) was shown to correlate with the presence of rotator cuff disease. The aim of this study was to investigate the regulatory effect of SNP on tendon regeneration and its molecular mechanism of action. Method: DEFB1 gene was constructed for rabbit tendon stem cell transfection. Differential binding proteins were screened for both different genotypes using DNA pull down and mass spectrometry, and the function of the binding proteins was further verified. Result: DEFB1-WT (GG) (wild-type DEFB1) and DEFB1-MUT (CC) (DEFB1 mutated) binding proteins were different, and the content of FUS RNA binding protein (FUS) protein in DEFB1-WT (GG) group was significantly higher than that in DEFB1-MUT (CC) group. Chromatin immunoprecipitation revealed that significantly more FUS protein was binding to DEFB1 (GG) than DEFB1 (CC). Subsequent validation tests at the cellular level showed that FUS gene significantly and positively regulated DEFB1 gene expression, and that after interfering with FUS expression, DEFB1 expression was significantly decreased, and tendon stem cell calcification and osteogenic differentiation were significantly enhanced. Conclusions: DEFB1-WT (GG) type might affect the prognostic repair of tendon rupture through the regulation of the non-tenogenic differentiation of tendon stem cells in the pathological state.