Microglia drive diurnal variation in susceptibility to inflammatory blood-brain barrier breakdown

The blood brain barrier (BBB) is critical for maintaining brain homeostasis but is susceptible to inflammatory breakdown. Permeability of the BBB to polar molecules shows circadian variation due to rhythmic transporter expression, while basal permeability to polar molecules in non-rhythmic. Whether daily timing influences BBB permeability in response to inflammation is unknown. Here, we induced systemic inflammation through repeated lipopolysaccharide (LPS) injections either in the morning (ZT1) or evening (ZT13) under standard lighting conditions, then examined BBB permeability to a polar molecule. We observed clear diurnal variation in inflammatory BBB permeability, with a striking increase in BBB breakdown specifically following evening LPS injection. Evening LPS led to persisting glia activation and inflammation in the brain that was not observed in the periphery. The exaggerated evening neuroinflammation and BBB disruption were suppressed by microglial depletion, were associated with enhanced expression of the Nos2 gene, and could be prevented in vivo by treatment with an iNOS inhibitor. Our data show that diurnal rhythms in microglial inflammatory responses to LPS drive daily variability in BBB breakdown, and reveals time-of-day as a key regulator of inflammatory BBB disruption. To account for the effect of time-of-day on neuroinflammatory responses, we conducted bulk RNAseq on cortex samples from mice treated with i.p. LPS (or PBS control) at ZT1 and ZT13

血脑屏障（blood brain barrier, BBB）是维持脑内稳态的关键结构，但易受炎症反应破坏。由于转运蛋白的节律性表达，血脑屏障对极性分子的通透性呈现昼夜节律变化，而非节律状态下其对极性分子的基础通透性则无此类节律波动。目前尚不清楚每日时间节点是否会影响炎症状态下血脑屏障的通透性。
本研究中，我们在标准光照条件下，于清晨（ZT1）或傍晚（ZT13）通过反复注射脂多糖（lipopolysaccharide, LPS）构建全身炎症模型，随后检测血脑屏障对极性分子的通透性。我们观察到炎症状态下血脑屏障通透性存在显著的昼夜节律差异，尤其在傍晚注射LPS后，血脑屏障的破坏程度显著升高。傍晚注射LPS会引发大脑中持续的胶质细胞激活与炎症反应，而这一现象在外周组织中并未出现。傍晚时加剧的神经炎症与血脑屏障破坏可通过小胶质细胞清除得到抑制，该过程与Nos2基因的表达上调相关，且可通过体内给予诱导型一氧化氮合酶（inducible nitric oxide synthase, iNOS）抑制剂加以阻断。本研究数据表明，小胶质细胞对LPS的炎症反应存在昼夜节律，这一节律驱动了血脑屏障破坏的每日差异，同时揭示了每日时间节点是调控炎症性血脑屏障破坏的关键因素。为阐明每日时间节点对神经炎症反应的调控作用，我们对分别于ZT1和ZT13接受腹腔注射LPS（或磷酸盐缓冲液PBS对照）的小鼠皮层样本进行了批量RNA测序（bulk RNAseq）。