Transcriptional profiling of murine oligodendrocyte precursor cells across the lifespan

Oligodendrocyte progenitor cells (OPCs) are highly dynamic, abundant glial cells of the central nervous system (CNS) that are responsible for generating myelinating oligodendrocytes during development. OPCs are also mobilized to form new myelin sheaths in the adult CNS in response to environmental and behavioral changes, and play a crucial role in regenerating myelin following demyelination (remyelination). However, the rates of OPC proliferation and differentiation decline dramatically with aging, impairing homeostasis, remyelination, and potentially adaptive myelination during learning. To determine how aging influences OPCs, we generated a novel transgenic mouse line that expresses EGFP under the endogenous promoter/enhancer of Matrilin-4 (Matn4-mEGFP), allowing OPCs to be purified apart from perivascular and mural cells. OPCs isolated from the cerebral cortex of Matn4-mEGFP mice were subjected to single-cell RNA sequencing, providing enhanced resolution of transcriptional changes during key transitions from quiescence to proliferation and differentiation. Comparative analysis of OPCs isolated from mice aged 30 to 720 days, revealed that aging induces distinct inflammatory transcriptomic changes in OPCs in different states, including enhanced activation of HIF-1 and Wnt pathways. Inhibition of these pathways in acutely isolated OPCs from aged mice promoted their differentiation, suggesting that this enhanced signaling may contribute to the decreased regenerative potential of OPCs with aging. This Matn4-mEGFP mouse line and single-cell mRNA datasets of cortical OPCs across ages serve as a valuable reference to help define the molecular changes guiding their behavior in various physiological and pathological contexts. Matn4-mEGFP OPCs from 1) 6x P30 cortex, 2) 4x P180 cortex, 3) 5x P360 cortex, 4) 5x P720 cortex to study the effect of aging on the transcriptome of OPCs in the mouse cortex少突胶质细胞前体细胞（Oligodendrocyte progenitor cells, OPCs）是中枢神经系统（central nervous system, CNS）中一类高度动态且分布广泛的胶质细胞，在个体发育过程中负责分化为成髓鞘少突胶质细胞。在成年中枢神经系统中，OPCs可被动员以响应环境与行为改变，生成新的髓鞘，并在脱髓鞘损伤后的髓鞘再生（remyelination）过程中发挥核心作用。然而，随着机体衰老，OPCs的增殖与分化速率会显著下降，破坏其稳态平衡，损害髓鞘再生能力，同时可能影响学习过程中的适应性髓鞘形成。 为阐明衰老对OPCs的调控机制，本研究构建了一种新型转基因小鼠品系：该小鼠在Matrilin-4（Matn4）的内源性启动子/增强子调控下表达增强型绿色荧光蛋白（Enhanced Green Fluorescent Protein, EGFP），可实现OPCs与血管周细胞及壁细胞的分离纯化。从该转基因小鼠大脑皮层中分离得到的OPCs被用于单细胞RNA测序（single-cell RNA sequencing），该技术显著提升了对OPCs从静息状态向增殖、分化关键过渡阶段转录组变化的解析分辨率。 通过对30至720日龄小鼠皮层OPCs的比较转录组分析，本研究发现衰老会在不同功能状态的OPCs中诱导独特的炎症相关转录组改变，包括缺氧诱导因子-1（Hypoxia-Inducible Factor 1, HIF-1）与Wnt信号通路（Wnt pathways）的过度激活。在从衰老小鼠体内急性分离的OPCs中抑制上述两条通路，可促进其分化，提示这种过度激活的信号通路可能是衰老导致OPCs再生能力下降的关键诱因。 本研究构建的Matn4-mEGFP转基因小鼠品系，以及不同年龄小鼠皮层OPCs的单细胞mRNA测序数据集，可为明确OPCs在多种生理与病理过程中的行为调控分子机制提供重要参考依据。本研究使用的样本包括：1）6组P30龄小鼠皮层来源的Matn4-mEGFP阳性OPCs，2）4组P180龄小鼠皮层来源的Matn4-mEGFP阳性OPCs，3）5组P360龄小鼠皮层来源的Matn4-mEGFP阳性OPCs，4）5组P720龄小鼠皮层来源的Matn4-mEGFP阳性OPCs，旨在探究衰老对小鼠皮层OPCs转录组的影响。