Maternal smoking during pregnancy aggravated muscle phenotype in FHL1-/y offspring mice similar to congenital clubfoot through P2RX7-mediated pyroptosis

Abstract

Congenital clubfoot (CCF) is a common birth defect. Maternal smoking during pregnancy increases the risk of CCF. In previous research, we found muscle phenotypes similar to CCF in four and a half LIM domain protein 1 (FHLI) offspring mice (FHL1^-/y). However, the role of P2RX7-mediated pyroptosis in the effect of cigarette smoke (CS) on the skeletal muscle of FHL1^-/y mice during pregnancy is unclear. In the present study, pregnant mice at 11 days of gestation were exposed to CS and male offspring of wild-type (WT) and FHL1^-/y mice were divided into four groups (Control-WT, Control-KO, CS-WT, CS-KO). The histomorphology of lower limb muscles was examined using hematoxylin and eosin (H&E) staining. P2RX7, indicators of pyroptosis (NLRP3, ASC, cleaved-caspase 1, IL-1β), and cytoskeletal proteins (MYBPC2, LDB3) were also detected using immunoblotting. CS exposure during pregnancy aggravated the muscle phenotype similar to CCF in FHL1^-/y offspring mice. FHL1 gene knockout (KO) or CS exposure during pregnancy each activated the expression of P2RX7, cell pyroptosis-related proteins (NLRP3, ASC, cleaved-caspase 1, IL-1β), a muscle injury marker (MYOD1), and cytoskeletal proteins (MYBPC2, LDB3); these two factors had an additive effect. The results showed maternal smoking during pregnancy aggravated muscle phenotype similar to CCF in FHL1^-/y offspring mice through P2RX7-mediated pyroptosis.

Keywords: Cigarette smoke; Congenital clubfoot; FHL1; P2RX7; Pyroptosis; Skeletal muscle.