TITLE:
M3 Muscarinic Acetylcholine Receptor Antagonist Darifenacin Protects against Pulmonary Fibrosis through ERK/NF-κB/miR-21 Pathway
AUTHORS:
Ying Liu, Yanan Jiang, Chao Wang, Haiying Zhang, Yan Liu
KEYWORDS:
Pulmonary Fibrosis, M3 Muscarinic Acetylcholine Receptor, Darifenacin
JOURNAL NAME:
American Journal of Molecular Biology,
Vol.12 No.2,
April
6,
2022
ABSTRACT: Idiopathic pulmonary
fibrosis is an untreatable lethal lung disease, which is
related to the aberrant proliferation of fibroblasts. M3 muscarinic
acetylcholine receptor (M3-mAChR) activation exerts proliferative
effect on various kinds of cells. However, whether M3-mAChR
inhibition has a protective effect on pulmonary fibrosis remains unexplored. A rat model of pulmonary fibrosis was established by
intratracheal instillation of bleomycin. Darifenacin was used to block M3-mAChR.
Histological changes were observed using Masson’s Trichrome and hematoxylin and
eosin (HE) staining. Hydroxyproline was measured by Hydroxyproline detection
kit. Transforming growth factor β1
(TGF-β1) and tumor necrosis factor-α (TNF-α) were measured by enzyme-linked immunosorbent assay (ELISA). In vitro, pulmonary fibroblasts were
isolated from lungs of neonatal rat. After treatment, the cell viability,
Hydroxyproline level was measured by MTT and Hydroxyproline detection kit
respectively. The expression level of extracellular signal-regulated kinase
(ERK), nuclear factor kappa-B (N-NF-κB), and
microRNA-21 (miR-21) was detected by western blot or quantitative real-time PCR
(qRT-PCR). Darifenacin relieved the fibrotic effects provoked by bleomycin. The
expression level of hydroxyproline, TGF-β1
and TNF-α level was all downregulated
after darifenacin treatment. In lung fibroblasts, darifenacin decreased cell
viability and hydroxyproline level induced by bleomycin. Besides,
phosphorylation-ERK and nuclear N-NF-κB protein level was downregulated, as
well as miR-21 level. M3-mAChR antagonist darifenacin
attenuates bleomycin-induced pulmonary fibrosis in rats, which may relate to
the ERK/NF-κB/miRNA-21 signaling pathway.