McKinnon Rebecca L, Lidington Darcy, Bolon Michael, Ouellette Yves, Kidder Gerald M, Tyml Karel
Lawson Health Research Institute, The Centre for Critical Illness Research, Victoria Research Laboratory, 6th Floor, 800 Commissioners Road East London, Ontario, Canada, N6C 2V5.
Cardiovasc Res. 2006 Jan;69(1):236-44. doi: 10.1016/j.cardiores.2005.09.003. Epub 2005 Oct 14.
Increased nitric oxide (NO) production in sepsis precipitates microcirculatory dysfunction. We aimed (i) to determine if NO is the key water-soluble factor in the recently discovered sepsis-induced deficit in arteriolar conducted vasoconstriction, (ii) to identify which nitric oxide synthase (NOS) isoforms account for this deficit, and (iii) to examine the potential role of connexin37 (Cx37, a hypothesized signaling target of NO) in arteriolar conduction.
Using intravital microscopy and the cecal ligation and perforation 24-h model of sepsis, arterioles in the cremaster muscle of male C57BL/6 wild-type (WT), iNOS-/-, eNOS-/-, nNOS-/- and Cx37-/- mice were locally stimulated with KCl to initiate conducted vasoconstriction. We used the ratio of conducted constriction (500 microm upstream) to local constriction as an index of conduction (CR500). NOS enzymatic activity and protein expression were determined in control and septic cremaster muscles.
Sepsis reduced CR500 in WT mice [from 0.77 +/- 0.05 to 0.20 +/- 0.02 (means +/- SE) independent of the site of stimulation along the arteriole], in iNOS-/- and eNOS-/- mice, but not in nNOS-/- mice. The nNOS inhibitor 7-nitroindazole or NO scavenger HbO2 restored CR500 in septic WT mice, but blockade of soluble guanylate cyclase had no effect. Sepsis increased cNOS (eNOS + nNOS) activity in WT mice (from 340 +/- 40 to 490 +/- 30 pmol/mg/h) and in eNOS-/-, but not in nNOS-/- mice (iNOS activity was negligible in all mice). Sepsis did not alter nNOS protein expression in WT mice. CR500 in non-septic Cx37-/- mice (0.15 +/- 0.1) was similar to that observed in septic WT mice.
Increased nNOS activity and the resultant increased NO production in the septic mouse cremaster muscle are the key factors responsible for the deficit in conducted vasoconstriction along the arteriole. Deletion of Cx37 results in reduced CR500, which is consistent with the hypothesis that Cx37 in the arteriole could be a target of NO signaling.
脓毒症时一氧化氮（NO）生成增加会促使微循环功能障碍。我们旨在：（i）确定NO是否是最近发现的脓毒症诱导的小动脉传导性血管收缩功能缺陷中关键的水溶性因子；（ii）确定哪种一氧化氮合酶（NOS）亚型导致了这种缺陷；（iii）研究连接蛋白37（Cx37，一种推测的NO信号转导靶点）在小动脉传导中的潜在作用。
利用活体显微镜以及盲肠结扎穿孔24小时脓毒症模型，对雄性C57BL/6野生型（WT）小鼠、诱导型一氧化氮合酶基因敲除（iNOS-/-）小鼠、内皮型一氧化氮合酶基因敲除（eNOS-/-）小鼠、神经元型一氧化氮合酶基因敲除（nNOS-/-）小鼠和连接蛋白37基因敲除（Cx37-/-）小鼠提睾肌中的小动脉局部给予氯化钾刺激以引发传导性血管收缩。我们将传导性收缩（上游500微米处）与局部收缩的比值作为传导指数（CR500）。测定对照和脓毒症提睾肌中的NOS酶活性和蛋白表达。
脓毒症使WT小鼠的CR500降低[从0.77±0.05降至0.20±0.02（均值±标准误），与沿小动脉的刺激部位无关]，iNOS-/-和eNOS-/-小鼠也如此，但nNOS-/-小鼠未出现这种情况。nNOS抑制剂7-硝基吲唑或NO清除剂血红蛋白氧合产物可使脓毒症WT小鼠的CR500恢复，但可溶性鸟苷酸环化酶阻断剂无效。脓毒症使WT小鼠以及eNOS-/-小鼠的组成型NOS（eNOS + nNOS）活性增加（从340±40增至490±30 pmol/mg/h），但nNOS-/-小鼠未增加（所有小鼠中iNOS活性可忽略不计）。脓毒症未改变WT小鼠中nNOS蛋白表达。非脓毒症Cx37-/-小鼠的CR500（0.15±0.1）与脓毒症WT小鼠中观察到的相似。
脓毒症小鼠提睾肌中nNOS活性增加及由此导致的NO生成增加是小动脉传导性血管收缩功能缺陷的关键因素。Cx37基因缺失导致CR500降低，这与小动脉中的Cx37可能是NO信号转导靶点的假说一致。
