Wang Yan, Liu Cuiying, Chen Yong, Chen Tiffany, Han Tao, Xue Lixiang, Xu Baohui
Institute of Medical Innovation and Research, Peking University Third Hospital, Beijing, China.
Medical Research Center, Peking University Third Hospital, Beijing, China.
Front Cardiovasc Med. 2022 May 6;9:876087. doi: 10.3389/fcvm.2022.876087. eCollection 2022.
Maclpil is a proinflammatory long non-coding RNA highly expressed on monocyte-derived macrophages in the ischemic brain. This study investigated the impact and the mechanisms of systemically delivering nanoparticle Maclpil short interfering RNA (siRNA) on experimental ischemic stroke in a mouse model.
Ischemic stroke (focal cerebral ischemia) was induced in male C57BL/6 mice through the middle cerebral artery occlusion. Three hours thereafter, mice were intravenously injected with Maclpil siRNA or scramble siRNA nanoparticles. Bone marrow cell-derived macrophages were transfected with Maclpil or scramble siRNA and subjected to oxygen glucose deprivation culture. The influence of silencing Maclpil on stroke outcomes, neuroinflammation, and macrophage fates was assessed histology, flow cytometry, Western blotting, and quantitative PCR analysis.
Three days following stroke induction, siRNA silencing Maclpil substantially reduced ischemic infarction size and improved neurological behaviors. Silencing Maclpil also markedly attenuated the accumulation of monocyte-derived macrophages, CD4 T cells, and CD8 T cells in the ischemic hemisphere without affecting microglia cellularity. Reciprocally, myeloid cells and both subsets of T cells were elevated in mouse peripheral blood following Maclpil siRNA treatment. Under oxygen glucose deprivation conditions that mimicked hypoxia and hypoglycemia , Maclpil siRNA silencing augmented macrophage apoptosis in conjunction with upregulation of proapoptotic Bax and caspase 3 expressions. siRNA knocking down Maclpil skewed macrophages from proinflammatory classical toward anti-inflammatory alternative activation as evidenced by increased arginase 1, Ym1, and Fizz1 and reduced inducible nitric oxide synthase, IL-1β, and TNF-α mRNA levels. Consistent with macrophage phenotype switching, silencing Maclpil by siRNA enhanced fatty acid oxidation as indicated by increased mRNA levels of 3 key metabolic enzymes (ACADM, ACADVL, and HADHA).
Systemically silencing Maclpil by siRNA nanoparticles attenuated experimental ischemic stroke by promoting macrophage apoptosis and anti-inflammatory alternative activation. Identifying and targeting Maclpil human homolog(s) may help develop a novel therapy for stroke clinical management.
Maclpil是一种促炎性长链非编码RNA，在缺血性脑内单核细胞衍生的巨噬细胞上高度表达。本研究在小鼠模型中探究了全身递送纳米颗粒Maclpil小干扰RNA（siRNA）对实验性缺血性中风的影响及机制。
通过大脑中动脉闭塞在雄性C57BL/6小鼠中诱导缺血性中风（局灶性脑缺血）。此后3小时，给小鼠静脉注射Maclpil siRNA或乱序siRNA纳米颗粒。用Maclpil或乱序siRNA转染骨髓细胞衍生的巨噬细胞，并进行氧糖剥夺培养。通过组织学、流式细胞术、蛋白质免疫印迹和定量PCR分析评估沉默Maclpil对中风结局、神经炎症和巨噬细胞命运的影响。
中风诱导后3天，沉默Maclpil的siRNA显著减小了缺血梗死面积并改善了神经行为。沉默Maclpil还显著减轻了缺血半球中单核细胞衍生的巨噬细胞、CD4 T细胞和CD8 T细胞的积聚，而不影响小胶质细胞数量。相反，Maclpil siRNA处理后小鼠外周血中的髓样细胞和两个T细胞亚群均升高。在模拟缺氧和低血糖的氧糖剥夺条件下，Maclpil siRNA沉默增加了巨噬细胞凋亡，同时上调了促凋亡蛋白Bax和半胱天冬酶3的表达。敲低Maclpil的siRNA使巨噬细胞从促炎性经典激活偏向抗炎性替代激活，这表现为精氨酸酶1、Ym1和Fizz1增加，以及诱导型一氧化氮合酶、IL-1β和TNF-α mRNA水平降低。与巨噬细胞表型转换一致，siRNA沉默Maclpil增强了脂肪酸氧化，这表现为3种关键代谢酶（ACADM、ACADVL和HADHA）的mRNA水平升高。
通过siRNA纳米颗粒全身沉默Maclpil可通过促进巨噬细胞凋亡和抗炎性替代激活来减轻实验性缺血性中风。鉴定并靶向Maclpil的人类同源物可能有助于开发一种用于中风临床治疗的新疗法。
