Single cell RNA sequencing analysis of vestibular schwannoma reveals functionally distinct macrophage subsets. Single cell RNA sequencing analysis of vestibular schwannoma reveals functionally distinct macrophage subsets

Background: Vestibular schwannomas (VS) remain a challenge due to their anatomical location and propensity to growth. Macrophages are present in VS but their roles in VS pathogenesis remains unknown. Objectives: Assess phenotypic and functional profile of macrophages in VS with single cell RNA sequencing (scRNAseq). Methods: scRNAseq was carried out in three VS samples to examine characteristics of macrophages in the tumor. RT-qPCR was carried out on ten VS samples for CD14, CD68 and CD163 and a panel of macrophage associated molecules. Results: scRNAseq revealed macrophages to be a major constituent of VS microenvironment with three distinct subclusters based on gene expression. The subclusters were also defined by expression of CD163, CD68 and IL-1β. AREG and PLAUR were expressed in the CD68+CD163+IL-1β+ subcluster, PLCG2 and NCKAP5 were expressed in CD68+CD163+IL-1β- subcluster and AUTS2 and SPP1 were expressed in the CD68+CD163-IL-1β+ subcluster. RT-qPCR showed expression of several macrophage markers in VS of which CD14, ALOX15, Interleukin-1β, INHBA and Colony Stimulating Factor-1R were found to have a high correlation with tumor volume. Conclusions: Macrophages form an important component of VS stroma. scRNAseq reveals three distinct subsets of macrophages in the VS tissue which may have differing roles in the pathogenesis of VS. Overall design: single cell RNA sequencing analysis of vestibular schwannoma

研究背景：前庭神经鞘瘤（Vestibular schwannomas，VS）因其特殊的解剖位置与生长特性，仍是临床诊疗的一大挑战。肿瘤组织内存在巨噬细胞，但其在前庭神经鞘瘤发病机制中的具体作用仍未明确。
研究目的：采用单细胞RNA测序（single cell RNA sequencing，scRNAseq）技术，分析前庭神经鞘瘤中巨噬细胞的表型与功能特征。
研究方法：对3例前庭神经鞘瘤样本开展单细胞RNA测序，以探究肿瘤内巨噬细胞的特征；同时对10例前庭神经鞘瘤样本进行逆转录实时定量聚合酶链反应（RT-qPCR）检测，检测目标涵盖CD14、CD68、CD163及一组巨噬细胞相关分子。
研究结果：单细胞RNA测序结果显示，巨噬细胞是前庭神经鞘瘤微环境的主要组成成分，基于基因表达特征可划分为3个独特的亚群；这些亚群可通过CD163、CD68及白细胞介素-1β（Interleukin-1β，IL-1β）的表达谱进行界定。其中，CD68+CD163+IL-1β+亚群表达AREG与PLAUR，CD68+CD163+IL-1β-亚群表达PLCG2与NCKAP5，CD68+CD163-IL-1β+亚群表达AUTS2与SPP1。逆转录实时定量聚合酶链反应检测发现，前庭神经鞘瘤组织中存在多种巨噬细胞标志物的表达，其中CD14、ALOX15、白细胞介素-1β、INHBA及集落刺激因子-1受体（Colony Stimulating Factor-1R，CSF-1R）的表达水平与肿瘤体积呈高度相关。
研究结论：巨噬细胞是前庭神经鞘瘤间质的重要组成成分；单细胞RNA测序揭示了前庭神经鞘瘤组织内巨噬细胞存在3个独特亚群，这些亚群可能在肿瘤发病机制中发挥不同作用。
实验整体设计：针对前庭神经鞘瘤开展单细胞RNA测序分析。