TGFBR2 but not SPP1 genotype modulates osteopontin expression in Duchenne muscular dystrophy muscle

A polymorphism (rs28357094) in the promoter region of the SPP1 gene coding for osteopontin (OPN) is a strong determinant of disease severity in Duchenne muscular dystrophy (DMD). The rare G allele of rs28357094 alters gene promoter function and reduces mRNA expression in transfected HeLa cells. To dissect the molecular mechanisms of increased disease severity associated with the G allele, we characterized SPP1 mRNA and protein in DMD muscle biopsies of patients with defined rs28357094 genotype. We did not find significant differences in osteopontin mRNA or protein expression between patients carrying the T (ancestral allele) or TG/GG genotypes at rs28357094. The G allele was significantly associated with reduced CD4(+) and CD68(+) cells on patient muscle biopsy. We also quantified transforming growth factor-β (TGFB) and TGFB receptor-2 (TGFBR2) mRNA in DMD muscle biopsies, given the ability of TGFB and TGFBR2 to activate SPP1 promoter region and their role in DMD pathogenesis. The amount of TGFB and TGFBR2 mRNA did not predict the amount of SPP1 mRNA or protein, while a polymorphism in the TGFBR2 gene (rs4522809) was found to be a strong predictor of SPP1 mRNA level. Our findings suggest that OPN mediates inflammatory changes in DMD and that TGFB signalling has a role in the complex regulation of osteopontin expression.