Abstract
Osteogenesis imperfecta (OI), also known as "brittle bone disease," is a rare inherited genetic disorder characterized by bone fragility and often associated with short stature. The mutation in WNT1 causes autosomal recessive OI (AR-OI) due to the key role of WNT/β-catenin signaling in bone formation. WNT1 mutations cause phenotypes in OI of varying degrees of clinical severity, ranging from moderate to progressively deforming forms. The nucleotide change c.677C > T is one of the recurrent variants in the WNT1 alleles in Chinese AR-OI patients. To explore the effects of mutation c.677C > T on WNT1 function, we evaluated the activation of WNT/β-catenin signaling, cell proliferation, osteoblast differentiation, and osteoclast differentiation in WNT1c.677C>T , WNT1c.884C>A , and wild type WNT1 transfected into MC3T3-E1 preosteoblasts. Plasmids containing wild type WNT1, WNT1c.677C>T , and WNT1c.884C>A cDNAs were constructed. Protein levels of phosphorylation at serine 9 of GSK-3β (p-GSK-3β), GSK-3β, nonphosphorylated β-catenin (non-p-β-catenin), and β-catenin were detected with western blot. Cell proliferation was determined using MTS. BMP-2 and RANKL mRNA and protein levels were detected by qPCR and western blot. Our results showed that WNT1c.677C>T failed to activate WNT/β-catenin signaling and impaired the proliferation of preosteoblasts. Moreover, compared to wild type WNT1, WNT1c.677C>T downregulated BMP-2 protein expression and was exhibited a diminished capacity to suppress the RANKL protein level. In conclusion, mutation c.677C > T hindered the ability of WNT1 to induce the WNT/β-catenin signaling pathway and it affected the WNT/β-catenin pathway which might potentially contribute to hampered bone homeostasis.