New genotype-phenotype correlations and transcriptomic findings in limb-girdle muscular dystrophy R9.

Objective: We aimed to comprehensively investigate the clinical, genetic, muscle imaging, and pathological characteristics of limb-girdle muscular dystrophy R9 (LGMDR9) in a large cohort of Chinese patients. In addition, we sought to delineate the muscle transcriptomic landscape of LGMDR9 which has not been investigated.

Methods: In total, 44 genetically confirmed Chinese LGMDR9 patients were enrolled. They underwent a detailed clinical, imaging, and pathological assessment, followed by customized bioinformatics analyses of genome-wide transcriptome data.

Results: LGMDR9 patients presented with heterogeneous clinical manifestations, including hyper-creatine kinase-emia without weakness (n = 15), as well as mild (n = 11), moderate (n = 7), and severe (n = 11) weakness subgroups determined by hierarchical analysis. Eleven of the 35 pathogenic FKRP variants identified in our cohort were novel, with the c.545A > G variant being the most common found in 72.7% (32/44) patients. Hierarchical analysis revealed that 15 patients harboring null variants (frameshift, nonsense, and large deletions) exhibited a more severe phenotype compared to those with missense/inframe variants in FKRP. Muscle biopsy showed a dystrophic pattern in 19 patients, necrotizing myopathy in 5 patients, and mild myopathic changes in 9 patients. Muscle magnetic resonance imaging analysis showed a concentric pattern of fatty infiltration in 60.7% (17/28) patients. Transcriptomic profiling of 12 muscle samples showed significant upregulation of genes related to inflammation/immune response and extracellular matrix remodeling (P < 0.05). Furthermore, weighted gene co-expression network analysis identified a "turquoise" module enriched in immune cell proliferation and inflammatory markers, which were strongly correlated with histopathological inflammatory scores validated by immunohistochemical staining.

Conclusion: Our findings indicate that null variants in FKRP may be associated with a severe phenotype. We also provide the first transcriptomic and experimental evidence of an immune-activated inflammatory microenvironment in LGMDR9 muscle tissue, which support the potential utility of immunomodulatory therapies in managing this condition.