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 NF-kappa B 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.
Using rat and mouse models of spinal cord injury, we discovered that pharmacological inhibition as well as gene suppression of SUR1-regulated NC (Ca-ATP) channels cause a striking reduction in progressive secondary hemorrhage and in hemorrhagic necrosis, and are associated with dramatic improvements in short-term neurological function. In this proposal, we will use gene suppression strategies in rat and mouse models of spinal cord injury to determine short and long-term consequences of SUR1 inhibition and to more fully characterize essential molecular principles governing NC (Ca-ATP) channel expression and function in SCI. These studies will form the basis for novel future therapies for spinal cord injury.
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