Using CXCR4 Antagonism and G-CSF to Induce Sustained Neutrophilia in Mice for Applications in Spinal Cord Injury Research
Abstract
In spinal cord injury, the immune response can affect the damage scope and functional recovery. However, without a comparable pre-clinical model of spinal cord injury, it is extremely difficult to research potential therapeutic treatments for inflammation-associated damage after spinal cord injury in humans. The goal of this project was to increase the abundance of circulating neutrophils in mice to a comparable human rate of 50-70% of total white blood cells to improve the murine pre-clinical model of spinal cord injury. To induce sustained neutrophilia, we assessed two independent methods. The first strategy utilized C-X-C chemokine receptor 4 (CXCR4) antagonism and we injected AMD3100 (Plerixafor), which is a drug known to target CXCR4. The second strategy we employed was to deliver G-CSF via a plasmid vector, injected directly into the bloodstream, to induce the proliferation of pre-neutrophil progenitor cells. To assess neutrophilia, we utilized flow cytometry and characterized neutrophil abundance out of total leukocytes in the peripheral blood and bone marrow, as well as neutrophil maturation. Using a 5 mg/kg dose of AMD3100, we did not observe an increase in neutrophils in the peripheral blood at 3 hours or 24 hours post-injection. Using a 10 mg/kg dose of AMD3100, we observed a significant increase in neutrophils in the peripheral blood at 3 hours that diminished by 24 hours post-injection. We also observed a significant decrease in the percentage of mature neutrophils in both the bone marrow and the peripheral blood 3 hours after the 10 mg/kg injection of AMD3100. Using the CSF3 (G-CSF) gene packaged in a non-viral vector and injected into the mouse retro-orbitally, we did not observe an increase in neutrophil abundance in the bone marrow or peripheral blood after a 1 or 2-week timepoint. We did, however, observe a marked increase in the number of cells in the bone marrow compared to a vehicle injection at both timepoints.
Subject
SCISpinal Cord Injury
Neutrophils
Mice
Immunology
CXCR4, AMD3100
G-CSF
pLIVE
Flow Cytometry
Sustained Neutrophilia
Citation
Maale, Marina Denise (2023). Using CXCR4 Antagonism and G-CSF to Induce Sustained Neutrophilia in Mice for Applications in Spinal Cord Injury Research. Master's thesis, Texas A&M University. Available electronically from https : / /hdl .handle .net /1969 .1 /199730.