Ramírez-Rodríguez Gerardo, Ortiz-López Leonardo, Benítez-King Gloria
Departamento de Neurofarmacología, Subdirección de Investigaciones Clínicas, Instituto Nacional de Psiquiatría Ramón de la Fuente Muñiz, Mexico City, Mexico.
J Pineal Res. 2007 Mar;42(2):180-90. doi: 10.1111/j.1600-079X.2006.00404.x.
Melatonin cyclically modifies water transport measured as dome formation in MDCK cells. An optimal increase in water transport, concomitant with elevated stress fiber (SF) formation, occurs at nocturnal plasma melatonin concentrations (1 nm) after 6 hr of incubation. Blockage in melatonin-elicited dome formation was observed with protein kinase C (PKC) inhibitors. Despite, this information on the precise mechanism by which melatonin increases SF formation involved in water transport is not known. Focal adhesion contacts (FAC) are cytoskeletal structures, which participate in MDCK membrane polarization. SF organization and vinculin phosphorylation are involved in FAC assembly and both processes are mediated by PKC, an enzyme stimulated by melatonin; in these processes also involved is Rho-associated kinase (ROCK). Thus, we studied FAC formation and the ROCK/PKC pathway as the mechanism by which melatonin increases SF formation and water transport. The results showed that 1 nM melatonin and the PKC agonist phorbol-12-miristate-13-acetate increased FAC. The PKC inhibitor GF109203x, and the ROCK inhibitor Y27632, blocked increased FAC caused by melatonin. ROCK and PKC activities, vinculin phosphorylation and FAC formation were increased with melatonin. The PKC inhibitor, GF109203x, abolished both melatonin stimulated FAC in whole cells and ROCK activity, indicating that ROCK is a downstream kinase in the melatonin-stimulated PKC pathway in MDCK cultured cells that causes an increase in SF and FAC formation. Data also document that melatonin modulates water transport through modifications of the cytoskeletal structure.
褪黑素可周期性地改变水转运，这种水转运以MDCK细胞中穹顶形成来衡量。在孵育6小时后，夜间血浆褪黑素浓度（1 nM）时会出现水转运的最佳增加，同时应激纤维（SF）形成增加。用蛋白激酶C（PKC）抑制剂可观察到褪黑素诱导的穹顶形成受阻。尽管如此，关于褪黑素增加参与水转运的SF形成的确切机制尚不清楚。粘着斑接触（FAC）是细胞骨架结构，参与MDCK膜极化。SF组织和纽蛋白磷酸化参与FAC组装，这两个过程均由PKC介导，PKC是一种受褪黑素刺激的酶；在这些过程中还涉及Rho相关激酶（ROCK）。因此，我们研究了FAC形成以及ROCK/PKC途径，以此作为褪黑素增加SF形成和水转运的机制。结果表明，1 nM褪黑素和PKC激动剂佛波醇-12-肉豆蔻酸酯-13-乙酸酯可增加FAC。PKC抑制剂GF109203x和ROCK抑制剂Y27632可阻断褪黑素引起的FAC增加。褪黑素可增加ROCK和PKC活性、纽蛋白磷酸化以及FAC形成。PKC抑制剂GF109203x消除了褪黑素在全细胞中刺激的FAC以及ROCK活性，表明ROCK是MDCK培养细胞中褪黑素刺激的PKC途径中的下游激酶，该途径会导致SF和FAC形成增加。数据还证明，褪黑素通过改变细胞骨架结构来调节水转运。
