Vitamin B12 enhances ZO-1 expression in HDM-treated human airway epithelial cells by down-regulating autophagy
LI Yuejiao
LAN Nan
WANG Xing
TANG Hongmei
WANG Zhibin
ZHANG Yun
YUAN Xiefang
WANG Xiaoyun
Abstract：Objective To investigate the effect of vitamin B12(VB12)on the expression of zonula occludens-1(ZO-1)in house dust mite(HDM)-treated human airway epithelial cell line(Beas-2b)and its underlying mechanism.Methods Beas-2b cells were cultured in DMEM high-glucose medium containing 10%fetal bovine serum.The cells were divided into four groups:control,VB12,HDM,and VB12+HDM.Beas-2b cells were trans-fected with lentiviruses carrying NC-siRNA,ATG5-siRNA,BECN1-siRNA,and mCherry-EGFP-LC3.After 12 hours of transfection(MOI=20),the medium was replaced with fresh medium,and stable transfected cell lines were selected using puromycin(1 µg/mL).Cells were stimulated with VB12(20 µg/mL)and HDM(50 µg/mL)for 24 hours.The protein levels of ZO-1,autophagy-related protein 5(ATG5),BECN1 and microtubule-associated protein light chain 3(LC3)were detected by immunofluorescence and Western blot.Autophagy in human airway epithelial cells was observed using confocal microscopy.Results Compared with the control group,the expression of ZO-1 in the HDM group was lower(P<0.05),while the expressions of ATG5,BECN1,and LC3 were higher(P<0.05).Compared with the HDM group,the VB12+HDM group showed increased ZO-1 expression(P<0.05),decreased expressions of ATG5,BECN1,and LC3(P<0.01),and reduced autophagosome formation(P<0.05).In ATG5-and BECN1-knockdown cell lines,ZO-1 expression increased after HDM treatment(P<0.05).Conclusion Vb12 can enhance ZO-1 expression in HDM-treated human airway epithelial cells by down-regulating autophagy,and its mechanism is associated with the ATG5 and BECN1 signaling pathways.
Keywords：vitamin b12zonula occludens-1autophagyATG5BECN1
Publication Date：2025-11-10
Online Publishing Date：2025-11-20(First online date of this platform, not the publication date of the document)
Pages：7( 3345-3351 )