Author ORCID Identifier

https://orcid.org/0000-0002-9466-606X

Date Available

12-2-2024

Year of Publication

2024

Document Type

Doctoral Dissertation

Degree Name

Doctor of Philosophy (PhD)

College

Medicine

Department/School/Program

Neuroscience

Advisor

Dr. Warren J. Alilain

Abstract

There are currently approximately 300,000 individuals living with a spinal cord injury (SCI) in the United States with an additional 17,000 injuries occurring each year. Perhaps of equal importance the median age at which an SCI occurs within the population has steadily increased since the 1970s to the 2010s from 29 years to 43 years and is still on the rise. Women, historically underrepresented in biomedical research, have also seen a slight increase in SCI incidence. This increase in age of injury onset means that more people with an SCI will be impacted by genetic factors associated with Alzheimer’s disease (AD). Therefore, it’s critical to examine not only the impact of the apolipoprotein E (APOE) gene typically associated with AD, but also sex as a biological variable in the context of SCI. Additionally, while the central nervous system (CNS) comprising of the brain and spinal cord has been extensively studied in the context of AD and SCI respectively, little is known about how SCI affects peripheral organs, including those involved in ApoE production such as the liver. Based on prior work conducted in our lab examining the impact high cervical injury on respiratory outcomes, including preliminary work investigating APOE, and the need for an injury rostral to the level of peripheral organ innervation we chose to utilize the left lateral C2 hemisection (C2Hx) to characterize our genotype and peripheral organ outcome measures. The APOE gene contains 3 distinct alleles, APOE2, E3, and E4. The E4 allele is the key genetic determinant for Alzheimer’s disease. The E2 genotype is considered protective although it is only found in 5-10 % of the population. For our initial experiments we compared targeted replacement homozygous E3 mice to homozygous E4 counterparts. We first focused on breathing function as measured by whole body plethysmography (WBP) in the absence of a spinal cord injury. We demonstrated that under normal breathing conditions, E3 mice exhibited more robust respiratory function compared to their E4 counterparts. However, when subjected to more severe respiratory challenges, such as those seen in sleep apnea, E4 carriers were able to match or even surpass the respiratory performance of E3 mice. Importantly, previous work from the Alilain lab had observed enhanced plasticity measures in female E3 carriers compared to E4 counterparts using electromyography. Building on our initial work characterizing respiratory responses without an SCI, we expanded our investigation to examine how C2Hx impacts measures of respiratory and motor recovery in our APOE mouse model as well as relevant spinal cord histopathology. First, we utilized WBP to evaluate the respiratory responses in E3 and E4 mice. We determined that even following an SCI, E3 mice breathe more robustly except when a more severe challenge is administered at which point E4 mice increase their breathing rate more than their E3 counterparts. Next, we examined measures of gait dynamics and locomotor coordination showing that at least in male mice, E4 animals displayed better recovery post injury. Finally, we performed immunohistochemistry on regions critical to respiratory and motor control to characterize differences in plasticity, inflammation, and myelination between our genotypes. Having further established the critical role of sex and APOE genotype in SCI recovery, we explored metabolomics as a technique that could expand our understanding of injury profiles and be integrated into future studies of our APOE mouse model. To determine how SCI impacts the peripheral organ metabolome, we gave a C2Hx to female rats and examined organs at acute timepoints post-injury as well as from a naïve group. For our peripheral organs, we chose the lungs, spleen, and liver due to their relevance to breathing, immune function, and ApoE production, respectively. We found significant upregulation of energy production substrates in the liver following injury. While changes in our other peripheral organs were less pronounced, the metabolomic shift in the liver presents an excellent target for investigation and intervention, particularly in the context of our APOE mouse model. Furthermore, our metabolomics work builds on preliminary studies from other groups initially defining a tertiary injury. This tertiary injury is characterized by changes in peripheral organs following the well-established secondary injury that occurs in the cord, which is characterized by CNS cell death and spinal cord degeneration subsequent to the initial traumatic primary injury. These findings further highlight the importance of APOE genotype and sex on respiratory and motor recovery following SCI. Furthermore, a better understanding of the impact of these factors on SCI recovery, as well as a more precise characterization of our tertiary injury hypothesis will aid in overcoming translational barriers in the treatment of SCI. This work holds the potential to uncover new therapeutic approaches for SCI treatment both in the context of individual genetics and the peripheral organ metabolome.

Digital Object Identifier (DOI)

https://doi.org/10.13023/etd.2024.499

Funding Information

This research was supported by grants awarded to Warren J Alilain PhD from the Craig H Neilsen Foundation (no 598741) , the Kentucky Spinal Cord and Head Injury Research Trust, University of Kentucky Spinal Cord and Brain Injury Research Center Endowment, the National Institutes of Health (R01 NS101105 WJA) (R21 NS121966 WJA) additional funding included (NIH AG065220 Lance A Johnson PhD) (NIH AG065220 LAJ) (NIH AG80589 LAJ) (NIH AG081421 LAJ) (NIH AG06653 Ramon C Sun PhD) (NIH CA266004 RCS) (NIH AG078702 RCS) (Alzheimer’s Association grant LAJ) (V-Scholar Grant RCS)

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