CSF-Exosomal miRNAs and Delayed Cerebral Ischemia: Insights into Pathophysiology but No Definitive Biomarkers

Aneurysmal subarachnoid hemorrhage (aSAH) is notoriously known for its high mortality and morbidity. Approximately one-third of the patients who survive aneurysm rupture are reported to develop delayed cerebral ischemia (DCI), which contributes to a poor clinical outcome. Currently, there are no biomarkers for identifying which aSAH patients are at risk of developing DCI. We aimed to determine the feasibility of cerebrospinal fluid (CSF) exosomal microRNAs (miRNAs) for predicting DCI post-aSAH.

aSAH patients were prospectively enrolled, and CSF samples were collected at two time points (<24 h and 72 h post-aSAH) from individuals undergoing external ventricular drainage. Exosomal miRNAs were isolated from the CSF for analysis. In the initial group of patients (discovery cohort), an exploratory analysis was conducted using a CSF panel containing 84 miRNAs, assessed by quantitative real-time PCR (RT-qPCR). Based on this analysis, 27 miRNAs were selected for further evaluation in a second group of patients (validation cohort). Among these, 10 miRNAs had previously been reported in SAH-related CSF studies, supporting their relevance for continued investigation.

In this study, RT-qPCR analysis of 84 miRNAs in CSF samples from aSAH patients (n = 10 DCI, n = 16 no DCI) and non-aSAH controls (n = 5) identified 9 upregulated and 13 downregulated miRNAs in the DCI group, and 7 upregulated and 18 downregulated miRNAs in the no-DCI group, compared to the controls. When comparing DCI to no-DCI patients, 13 miRNAs were found to be upregulated in the DCI group. Additionally, seven miRNAs showed temporal upregulation in DCI patients between early (<24 h/T1) and later (72 h/T3) time points across both discovery and validation cohorts. However, no miRNAs were uniquely expressed in either DCI or no-DCI groups, limiting their potential as specific biomarkers for DCI.

Despite analyses in both the discovery and validation phases, no miRNAs emerged as consistent and reliable biomarkers for distinguishing DCI from no-DCI patients. However, the identified miRNAs are involved in the key KEGG pathways that regulate vascular integrity, neuronal survival, and inflammatory processes central to DCI pathophysiology. These findings highlight the complexity of miRNA regulation following aSAH, as reflected by the variability in differentially expressed miRNAs between cohorts. This variability may be influenced by factors such as limited sample size, patient heterogeneity, individual biological differences, and experimental variability. Comprehensive profiling using larger, well-characterized cohorts, along with rigorous validation, is essential to determine the predictive value and mechanistic significance of candidate miRNAs in DCI.