Division of Spine Surgery, Department of Orthopaedics, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, Guangdong, China.
International Collaboration on Repair Discoveries (ICORD), Blusson Spinal Cord Centre, University of British Columbia, Vancouver, Canada.
Lipids Health Dis. 2024 Jun 22;23(1):194. doi: 10.1186/s12944-024-02185-y.
Lipid droplet (LD)-laden microglia is a key pathological hallmark of multiple sclerosis. The recent discovery of this novel microglial subtype, lipid-droplet-accumulating microglia (LDAM), is notable for increased inflammatory factor secretion and diminished phagocytic capability. Lipophagy, the autophagy-mediated selective degradation of LDs, plays a critical role in this context. This study investigated the involvement of microRNAs (miRNAs) in lipophagy during demyelinating diseases, assessed their capacity to modulate LDAM subtypes, and elucidated the potential underlying mechanisms involved.
C57BL/6 mice were used for in vivo experiments. Two weeks post demyelination induction at cervical level 4 (C4), histological assessments and confocal imaging were performed to examine LD accumulation in microglia within the lesion site. Autophagic changes were observed using transmission electron microscopy. miRNA and mRNA multi-omics analyses identified differentially expressed miRNAs and mRNAs under demyelinating conditions and the related autophagy target genes. The role of miR-223 in lipophagy under these conditions was specifically explored. In vitro studies, including miR-223 upregulation in BV2 cells via lentiviral infection, validated the bioinformatics findings. Immunofluorescence staining was used to measure LD accumulation, autophagy levels, target gene expression, and inflammatory mediator levels to elucidate the mechanisms of action of miR-223 in LDAM.
Oil Red O staining and confocal imaging revealed substantial LD accumulation in the demyelinated spinal cord. Transmission electron microscopy revealed increased numbers of autophagic vacuoles at the injury site. Multi-omics analysis revealed miR-223 as a crucial regulatory gene in lipophagy during demyelination. It was identified that cathepsin B (CTSB) targets miR-223 in autophagy to integrate miRNA, mRNA, and autophagy gene databases. In vitro, miR-223 upregulation suppressed CTSB expression in BV2 cells, augmented autophagy, alleviated LD accumulation, and decreased the expression of the inflammatory mediator IL-1β.
These findings indicate that miR-223 plays a pivotal role in lipophagy under demyelinating conditions. By inhibiting CTSB, miR-223 promotes selective LD degradation, thereby reducing the lipid burden and inflammatory phenotype in LDAM. This study broadens the understanding of the molecular mechanisms of lipophagy and proposes lipophagy induction as a potential therapeutic approach to mitigate inflammatory responses in demyelinating diseases.
脂滴（LD）负荷的小胶质细胞是多发性硬化症的一个关键病理标志。最近发现的这种新型小胶质细胞亚型，即脂滴积累小胶质细胞（LDAM），其特点是炎症因子分泌增加和吞噬能力降低。脂噬作用，即自噬介导的 LD 选择性降解，在这种情况下起着关键作用。本研究调查了微小 RNA（miRNA）在脱髓鞘疾病中的脂噬作用中的作用，评估了它们调节 LDAM 亚型的能力，并阐明了涉及的潜在机制。
使用 C57BL/6 小鼠进行体内实验。在颈 4 水平（C4）诱导脱髓鞘后 2 周，进行组织学评估和共聚焦成像，以检查病变部位小胶质细胞中的 LD 积累。使用透射电子显微镜观察自噬变化。miRNA 和 mRNA 多组学分析鉴定了脱髓鞘条件下差异表达的 miRNA 和 mRNAs 以及相关的自噬靶基因。特别探讨了 miR-223 在这些条件下的脂噬作用。通过慢病毒感染上调 BV2 细胞中的 miR-223 的体外研究，验证了生物信息学发现。免疫荧光染色用于测量 LD 积累、自噬水平、靶基因表达和炎症介质水平，以阐明 miR-223 在 LDAM 中的作用机制。
油红 O 染色和共聚焦成像显示脱髓鞘脊髓中大量 LD 积累。透射电子显微镜显示损伤部位自噬小体数量增加。多组学分析表明，miR-223 是脱髓鞘过程中脂噬作用的关键调节基因。研究发现，组织蛋白酶 B（CTSB）是自噬中 miR-223 的靶点，整合了 miRNA、mRNA 和自噬基因数据库。在体外，miR-223 的上调抑制了 BV2 细胞中 CTSB 的表达，增强了自噬，减轻了 LD 积累，并降低了炎症介质 IL-1β的表达。
这些发现表明，miR-223 在脱髓鞘条件下的脂噬作用中起着关键作用。通过抑制 CTSB，miR-223 促进了选择性 LD 降解，从而减轻了 LDAM 中的脂质负担和炎症表型。本研究拓宽了对脂噬作用分子机制的理解，并提出了诱导脂噬作用作为减轻脱髓鞘疾病炎症反应的潜在治疗方法。
