Spinal cord injury, progressive hemorrhagic necrosis and the NC(Ca-ATP) channel

摘要 脊髓损伤（SCI）导致称为“进行性”的自我破坏过程 出血性坏死”（PHN），导致脊髓组织的毁灭性损失。两 PHN的关键成分是：1）进行性继发性出血; 2）坏死细胞 死亡我们最近发现，新表达的SUR 1调节的NC（Ca-ATP） 通道与坏死细胞死亡和继发性出血密切相关， SCI.在这里，我们将进一步表征SUR 1调节的NC（Ca-ATP）通道的作用， 在SCI。我们的总体假设是，NF κ B信号的激活是一个关键， 在内皮细胞中SUR 1调节的NC（Ca-ATP）通道的从头表达中的作用， 神经元和少突胶质细胞，以及随后ATP- 耗尽导致毛细血管的灾难性失效，形成瘀点 以及神经元和少突胶质细胞的坏死性死亡，这反过来又引起氧化应激。 压力和炎症，它们共同助长PHN。我们在小鼠和大鼠模型中的数据 创伤性脊髓损伤表明， 使用格列本脲阻断SUR 1，通过使用反义基因抑制SUR 1 寡脱氧核苷酸（AS-ODN），其优先靶向半暗带毛细血管，和 在转基因SUR 1-KO（SUR 1-/-）小鼠中， 治疗或条件与短期内的显着改善有关， 神经行为功能在特定目标（SA）1中，使用基因抑制策略 针对SCI小鼠和大鼠模型中的SUR 1，我们将评估进行性 短期后遗症继发出血，包括炎症和氧化 压力以及长期后遗症，包括组织病理学和神经行为 功能其他初步数据表明，PHN中最关键的细胞是 毛细血管内皮细胞、神经元和少突胶质细胞。在SA 2中，使用原代培养物 小鼠脊髓微血管内皮细胞、神经元和少突胶质细胞 从野生型（WT）与SUR 1-KO小鼠，我们将证实每种细胞类型可以 上调SUR 1调节的NC（Ca-ATP）通道，我们将表征新诱导的 通道，确定其生理调节pH值和它们在细胞死亡中的作用。 其他初步数据表明，NF κ B，这是已知的显着 参与脊髓损伤，可能是一个重要的转录激活剂，在从头 NC（Ca-ATP）通道的表达。在SA 3中，使用培养的细胞和脐带组织， SCI中，我们将确定NF κ B刺激的转录在从头转录中的作用。 SUR 1的表达和功能性SUR 1调节的NC（Ca-ATP）的从头表达 渠道这些研究将使我们更深入地了解NC（Ca-2+）的作用。 ATP）通道的研究将为我们提供新的分子见解和重要的新治疗方法 造成了如此严重的人类状况

ABSTRACT Spinal cord injury (SCI) results in the autodestructive process termed "progressive hemorrhagic necrosis" (PHN), which leads to devastating loss of spinal cord tissue. Two critical components of PHN are: 1) progressive secondary hemorrhage; 2) necrotic cell death. We recently discovered that newly expressed SUR1-regulated NC(Ca-ATP) channels are critically involved in necrotic cell death and in secondary hemorrhage post- SCI. Here, we will further characterize the role of SUR1-regulated NC(Ca-ATP) channels in SCI. Our overarching hypothesis is that activation of NFkappaB signaling plays a key role in de novo expression of SUR1-regulated NC(Ca-ATP) channels in endothelium, neurons and oligodendrocytes, and that subsequent opening of the channels by ATP- depletion results in catastrophic failure of capillaries, formation of petechial hemorrhages and necrotic death of neurons and oligodendrocytes, which in turn provokes oxidative stress and inflammation, which together fuel PHN. Our data in mouse and rat models of contusion SCI demonstrate that hemorrhage is dramatically reduced by pharmacological block of SUR1 using glibenclamide, by gene suppression of SUR1 using antisense oligodeoxynucleotide (AS-ODN), which preferentially targets penumbral capillaries, and by gene suppression in transgenic SUR1-KO (SUR1-/-) mice, and that these 3 treatments or conditions are associated with dramatic improvements in short-term neurobehavioral function. In specific aim (SA) 1, using gene suppression strategies targeting SUR1 in mouse and rat models of SCI, we will assess the role of progressive secondary hemorrhage on short-term sequelae, including inflammation and oxidative stress, and on long-term sequelae, including histopathology and neurobehavioral function. Other Preliminary Data indicate that the cells most critically involved in PHN are capillary endothelial cells, neurons and oligodendrocytes. In SA2, using primary cultures of murine spinal cord microvascular endothelial cells, neurons and oligodendrocytes from wild-type (WT) vs. SUR1-KO mice, we will confirm that each cell type can upregulate SUR1-regulated NC(Ca-ATP) channels, we will characterize newly induced channels, determine their physiological regulation by pH and their role in cell death. Other Preliminary Data demonstrate that NFkappaB, which is known to be prominently involved in SCI, is likely to act as an important transcriptional activator in de novo expression of NC(Ca-ATP) channels. In SA3, using cultured cells and cord tissues post- SCI, we will determine the role of NFkappaB-stimulated transcription in de novo expression of SUR1 and de novo expression of functional SUR1-regulated NC(Ca-ATP) channels. These studies will yield a more thorough understanding of the role of NC(Ca- ATP) channels in SCI will lead to novel molecular insights and significant new treatments for this devastating human condition.