Department of Nutrition, The Affiliated Hospital of Qingdao University, Qingdao, China.
Center for Medical Research, The Affiliated Hospital of Qingdao University, Qingdao, China.
Dig Dis Sci. 2021 Dec;66(12):4251-4262. doi: 10.1007/s10620-020-06817-y. Epub 2021 Feb 2.
Whether 7,8-dihydroxyflavone (7,8-DHF), a tyrosine kinase receptor B (TrkB) agonist, modulates colonic smooth muscle motility and/or alleviates constipation has not yet been studied.
Here, we aimed to determine how 7,8-DHF influences carbachol (CCh)-stimulated contraction of colonic strips and the in vivo effect of 7,8-DHF on constipation.
Muscle strips were isolated from rat colons for recording contractile tension and performing western blotting. Constipation was induced in rats with loperamide.
Although it specifically activated TrkB, 7,8-DHF applied alone neither activated PLCγ1 in the colonic strips nor induced colonic strip contraction. However, 7,8-DHF enhanced CCh-stimulated PLCγ1 activation and strip contraction. The PLCγ1 antagonist U73122 suppressed both CCh-stimulated and 7,8-DHF-enhanced/CCh-stimulated contraction. While clarifying the underlying mechanism, we revealed that 7,8-DHF augmented muscarinic M3 receptor expression in the colonic strips. The M3-selective antagonist tarafenacin specifically inhibited the 7,8-DHF-enhanced/CCh-stimulated contraction of the colonic strips. Since 7,8-DHF increased Akt phosphorylation, and LY294002 (an antagonist of PI3K upstream of Akt) dramatically inhibited both 7,8-DHF-augmented M3 expression and 7,8-DHF-enhanced/CCh-stimulated contractions, we assumed that 7,8-DHF/TrkB/Akt was associated with the modulation of M3 expression in the colonic strips. ANA-12, a specific TrkB antagonist, not only inhibited TrkB activation by 7,8-DHF but also suppressed 7,8-DHF-enhanced cholinergic contraction, 7,8-DHF/CCh-mediated activation of PLCγ1/Akt, and M3 overexpression in colonic strips. In vivo 7,8-DHF, also by promoting intestinal motility and M3 expression, significantly alleviated loperamide-induced functional constipation in rats.
Our results suggest that 7,8-DHF regulates colonic motility possibly via a TrkB/Akt/M3 pathway and may be applicable for alleviating constipation.
7,8-二羟基黄酮（7,8-DHF）是一种酪氨酸激酶受体 B（TrkB）激动剂，它是否调节结肠平滑肌运动和/或缓解便秘尚未研究。
本研究旨在确定 7,8-DHF 如何影响乙酰胆碱（CCh）刺激的结肠带收缩，以及 7,8-DHF 对便秘的体内作用。
从大鼠结肠中分离肌肉条，用于记录收缩张力和进行 Western blot 分析。用洛哌丁胺诱导大鼠便秘。
虽然 7,8-DHF 特异性激活了 TrkB，但单独应用 7,8-DHF 既不能激活结肠带中的 PLCγ1，也不能诱导结肠带收缩。然而，7,8-DHF 增强了 CCh 刺激的 PLCγ1 激活和带收缩。PLCγ1 拮抗剂 U73122 抑制了 CCh 刺激和 7,8-DHF 增强/CCh 刺激的收缩。在阐明潜在机制的过程中，我们发现 7,8-DHF 增加了结肠带中的毒蕈碱 M3 受体表达。M3 选择性拮抗剂塔法那辛特异性抑制了 7,8-DHF 增强/CCh 刺激的结肠带收缩。由于 7,8-DHF 增加了 Akt 磷酸化，而 LY294002（Akt 的上游 PI3K 拮抗剂）显著抑制了 7,8-DHF 增强的 M3 表达和 7,8-DHF 增强/CCh 刺激的收缩，我们假设 7,8-DHF/TrkB/Akt 与结肠带中 M3 表达的调节有关。特异性 TrkB 拮抗剂 ANA-12 不仅抑制了 7,8-DHF 对 TrkB 的激活，还抑制了 7,8-DHF 增强的胆碱能收缩、7,8-DHF/CCh 介导的 PLCγ1/Akt 激活以及结肠带中 M3 的过度表达。体内 7,8-DHF 通过促进肠道蠕动和 M3 表达，显著缓解了洛哌丁胺诱导的大鼠功能性便秘。
我们的结果表明，7,8-DHF 通过 TrkB/Akt/M3 途径调节结肠运动，可能适用于缓解便秘。
