Primary cortical cell tri-culture to study effects of amyloid-β on microglia function and neuroinflammatory response.

IF 3.4 3区 医学 Q2 NEUROSCIENCES
Hyehyun Kim, Bryan Le, Noah Goshi, Kan Zhu, Ana Cristina Grodzki, Pamela J Lein, Min Zhao, Erkin Seker
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引用次数: 0

Abstract

Background: Microglia play a critical role in neurodegenerative disorders, such as Alzheimer's disease, where alterations in microglial function may result in pathogenic amyloid-β (Aβ) accumulation, chronic neuroinflammation, and deleterious effects on neuronal function. However, studying these complex factors in vivo, where numerous confounding processes exist, is challenging, and until recently, in vitro models have not allowed sustained culture of critical cell types in the same culture.

Objective: We employed a rat primary tri-culture (neurons, astrocytes, and microglia) model and compared it to co-culture (neurons and astrocytes) and mono-culture (microglia) to study microglial function (i.e., motility and Aβ clearance) and proteomic response to exogenous Aβ.

Methods: The cultures were exposed to fluorescently-labeled Aβ (FITC-Aβ) particles for varying durations. Epifluorescence microscopy images were analyzed to quantify the number of FITC-Aβ particles and assess cytomorphological features. Cytokine profiles from conditioned media were obtained. Live-cell imaging was employed to extract microglia motility parameters.

Results: FITC-Aβ particles were more effectively cleared in the tri-culture compared to the co-culture. This was attributed to microglia engulfing FITC-Aβ particles, as confirmed via epifluorescence and confocal microscopy. FITC-Aβ treatment significantly increased microglia size, but had no significant effect on neuronal surface coverage or astrocyte size. Upon FITC-Aβ treatment, there was a significant increase in proinflammatory cytokines in tri-culture, but not in co-culture. Aβ treatment altered microglia motility evident as a swarming-like motion.

Conclusions: The results suggest that neuron-astrocyte-microglia interactions influence microglia function and highlight the utility of the tri-culture model for studies of neuroinflammation, neurodegeneration, and cell-cell communication.

原代皮质细胞三层培养法研究淀粉样蛋白-β对小胶质细胞功能和神经炎症反应的影响。
背景:小胶质细胞在阿尔茨海默病等神经退行性疾病中发挥着关键作用,小胶质细胞功能的改变可能导致致病性淀粉样蛋白-β(Aβ)积累、慢性神经炎症以及对神经元功能的有害影响。然而,在体内研究这些复杂的因素具有挑战性,因为体内存在许多混杂的过程,而且直到最近,体外模型还不能在同一培养物中持续培养关键的细胞类型:我们采用了大鼠原代三培养(神经元、星形胶质细胞和小胶质细胞)模型,并将其与共培养(神经元和星形胶质细胞)和单培养(小胶质细胞)进行了比较,以研究小胶质细胞的功能(即运动性和 Aβ 清除率)以及蛋白质组对外源 Aβ 的反应:方法:培养物暴露于荧光标记的 Aβ(FITC-Aβ)颗粒的时间长短不一。对荧光显微镜图像进行分析,以量化 FITC-Aβ 颗粒的数量并评估细胞形态特征。从条件培养基中获得细胞因子图谱。活细胞成像用于提取小胶质细胞的运动参数:结果:与共培养相比,FITC-Aβ颗粒在三培养基中被更有效地清除。这归因于小胶质细胞吞噬了 FITC-Aβ 颗粒,并通过荧光显微镜和共聚焦显微镜得到了证实。FITC-Aβ 处理会明显增加小胶质细胞的体积,但对神经元表面覆盖率或星形胶质细胞的体积没有明显影响。经 FITC-Aβ 处理后,促炎细胞因子在三细胞培养中明显增加,但在共培养中没有增加。Aβ 处理会改变小胶质细胞的运动,表现为蜂拥般的运动:结果表明,神经元-胃细胞-小胶质细胞之间的相互作用会影响小胶质细胞的功能,并凸显了三细胞培养模型在神经炎症、神经变性和细胞-细胞通讯研究中的实用性。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
Journal of Alzheimer's Disease
Journal of Alzheimer's Disease 医学-神经科学
CiteScore
6.40
自引率
7.50%
发文量
1327
审稿时长
2 months
期刊介绍: The Journal of Alzheimer''s Disease (JAD) is an international multidisciplinary journal to facilitate progress in understanding the etiology, pathogenesis, epidemiology, genetics, behavior, treatment and psychology of Alzheimer''s disease. The journal publishes research reports, reviews, short communications, hypotheses, ethics reviews, book reviews, and letters-to-the-editor. The journal is dedicated to providing an open forum for original research that will expedite our fundamental understanding of Alzheimer''s disease.
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