CHARACTERIZATION OF THE ACUTE GENE RESPONSE IN ZEBRAFISH DEMYELINATION MODEL

Zebrafish possess an extensive capacity for CNS regeneration, providing a unique model to identify the mechanisms of successful repair, unlike mammals which own limited regenerative capacity. To investigate the molecular mechanisms of CNS demyelination in zebrafish, we established a focal myelin lesion model utilizing lysophosphatidylcholine (LPC) microinjection into the spinal cord. To investigate cellular and molecular changes we used high-resolution live imaging and RNA sequencing.
We used double-transgenic Tg(cldk:mem-tdTomato; mpeg1.1:GFP) zebrafish larvae for visualizing myelin structure and microglia, respectively. High-resolution fluorescent live imaging at 5dpi, confirmed robust myelin sheath fragmentation concurrent with rapid microglial infiltration into the lesion site at 1 dpi. Next, we investigated transcriptomic profiling of peak injury window by RNA sequencing.
Results revealed a profound functional reprogramming of the lesion microenvironment. We observed significant downregulation of myelin related gene expression accompanied with upregulation of microglia related genes, supporting imaging results. Further, we observed a robust pro-inflammatory signature marked by the significant upregulation of certain TNF-alpha and NF-kB regulators, other master regulators and cytokine receptors. This activation is further evidenced by increased expression of mpeg1.2 and defined chemokines, while homeostatic regulators were generally downregulated. Crucially, this immune response is coupled with a lipid-clearance program: significant upregulation of apolipoproteins and metabolic enzymes, suggesting increased microglial lipid processing. These findings indicate that the zebrafish myelin injury response prioritizes the rapid conversion of microglia into "foamy" phagocytes to actively catabolize myelin debris, establishing the necessary debris-free niche required for subsequent remyelination, which will be further investigated