RS4651 suppresses lung fibroblast activation via the TGF-β1/SMAD signalling pathway

https://doi.org/10.1016/j.ejphar.2021.174135Get rights and content
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Abstract

Background

Idiopathic pulmonary fibrosis (IPF) is a progressive disease resulting in respiratory failure with no efficient treatment options. We investigated the protective effect of RS4651 on pulmonary fibrosis in mice and the mechanism.

Methods

Intratracheal injection of bleomycin (BLM) was used to induce pulmonary fibrosis in mice. RS4561 was administered intraperitoneally at different doses. Histopathological changes were observed. The level of alpha-smooth muscle actin (α-SMA) were also tested. In vitro, the proliferation and migratory effects of RS4651 treatment on MRC-5 cells pre-treated with transforming growth factor (TGF-β1) were examined. RNA-sequencing was used to detect differentially expressed target genes. Then, the expression of α-SMA, pSMAD2 and SMAD7 were analysed during RS4651 treatment of MRC-5 cells with or without silencing by SMAD7 siRNA.

Results

Histopathological staining results showed decreased collagen deposition in RS4651 administered mice. Additionally, a lower level of α-SMA was also observed compared to the BLM group. The results of in vitro studies confirmed that RS4651 can inhibit the proliferation and migration, as well as α-SMA and pSMAD2 expression in MRC-5 cells treated with TGF-β1. RNA-sequencing data identified the target gene SMAD7. We found that RS4651 could upregulate SMAD7 expression and inhibit the proliferation and migration of MRC-5 cells via SMAD7, and RS4651 inhibition of α-SMA and pSMAD2 expression was blocked in SMAD7-siRNA MRC-5 cells. In vivo studies further confirmed that RS4651 could upregulate SMAD7 expression in BLM-induced lung fibrosis in mice.

Conclusions

Our data suggest that RS4651 alleviates BLM-induced pulmonary fibrosis in mice by inhibiting the TGF-β1/SMAD signalling pathway.

Keywords

Pulmonary fibrosis
RS4651
α-SMA
TGF-β1
SMAD7

Abbreviations

IPF
idiopathic pulmonary fibrosis
TGF
transforming growth factor
ECM
extracellular matrix
H&E
haematoxylin and eosin
BLM
bleomycin
IOD
integrated optical density
α-SMA
alpha-smooth muscle actin
MEM
minimum essential medium
FBS
foetal bovine serum
SD
standard deviation
siSMAD7
SMAD7 -siRNA
I.T
intratracheal
MFBs
myofibroblasts
RS
RS4651
PCR
polymerase chain reaction
RT-PCR
real-time PCR
IHC
immunohistochemistry
siRNA
small interfering RNA

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