Targeting vascular contractile cells in capillary stasis after cardiac arrest

摘要 在美国，每年有超过50万例心脏骤停。虽然及时复苏通常是 在恢复心脏功能方面很有效，许多幸存者患上了脑损伤。从心脏复苏后 停搏，脑微血管障碍会产生继发性神经元损伤，加重神经学 结果。我们最近在一个临床相关的模型中发现了重要的皮质微血管改变。 窒息心脏骤停的原因：皮质毛细血管多灶性淤滞和小动脉血管收缩。 初步数据还表明，周细胞和平滑肌细胞是毛细血管停滞的原因之一。此外，一个 在我们的模型中，微血管靶向策略部分改善了皮质血流灌注和神经学结果。 为了充分阐明血管收缩细胞在毛细血管淤滞和神经预后中的作用，我们建议 在小鼠窒息心脏骤停模型中使用高度特异的化学遗传学方法。使用病毒 转导，我们将诱导由Designer独有激活的抑制性Designer受体的表达 药物(DREADD)在PDGFRβ-creer小鼠的周细胞内，以及在NG2-CRER小鼠的周细胞和平滑肌细胞中。 克里尔老鼠。抑制性DREADD的激活将导致周细胞和平滑肌细胞的松弛 心脏骤停。目标1将全面定义周细胞和平滑肌细胞松弛对 表达抑制性hM4Di化学发生受体的小鼠心脏骤停导致毛细血管停滞。这个 将抑制性DREADD AAV-EF1a-DIO-hM4D(GI)-mCherry注射到PDGFRβ-Creer的运动皮质 和NG2-Creer小鼠。我们将评估周细胞松弛的单独作用以及联合作用 周细胞和小动脉平滑肌细胞在毛细血管灌流时松弛。目标2将决定是否诱导 心脏骤停后周细胞松弛可改善神经学和组织学结果。为此，我们 将抑制DREADDhM4Di通过用R26-LSL-R26-LSL饲养PDGFRβ小鼠在所有周细胞中表达 HM4Di/mCitine小鼠。我们将在心脏骤停后诱导脑周细胞松弛，然后是全局周细胞松弛 评估神经学和组织学结果。 通过这一新的和特定的方法，我们将表征周细胞和平滑肌细胞在 介导心脏骤停后的微血管停滞。如果在改善神经学结果方面取得成功，这 微血管靶向策略可能为预防继发性脑损伤提供新的治疗机会。

ABSTRACT There are more than 500,000 cases of cardiac arrest in the US annually. While timely resuscitation is often effective at restoring cardiac function, many survivors develop brain injury. After resuscitation from cardiac arrest, cerebral microvascular disturbances produce a secondary neuronal injury that worsens neurological outcome. We have recently uncovered important cortical microvascular alterations in a clinically relevant model of asphyxial cardiac arrest: multifocal capillary stasis in cortical capillaries and arteriolar vasoconstriction. Preliminary data also suggest that pericytes and smooth muscle cells contribute to capillary stasis. Moreover, a microvascular-targeted strategy partially improved cortical perfusion and neurological outcome in our model. To fully elucidate the role of vascular contractile cells in capillary stasis and neurological outcome, we propose to use a highly specific chemogenetic approach in a model of asphyxial cardiac arrest in mice. Using viral transduction, we will induce the expression of inhibitory Designer Receptors Exclusively Activated by Designer Drugs (DREADD) in pericytes in PDGFRβ-CreER mice, and in both pericytes and smooth muscle cells in NG2- CreER mice. Activation of inhibitory DREADDs will induce relaxation of pericytes and smooth muscle cells after cardiac arrest. Aim 1 will define the full scope of the effect of pericyte and smooth muscle cells relaxation on cardiac arrest-induced capillary stasis in mice expressing the inhibitory hM4Di chemogenetic receptor. The inhibitory DREADD AAV-EF1a-DIO-hM4D(Gi)-mCherry will be injected in the motor cortex of PDGFRβ-CreER and NG2-CreER mice. We will assess the isolated effect of pericyte relaxation and the combined effect of pericyte and arteriolar smooth muscle cells relaxation on capillary perfusion. Aim 2 will determine if inducing relaxation of pericytes post-cardiac arrest improves neurological and histological outcomes. For this aim, we will express the inhibitory DREADD hM4Di in all pericytes by breeding PDGFRβ mice with R26-LSL- hM4Di/mCitrine mice. We will induce cerebral and then global pericyte relaxation after cardiac arrest and assess neurological and histological outcomes. Through this novel and specific approach, we will characterize the role of pericytes and smooth muscle cells in mediating microvascular stasis after cardiac arrest. If successful in improving neurological outcome, this microvascular targeted strategy could lead to a novel therapeutic opportunity to prevent secondary brain injury.