Use of network pharmacology to analyze compound reserpine and triamterene tablets in the treatment of hypertension / 药学学报

Compound reserpine and triamterene tablets (CRTT), a compound antihypertensive drug developed by Chinese scientists, is still widely used in clinical practice. However, the mechanisms by which CRTT treats hypertension remain to be fully understood. This study used network pharmacology to analyze CRTT's antihypertensive mechanisms with in vitro experiments. The targets of the four chemical components of CRTT were collected from the Swiss Target Prediction database; 1 828 protein targets related to hypertension were collected from the Therapeutic Target Database (TTD) and Online Mendelian Inheritance in Man (OMIM) database. The CRTT-hypertension network model was constructed using a search tool for recurring instances of neighbouring genes (STRING). Gene ontology (GO) and pathway enrichment analysis of targets of interest was conducted with the Metascape database. In the in vitro study, human umbilical vein endothelial cells (HUVEC) and vascular smooth muscle cells (VSMC) were treated with 1 μmol·L-1 angiotensin Ⅱ (AngⅡ) and CRTT was administered at concentrations of 0.01, 0.1, and 1 μmol·L-1. Changes in the phosphatidylinositol-3-kinase/protein serine threonine kinase/endothelial nitric oxide synthase (PI3K/Akt/eNOS) pathway in HUVEC and the cyclic guanosine monophosphate/cGMP-dependent protein kinase (cGMP/PKG) pathway in VSMC were determined by Western blot. Network pharmacological analysis revealed that the antihypertensive effect of CRTT is closely associated with biological pathways such as vascular tone regulation, adrenergic receptor activation, protein kinase activity and signaling pathways such as the cGMP/PKG signaling pathway, vascular smooth muscle contraction, neuroactive ligand-receptor interaction, adrenergic signaling in cardiomyocytes and calcium signaling pathways. The in vitro study confirmed that CRTT increased the levels of phosphorylated phosphatidylinositol-3-kinase (p-PI3K), phosphorylated protein serine threonine kinase (p-Akt), phosphorylated endothelial nitric oxide synthase (p-eNOS) in HUVEC and the levels of eNOS, phosphorylated vasodilator-stimulated phosphoprotein (p-VASP), and PKG in VSMC through multiple targets and pathways. These results suggest that the activation of PI3K/Akt/eNOS pathway and endothelial-dependent NO/cGMP signaling may be involved in the CRTT-mediated hypotensive effect.

Possible mechanisms involved in the vasorelaxant effect produced by clobenzorex in aortic segments of rats

Clobenzorex is a metabolic precursor of amphetamine indicated for the treatment of obesity. Amphetamines have been involved with cardiovascular side effects such as hypertension and pulmonary arterial hypertension. The aim of the present study was to investigate whether the direct application of 10-9-10-5 M clobenzorex on isolated phenylephrine-precontracted rat aortic rings produces vascular effects, and if so, what mechanisms may be involved. Clobenzorex produced an immediate concentration-dependent vasorelaxant effect at the higher concentrations (10-7.5-10-5 M). The present outcome was not modified by 10-6 M atropine (an antagonist of muscarinic acetylcholine receptors), 3.1×10-7 M glibenclamide (an ATP-sensitive K+ channel blocker), 10-3 M 4-aminopyridine (4-AP; a voltage-activated K+ channel blocker), 10-5 M indomethacin (a prostaglandin synthesis inhibitor), 10-5 M clotrimazole (a cytochrome P450 inhibitor) or 10-5 M cycloheximide (a general protein synthesis inhibitor). Contrarily, the clobenzorex-induced vasorelaxation was significantly attenuated (P<0.05) by 10-5 M L-NAME (a direct inhibitor of nitric oxide synthase), 10-7 M ODQ (an inhibitor of nitric oxide-sensitive guanylyl cyclase), 10-6 M KT 5823 (an inhibitor of protein kinase G), 10-2 M TEA (a Ca2+-activated K+ channel blocker and non-specific voltage-activated K+ channel blocker) and 10-7 M apamin plus 10-7 M charybdotoxin (blockers of small- and large-conductance Ca2+-activated K+ channels, respectively), and was blocked by 8×10-2 M potassium (a high concentration) and removal of the vascular endothelium. These results suggest that the direct vasorelaxant effect by clobenzorex on phenylephrine-precontracted rat aortic rings involved stimulation of the NO/cGMP/PKG/Ca2+-activated K+ channel pathway.

Pharmacological study of the mechanisms involved in the vasodilator effect produced by the acute application of triiodothyronine to rat aortic rings

A relationship between thyroid hormones and the cardiovascular system has been well established in the literature. The present in vitro study aimed to investigate the mechanisms involved in the vasodilator effect produced by the acute application of 10-8–10-4 M triiodothyronine (T3) to isolated rat aortic rings. Thoracic aortic rings from 80 adult male Wistar rats were isolated and mounted in tissue chambers filled with Krebs-Henseleit bicarbonate buffer in order to analyze the influence of endothelial tissue, inhibitors and blockers on the vascular effect produced by T3. T3 induced a vasorelaxant response in phenylephrine-precontracted rat aortic rings at higher concentrations (10-4.5–10-4.0 M). This outcome was unaffected by 3.1×10-7 M glibenclamide, 10-3 M 4-aminopyridine (4-AP), 10-5 M indomethacin, or 10-5 M cycloheximide. Contrarily, vasorelaxant responses to T3 were significantly (P<0.05) attenuated by endothelium removal or the application of 10-6 M atropine, 10-5 M L-NG-nitroarginine methyl ester (L-NAME), 10-7 M 1H-(1,2,4)oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), 10-6 M (9S,10R,12R)-2,3,9,10,11,12-Hexahydro-10-methoxy-2,9-dimethyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3′,2′,1′-kl]pyrrolo[3,4-i](1,6)benzodiazocine-10-carboxylic acid, methyl ester KT 5823, 10-2 M tetraethylammonium (TEA), or 10-7 M apamin plus 10-7 M charybdotoxin. The results suggest the involvement of endothelial mechanisms in the vasodilator effect produced by the acute in vitro application of T3 to rat aortic rings. Possible mechanisms include the stimulation of muscarinic receptors, activation of the NO-cGMP-PKG pathway, and opening of Ca2+-activated K+ channels.