Two novel BMP-2 variants identified in patients with thoracic ossification of the ligamentum flavum

Thoracic ossification of the ligamentum flavum (TOLF)is a common cause of thoracic spinal canal stenosis and has been reported almost exclusively in East Asian countries. In this study, we established a relationship between bone morphogenic protein 2 (BMP-2) and TOLF. We divided patients into two groups according to severity of ossification and identified susceptible loci through exome sequencing. We identified 39 novel likely pathogenic variants in 29 genes in the transforming growth factor-beta (TGF-β) superfamily or TGF-β/BMPs signaling pathway, including two missense variants in BMP-2 (NM_001200.3) exon region, c.460C>G:p.(R154G) and c.584G>T:p.(R195M). Further Sanger sequencing and genotyping suggested the variants were only found in patients with long regional OLF. Bioinformatic assays predicted the two BMP-2 variants to cause significant alterations to gene and protein expression. Functional assays showed upregulation of BMP-2 expression, increased osteogenic marker expression, and enhanced osteogenic differentiation. Collectively, these results suggest a genetic contribution to the pathogenesis of TOLF, particularly in patients with long segment disease, and that nucleotide substitutions associated with increased BMP-2 expression may be involved in TOLF pathogenesis.