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An integrative correlation of myopathology, phenotype and genotype in late onset Pompe disease
(2019)
Aims: Pompe disease is caused by pathogenic mutations in the alpha 1,4‐glucosidase (GAA) gene and in patients with late onset Pome disease (LOPD), genotype–phenotype correlations are unpredictable. Skeletal muscle pathology includes glycogen accumulation and altered autophagy of various degrees. A correlation of the muscle morphology with clinical features and the genetic background in GAA may contribute to the understanding of the phenotypic variability.
Methods: Muscle biopsies taken before enzyme replacement therapy were analysed from 53 patients with LOPD. On resin sections, glycogen accumulation, fibrosis, autophagic vacuoles and the degree of muscle damage (morphology‐score) were analysed and the results were compared with clinical findings. Additional autophagy markers microtubule‐associated protein 1A/1B‐light chain 3, p62 and Bcl2‐associated athanogene 3 were analysed on cryosections from 22 LOPD biopsies.
Results: The myopathology showed a high variability with, in most patients, a moderate glycogen accumulation and a low morphology‐score. High morphology‐scores were associated with increased fibrosis and autophagy highlighting the role of autophagy in severe stages of skeletal muscle damage. The morphology‐score did not correlate with the patient's age at biopsy, disease duration, nor with the residual GAA enzyme activity or creatine‐kinase levels. In 37 patients with LOPD, genetic analysis identified the most frequent mutation, c.‐32‐13T>G, in 95%, most commonly in combination with c.525delT (19%). No significant correlation was found between the different GAA genotypes and muscle morphology type.
Conclusions: Muscle morphology in LOPD patients shows a high variability with, in most cases, moderate pathology. Increased pathology is associated with more fibrosis and autophagy.
Highlights
• Pediatric patients with RASopathies exhibit an increased heart mass.
• Cardiomyocyte size and volume are reduced in pediatric patients with RASopathies.
• Cardiomyocyte numbers per heart are increased in pediatric RASopathy patients.
• Cardiomyocytes in juvenile RASopathy have an immature state.
• Elevated LV mass in RASopathy patients is mainly due to cardiomyocyte hyperplasia.
Abstract
Aims: RASopathies are caused by mutations in genes that alter the MAP kinase pathway and are marked by several malformations with cardiovascular disorders as the predominant cause of mortality. Mechanistic insights in the underlying pathogenesis in affected cardiac tissue are rare. The aim of the study was to assess the impact of RASopathy causing mutations on the human heart.
Methods and results: Using single cell approaches and histopathology we analyzed cardiac tissue from children with different RASopathy-associated mutations compared to age-matched dilated cardiomyopathy (DCM) and control hearts. The volume of cardiomyocytes was reduced in RASopathy conditions compared to controls and DCM patients, and the estimated number of cardiomyocytes per heart was ∼4–10 times higher. Single nuclei RNA sequencing of a 13-year-old RASopathy patient (carrying a PTPN11 c.1528C > G mutation) revealed that myocardial cell composition and transcriptional patterns were similar to <1 year old DCM hearts. Additionally, immaturity of cardiomyocytes is shown by an increased MYH6/MYH7 expression ratio and reduced expression of genes associated with fatty acid metabolism. In the patient with the PTPN11 mutation activation of the MAP kinase pathway was not evident in cardiomyocytes, whereas increased phosphorylation of PDK1 and its downstream kinase Akt was detected.
Conclusion: In conclusion, an immature cardiomyocyte differentiation status appears to be preserved in juvenile RASopathy patients. The increased mass of the heart in such patients is due to an increase in cardiomyocyte number (hyperplasia) but not an enlargement of individual cardiomyocytes (hypertrophy).