Ursodeoxycholic Acid Is Conjugated with Taurine to Promote Secretin-Stimulated Biliary Hydrocholeresis in the Normal Rat

Background & AimsSecretin induces bicarbonate-rich hydrocholeresis in healthy individuals, but not in untreated patients with primary biliary cirrhosis (PBC). Ursodeoxycholic acid (UDCA) – the first choice treatment for PBC – restores the secretin response. Compared with humans, secretin has poor effect in experimental normal-rat models with biliary drainage, although it may elicit hydrocholeresis when the bile-acid pool is maintained. In view of the benefits of UDCA in PBC, we used normal-rat models to unravel the acute contribution of UDCA (and/or taurine-conjugated TUDCA) for eliciting the biliary secretin response. MethodsIntravascular and/or intrabiliary administration of agonists and inhibitors was performed in normal rats with biliary monitoring. Secretin/bile-acid interplay was analyzed in 3D cultured rat cholangiocytes that formed expansive cystic structures with intralumenal hydroionic secretion. ResultsIn vivo, secretin stimulates hydrocholeresis upon UDCA/TUDCA infusion, but does not modify the intrinsic hypercholeretic effect of dehydrocholic acid (DHCA). The former effect is dependent on microtubule polymerization, and involves PKCα, PI3K and MEK pathways, as shown by colchicine (i.p.) and retrograde biliary inhibitors. In vitro, while secretin alone accelerates the spontaneous expansion of 3D-cystic structures, this effect is enhanced in the presence of TUDCA, but not UDCA or DHCA. Experiments with inhibitors and Ca2+-chelator confirmed that the synergistic effect of secretin plus TUDCA involves microtubules, intracellular Ca2+, PKCα, PI3K, PKA and MEK pathways. Gene silencing also demonstrated the involvement of the bicarbonate extruder Ae2. ConclusionsUDCA is conjugated in order to promote secretin-stimulated hydrocholeresis in rats through Ae2, microtubules, intracellular Ca2+, PKCα, PI3K, PKA, and MEK.研究背景与目的 促胰液素（Secretin）可在健康个体中诱导产生富含碳酸氢盐的水利胆作用，但对未接受治疗的原发性胆汁性胆管炎（PBC）患者无此效果。熊去氧胆酸（UDCA）作为PBC的一线治疗药物，可恢复患者对促胰液素的应答反应。与人类相比，促胰液素在伴有胆汁引流的正常大鼠实验模型中作用微弱，仅在胆汁酸池维持稳定的情况下可诱发水利胆作用。鉴于UDCA对PBC的治疗益处，本研究采用正常大鼠模型，旨在阐明UDCA（及/或牛磺酸结合型牛磺熊去氧胆酸TUDCA）在诱发胆汁促胰液素应答反应中的急性作用。 研究方法 本研究对开展胆汁监测的正常大鼠实施了激动剂与抑制剂的血管内及/或胆道内给药。本研究对形成具有腔内水离子分泌特性的扩张性囊状结构的三维培养大鼠胆管上皮细胞，开展了促胰液素与胆汁酸相互作用的相关分析。 研究结果 体内实验结果显示，在输注UDCA或TUDCA后，促胰液素可发挥水利胆作用，但不会改变脱氢胆酸（DHCA）固有的强利胆效应。如秋水仙碱（腹腔注射）及胆道逆行给药抑制剂实验所示，该效应依赖于微管聚合过程，并涉及蛋白激酶Cα（PKCα）、磷脂酰肌醇3-激酶（PI3K）及丝裂原活化蛋白激酶激酶（MEK）信号通路。体外实验中，尽管单独使用促胰液素即可加速三维囊状结构的自发性扩张，但该效应在TUDCA存在时会进一步增强，而UDCA或DHCA则无此效果。采用抑制剂及钙离子螯合剂开展的实验证实，促胰液素与TUDCA的协同效应涉及微管、细胞内钙离子、PKCα、PI3K、蛋白激酶A（PKA）及MEK信号通路。基因沉默实验同样证实，碳酸氢盐外排载体Ae2参与了该调控过程。 研究结论 UDCA需经结合修饰，方可通过Ae2、微管、细胞内钙离子、PKCα、PI3K、PKA及MEK信号通路，促进大鼠体内促胰液素介导的水利胆作用。