Department of Physiology, The University of Texas Southwestern Medical Center at Dallas, Dallas, Texas, USA.
Glia. 2012 Feb;60(2):189-202. doi: 10.1002/glia.21254. Epub 2011 Oct 11.
The exact biological role of the cytokine tumor necrosis factor (TNF) in the central nervous system (CNS) is not well understood; but overproduction of TNF by activated microglia has been implicated in neuronal death, suggesting that TNF inhibition in the CNS may be a viable neuroprotective strategy. We investigated the role of TNF signaling in regulation of microglia effector functions using molecular, cellular, and functional analyses of postnatal and adult microglia populations in the CNS. No differences were found by flow cytometric analyses in the basal activation state between TNF-null and wild-type mice. Although TNF-null microglia displayed an atypical morphology with cytoplasmic vacuoles in response to stimulation with lipopolysaccharide (LPS), the phagocytic response of TNF-null microglia to Escherichia coli particles in vitro was normal and there were no signs of enhanced caspase 3 activation or apoptosis. Functionally, conditioned media from LPS-stimulated TNF-null microglia was found to have significantly reduced levels of IL-10, IL-6, IL-1β, IL-12, and CXCL1 relative to wild-type microglia and exerted no cytotoxic effects on neurally differentiated dopaminergic (DA) MN9D cells. In contrast, incubation of wild-type microglia with TNF inhibitors selectively depleted the levels of soluble TNF and its cytotoxicity on MN9D cells. To distinguish whether reduced cytotoxicity by LPS-activated TNF-null microglia could be attributed to deficient autocrine TNF signaling, we employed primary microglia deficient in one or both TNF receptors (TNFR1 and TNFR2) in co-culture with MN9D cells and found that neither receptor is required to elicit LPS-evoked TNF production and cytotoxicity on DA cells.
细胞因子肿瘤坏死因子 (TNF) 在中枢神经系统 (CNS) 中的确切生物学作用尚不清楚；但是，活化的小胶质细胞中 TNF 的过度产生与神经元死亡有关，这表明 CNS 中 TNF 的抑制可能是一种可行的神经保护策略。我们使用分子、细胞和功能分析方法研究了 TNF 信号转导在调节小胶质细胞效应功能中的作用，这些方法涉及 CNS 中的出生后和成年小胶质细胞群体。通过流式细胞术分析，在基础激活状态下，TNF 缺失型和野生型小鼠之间没有发现差异。尽管 TNF 缺失型小胶质细胞在受到脂多糖 (LPS) 刺激时表现出典型的形态，细胞质中出现空泡，但 TNF 缺失型小胶质细胞对大肠杆菌颗粒的体外吞噬反应是正常的，没有迹象表明 caspase 3 激活或凋亡增强。功能上，发现 LPS 刺激的 TNF 缺失型小胶质细胞条件培养基中 IL-10、IL-6、IL-1β、IL-12 和 CXCL1 的水平明显低于野生型小胶质细胞，并且对神经分化的多巴胺能 (DA) MN9D 细胞没有细胞毒性作用。相比之下，用 TNF 抑制剂孵育野生型小胶质细胞可选择性地耗尽可溶性 TNF 的水平，并对 MN9D 细胞产生细胞毒性。为了区分 LPS 激活的 TNF 缺失型小胶质细胞降低细胞毒性是否归因于缺乏自分泌 TNF 信号，我们使用在与 MN9D 细胞共培养中缺乏一种或两种 TNF 受体 (TNFR1 和 TNFR2) 的原代小胶质细胞进行实验，发现这两种受体都不需要引发 LPS 诱导的 TNF 产生和对 DA 细胞的细胞毒性。
