Tauroursodeoxycholic acid (TUDCA)-induced increase in ectopic muscle mineralization occurs exclusively in dystrophic muscles and is independent of endoplasmic reticulum stress

Calcification of dystrophic skeletal muscles was described previously and attributed, among others, to ER-stress, elevated phosphate concentration and chronic inflammation. Tauroursodeoxycholic acid (TUDCA) has been broadly tested as a potential drug in therapy of various diseases. In particular it is considered an artificial chaperone protecting cells against ER-stress which could prevent an ectopic mineralisation of soft tissues . We hypothesised that TUDCA treatment should reduce muscle mineral deposits. The goal of this study was to test this concept using two mouse models of DMD. Four-week old mdx, mdx ^betageo and w/t mice were administered TUDCA in drinking water for 4 weeks. Then, following tissue-clearing and calcium minerals staining with alizarin, animal bodies were evaluated using whole body scanning Additionally, isolated skeletal muscles were analysed by Western blotting for ER-stress and calcification markers, and using various microscopic methods. Enzymatic activity of alkaline phosphatase was also assayed. Unexpectedly, TUDCA enhanced calcification of dystrophic but not dystrophin-positive muscles. TUDCA did not affect the elevated ER-stress markers found in dystrophic muscles nor impact pro-calcifying proteins RUNX2, Osterix and BMP2/4, which were overexpressed in dystrophic muscles. The alkaline phosphatase levels, which were reduced in dystrophic muscles, were not affected by this treatment. The increase in ectopic calcification in dystrophic muscles induced by TUDCA is specific to muscles lacking dystrophin. This effect is not linked to the alleviation of ER stress or the overexpression of proteins directly involved in calcium mineral accumulation, both of which are elevated in dystrophic muscles.