内皮源性表氧化二十烷烯酸在门脉高压症内脏动脉扩张的作用及机制

Role of epoxyeicosatrienoic acid in splanchnic vasodilation in portal hypertension

目的观察肠系膜血管血浆表氧化二十烷烯酸(EETs)水平在门静脉高压症(PHT)鼠的变化,探讨其表达与肠系膜小动脉血管张力及肌球蛋白轻链磷酸酶(MLCP)活性、肌球蛋白轻链(MLC20)的磷酸化水平之间关系。方法采用CCl4导致肝硬化门静脉高压症鼠为模型,正常对照组(20只)、PHT模型组(40只)。于25、45、65 d取标本,酶联免疫吸附试验(ELISA)法检测肠系膜动脉血浆EETs水平;离体肠系膜小动脉对NE、Ca^2+梯度浓度的血管反应性、钙敏感性及MLCP活性、MLC20的磷酸化水平改变,以及抑制剂indo、L-NAME、ODQ、IbTx、Glibenclamide、miconazole在其中的影响。结果PHT模型组鼠肠系膜动脉血浆内EETs的产生明显增高,于45 d明显增高,65 d达到高峰;同步离体肠系膜小动脉对NE、Ca^2+梯度浓度的血管反应性、钙敏感性明显降低,且MLCP活性增强、MLC20的磷酸化水平下降;给予EETs抑制剂miconazole肠系膜小动脉MLCP活性降低、MLC20磷酸化水平提高,差异有统计学意义(P<0.05);相关分析PHT肠系膜动脉血浆内EETs水平与NE-pD2、Ca^2+-pD2及MLCP活性、MLC20磷酸化水平变化呈正相关(r=0.751,P<0.01;r=0.772,P<0.01;r=0.641,P<0.01;r=0.561,P<0.01)。结论内皮源性EETs通过肠系膜小动脉平滑肌血管反应性、钙敏感性下降,引起内脏动脉扩张,参与内脏动脉高动力循环。

Objective To evaluate the role of epoxyeicosatrienoic acid (EETs) in the haemodynamic alterations of experimental cirrhosis, and its relationship with vascular contractile reactivity and calcium sensitivity of mesenteric arteries. Methods Twenty control rats and 40 rats with carbon tetrachloride (CCl4)-induced cirrhosis were studied. The plasma EETs in the mesenteric arteries was evaluated by enzyme linked immunosorbent assay (ELISA). Concentration-response curves to Norepinephrine (NE) and calcium (Ca2+) were evaluated in mesenteric arteries pre-incubated with indomethacin (indo), N (G)-nitro-L-arginine-methyl-ester (L-NAME), 1H-[1, 2, 4]oxadiazolo[4, 3-a]quinoxalin-1-one (ODQ), ibetiotoxin (IbTx) and Glibenclamide before and after the epoxygenase inhibitor miconazole. pD2 was calculated. the activity of myosin light chain phosphatase (MLCP) and myosin light chain (MLC20) phosphorylation were observed. Results The plasma EETs in mesenteric arteries were increased more than that in control rats (P<0.01), and the activity of MLCP was increased, the MLC20 phosphorylation decreased;The NE and Ca2+ response were decreased in small mesenteric arteries. Indo, L-NAME, ODQ, IbTx and Glibenclamide blunted the response to NE and Ca^2+ more in cirrhotic than that in control rats (P<0.05). Concentration-response curves to miconazole showed an increased NE and Ca^2+ response in cirrhotic rats (P<0.05);the activity of MLCP was decreased, the MLC20 phosphorylation was increased by using miconazole. An increased plasma EETs were significantly correlated with NE-pD2, Ca^2+-pD2, the activity of MLCP and the MLC20 phosphorylation in mesenteric arteries. Conclusion The increased plasma EETs play a role in splanchnic vasodilation of PHT through regulating the response to NE and Ca^2+ in mesenteric arteries.