Tombola F, Morbiato L, Del Giudice G, Rappuoli R, Zoratti M, Papini E
Centro Consiglio Nazionale delle Ricerche Biomembrane, Department of Biomedical Sciences, University of Padova, Padova, Italy.
J Clin Invest. 2001 Sep;108(6):929-37. doi: 10.1172/JCI13045.
Urease and the cytotoxin VacA are two major virulence factors of the human pathogen Helicobacter pylori, which is responsible for severe gastroduodenal diseases. Diffusion of urea, the substrate of urease, into the stomach is critically required for the survival of infecting H. pylori. We now show that VacA increases the transepithelial flux of urea across model epithelia by inducing an unsaturable permeation pathway. This transcellular pathway is selective, as it conducts thiourea, but not glycerol and mannitol, demonstrating that it is not due to a loosening of intercellular junctions. Experiments performed with different cell lines, grown in a nonpolarized state, confirm that VacA permeabilizes the cell plasma membrane to urea. Inhibition studies indicate that transmembrane pores formed by VacA act as passive urea transporters. Thus, their inhibition by the anion channel blocker 5-nitro-2-(3-phenylpropylamino) benzoic acid significantly decreases toxin-induced urea fluxes in both polarized and nonpolarized cells. Moreover, phloretin, a well-known inhibitor of eukaryotic urea transporters, blocks VacA-mediated urea and ion transport and the toxin's main biologic effects. These data show that VacA behaves as a low-pH activated, passive urea transporter potentially capable of permeabilizing the gastric epithelium to urea. This opens the novel possibility that in vivo VacA may favor H. pylori infectivity by optimizing urease activity.
脲酶和细胞毒素VacA是人类病原体幽门螺杆菌的两个主要毒力因子，幽门螺杆菌可导致严重的胃十二指肠疾病。尿素作为脲酶的底物，其向胃内的扩散对于感染的幽门螺杆菌的存活至关重要。我们现在表明，VacA通过诱导一条不饱和渗透途径增加了尿素跨模型上皮的跨上皮通量。这种跨细胞途径具有选择性，因为它能传导硫脲，但不能传导甘油和甘露醇，这表明它不是由于细胞间连接的松弛。对以非极化状态生长的不同细胞系进行的实验证实，VacA使细胞质膜对尿素具有通透性。抑制研究表明，VacA形成的跨膜孔作为被动尿素转运体起作用。因此，阴离子通道阻滞剂5-硝基-2-(3-苯基丙基氨基)苯甲酸对它们的抑制作用显著降低了毒素诱导的极化和非极化细胞中的尿素通量。此外，根皮素是一种著名的真核生物尿素转运体抑制剂，它能阻断VacA介导的尿素和离子转运以及毒素的主要生物学效应。这些数据表明，VacA表现为一种低pH激活的被动尿素转运体，可能能够使胃上皮对尿素具有通透性。这开启了一种新的可能性，即体内VacA可能通过优化脲酶活性来促进幽门螺杆菌的感染性。
