Durbin Adam, Nadir Nur-Ain, Rosenthal Alan, Gotlieb Avrum I
Toronto General Research Institute, Toronto, ON, Canada.
Cardiovasc Pathol. 2005 Jan-Feb;14(1):12-8. doi: 10.1016/j.carpath.2004.11.004.
The cell and molecular biology of heart valve wound repair is not well understood. Valve interstitial cells (IC) are thought to play an important role in valvular wound repair. Because nitric oxide (NO) has been implicated in wound repair, we tested the hypothesis that NO promotes valvular wound repair by examining the presence of the inducible form of nitric oxide synthase (iNOS) in wounded IC monolayers, in vitro.
Linear denuding wounds were made in confluent monolayers of porcine mitral valve IC plated on glass coverslips. Cultures were fixed at various times (0 to 48 h postwounding), and iNOS was localized in the cells by immunofluorescence microscopy. Cultures were also incubated with iNOS inhibitors L-N(G)-nitroarginine methyl ester (L-NAME) and N-(3-(Aminomethyl)benzyl)acetamidine (1400W), and the extent of wound closure with and without inhibitor was measured at 24, 48 and 72 h postwounding.
From 6 to 24 h postwounding, iNOS localization was increased at the wound edge. At 48 h, iNOS was localized beyond the wound edge, into the monolayer, where the intensity of the signal gradually diminished until it was virtually imperceptible. At 24 and 48 h, the inhibition of iNOS with both L-NAME and 1400W resulted in a significant delay in wound closure.
NO promotes valve wound repair through an effect on IC migration.
心脏瓣膜伤口修复的细胞和分子生物学尚未完全明晰。瓣膜间质细胞（IC）被认为在瓣膜伤口修复中发挥重要作用。由于一氧化氮（NO）与伤口修复有关，我们通过在体外检测受伤IC单层中诱导型一氧化氮合酶（iNOS）的存在，来验证NO促进瓣膜伤口修复这一假说。
在接种于玻璃盖玻片上的猪二尖瓣IC汇合单层上制造线性剥脱伤口。在不同时间点（受伤后0至48小时）固定培养物，并通过免疫荧光显微镜在细胞中定位iNOS。培养物还与iNOS抑制剂L-N(G)-硝基精氨酸甲酯（L-NAME）和N-(3-(氨甲基)苄基)脒（1400W）一起孵育，并在受伤后24、48和72小时测量有无抑制剂时的伤口闭合程度。
受伤后6至24小时，伤口边缘的iNOS定位增加。在48小时时，iNOS定位超出伤口边缘，进入单层，信号强度逐渐减弱直至几乎无法察觉。在24和48小时时，L-NAME和1400W对iNOS的抑制导致伤口闭合显著延迟。
NO通过影响IC迁移促进瓣膜伤口修复。
