Inhibition of Na+ Transport in Lung Epithelial Cells by Respiratory Syncytial Virus Infection
Inhibition of Na+ Transport in Lung Epithelial Cells by Respiratory Syncytial Virus Infection
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DOI：
10.1165/rcmb.2008-0034oc
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发表时间：
2009-05-01
影响因子：
6.4
通讯作者：
Matalon, Sadis
中科院分区：
文献类型：
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作者：
Chen, Lan;Song, Weifeng;Matalon, Sadis
我们研究了呼吸道合胞病毒（RSV）感染降低呼吸道上皮细胞Na+转运的机制。将来自BALB/c或C57 BL/6小鼠的小鼠气管上皮（MTE）细胞和人气道H441细胞在气-液界面下的半透性支持物上生长。用RSV-A2感染细胞并将其固定在用于测量短路电流（If）的Ussing室中。用RSV感染24小时（感染复数= 1）导致在少于10%的MTE或H441细胞中RSV抗原的阳性免疫荧光。尽管感染的细胞数量有限，但RSV使MTE和H441细胞中的基础和阿米洛利敏感性I-sc均降低约50%，而不引起跨上皮阻力的伴随降低。增加细胞内CAMP的试剂（毛喉素、cpt-CAMP和IBMX）主要增加MTE细胞中的Cl-分泌和H441细胞中的Na+吸收。RSV感染24小时使这两个变量减弱。相比之下，哇巴因敏感的I-sc，在顶部透化的H441单层测量，保持不变。H441细胞裂解物的蛋白质印迹分析证明在RSV感染后24小时α-而非γ-ENaC亚基蛋白水平降低。通过用从头嘧啶或嘌呤合成抑制剂（A77-1726和6-MP，分别为50 μ M）预处理，可防止H441细胞中阿米洛利敏感性I-sc的减少。我们的研究结果表明，感染RSV的小鼠和人呼吸道上皮细胞通过核苷酸释放抑制载体Na+转运。这些发现与我们先前的研究一致，表明BALB/c小鼠RSV感染后肺泡液清除率降低。
We investigated the mechanisms by which respiratory syncytial virus (RSV) infection decreases vectorial Na+ transport across respiratory epithelial cells. Mouse tracheal epithelial (MTE) cells from either BALB/c or C57BL/6 mice and human airway H441 cells were grown on semipermeable supports under an air-liquid interface. Cells were infected with RSV-A2 and mounted in Ussing chambers for measurements of short-circuit currents (If,). Infection with RSV for 24 hours (multiplicity of infection = 1) resulted in positive immunofluorescence for RSV antigen in less than 10% of MTE or H441 cells. In spite of the limited number of cells infected, RSV reduced both basal and amiloride-sensitive I-sc in both MTE and H441 cells by approximately 50%, without causing a concomitant reduction in transepithelial resistance. Agents that increased intracellular CAMP (forskolin, cpt-CAMP, and IBMX) increased mainly CI- secretion in MTE cells and Na+ absorption in H441 cells. RSV infection for 24 hours blunted both variables. In contrast, ouabain sensitive I-sc, measured across apically permeabilized H441 monolayers, remained unchanged. Western blot analysis of H441 cell lysates demonstrated reductions in alpha- but not gamma-ENaC subunit protein levels at 24 hours after RSV infection. The reduction in amiloride-sensitive I-sc in H441 cells was prevented by pretreatment with inhibitors of de novo pyrimidine or purine synthesis (A77-1726 and 6-MP, respectively, 50 mu M). Our results suggest that infection of both murine and human respiratory epithelial cells with RSV inhibits vectorial Na+ transport via nucleotide release. These findings are consistent with our previous studies showing reduced alveolar fluid clearance after RSV infection of BALB/c mice.