Abstract Body: Introduction: Subarachnoid hemorrhage (SAH) is associated with elevated morbidity and mortality. We previously showed that the immunomodulatory agent FTY720 improves neurological outcome in rats subjected to SAH.
Hypothesis: FTY720 protects against SAH-associated neuronal cell death and white matter injury (WMI).
Methods: We used the endovascular perforation model. Animals were divided into Sham, SAH-vehicle, and SAH-FTY720 groups. The SAH-FTY720 group received a single dose of FTY720 (0.5 mg/kg) intraperitoneally 3 hours after surgery. Brains were isolated and analyzed on day 7 post-injury. Neuroinflammation was ascertained by immunostaining for astrocyte (GFAP) and microglia (Iba-1) cell and the expression of proinflammatory mediators inducible nitric-oxide synthase (iNOS) through the NF-κB signaling pathway and TLR4. WMI was determined by quantifying the expression of myelin basic protein (MBP) and oligodendrocyte precursor cells (NG2). Neuronal death was determined by NeuN staining. Statistical analyses were performed with one-way ANOVA followed by Tukey’s post-hoc test using Graph Prism 10. Data are represented as mean±SD. Statistical significance was determined at the level of p<0.05.
Results: 5 rats were included per group. Relative to the Sham group, SAH-vehicle animals had a decreased number of NeuN(+) cells but an increased number of Iba-1(+) (p<0.001) and GFAP(+) cells (p<0.001) in the ipsilateral cortex and CA1 region of the hippocampus. FTY720 after SAH normalized the number of NeuN(+), Iba-1(+), and GFAP(+) cells in both regions (p<0.001 vs SAH-FTY720). These findings were also confirmed by Western blot analysis for GFAP and Iba1. The SAH-vehicle group also had an increased expression of iNOS and TLR4 in cortex and hippocampus (p<0.001 vs Sham) which colocalized with Iba-1 and GFAP. Furthermore, SAH increased the expression of activated NF-κB (Pp65) but decreased the expression of the NF-kB inhibitor, IkBα in the cortex and hippocampus as compared to the Sham group (p<0.001 vs Sham). The expressions of iNOS, TLR4, IkBα, and Pp65 were normalized in the SAH-FTY720 group. The number of MBP(+) and NG2 (+) cells was decreased in the ipsilateral cortex and corpus callosum of SAH-vehicle animals as compared to Sham group (p<0.001 vs Sham). Conversely, the expression of both markers in the SAH-FTY720 group was similar to the Sham group in both regions.
Conclusion: FTY720 reduces neuroinflammation, neuronal death, and WMI after SAH.