Activation of ERK during ischemia-reperfusion in lung transplantation.

1.在大鼠肺移植模型中的实验在人肺移植的缺血再灌注过程中，发现蛋白酪氨酸磷酸化的显著变化。本研究的目的是确定这些变化是否存在于大鼠单肺移植模型。冷缺血保存6小时后进行肺移植。在同种和同种异体移植物中，移植物的肺功能保存得非常好。蛋白酪氨酸磷酸化、蛋白酪氨酸激酶和蛋白酪氨酸磷酸酶活性在再灌注2 h后显著降低。再灌注2小时后，Src蛋白水平和p38磷酸化水平降低。人肺移植中MAPK的活化本实验检测了人肺移植缺血再灌注过程中MAPK的磷酸化状态。这些移植在加拿大的多伦多综合医院进行。对15例接受移植的患者进行肺组织活检：冷缺血保存后、热缺血（植入）后和再灌注1 h或2 h后。Western blotting检测ERK、p38-MAPK、INK和MEK的磷酸化状态。在较低水平下，INK的磷酸化也显著增加。相反，p38的磷酸化没有显著变化。ERK磷酸化的增加与ERK的上游激酶MEK的激活无关。由于MKP-3蛋白水平与其活性相对应，我们检查了这些样品中MKP-3的蛋白表达。令人惊讶的是，MKP-3蛋白水平在再灌注后显著增加。因此，ERK磷酸化的显著增加不是由于MKP-3的表达受到抑制。

1.Experiment in a rat lung transplant model.Dramatic alterations of protein tyrosine phosphorylation have been found during the ischemia-reperfusion period of human lung transplantation. The object of the present study was to determine whether these changes exist in a rat single-lung transplant model. Lung transplantations were performed after 6 hours of cold ischemic preservation. In both iso-and allografts, the lung function of transplants was very well preserved. Protein tyrosine phosphorylation, protein tyrosine kinase and protein tyrosine phosphatase activities were decreased significantly after 2 hours of reperfusion. Src protein level and phosphorylation of p38 were reduced after 2 hours of reperfusion. This lung transplantation model is useful for studying ischemia-reperfusion injury.MAPK activation in human lung transplantThe phosphorylation status of MAPK during ischemia-reperfusion in human lung transplant was examined. These transplantations were performed in Toronto General Hospital in Canada. Lung tissue biopsy was performed on 15 patients undergoing transplantation: after the cold ischemic preservation, after the warm ischemia (implantation) and after 1 h or 2 h reperfusion. The phosphorylation status of ERK, p38-MAPK, INK and MEK was examined with Western blotting. Phosphorylation of INK also significantly increased at lower levels. In contrast, phosphorylation of p38 had no significant changes. The increase in ERK phosphotylation was not associated with the activation of MEK, the up-stream kinase for ERK. Since MKP-3 protein levels correspond to its activity, we examined the protein expression of MKP-3 in these samples. Surprisingly, the MKP-3 protein level significantly increased after reperfusion. Therefore, the dramatic increase in ERK phosphorylation is not due to the inhibited expression of MKP-3.