Toll-like receptor 2 regulates the barrier function of human bronchial epithelial monolayers through atypical protein kinase C zeta, and an increase in expression of claudin-1.
Toll-like receptor 2 regulates the barrier function of human bronchial epithelial monolayers through atypical protein kinase C zeta, and an increase in expression of claudin-1.
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DOI：
10.4161/tisb.29166
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发表时间：
2014
期刊：
影响因子：
3.1
通讯作者：
Borchard G
中科院分区：
文献类型：
--
作者：
Ragupathy S;Esmaeili F;Paschoud S;Sublet E;Citi S;Borchard G
我们使用人支气管上皮细胞系Calu-3研究了Toll样受体（TLR）2在维持气道上皮屏障完整性中的作用。TLR 2通过其配体Pam 3CysSK 4和肽聚糖的活化显示了上皮屏障功能的浓度依赖性增加，如通过跨上皮电阻（TEER）测量的。这通过荧光素钠的细胞旁通量的减少来证实。这种TLR 2诱导的TEER增加通过用多克隆抗人TLR 2中和抗体预处理而显著降低。TLR 2刺激Calu-3细胞单层导致紧密连接蛋白claudin-1和ZO-1的表达增加，occludin的表达减少，在mRNA和蛋白水平。PKC β的假底物抑制剂显著阻止了TLR 2介导的屏障功能的增加。它还以浓度依赖性方式阻止claudin-1的增加，直至1 µM。TLR 2刺激导致非典型PKC β的磷酸化增加，假底物抑制剂以浓度依赖性方式阻止这种增加。总之，我们的观察结果支持一种模型，即通过增加claudin-1的表达和通过调节PKC β活性来增加由TLR 2激活诱导的紧密连接屏障功能。
We investigated the role of Toll-like receptor (TLR) 2 in maintaining the integrity of the airway epithelial barrier using the human bronchial epithelial cell line Calu-3. Activation of TLR2 by its ligands, Pam3CysSK4 and Peptidoglycan showed a concentration dependent increase in epithelial barrier function, as measured by transepithelial electrical resistance (TEER). This was confirmed by a decrease in paracellular flux of fluorescein sodium. This TLR2 induced increase in TEER was significantly reduced by pretreatment with polyclonal anti-human TLR2-neutralizing antibody. TLR2 stimulation in Calu-3 cell monolayers resulted in an increased expression of the tight junction proteins claudin-1 and ZO-1, and a decreased expression of occludin, at both the mRNA and protein levels. A pseudosubstrate inhibitor to PKCζ significantly prevented the TLR2 mediated increase in barrier function. It also prevented the increase in claudin-1 in a concentration dependent manner up to 1 µM. TLR2 stimulation led to an increase in phosphorylation of atypical PKC ζ, which was prevented by the pseudosubstrate inhibitor in a concentration dependent manner. Taken together, our observations support a model whereby increased tight junction barrier function induced by activation of TLR2 occurs through increased expression of claudin-1, and through modulation of PKC ζ activity.