SMART-seq of Microglia in a Mouse Subarachnoid Hemorrhage Model
AbstractSubarachnoid hemorrhage (SAH) is a devastating form of stroke with high mortality and morbidity. The inflammatory response, particularly the activation of microglia, the resident immune cells of the brain, is a critical contributor to early brain injury and secondary complications following SAH. However, the precise transcriptomic changes in microglia during the acute phase of SAH remain incompletely characterized, in part due to the technical challenge of obtaining sufficient high-quality RNA from these acutely isolated cells. This study employs the SMART-seq2 protocol, a full-length RNA-seq method optimized for low-input samples, to comprehensively map the transcriptome of microglia acutely isolated from the brains of wild-type mice following experimental SAH.Experimental Design and MethodsWe induced SAH in adult wild-type C57BL/6 mice using a standardized prechiasmatic injection model. Sham-operated animals served as controls. At 24 hours post-operation, a critical time point for acute neuroinflammation, microglia were acutely isolated from the whole brain using a combination of mechanical and enzymatic dissociation, followed by CD11b-positive selection. Due to the limited cell yield from this acute isolation protocol, total RNA was extracted and amplified using the SMART-seq2 method to construct sequencing libraries. This approach allows for high-sensitivity, full-length transcript coverage from minute amounts of input RNA. Sequencing was performed on the Illumina NovaSeq 6000 platform to generate 150 bp paired-end reads. The dataset includes a total of 8 samples: 3 biological replicates for the sham group and 5 biological replicates for the SAH group.Data Content and Potential ApplicationsThis dataset provides a detailed and quantitative view of the gene expression changes in microglia during the acute phase of SAH, specifically enabled by a low-input RNA-seq technology. The raw sequencing data (FASTQ files) are deposited here. We anticipate that this resource will be invaluable for:Identifying differentially expressed genes (DEGs) and key signaling pathways involved in the acute microglial response to SAH with high transcript detection sensitivity.Gaining insights into the molecular mechanisms underlying microglia-mediated neuroinflammation and neural damage.Discovering novel transcriptional regulators and potential therapeutic targets for mitigating early brain injury after SAH.Serving as a high-quality reference for studying microglial biology in conditions where cell numbers are limited.Enabling alternative splicing and isoform-level analysis due to the full-length transcript coverage provided by the SMART-seq2 method.