中文摘要：

【摘要】目的探讨盐酸（HCl）刺激与机械牵张诱导人肺上皮细胞向间质转分化以及对透明质酸（HA）分泌的影响。方法体外培养人肺上皮细胞株BEAS-2B，取对数生长期的细胞分为磷酸盐缓冲液（PBS）＋静止组、HCl＋静止组、PBS＋牵张组、HCl＋牵张组。采用FX-5000T细胞应力加载系统向两个牵张组BEAS-2B细胞持续48h施加20％振幅、频率0．33Hz、正弦波的机械牵张。分别于牵张前后用倒置显微镜观察细胞形态变化；用蛋白质免疫印迹试验（WesternBlot）检测细胞上皮标志物E-钙黏蛋白和细胞角蛋白-8（CK-8）以及间质标志物波形蛋白和α-平滑肌肌动蛋白（α-SMA）的蛋白表达；用酶联免疫吸附试验（ELISA）检测细胞培养上清中HA分泌量。结果①镜下观察显示：PBS＋静止组BEAS-2B细胞呈卵圆形，且细胞紧密相连，细胞间存在极性；单纯HCl刺激后细胞形态无明显变化；而单纯给予机械牵张48h后细胞由卵圆形变成长梭形，细胞间隙明显增大；给予HCl刺激和机械牵张双重打击后细胞形态改变更为显著。②WesternBlot显示：与PBS＋静止组比较，单纯或联合HCl和机械牵张48h后BEAS-2B细胞E-钙黏蛋白、CK-8表达均下调，波形蛋白、α-SMA表达均上调，以HCl＋牵张组表达变化更为明显[以PBS＋静止组表达量（灰度值）为基数1，E-钙黏蛋白为0．16±0．08比1，CK-8为0．10±0．03比1，波形蛋白为3．35±0．38比1，α-SMA为3．10±0．45比1，均P〈0．01]。③ELISA试验显示：与PBS＋静止组比较，单纯HCl刺激或机械牵张均可诱导BEAS-2B细胞分泌HA（μg／L：55．763±0．687、63．005±0．493比49．876±1．867）；而HCl与机械牵张联合刺激后细胞培养上清液中HA分泌量增加更为显著（μg／L：78．220±1．085比49．876±1．867，P〈0．01）。结论HCl或机械牵张均能诱导人肺上皮细胞发生上皮向间质转分化，同时增加HA的分?

英文摘要：

Objective To investigate the effect of hydrochloric acid （HCl） stimulation and mechanical stretch on epithelial-mesenchymal transition （EMT） and hyaluronan （HA） production in human lung epithelial ceils. Methods Human lung epithelial cell line BEAS-2B was cultured in vitro, which was divided into phosphate-buffer saline （PBS） ± static group, HCl ± static group, PBS ± stretch group, and HCl ± stretch group respectively in the logarithmic phase. The BEAS-2B cells in two stretching groups were challenged by cyclic stretch with 20% amplitude, frequency of 0.33 Hz, sine wave of the FX-5000T system for 48 hours. The morphology changes in cells before and after stretch were observed with inverted microscope. The protein expressions of epithelial markers E-cadherin and cytokeratin-8 （CK-8） as well as mesenchymal markers vimentin and α-smooth muscle actin （α-SMA） were determined by Western Blot. The secretion of HA was determined by enzyme linked immunosorbent assay （ELISA）.Results ① It was shown by microscopic observation that BEAS-2B cells displayed cobblestone morphology, linked closely and cell polarity in PBS ＋ static group, which did not change obviously after HCl stimulation alone. Given purely mechanical stretch after 48 hours, the cells morphology changed from cobblestone shape into long spindle, and increased intercellular space obviously. Double hit of HCl and stretch changed the cells morphology more significantly. ② It was shown by Western Blot that compared with the PBS ＋ static group, HCl alone or combined with purely mechanical stretch after 48 hours, the expressions of E-cadherin and CK-8 were decreased, while those of vimentin and α -SMA were increased, and it was more pronounced in HCl ± stretch group [the expression quantity （gray value） as base 1 in PBS ± static group, E-cadherin： 0.16±0.08 vs. 1, CK-8：0.10±0.03 vs. 1, vimentin： 3.35±0.38 vs. 1, α-SMA： 3.10 ± 0.45 vs. 1, all P 〈 0,01]. ③ It was shown by ELISA that both HCl stimulation a