Morpholino Treatment Improved Muscle Function and Pathology in Pitx1 Transgenic Mice
Open Access"Morpholino Treatment Improved Muscle Function and Pathology in Pitx1 Transgenic Mice"Paired-like homeodomain transcription factor 1 (PITX1) was proposed to be critically involved in facioscapulohumeral muscular dystrophy (FSHD). We generated a tet-repressible muscle-specific Pitx1 transgenic mouse model in which expression of PITX1can be induced in skeletal muscle. In this study, we attempted to knock-down PITX1 expression using morpholino molecules designed to block translation of the Pitx1 mRNA.Three groups of the Pitx1 transgenic mice (n=5) received weekly intravenous injections of phosphorodiamidatemorpholino oligomers (PMO) (100mg/kg), octaguanidiniumdendrimer-conjugated Morpholino(Vivo-morpholinos)(10mg/kg), or phosphate buffered saline (PBS) for six weeks. A group of control mice were used to provide baseline expression level of PITX1. The mice received the first injection of morpholinos when the oral doxycycline was discontinued to induce PITX1 expression, followed by weekly injections for six weeks. Four weeks after PITX1 induction, muscle function of the mice was evaluated by grip strength and rotarod testes. Triceps and quadriceps muscles were collected for H&E;, immunohistochemistry and immunoblotting at the end of the 6th week.Grip strength data showed that treatment with Vivo-morpholinos significantly (p<0.05) improved muscle function of the transgenic mice over-expressing Pitx1 while the PMO treatment did not improve the muscle function. Immunohistochemistry and immunoblotting data showed that PITX1 protein expression in the triceps and quadricepswas reduced 2.7 (p<0.05) and 2.2 (p=0.06) folds respectively by the Vivo-morpholinostreatment but not the PMO treatment. The pathology of the muscles of mice treated with Vivo-morpholinos was improved by showing significantly fewer angular atrophic myofibers (p< 0.05).Pitx1 transgenic mice showed significant improvement in both muscle function and pathology after receiving Vivo-morpholinos injections, while the treatment of PMO didn't improve the phenotype. The results suggest that morpholinos can be a potential option for treatment development for FSHD.
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