Potter Elizabeth A, Stewart Gavin, Smith Craig P
Faculty of Life Sciences, University of Manchester, G.38, Stopford Bldg., Oxford Road, Manchester, M13 9PT, UK.
Am J Physiol Renal Physiol. 2006 Jul;291(1):F122-8. doi: 10.1152/ajprenal.00423.2005. Epub 2006 Jan 31.
In this study, we engineered a Madin-Darby canine kidney (MDCK) type I cell line to stably express the mouse urea transporter UT-A2. Monolayers of MDCK-mUT-A2 cells had a basal phloretin-inhibitable urea permeability of 8.4x10(-6)+/-0.3 cm/s. Treatment of MDCK-mUT-A2 monolayers with AVP led to a rapid dose-dependent increase in trans-monolayer phloretin-inhibitable urea flux. The temporal pattern of response was markedly different from that observed for MDCK cells expressing rat UT-A1. Exposure of MDCK-mUT-A2 cells to either 10 microM forskolin or 250 microM 8-bromo cAMP also increased urea flux rate. Inclusion of the PKA inhibitor H89 (10 microM) had no effect on the forskolin-stimulated increase in urea flux across MDCK-mUT-A2 monolayers. Treatment with either 10 microM CPA or 1 mM ATP also caused an increase in UT-A2-mediated urea flux, although these responses where transient compared with those induced by AVP or elevated cAMP. Taken together, these results show for the first time that UT-A2 is acutely sensitive to AVP, cAMP, or increased intracellular calcium.
在本研究中，我们构建了一种稳定表达小鼠尿素转运体UT - A2的I型马-达二氏犬肾（MDCK）细胞系。MDCK - mUT - A2细胞单层的根皮素抑制性尿素基础通透性为8.4×10⁻⁶±0.3 cm/s。用抗利尿激素（AVP）处理MDCK - mUT - A2细胞单层导致跨细胞单层的根皮素抑制性尿素通量迅速呈剂量依赖性增加。其反应的时间模式与表达大鼠UT - A1的MDCK细胞所观察到的明显不同。将MDCK - mUT - A2细胞暴露于10微摩尔的福斯高林或250微摩尔的8 - 溴环磷酸腺苷（8 - bromo cAMP）也会增加尿素通量率。加入蛋白激酶A（PKA）抑制剂H89（10微摩尔）对福斯高林刺激的MDCK - mUT - A2细胞单层尿素通量增加没有影响。用10微摩尔的环磷腺苷（CPA）或1毫摩尔的三磷酸腺苷（ATP）处理也会导致UT - A2介导的尿素通量增加，尽管与AVP或升高的环磷酸腺苷（cAMP）诱导的反应相比，这些反应是短暂的。综上所述，这些结果首次表明UT - A2对AVP、环磷酸腺苷（cAMP）或细胞内钙增加具有急性敏感性。
