Camostat mesilate inhibits pro-inflammatory cytokine secretion and improves cell viability by regulating MFGE8 and HMGN1 in lipopolysaccharide-stimulated DF-1 chicken embryo fibroblasts.

Lin Yuan,Zhishuai Zhang,Mengjie Li,Wei Jin,Wanli Li,Mingfa Wang

doi:10.7717/peerj.12053

Abstract

Camostat mesilate (CM) possesses potential anti-viral and anti-inflammatory activities. However, it remains unknown whether CM is involved in lipopolysaccharide (LPS)-mediated inflammatory responses and cell injury. In this project, differentially expressed proteins (DEPs, fold change ≥ 1.2 or ≤ 0.83 and Q value ≤ 0.05) in response to LPS stimulation alone or in combination with CM were identified through tandem mass tags (TMT)/mass spectrometry (MS)-based proteomics analysis in DF-1 chicken embryo fibroblasts. The mRNA expression levels of filtered genes were determined by RT-qPCR assay. The results showed that CM alleviated the detrimental effect of LPS on cell viability and inhibited LPS-induced TNF-α and IL-6 secretions in DF-1 chicken embryo fibroblasts. A total of 141 DEPs that might be involved in mediating functions of both LPS and CM were identified by proteomics analysis in DF-1 chicken embryo fibroblasts. LPS inhibited milk fat globule EGF and factor V/VIII domain containing (MFGE8) expression and induced high mobility group nucleosome binding domain 1 (HMGN1) expression, while these effects were abrogated by CM in DF-1 chicken embryo fibroblasts. MFGE8 knockdown facilitated TNF-α and IL-6 secretions , reduced cell viability, stimulated cell apoptosis in DF-1 chicken embryo fibroblasts co-treated with LPS and CM. HMGN1 loss did not influence TNF-α and IL-6 secretions, cell viability, and cell apoptosis in DF-1 chicken embryo fibroblasts co-treated with LPS and CM. In conclusion, CM exerted anti-inflammatory and pro-survival activities by regulating MFGE8 in LPS-stimulated DF-1 chicken embryo fibroblasts, deepening our understanding of the roles and molecular basis of CM in protecting against Gram-negative bacteria.

Highlights

Camostat mesilate (CM, FoipanTM), a protease inhibitor, has been used for the treatment of multiple diseases such as chronic pancreatitis and recessive dystrophic epidermolysis bullosa in human (McKee et al, 2020; Ramsey, Nuttall & Hart, 2019; Ikeda et al, 1988)
Our study demonstrated that CM dose-dependently inhibited the secretions of tumor necrosis factor-α (TNF-α) and IL-6 in the LPS (10 μg/ml)-exposed DF-1 cell model (Figs. 1D and 1E)
CM increased cell viability in a time- and concentration-dependent manner in LPS-treated DF-1 chicken embryo fibroblasts (Fig. 1F). These data suggested that CM could protect DF-1 chicken embryo fibroblasts from LPS-induced inflammation and cell injury

Summary

Introduction

Camostat mesilate (CM, FoipanTM), a protease inhibitor, has been used for the treatment of multiple diseases such as chronic pancreatitis and recessive dystrophic epidermolysis bullosa in human (McKee et al, 2020; Ramsey, Nuttall & Hart, 2019; Ikeda et al, 1988). Some studies have proposed that CM (an inhibitor of TMPRSS2 serine protease) might act as a candidate drug against SARS-CoV-2 and COVID-19 given its inhibitory effect on host cell entry of SARS-CoV-2 and its protective effects against SARS-CoV-induced mouse death (McKee et al, 2020; Uno, 2020; Hoffmann et al, 2020). Previous in vitro and in vivo studies have demonstrated that CM exerts anti-inflammatory activity in LPS-activated rat monocytes and chronic pancreatitis and genetically diabetic rat models (Gibo et al, 2005; Jia, Taguchi & Otsuki, 2005). Few studies have been performed currently to investigate the association of CM and Gram-negative bacteria-mediated inflammatory responses and cell injury

Methods

Results

Conclusion