Li J, Cong X, Zhang Y, Xiang R L, Mei M, Yang N Y, Su Y C, Choi S, Park K, Zhang L W, Wu L L, Yu G Y
Department of Oral and Maxillofacial Surgery, Peking University School and Hospital of Stomatology, Beijing, China.
Department of Physiology and Pathophysiology, Peking University Health Science Center, Key Laboratory of Molecular Cardiovascular Sciences, Ministry of Education and Beijing Key Laboratory of Cardiovascular Receptors Research, Beijing, China.
J Dent Res. 2015 Dec;94(12):1748-56. doi: 10.1177/0022034515609268. Epub 2015 Oct 13.
The tight junction-based paracellular pathway plays an important role in saliva secretion. Zonula occludens (ZO) proteins are submembranous proteins of tight junction complex; however, their function in salivary epithelium is poorly understood. Here, we found that activation of transient receptor potential vanilloid subtype 1 (TRPV1) by capsaicin increased rat saliva secretion both in vivo and ex vivo. Meanwhile, TRPV1 activation enlarged the width of tight junctions between neighboring acinar cells, increased the paracellular flux of 4-kDa fluorescein isothiocyanate (FITC)-dextran in submandibular gland (SMG) tissues, and decreased transepithelial electric resistance (TER) in SMG-C6 cells. ZO-1, -2, and -3 were distributed principally to the apical lateral region of acinar cells in SMG tissues and continuously encircled the peripheries of SMG-C6 cells in the untreated condition. TRPV1 activation obviously diminished ZO-1 and -2 staining, but not ZO-3 or β-catenin, at the cell-cell contacts ex vivo and in vitro. Moreover, in untreated SMG-C6 cells, ZO-1 and -2 single or double knockdown by small interfering RNA (siRNA) increased the paracellular flux of 4-kDa FITC-dextran. In capsaicin-treated cells, ZO-1 and -2 single or double knockdown abolished, whereas their re-expression restored, the capsaicin-induced increase in paracellular permeability. Furthermore, TRPV1 activation increased RhoA activity, and inhibition of either RhoA or Rho kinase (ROCK) abolished the capsaicin-induced TER decrease as well as ZO-1 and -2 redistribution. These results indicate that ZO-1 and -2 play crucial roles in both basal salivary epithelial barrier function and TRPV1-modulated paracellular transport. RhoA-ROCK signaling pathway is responsible for TRPV1-modulated paracellular permeability as well as ZO-1 and -2 redistribution.
基于紧密连接的细胞旁途径在唾液分泌中起重要作用。闭合蛋白（ZO）是紧密连接复合物的膜下蛋白；然而，它们在唾液上皮中的功能尚不清楚。在此，我们发现辣椒素激活瞬时受体电位香草酸亚型1（TRPV1）可在体内和体外增加大鼠唾液分泌。同时，TRPV1激活扩大了相邻腺泡细胞间紧密连接的宽度，增加了下颌下腺（SMG）组织中4 kDa异硫氰酸荧光素（FITC）-葡聚糖的细胞旁通量，并降低了SMG-C6细胞的跨上皮电阻（TER）。ZO-1、-2和-3主要分布于SMG组织腺泡细胞的顶端外侧区域，在未处理条件下持续环绕SMG-C6细胞的周边。TRPV1激活在体内外细胞间接触处明显减少了ZO-1和-2的染色，但不影响ZO-3或β-连环蛋白。此外，在未处理的SMG-C6细胞中，小干扰RNA（siRNA）单独或双重敲低ZO-1和-2可增加4 kDa FITC-葡聚糖的细胞旁通量。在辣椒素处理的细胞中，ZO-1和-2单独或双重敲低可消除辣椒素诱导的细胞旁通透性增加，而它们的重新表达可恢复这种增加。此外，TRPV1激活增加了RhoA活性，抑制RhoA或Rho激酶（ROCK）可消除辣椒素诱导的TER降低以及ZO-1和-2的重新分布。这些结果表明，ZO-1和-2在基础唾液上皮屏障功能和TRPV1调节的细胞旁转运中均起关键作用。RhoA-ROCK信号通路负责TRPV1调节的细胞旁通透性以及ZO-1和-2的重新分布。
