UOhio State University (Columbus)
Dr. Michael Beattie

TNFa and Cell Death in Rat Spinal Cord White Matter

The spinal cord is not thought to grossly repair itself after injury, and in fact, after SCI, it undergoes substantial secondary injury resulting in larger, more debilitating lesions. The objective of this research is to dissect the mechanisms of secondary cell death in SCI by isolating excitotoxic death pathways from traumatic mechanical death pathways. An understanding of what causes secondary cell death and lesion spread after SCI will lead to prevention of the paralyzing cell loss.

Tumor necrosis factor alpha (TNFα) and glutamate – both molecules released in high concentrations after SCI – appear to act synergistically in bringing about rapid cell death in the CNS. Recent work suggests that TNFα upregulates AMPA-type glutamate receptors on the cell surface. We hypothesize that: 1) TNFα potentiates the excitotoxic effect of glutamate-agonists on oligodendrocytes, 2) pharmacologically blocking glutamate receptor pathways will rescue oligodendrocytes from TNFα-potentiated excitotoxic cell death, and 3) TNFα induces p75 or FAS “death receptor” expression in white matter after SCI, making oligodendrocytes more likely to undergo apoptosis.

We will test these hypotheses in two specific aims. In Specific Aim 1 we will test the role of TNFα in white matter excitotoxicity by nanoinjection of TNFα alone, kainic acid (KA) alone, TNFα plus KA, or albumin controls into the dorsal funiculus. We will also attempt to block the excitotoxicity via injection, and serial sections of the spinal cord will be immunostained and examined for cell-type specific cell loss.

In Specific Aim 2 will test the role of TNFα in white matter apoptosis over a time-course of two weeks. Animals injected as in specific aim 1 will be sacrificed at 16 hours, 2 day, 7 days and 14 days post-injection, and serial sections will be immunostained for p75 or FAS receptors, processes for TUNEL or Hoechst labeling of apoptosis, or processes for in situ hybridization labeling of p75 or FAS mRNA expression.

Secondary cell death and the spread of the lesion via apoptosis add considerably more CNS damage to the initial spinal cord injury. This study aims to identify factors involved in apoptosis and secondary injury after SCI. As these factors involved in secondary injury, their mechanism of action, and their downstream effects are identified, treatments that combat secondary injury will also become more identifiable. If secondary injury is prevented, spinal cord tissue will be spared, allowing for less severe lesions and increased chances for recovery.