Transmigration of neutrophils through epithelium

我们评估了来自内皮细胞的一氧化氮（NO）在中性粒细胞跨内皮迁移（TEM）中的作用。NO合成酶抑制剂ng -硝基l -精氨酸甲酯盐酸盐（L-NAME）或ng -单甲基l -精氨酸（L-NMMA）预处理人脐静脉内皮细胞（HUVECs），可增强中性粒细胞TEM。在NO清除剂2-（4-羧基苯基）-4,4,5,5-四甲基咪唑啉-1-氧-3-氧化物（羧基PTIO）的存在下，TEM也有类似的增强。中性粒细胞TEM在L-NAME或l - nmma处理的HUVECs中被NO供体持续供应的NO所抑制。这些发现支持内皮细胞持续产生NO抑制中性粒细胞TEM的观点。流式细胞术分析显示，NO在中性粒细胞与产生NO的HUVECs共培养中积累。与未处理的HUVECs共培养的中性粒细胞相比，l - name处理的HUVECs共培养的中性粒细胞中NO含量降低。可溶性胍基环化酶（sGC）是中性粒细胞中NO的重要靶点之一。3-（53-羟基甲基-23呋喃基）-1-苄基茚唑（YC-1）是sGC的活化剂，抑制l- name诱导的中性粒细胞TEM。有趣的是，1- h[1,2,4-]恶二唑[4,3-a]喹诺沙林-1-1 （ODQ）对中性粒细胞sGC的抑制足以增强TEM。这些结果表明，来自HUVECs的NO作用于中性粒细胞抑制TEM，至少部分是通过激活sGC。我们的研究结果表明，在生理条件下，由HUVECs产生的一氧化氮在防止中性粒细胞过度外溢和不必要的组织损伤方面的作用。

We evaluated the roles of nitric oxide(NO) derived from endothelial cells in neutrophil transendothelial migration(TEM). Pretreatment of human umbilical vein endothelial cells (HUVECs) with NG-nitro-L-arginine methyl ester hydrochloride(L-NAME) or NG-monomethyl L-arginine (L-NMMA), which are inhibitors of NO synthases, enhanced neutrophil TEM. Similar augmentation of TEM was observed in the presence of an NO scavenger, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (carboxy PTIO). Neutrophil TEM across L-NAME- or L-NMMA-treated HUVECs was inhibited by continuous NO supply by NO donors. These findings support the suggestion that continuous production of NO by endothelial cells suppresses neutrophil TEM. Flow cytometric analyses revealed that NO accumulates in neutrophils co-cultured with NO-producing HUVECs. A decreased amount of NO was detected in neutrophils co-cultured with L-NAME-treated HUVECs compared with neutrophils co-cultured with untreated HUVECs. Soluble guanylyl cyclase (sGC) is known as one of the most important targets of NO in neutrophils. 3-(53-Hydroxymethyl-23furyl)-1-benzyl indazole (YC-1), an activator of sGC, inhibited L-NAME-induced neutrophil TEM. It was interesting that inhibition of neutrophil sGC with 1-H[1,2,4-]oxadiazolo[4,3-a]quinoxalin-1-1 (ODQ) was sufficient to enhance TEM. These results suggest that NO derived from HUVECs acts on neutrophils to inhibit TEM, at least in part by activating sGC. Our findings imply the role of NO constitutively generated by HUVECs in protection against excessive neutrophil extravasation and unnecessary tissue damage under physiological conditions.