Ragm area with degenerating fibers ?gastroc Centralized nuclei fiber-gastroc* Centralized nuclei fiber-diaphragm * apoptosis nuclei per field*6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6*Non-parametric comparison of medians; data expressed as mean 6 SD; median (range). Abbreviations: FS ?percent fractional shortening, EF- percent ejection fraction, BPM- beats per minute, bpm ?breaths per minute, SD ?standard deviation, PA ?pulmonary artery, AO ?aortic, E/A ?ratio of mitral valve E and A wave velocities, GSM ?grip strength meter, BW- body weight, Gastroc 10457188 ?gastrocnemius, TA ?tibialis anterior, KGF ?kilogram-force. doi:10.1371/journal.pone.0065468.tfound decreased apoptosis in the tibialis anterior muscle of omigapil treated dy2J mice. (Figure 2) Apoptosis is a known pathologic pathway in congenital muscular dystrophy patients. [4]. Erb et al. (2009) also measured manual recordings of mouse activity in a new cage environment and showed omigapil treated mice had significantly increased activity compared to vehicle treated mice at 5? weeks of age. This significance was lost at 10 weeks of age, but a trend continued. In the milder phenotype of the dy2J mice, this study showed significantly increased movement times and decreased rest times in mice treated with 0.1 mg/kg. So, in the more severe model, an improvement was demonstrated early and lost over time, while in this milder phenotypic model, the improvements were beginning to show and likely require a longer treatment period to fully develop. Erb et al. (2009) also Title Loaded From File presented histological data showing the muscle fiber size distribution normalized by reducing the proportion of small caliber and increasing the proportion of large caliber muscle fibers in the triceps brachii of dyW mice treated with0.1 mg/kg omigapil. The current study did not measure fiber size, but we did see a significant decrease in percent centralized nuclei per fiber (a measure of total regeneration) between omigapil treatment and vehicle control groups in the gastrocnemius. We also showed significantly decreased percent in areas of degenerating fibers in the gastrocnemius in the omigapil treated mice. A decrease in degeneration leads to less regeneration and preservation of larger fibers, a similar observation as reported by Erb et al. dy2J mice showed significantly increased respiratory rates in omigapil treated mice at the end of the trial compared to vehicle treated. These increased rates were similar to wild type controls. This in vivo functional measure could reflect improved diaphragm function. This finding is quite important since clinically many of the El of phospho-JNK was not affected by HLJDT treatment (P.0.05, Fig. affected patients suffer significant respiratory insufficiency and this is a leading cause of death. Any effective therapy needs to demonstrate improvements in respiratory function and these changes support a putative role for omigapil.Omigapil Treatment in dy2J MiceFigure 1. Histological analysis of gastrocnemius and diaphragm with H E (top two rows) and gastrocnemius with picrosirius red (bottom row) show increased fibrosis and centralized nuclei in dy2J mice. BL6 control mice are shown in column A. dy2J mice treated with 0.1 mg/kg omigapil (Column B) showed markedly less fibrosis compared to dy2J mice treated with 1 mg/kg omigapil (Column C) or vehicle (Column D). doi:10.1371/journal.pone.0065468.gEchocardiographic analysis found increased heart rates in dy2J mice. This is a consistent finding in other dystrophic mouse models and could reflec.Ragm area with degenerating fibers ?gastroc Centralized nuclei fiber-gastroc* Centralized nuclei fiber-diaphragm * apoptosis nuclei per field*6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6*Non-parametric comparison of medians; data expressed as mean 6 SD; median (range). Abbreviations: FS ?percent fractional shortening, EF- percent ejection fraction, BPM- beats per minute, bpm ?breaths per minute, SD ?standard deviation, PA ?pulmonary artery, AO ?aortic, E/A ?ratio of mitral valve E and A wave velocities, GSM ?grip strength meter, BW- body weight, Gastroc 10457188 ?gastrocnemius, TA ?tibialis anterior, KGF ?kilogram-force. doi:10.1371/journal.pone.0065468.tfound decreased apoptosis in the tibialis anterior muscle of omigapil treated dy2J mice. (Figure 2) Apoptosis is a known pathologic pathway in congenital muscular dystrophy patients. [4]. Erb et al. (2009) also measured manual recordings of mouse activity in a new cage environment and showed omigapil treated mice had significantly increased activity compared to vehicle treated mice at 5? weeks of age. This significance was lost at 10 weeks of age, but a trend continued. In the milder phenotype of the dy2J mice, this study showed significantly increased movement times and decreased rest times in mice treated with 0.1 mg/kg. So, in the more severe model, an improvement was demonstrated early and lost over time, while in this milder phenotypic model, the improvements were beginning to show and likely require a longer treatment period to fully develop. Erb et al. (2009) also presented histological data showing the muscle fiber size distribution normalized by reducing the proportion of small caliber and increasing the proportion of large caliber muscle fibers in the triceps brachii of dyW mice treated with0.1 mg/kg omigapil. The current study did not measure fiber size, but we did see a significant decrease in percent centralized nuclei per fiber (a measure of total regeneration) between omigapil treatment and vehicle control groups in the gastrocnemius. We also showed significantly decreased percent in areas of degenerating fibers in the gastrocnemius in the omigapil treated mice. A decrease in degeneration leads to less regeneration and preservation of larger fibers, a similar observation as reported by Erb et al. dy2J mice showed significantly increased respiratory rates in omigapil treated mice at the end of the trial compared to vehicle treated. These increased rates were similar to wild type controls. This in vivo functional measure could reflect improved diaphragm function. This finding is quite important since clinically many of the affected patients suffer significant respiratory insufficiency and this is a leading cause of death. Any effective therapy needs to demonstrate improvements in respiratory function and these changes support a putative role for omigapil.Omigapil Treatment in dy2J MiceFigure 1. Histological analysis of gastrocnemius and diaphragm with H E (top two rows) and gastrocnemius with picrosirius red (bottom row) show increased fibrosis and centralized nuclei in dy2J mice. BL6 control mice are shown in column A. dy2J mice treated with 0.1 mg/kg omigapil (Column B) showed markedly less fibrosis compared to dy2J mice treated with 1 mg/kg omigapil (Column C) or vehicle (Column D). doi:10.1371/journal.pone.0065468.gEchocardiographic analysis found increased heart rates in dy2J mice. This is a consistent finding in other dystrophic mouse models and could reflec.
