Hardison W G, Lowe P J, Shanahan M
Department of Medicine, Veterans Administration Medical Center, San Diego, California 92161.
Hepatology. 1989 Jun;9(6):866-71. doi: 10.1002/hep.1840090613.
The permeability pathway into the biliary tree for small inert molecules exhibits a charge selectivity. Using a method which distinguishes trans- from paracellular access, we have examined the charge selectivity of biliary access pathways for the 40-kD protein horseradish peroxidase (pI 7.5), which was derivatized to strongly anionic (pI less than 3.5) and strongly cationic (pI greater than 9.5) isoenzymes. Each isoenzyme was injected as a bolus into the perfusate of an isolated rat liver perfused in situ with a nonrecirculating Krebs-Ringer buffer. Bile was collected at intervals and horseradish peroxidase activity was measured. Its appearance allowed differentiation of paracellular from transcellular access, and the amount entering via each pathway was quantified. The species of enzyme entering bile was the same as that injected as determined by cation-exchange high-performance liquid chromatography of biliary horseradish peroxidase. Paracellular biliary access of anionic horseradish peroxidase was less than 50% that of neutral and cationic horseradish peroxidase both in the control state and when paracellular entry was augmented with 10(-10) M vasopressin. Transcellular access of anionic horseradish peroxidase was similarly restricted. To determine whether this restriction of anionic transcellular access was brought about by diminished hepatocellular uptake or augmented catabolism, we studied these parameters in 4-hr primary hepatocyte cultures. The uptake rates of all species were similar. Little or no degradation or efflux of any horseradish peroxidase species occurred over 30 min in the cultured cells. We conclude that access is charge selective for macromolecules and that this selectivity holds for trans- as well as for paracellular pathways.(ABSTRACT TRUNCATED AT 250 WORDS)
小的惰性分子进入胆管树的渗透途径表现出电荷选择性。我们采用一种能区分跨细胞途径与细胞旁途径的方法，研究了40kD蛋白辣根过氧化物酶（pI 7.5）进入胆管途径的电荷选择性，该蛋白被衍生化为强阴离子型（pI小于3.5）和强阳离子型（pI大于9.5）同工酶。将每种同工酶以团注形式注入原位灌注的分离大鼠肝脏的灌流液中，灌流液为非循环的 Krebs-Ringer 缓冲液。每隔一段时间收集胆汁并测量辣根过氧化物酶活性。其出现可区分细胞旁途径与跨细胞途径，且可对通过每条途径进入的量进行定量。通过胆汁中辣根过氧化物酶的阳离子交换高效液相色谱法测定，进入胆汁的酶种类与注入的相同。在对照状态下以及当细胞旁进入因10(-10)M血管加压素而增加时，阴离子型辣根过氧化物酶的细胞旁胆管进入量均不到中性和阳离子型辣根过氧化物酶的50%。阴离子型辣根过氧化物酶的跨细胞途径进入同样受限。为确定这种阴离子跨细胞进入的限制是由肝细胞摄取减少还是分解代谢增强引起的，我们在原代肝细胞4小时培养物中研究了这些参数。所有种类的摄取率相似。在培养细胞中，30分钟内任何辣根过氧化物酶种类几乎没有降解或流出。我们得出结论，大分子的进入具有电荷选择性，且这种选择性在跨细胞途径和细胞旁途径中均成立。（摘要截短至250字）
