The role of the Cav3.1 subunit of the T-type calcium channel in the heart after injury using a loss-of-function model

 描述(申请人提供)：T型钙通道在发育过程中在整个心脏中高度表达，但其表达仅限于出生后的起搏细胞。在心脏受到损伤后，如心肌梗塞，这些通道在成人的心室肌细胞中重新表达。通过这些通道的钙内流与多种细胞类型的增殖增加有关。这些通道的重新表达可能在心肌梗死后起到有益的作用。我们将使用全球基因敲除模型，确定T型钙通道Cav3.1亚单位的缺失是否会改变心肌梗死后发生的结构、功能和分子适应。野生型和Cav 3.1基因敲除小鼠心肌梗死后的结构变化将通过超声心动图和组织学进行评估。功能变化将通过超声心动图、血流动力学、心电图和电生理学进行评估。分子变化将使用qRT-PCR和蛋白质印迹进行评估。我们还将通过分离心脏干细胞并在培养中生长来确定Cav3.1表达的缺失如何改变驻留的心脏干细胞的特性。与WT细胞相比，这些细胞的表面标志表达、形态、增殖率、衰老率、凋亡率和旁分泌因子的分泌将是其特征。体内研究将探索Cav3.1 KO对MI后血管生成的影响，因为CDC最终会产生新的血管，我们将通过将Cav3.1 KO骨髓移植到野生型小鼠的骨髓中，确定MI后渗入心脏的骨髓炎性细胞中Cav3.1表达的缺失是否有助于改变MI后的重塑。

 DESCRIPTION (provided by applicant): T-type calcium channels are highly expressed throughout the heart during development, but their expression is limited to pacemaker cells after birth. After an injury to the heart, such as MI, these channels are re-expressed in the adult ventricular myocytes. Calcium influx through these channels has been linked to increased proliferation in a number of cell types. The re-expression of these channels may be serving a beneficial purpose after MI. We will determine if loss of the Cav3.1 subunit of the T-Type calcium channel, using a global knockout model, alters the structural, functional, and molecular adaptions occurring in the myocardium after MI. The structural changes occurring after MI in wild type and Cav 3.1 knockout mice will be assessed using echocardiography and histology. Functional changes will be assessed via echocardiography, hemodynamics, electrocardiogram, and electrophysiology. Molecular changes will be assessed using qRT-PCR and western blot. We will also determine how loss of Cav3.1 expression alters the properties of resident cardiac stem cells by isolating cardiac stem cells and growing them in culture. These cells will be characterized by their surface marker expression, morphology, proliferation rate, rate of senescence, percent of apoptosis, and paracrine factor secretion in comparison to WT cells. In-vivo studies will explore the effects of Cav3.1 KO on post MI angiogenesis, since CDCs are known to generate new blood vessels Finally, we will determine if loss of Cav3.1 expression in bone marrow derived inflammatory cells that infiltrate the heart after MI contributes to altered post MI remodeling via bone marrow transplantation of Cav3.1 KO bone marrow into wild type mice.