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Prevention of exercised induced cardiomyopathy following Pip-PMO treatment in dystrophic mdx mice.

Betts CA, Saleh AF, Carr CA, Hammond SM, Coenen-Stass AM, Godfrey C, McClorey G, Varela MA, Roberts TC, Clarke K, Gait MJ, Wood MJ - Sci Rep (2015)

Bottom Line: Pip peptide-AOs demonstrate high activity in cardiac muscle.Repeated peptide-AO treatments resulted in high levels of cardiac dystrophin protein, which prevented the exercised induced progression of cardiomyopathy, normalising heart size as well as stabilising other cardiac parameters.Treated mice also exhibited significantly reduced cardiac fibrosis and improved sarcolemmal integrity.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, South Parks Road, Oxford, UK, OX1 3QX.

ABSTRACT
Duchenne muscular dystrophy (DMD) is a fatal neuromuscular disorder caused by mutations in the Dmd gene. In addition to skeletal muscle wasting, DMD patients develop cardiomyopathy, which significantly contributes to mortality. Antisense oligonucleotides (AOs) are a promising DMD therapy, restoring functional dystrophin protein by exon skipping. However, a major limitation with current AOs is the absence of dystrophin correction in heart. Pip peptide-AOs demonstrate high activity in cardiac muscle. To determine their therapeutic value, dystrophic mdx mice were subject to forced exercise to model the DMD cardiac phenotype. Repeated peptide-AO treatments resulted in high levels of cardiac dystrophin protein, which prevented the exercised induced progression of cardiomyopathy, normalising heart size as well as stabilising other cardiac parameters. Treated mice also exhibited significantly reduced cardiac fibrosis and improved sarcolemmal integrity. This work demonstrates that high levels of cardiac dystrophin restored by Pip peptide-AOs prevents further deterioration of cardiomyopathy and pathology following exercise in dystrophic DMD mice.

No MeSH data available.


Related in: MedlinePlus

Reduction in cardiac pathology as determined by the detection of fibrosis and sarcolemmal damage in the hearts of exercised mdx mice following Pip6f-PMO treatment.(A) RT-qPCR analysis of miRNA-21 in heart tissue normalised to C57BL/10 cohort. (B) Quantification of Masson's trichrome staining in hearts of exercised cohorts. (C) Evans blue dye infiltration into heart following exercise. (D) Masson's trichrome images of worst areas of collagen deposition. miRNA-21 expression and Evans blue dye leakage of Pip6f-PMO hearts is normalised in contrast to untreated control. In addition Masson's trichrome staining is reduced and the representative images indicate less fibrosis then the untreated mdx cohort. Statistical significance was determined using ANOVA followed by Tukey post-hoc test (*** = P < 0.001, ** = P < 0.01, * = P < 0.05).
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f4: Reduction in cardiac pathology as determined by the detection of fibrosis and sarcolemmal damage in the hearts of exercised mdx mice following Pip6f-PMO treatment.(A) RT-qPCR analysis of miRNA-21 in heart tissue normalised to C57BL/10 cohort. (B) Quantification of Masson's trichrome staining in hearts of exercised cohorts. (C) Evans blue dye infiltration into heart following exercise. (D) Masson's trichrome images of worst areas of collagen deposition. miRNA-21 expression and Evans blue dye leakage of Pip6f-PMO hearts is normalised in contrast to untreated control. In addition Masson's trichrome staining is reduced and the representative images indicate less fibrosis then the untreated mdx cohort. Statistical significance was determined using ANOVA followed by Tukey post-hoc test (*** = P < 0.001, ** = P < 0.01, * = P < 0.05).

Mentions: MicroRNA (miRNA) 21 is another biomarker of cardiac injury, specifically, it has been directly linked to myocardial fibrosis during hypertrophy40. Mdx mice showed an increase in miR-21 expression over wild type mice suggesting an increase in interstitial fibrosis (P < 0.001; Fig. 4A). Treatment with Pip6f-PMO partially normalised miR-21 expression (P < 0.05) over the untreated counterpart. The quantification of Masson's trichrome staining, which stains for collagen deposition, confirmed this observation of significantly reduced interstitial fibrosis in Pip6f-PMO treated mdx hearts (Fig. 4B). Again the untreated mdx hearts exhibited greater collagen deposition compared to C57BL/10 (P < 0.01) and Pip6f-PMO (P < 0.001) cohorts. Images of cardiac fibrosis strongly indicated the greater collagen deposition in untreated mdx hearts that was significantly reduced by treatment (Fig. 4D). Moreover, infiltration of Evans blue dye in the hearts of Pip6f-treated mice was also significantly reduced compared to untreated counterparts (P < 0.05; Fig. 4C). This showed that Pip6f-PMO treatment prevented sarcolemmal damage following exercise. These results denote a significant reduction in cardiac pathology in the treated mouse cohort.


Prevention of exercised induced cardiomyopathy following Pip-PMO treatment in dystrophic mdx mice.

Betts CA, Saleh AF, Carr CA, Hammond SM, Coenen-Stass AM, Godfrey C, McClorey G, Varela MA, Roberts TC, Clarke K, Gait MJ, Wood MJ - Sci Rep (2015)

Reduction in cardiac pathology as determined by the detection of fibrosis and sarcolemmal damage in the hearts of exercised mdx mice following Pip6f-PMO treatment.(A) RT-qPCR analysis of miRNA-21 in heart tissue normalised to C57BL/10 cohort. (B) Quantification of Masson's trichrome staining in hearts of exercised cohorts. (C) Evans blue dye infiltration into heart following exercise. (D) Masson's trichrome images of worst areas of collagen deposition. miRNA-21 expression and Evans blue dye leakage of Pip6f-PMO hearts is normalised in contrast to untreated control. In addition Masson's trichrome staining is reduced and the representative images indicate less fibrosis then the untreated mdx cohort. Statistical significance was determined using ANOVA followed by Tukey post-hoc test (*** = P < 0.001, ** = P < 0.01, * = P < 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4355666&req=5

f4: Reduction in cardiac pathology as determined by the detection of fibrosis and sarcolemmal damage in the hearts of exercised mdx mice following Pip6f-PMO treatment.(A) RT-qPCR analysis of miRNA-21 in heart tissue normalised to C57BL/10 cohort. (B) Quantification of Masson's trichrome staining in hearts of exercised cohorts. (C) Evans blue dye infiltration into heart following exercise. (D) Masson's trichrome images of worst areas of collagen deposition. miRNA-21 expression and Evans blue dye leakage of Pip6f-PMO hearts is normalised in contrast to untreated control. In addition Masson's trichrome staining is reduced and the representative images indicate less fibrosis then the untreated mdx cohort. Statistical significance was determined using ANOVA followed by Tukey post-hoc test (*** = P < 0.001, ** = P < 0.01, * = P < 0.05).
Mentions: MicroRNA (miRNA) 21 is another biomarker of cardiac injury, specifically, it has been directly linked to myocardial fibrosis during hypertrophy40. Mdx mice showed an increase in miR-21 expression over wild type mice suggesting an increase in interstitial fibrosis (P < 0.001; Fig. 4A). Treatment with Pip6f-PMO partially normalised miR-21 expression (P < 0.05) over the untreated counterpart. The quantification of Masson's trichrome staining, which stains for collagen deposition, confirmed this observation of significantly reduced interstitial fibrosis in Pip6f-PMO treated mdx hearts (Fig. 4B). Again the untreated mdx hearts exhibited greater collagen deposition compared to C57BL/10 (P < 0.01) and Pip6f-PMO (P < 0.001) cohorts. Images of cardiac fibrosis strongly indicated the greater collagen deposition in untreated mdx hearts that was significantly reduced by treatment (Fig. 4D). Moreover, infiltration of Evans blue dye in the hearts of Pip6f-treated mice was also significantly reduced compared to untreated counterparts (P < 0.05; Fig. 4C). This showed that Pip6f-PMO treatment prevented sarcolemmal damage following exercise. These results denote a significant reduction in cardiac pathology in the treated mouse cohort.

Bottom Line: Pip peptide-AOs demonstrate high activity in cardiac muscle.Repeated peptide-AO treatments resulted in high levels of cardiac dystrophin protein, which prevented the exercised induced progression of cardiomyopathy, normalising heart size as well as stabilising other cardiac parameters.Treated mice also exhibited significantly reduced cardiac fibrosis and improved sarcolemmal integrity.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, South Parks Road, Oxford, UK, OX1 3QX.

ABSTRACT
Duchenne muscular dystrophy (DMD) is a fatal neuromuscular disorder caused by mutations in the Dmd gene. In addition to skeletal muscle wasting, DMD patients develop cardiomyopathy, which significantly contributes to mortality. Antisense oligonucleotides (AOs) are a promising DMD therapy, restoring functional dystrophin protein by exon skipping. However, a major limitation with current AOs is the absence of dystrophin correction in heart. Pip peptide-AOs demonstrate high activity in cardiac muscle. To determine their therapeutic value, dystrophic mdx mice were subject to forced exercise to model the DMD cardiac phenotype. Repeated peptide-AO treatments resulted in high levels of cardiac dystrophin protein, which prevented the exercised induced progression of cardiomyopathy, normalising heart size as well as stabilising other cardiac parameters. Treated mice also exhibited significantly reduced cardiac fibrosis and improved sarcolemmal integrity. This work demonstrates that high levels of cardiac dystrophin restored by Pip peptide-AOs prevents further deterioration of cardiomyopathy and pathology following exercise in dystrophic DMD mice.

No MeSH data available.


Related in: MedlinePlus