Abstract
Purpose :
We previously reported on the neuroprotective activity of the lipid mediators, lipoxin A4 and B4 (LXA4 and LXB4). However, the downstream signaling cascades induced by these molecules in neuronal cells are still unclear. Here we aimed to develop a preliminary molecular signature describing LXA4 and LXB4 transcriptomic changes following treatment in neuronal cells.
Methods :
Using next-generation sequencing, 4 treatment groups were analyzed: LXA4, LXB4,15-HETE (an inactive LXB4 precursor) and Vehicle. The 15-HETE and Vehicle groups served as controls to identify genes specifically induced by each lipoxin treatment. HT22 neuronal cells were plated at 1x106 cells per well in supplemented high glucose DMEM media and seeded overnight at 37°C. The following day, cells were treated with LXB4, LXA4, 15-HETE and Vehicle at 1μM in 4 biological replicates. Cell lysates were collected 1 hour after treatment, and total RNA was purified and seqeunced using an Illumina platform. Following sequencing, the data was bioinformatically sorted for differential gene expression analysis. Treatment groups were then fruther comparatively analyzed in R to identify lipoxin-specific transcripts.
Results :
In comparison to vehicle 242 genes were identified specific to LXB4 treatment, 539 for LXA4, and 440 genes for 15-HETE treatment (p<0.01). Interestingly, only 23 genes were common between LXA4 and LXB4. In further analyses 15-HETE-specific genes were matched against LXA4 and LXB4 and common reads were removed to generate a molecular signature specific to each lipoxin treatment. The final unique gene counts were 185 and 315 genes with altered expression for LXB4 and LXA4 treatment, respectively.
Conclusions :
A preliminary neuronal molecular signature for LXA4 and LXB4 signaling was identified, consisting of altered transcripts unique to each treatment. Next steps include ongoing characterization of the top hits under a variety of conditions and timepoints.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.