Genetic deletion of the purinergic receptor P2rx7 worsens the phenotype of α‑sarcoglycan muscular dystrophy

Limb-girdle muscular dystrophy R3 (LGMDR3), a rare genetic disorder characterized by progressive impairment of limb, diaphragmatic, and respiratory muscles, is caused by loss-of-function mutations in the α-sarcoglycan gene (SGCA) and aggravated by immune-mediated damage and fibrotic tissue replacement. Pharmacological inhibition of purinergic receptor P2X7 (P2X7R) reduced inflammation and fibrosis in Sgca(-/-) mice. To further define the role of P2X7R, we generated a double knockout mouse model Sgca(-/-) P2rx7(-/-). We compared diaphragms isolated from 24-week-old Sgca(-/-) P2rx7(+/+) and Sgca(-/-) P2rx7(-/-) mice since the diaphragmatic muscle is early and severely damaged by Sgca genetic loss-of-function. Unexpectedly, Sgca(-/-) P2rx7(-/-) mice displayed increased extracellular matrix deposition and augmented cellularity in fibrotic areas, in particular, a higher number of CD3(+) lymphocytes and Iba1(+) macrophages compared to Sgca(-/-) P2rx7(+/+) mice. Moreover, intense P2X4R signal colocalized with CD3(+) and Iba1(+) cells, confirming its expression by these infiltrating immune cells. Absence of an improvement of the dystrophic phenotype was histologically confirmed in Sgca(-/-) P2rx7(-/-) quadriceps, although the fibrotic reaction was milder than that in diaphragms, suggesting a differential influence of the tissue microenvironment on the receptor functions. Flow cytometric analysis of limb muscle-infiltrating immune cells revealed a decrease in NK cells. Motor performance tests did not reveal any difference between the two genotypes. In conclusion, this study identified a divergent outcome of genetic deletion of the P2rx7 gene as compared to P2X7R blockade in α-sarcoglycan dystrophic tissue, suggesting that pharmacological interventions targeting the P2X7R in dystrophic immune-mediated damage require careful definition of a precise time window and dosage.