Kangfuxin Liquid Ameliorates LPS-induced Inflammatory Injury in Oral Mucosal Epithelial Cells via the miR-449a/Notch1 Axis

doi:10.3870/j.issn.1672-0741.25.10.008

Abstract

Abstract: Objective To investigate the effect of Kangfuxin liquid(KFX)on lipopolysaccharide(LPS)-induced inflammatory injury in oral mucosal epithelial cells(OMECs)and its underlying mechanism. Methods Mouse OMECs(mOMECs)were isolated,identified,and cultured ex vivo,then treated with various concentrations of KFX(0,0.078%,0.156%,0.313%,0.625%,1.25%,2.5%,5%,10%)or LPS(0,0.1,1,10 μg/mL)for 48 h.Cell proliferation and apoptosis were assessed by CCK-8 assay and flow cytometry,respectively,to determine the optimal concentrations of KFX and LPS for intervention.The mOMECs were then divided into the following groups:Control,LPS(1 μg/mL LPS),KFX(0.625% KFX),KFX+negative control inhibitor(KFX+inhibitor-NC),and KFX+miR-449a inhibitor.Cell proliferation was assessed by CCK-8 assay,and apoptosis by flow cytometry.The levels of TNF-α and IL-1β in the supernatant were measured by ELISA.The expression of miR-449a and Notch1 mRNA was determined by qRT-PCR.The protein expression levels of Notch1,cleaved-Caspase-3,Bcl-2,and Bax were detected by Western blotting.Potential binding sites between miR-449a and Notch1 were predicted using bioinformatics tools.The targeting relationship between miR-449a and Notch1 was verified by dual-luciferase reporter assay,qRT-PCR,and Western blotting. Results Compared with the Control group,the LPS group showed significantly decreased cell proliferation,miR-449a expression,and Bcl-2 protein levels(all P<0.05),along with increased apoptosis,TNF-α and IL-1β levels,Notch1 mRNA expression,and Notch1,cleaved-Caspase-3,and Bax protein levels(all P<0.05).Compared with the LPS group,the KFX group exhibited increased cell proliferation,miR-449a expression,and Bcl-2 protein levels(all P<0.05),and decreased apoptosis,TNF-α and IL-1β levels,Notch1 mRNA expression,and Notch1,cleaved-Caspase-3,and Bax protein levels(all P<0.05).Compared with the KFX group,the KFX+miR-449a inhibitor group showed reversed trends in all aforementioned indicators(all P<0.05),whereas no significant changes were observed in the KFX+inhibitor-NC group(P>0.05).Dual-luciferase reporter assay,qRT-PCR,and Western blotting confirmed that miR-449a targeted and inhibited Notch1 expression. Conclusion KFX may ameliorate LPS-induced inflammatory injury in mOMECs via the miR-449a/Notch1 axis.

Key words: Kangfuxin liquid, miR-449a/Notch1 axis, lipopolysaccharide, oral mucosal epithelial cells, inflammatory injury

CLC Number:

R781.05

Wang Zhaoxin, Xiang Guolin. Kangfuxin Liquid Ameliorates LPS-induced Inflammatory Injury in Oral Mucosal Epithelial Cells via the miR-449a/Notch1 Axis[J]. Acta Medicinae Universitatis Scientiae et Technologiae Huazhong, 2026, 55(2): 226-234.

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