Abstract
Mesenchymal stem cells (MSCs) are capable of homing injury sites to exert anti-inflammatory as well as anti-damage effects and can be used as a vehicle for gene therapy. Angiotensin-converting enzyme 2 (ACE2) plays an important role in numerous inflammatory diseases, but fewer studies have been reported in animal mastitis. We hypothesized that MSCs overexpressing ACE2 is more effective in ameliorating lipopolysaccharide (LPS)-induced inflammatory injury in mammary epithelial cells compared to MSCs alone. The results showed that MSC-ACE2 inhibited the LPS induction by upregulation of TNF-α, IL-Iβ, IL-6, and iNOS mRNA expression levels in EpH4-Ev cells compared with MSCs. Furthermore, results showed that both MSC and MSC-ACE2 were significantly activated IL-10/STAT3/SOCS3 signaling pathway as well as inhibited TLR4/NF-κB and MAPK signaling pathways, but MSC-ACE2 had more significant effects. Meanwhile, MSC-ACE2 promoted the expression of proliferation-associated proteins and inhibited the expression of the apoptosis-associated proteins in EpH4-Ev cells. In addition, MSC and MSC-ACE2 reversed the LPS-induced downregulation expression levels of the tight junction proteins in mammary epithelial cells, indicating that both MSC as well as MSC-ACE2 could promote blood-milk barrier repair, and MSC-ACE2 was more effective. These results suggested that MSCs overexpressing ACE2 were more anti-inflammatory as well as anti-injurious action into LPS-induced inflammatory injury in the EpH4-Ev cells. Thus, MSCs overexpressing ACE2 is expected to serve as a potential strategy for mastitis treatment.
Highlights
Mastitis is an inflammatory disease caused by a variety of pathogens, the highest incidence of which is Staphylococcus aureus, E. coli, streptococcus uberis, etc. [1]
The results (Figure 2) showed that NAGase activity in the cell culture supernatant was significantly increased by 1 mg/mL LPS treatment of EpH4-Ev cells for 9 h (*P < 0.05 vs. EpH4-Ev), while NAGase activity was significantly decreased by co-culture with Mesenchymal stem cells (MSCs) or MSC-GFP group (#P < 0.05 vs. LPS)
The inhibition effect was stronger in MSC-Angiotensin-converting enzyme 2 (ACE2) group over in MSC as well as MSCs were transfected with carrying GFP (MSCGFP) groups ($P < 0.05 vs. MSC; &P < 0.05 vs. MSC-GFP). These results suggested that MSC-ACE2 can better inhibit the stimulation of toll receptor-4 (TLR4)/MAPK/nuclear factor-kB (NF-kB) signaling pathway to resist LPS-induced inflammation in EpH4-Ev cells
Summary
Mastitis is an inflammatory disease caused by a variety of pathogens, the highest incidence of which is Staphylococcus aureus, E. coli, streptococcus uberis, etc. [1]. Mastitis is an inflammatory disease caused by a variety of pathogens, the highest incidence of which is Staphylococcus aureus, E. coli, streptococcus uberis, etc. Lipopolysaccharide (LPS), a major component of the cell wall of Gram-negative bacteria, is often used to model mastitis. In this study, we will use LPS to establish a typical model of inflammation in EpH4-Ev cells (Mouse mammary epithelial cells). Li et al discovered that ACE2/Ang1-7/Mas prohibited LPSprompted apoptosis of microvascular pulmonary endothelial cells by hindering JNK/NF-kB pathway [6]. Wang et al reported that ACE2/Ang1-7/Mas might considerably restrain pancreatitis and decrease the discharge of inflammatory cytokines [7]. Our previous study showed that LPS treatment of EpH4Ev cells for 9 h showed a trend of ACE2 first increasing and decreasing, and was involved in LPS-induced injury to EpH4-Ev cells [10]
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