Simplicio Janaina Ap, Pernomian Larissa, Simão Marilia R, Carnio Evelin C, Batalhão Marcelo E, Ambrosio Sergio R, Tirapelli Carlos R
Eur J Pharmacol. 2014 Mar 5;726:66-76. doi: 10.1016/j.ejphar.2014.01.018.
We investigated the mechanisms underlying the vasorelaxant and hypotensive actions of the labdane-type diterpene ent-3-acetoxy-labda-8(17),13-dien-15-oic acid (labda-15-oic acid). Vascular reactivity experiments were performed in aortic rings isolated from male Wistar rats. cAMP and cGMP were measured by enzyme immunoassay (EIA) whereas nitrate measurement was performed by chemiluminescence. Nitric oxide (NO) concentration ([NO]c) was measured in endothelial cells by flow cytometry. The cytosolic calcium concentration ([Ca2+]c) in vascular smooth muscle cells (VSMC) was measured by confocal microscopy. Blood pressure measurements were performed in conscious rats. Labda-15-oic acid inhibited the contraction induced by phenylephrine and serotonin in either endothelium-intact or endothelium-denuded rat aortic rings. The labdane significantly reduced CaCl2-induced contraction in a Ca2+-free solution containing KCl or phenylephrine. Labda-15-oic acid (0.1–300 μmol/l) concentration-dependently relaxed endothelium-intact and endothelium-denuded aortas pre-contracted with either phenylephrine or KCl. In endothelium-intact rings, the relaxation induced by labda-15-oic acid was affected by L-NAME, 7-nitroindazole, ODQ, hemoglobin, Rp-8-Br-Pet-cGMPS and thapsigargin. Blockade of K+ channels with 4-aminopyridine, apamin, charybdotoxin and glibenclamide affected the relaxation induced by labda-15-oic acid. The labdane increased cGMP and nitrate levels but did not affect cAMP levels in endothelium-intact aortas. Labda-15-oic acid increased [NO]c in endothelial cells and decreased [Ca2+]c in VSMC. The hypotension induced by intravenous administration of labda-15-oic acid (0.3–3 mg/kg) was partially reduced by L-NAME. In conclusion, the mechanisms underlying the cardiovascular actions of the labdane involve the activation of the endothelial NO-cGMP pathway, the opening of K+ channels and the alteration on Ca2+ mobilization.
我们研究了半日花烷型二萜ent-3-乙酰氧基-半日花-8(17),13-二烯-15-酸（半日花-15-酸）的血管舒张和降压作用的潜在机制。在从雄性Wistar大鼠分离的主动脉环中进行血管反应性实验。通过酶免疫测定法（EIA）测量cAMP和cGMP，而通过化学发光法进行硝酸盐测量。通过流式细胞术测量内皮细胞中的一氧化氮（NO）浓度（[NO]c）。通过共聚焦显微镜测量血管平滑肌细胞（VSMC）中的胞质钙浓度（[Ca2+]c）。在清醒大鼠中进行血压测量。半日花-15-酸抑制苯肾上腺素和5-羟色胺在完整内皮或去内皮大鼠主动脉环中诱导的收缩。在含有氯化钾或苯肾上腺素的无钙溶液中，半日花烷显著降低氯化钙诱导的收缩。半日花-15-酸（0.1 - 300μmol/L）浓度依赖性地舒张预先用苯肾上腺素或氯化钾预收缩的完整内皮和去内皮主动脉。在完整内皮环中，半日花-15-酸诱导的舒张受L-硝基精氨酸甲酯（L-NAME）、7-硝基吲唑、1H-[1,2,4]恶二唑并[4,3-a]喹喔啉-1-酮（ODQ）、血红蛋白、Rp-8-溴-Pet-cGMPS和毒胡萝卜素影响。用4-氨基吡啶、蜂毒明肽、大蝎毒素和格列本脲阻断钾通道影响半日花-15-酸诱导的舒张。半日花烷增加完整内皮主动脉中的cGMP和硝酸盐水平，但不影响cAMP水平。半日花-15-酸增加内皮细胞中的[NO]c并降低VSMC中的[Ca2+]c。静脉注射半日花-15-酸（0.3 - 3mg/kg）诱导的低血压被L-NAME部分降低。总之，半日花烷心血管作用的潜在机制涉及内皮NO-cGMP途径的激活、钾通道的开放以及钙动员改变。
