Role of carbonic anhydrase IV in corneal endothelial HCO3- transport.
Role of carbonic anhydrase IV in corneal endothelial HCO3- transport.
复制标题
碳酸酐酶 IV 在角膜内皮 HCO3- 转运中的作用。
DOI：
10.1167/iovs.07-1188
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发表时间：
2008
影响因子：
4.4
通讯作者：
Bonanno,JosephA
中科院分区：
文献类型：
--
作者：
Sun,XingCai;Li,Jinhua;Cui,Miao;Bonanno,JosephA
目的.碳酸酐酶活性在角膜内皮功能中具有核心作用。作者研究了碳酸酐酶IV（CAIV）在促进CO2通量、HCO 3−渗透性和HCO 3−通量穿过顶膜中的作用。牛角膜内皮细胞的原代培养物建立在膜渗透性过滤器上。通过苯甲酰胺或siRNA敲低CAIV抑制顶端CAIV。顶端CO2通量和HCO 3−渗透性通过测量pH i变化来确定，pH i变化是对改变跨顶端膜的CO2或HCO 3−梯度的响应。通过在基底外侧富含碳酸氢盐的林格氏液存在下测量弱缓冲顶浴的pH值，测定基底外侧至顶（B至A）HCO 3−通量。此外，在DMEM中孵育4小时后，测量苯甲酰胺和CAIV敲低对稳态ΔpH（顶侧-基底侧室pH）的影响。CAIV表达得到证实，CAIV仅定位于顶端膜共聚焦显微镜。10 μM苯甲酰胺和CAIV siRNA均使表观顶端CO2通量降低约20%;然而，它们对HCO 3−渗透性或HCO 3−通量没有影响。稳定状态的顶端-基底侧pH梯度在4小时分别减少了0.12和0.09 pH单位在苯甲酰胺和siRNA处理的细胞，不一致的净细胞到顶端室CO2通量。CAIV不促进稳态细胞至心尖CO2通量、心尖HCO 3−渗透性或B至A HCO 3−通量。然而，稳态pH值的变化表明，CAIV可能有缓冲顶端表面的作用。
purpose. Carbonic anhydrase activity has a central role in corneal endothelial function. The authors examined the role of carbonic anhydrase IV (CAIV) in facilitating CO 2 flux, HCO 3− permeability, and HCO 3− flux across the apical membrane.methods. Primary cultures of bovine corneal endothelial cells were established on membrane-permeable filters. Apical CAIV was inhibited by benzolamide or siRNA knockdown of CAIV. Apical CO 2 fluxes and HCO 3− permeability were determined by measuring pH i changes in response to altering the CO 2 or HCO 3− gradient across the apical membrane. Basolateral to apical (B-to-A) HCO 3− flux was determined by measuring the pH of a weakly buffered apical bath in the presence of basolateral bicarbonate-rich Ringer solution. In addition, the effects of benzolamide and CAIV knockdown on steady state ΔpH (apical-basolateral compartment pH) after 4-hour incubation in DMEM were measured.results. CAIV expression was confirmed, and CAIV was localized exclusively to the apical membrane by confocal microscopy. Both 10 μM benzolamide and CAIV siRNA reduced apparent apical CO 2 flux by approximately 20%; however, they had no effect on HCO 3− permeability or HCO 3− flux. The steady state apical-basolateral pH gradient at 4 hours was reduced by 0.12 and 0.09 pH units in benzolamide-and siRNA-treated cells, respectively, inconsistent with a net cell-to-apical compartment CO 2 flux.conclusions. CAIV does not facilitate steady state cell-to-apical CO 2 flux, apical HCO 3− permeability, or B-to-A HCO 3− flux. Steady state pH changes, however, suggest that CAIV may have a role in buffering the apical surface.