Department of Obstetrics and Gynecology, Guangdong Provincial Key Laboratory of Major Obstetric Diseases, Guangdong Provincial Clinical Research Center for Obstetrics and Gynecology, Guangdong-Hong Kong-Macao Greater Bay Area Higher Education Joint Laboratory of Maternal-Fetal Medicine, The Third Affiliated Hospital of Guangzhou Medical University, Guangzhou, China.
Department of Neurology, Tianjin Neurological Institute, Tianjin Institute of Immunology, State Key Laboratory of Experimental Hematology, Haihe Laboratory of Cell Ecosystem, Tianjin Medical University General Hospital, Tianjin, China.
Aging (Albany NY). 2024 Aug 7;16(15):11656-11667. doi: 10.18632/aging.206045.
Several aberrant alternative splicing (AS) events and their regulatory mechanisms are widely recognized in multiple sclerosis (MS). Yet the cell-type specific AS events have not been extensively examined. Here we assessed the diversity of AS events using web-based RNA-seq data of sorted CD15CD11b microglia in white matter (WM) region from 10 patients with MS and 11 control subjects. The GSE111972 dataset was downloaded from GEO and ENA databases, aligned to the GRCh38 reference genome from ENSEMBL via STAR. rMATS was used to assess five types of AS events, alternative 3'SS (A3SS), alternative 5'SS (A5SS), skipped exon (SE), retained intron (RI) and mutually exclusive exons (MXE), followed by visualizing with rmats2sashimiplot and maser. Differential genes or transcripts were analyzed using the limma R package. Gene ontology (GO) analysis was performed with the clusterProfiler R package. 42,663 raw counts of AS events were identified and 132 significant AS events were retained based on the filtered criteria: 1) average coverage >10 and 2) delta percent spliced in (ΔPSI) >0.1. SE was the most common AS event (36.36%), followed by MXE events (32.58%), and RI (18.94%). Genes related to telomere maintenance and organization primarily underwent SE splicing, while genes associated with protein folding and mitochondrion organization were predominantly spliced in the MXE pattern. Conversely, genes experiencing RI were enriched in immune response and immunoglobulin production. In conclusion, we identified microglia-specific AS changes in the white matter of MS patients, which may shed light on novel pathological mechanisms underlying MS.
多种多发性硬化症（MS）中存在广泛的异常剪接（AS）事件及其调控机制。然而，细胞类型特异性的 AS 事件尚未得到广泛研究。在此，我们使用 10 名 MS 患者和 11 名对照的白质（WM）区 CD15+CD11b+小胶质细胞的基于网络的 RNA-seq 数据评估 AS 事件的多样性。从 GEO 和 ENA 数据库下载 GSE111972 数据集，通过 STAR 与 ENSEMBL 的 GRCh38 参考基因组进行比对。使用 rMATS 评估五种 AS 事件，包括选择性 3'剪接位点（A3SS）、选择性 5'剪接位点（A5SS）、外显子跳跃（SE）、内含子保留（RI）和互斥外显子（MXE），然后使用 rmats2sashimiplot 和 maser 可视化。使用 limma R 包分析差异基因或转录本。使用 clusterProfiler R 包进行基因本体（GO）分析。共鉴定出 42663 个 AS 事件的原始计数，基于过滤标准保留了 132 个显著的 AS 事件：1）平均覆盖度>10 和 2）剪接百分比变化（ΔPSI）>0.1。SE 是最常见的 AS 事件（36.36%），其次是 MXE 事件（32.58%）和 RI 事件（18.94%）。与端粒维持和组织相关的基因主要发生 SE 剪接，而与蛋白质折叠和线粒体组织相关的基因主要以 MXE 模式剪接。相反，经历 RI 的基因在免疫反应和免疫球蛋白产生中富集。总之，我们在 MS 患者的 WM 中鉴定出小胶质细胞特异性的 AS 变化，这可能为 MS 的新病理机制提供线索。
