Mendoza Carmen, Matheus Nyurky, Latorre Eva, Castro Marta, Mesonero José Emilio, Alcalde Ana Isabel
Department of Pharmacology and Physiology, Faculty of Veterinary Sciences, University of Zaragoza, Zaragoza, Spain.
Cell Physiol Biochem. 2012;30(1):187-98. doi: 10.1159/000339057. Epub 2012 Jun 18.
Serotonin, a neurotransmitter/autocrineagent mainly synthesized by intestinal enterochromaffin cells, regulates the whole intestinal physiology. Toll-like receptor 3 (TLR3) also contributes to the intestinal physiology by modulating intestinal innate immunity responses. Both serotonin and TLR3 are involved in intestinal inflammatory processes; however, the role of TLR3 in the regulation of intestinal 5-HT availability remains unexplored. The present study analyzes the effect of TLR3 activation on serotonin transporter (SERT) activity in Caco-2 cells. Treatment with poly(I:C), dsRNA synthetic analogue and TLR3 ligand, was assayed and SERT activity determined by 5-HT uptake and transepithelial flux. SERT expression was analyzed by qRT-PCR and western blotting. Poly(I:C) short-term treatment inhibited SERT activity in the apical and basal membrane of epithelial cells and diminished SERT protein content in the membrane. SERT total protein and mRNA levels were not affected by poly(I:C), suggesting a post-translational alteration of SERT. The poly(I:C) effect on SERT activity did not appear to be mediated by PKC, cAMP, PKR or JNK signaling pathways; however, the p38 MAPK pathway seemed to be involved. Our results demonstrate that TLR3 inhibits SERT activity, which may increase 5-HT extracellular levels and contribute to the inflammatory response; however, 5-HT treatment did not affect TLR3 expression.
血清素是一种主要由肠道肠嗜铬细胞合成的神经递质/自分泌因子，可调节整个肠道生理功能。Toll样受体3（TLR3）也通过调节肠道固有免疫反应对肠道生理功能发挥作用。血清素和TLR3均参与肠道炎症过程；然而，TLR3在调节肠道5-羟色胺（5-HT）可用性方面的作用仍未得到探索。本研究分析了TLR3激活对Caco-2细胞中5-羟色胺转运体（SERT）活性的影响。采用双链RNA合成类似物及TLR3配体聚肌苷酸-聚胞苷酸（poly(I:C)）进行处理，并通过5-HT摄取和跨上皮通量测定SERT活性。通过定量逆转录聚合酶链反应（qRT-PCR）和蛋白质免疫印迹法分析SERT表达。Poly(I:C)短期处理可抑制上皮细胞顶端和基底膜中的SERT活性，并降低膜中SERT蛋白含量。Poly(I:C)不影响SERT总蛋白和mRNA水平，提示SERT存在翻译后改变。Poly(I:C)对SERT活性的影响似乎不是由蛋白激酶C（PKC）、环磷酸腺苷（cAMP）、蛋白激酶R（PKR）或应激活化蛋白激酶（JNK）信号通路介导的；然而，p38丝裂原活化蛋白激酶（p38 MAPK）通路似乎参与其中。我们的结果表明，TLR3抑制SERT活性，这可能会增加细胞外5-HT水平并促进炎症反应；然而，5-HT处理不影响TLR3表达。
