Transcriptome studies have used 3’-biased microarrays to study regulated genes and pathways in the blood of ischemic stroke (IS) patients, missing a majority of transcript isoforms due to alternative splicing (AS). AS is implicated in many diseases but is unexplored with respect to IS and intracerebral hemorrhage (ICH). Thus, we hypothesized AS differs between 1) IS, ICH and vascular risk factor control (VRFC) subjects; 2) three main causes of IS (Cardioembolic, Large Vessel and Lacunar), ICH and VRFC.
For Aim1, differential alternative splicing (DAS) in whole blood of IS vs ICH vs VRFC males was studied on Affymetrix exon junction HTA 2.0 arrays (n=36, 12 per group). For Aim 2, we investigated DAS between Cardioembolic, Large Vessel and Lacunar IS etiologies, and ICH and VRFC using paired-end RNA-Seq (Illumina Solexa, 200M 2X100bp reads per sample) on 20 whole blood RNA samples (n=4 males per group). DAS was assessed with 1-way-ANOVA (FDR p<0.05). Significant differential exon-usage was calculated between each group (p<0.0005; fold change> |1.2|). Differential usage of 90 exons separated the IS, ICH and VRFC groups. Affected genes included HGF (hepatic growth factor), involved in thrombosis and angiogenesis, and NOS1 (nitric oxide synthase 1), implicated in stroke risk. DAS among ICH, three main causes of IS, and VRFC was displayed by 412 genes. Differential expression of 308 exons separated these 5 groups. Several genes with DAS belonged to NF-kB and PPAR signaling pathways, SRA1, steroid receptor RNA activator 1, and Akt1, all of which have been implicated in experimental ICH pathogenesis. ICH activates NF-kB transcription factor which perpetuates inflammation. PPAR-gamma, via inhibition of NF-kB and induction of antioxidative pathways, reduces inflammation and oxidative stress and stimulates phagocytosis-mediated hematoma cleanup. In conclusion, our results provide first evidence for DAS in whole blood in ICH vs IS, and vs different IS etiologies. These data indicate disease specific immune response and could yield biomarkers for ICH and various IS causes. Future studies in a larger cohort are needed to validate results.