抗氧化蛋白Prx II通过蛋白磷酸化酶1调控PLN磷酸化和心脏泵功能的实验研究

Prx II是过氧化氢过氧化物酶超家族成员之一，我们前期研究发现：PrxII在心衰病人的心肌表达量明显下降，而在两个心肌收缩功能增强的小鼠模型：phospholamban(PLN)基因敲除和蛋白磷酸化酶1（PP1）抑制因子1过表达，其表达量上调；但尚不清楚PrxII是否通过PLN调控心肌收缩，本项目旨在探讨：1）腺病毒中介的急性PrxII过表达和表达下调对心肌细胞收缩和肌浆网钙转运的调控；2）慢性PrxII心肌特异性过表达和3）Cre-LoxP中介的条件性PrxII基因敲除对心肌收缩和肌浆网钙转运以及PLN磷酸化的调控；4）将上述PrxII单基因修饰小鼠与PLN基因敲除小鼠交配获得双基因修饰小鼠并检测其心功能；5）确定PrxII与PLN， SERCA2和PP1等收缩蛋白直接相互作用及其6)对病理性心肌肥厚和心衰进程的影响；通过上述研究阐明PrxII可否作为新的心肌收缩调控因子的假说。

Peroxiredoxin II(PrxII) is one of the members of 2-Cys peroxide peroxidase superfamily in most mammalian cells, which reduces H2O2 or the intracellular peroxides to alcohol and water. Previously, we found that the expression levels of PrxII in the failing human hearts were significantly decreased. In the meantime, the levels were increased in the two functionally hyperdynamic gene engieered mouse hearts, with phospholamban(PLN) knockout or protein phosphostase 1 inhibitor 1 overexpression. However, it remains unclear that the functional significance of PrxII in cardiac contractility and sarcoplasmic reticulum(SR) calcium handling as well as the involvment of PLN. In the current project, we are aiming to evalute the following questions: 1) The role of acute enhanced or reduced expression of PrxII, induced by adenoviral infections, in rat cardiomyocyte contractility and calcium transient and the molecular mechanisms; 2) A transgenic mouse with cardiac specific PrxII overexpression and 3) a Cre-LoxP-induced cardiac specific conditional PrxII deficient mouse will be generated. The cardiac contractile function and calcium transient as well as the SR calcium handing in those two PrxII gene engieered mice will be investigated, under the whole animal, organ and single myocyte levels. 4)The two indicated mice in 2)and 3) will be crossed with PLN knockout mice to generate a double gene altered mouse. The involved cardiac function and SR calcium handling in those animal models will also be determined. 5)In vitro immunoprecipitating study and confocal microscope will be used to further evaluate the interaction between PrxII and several calcium handling proteins, including: PLN, SERCA2 and PP1. 6)Furthermore, the role of the cardiac specific decreased or enhanced expression of PrxII in transaortic banding-induced hypertrophic hearts and the development to heart failure will also be investigated. In summary, the goal of this project is to demonstrate the novel role of PrxII as a regulator for cardiac contractile performance and its pathophysiologic involvement in the progress of heart failure.