Lipopolysaccharide-induced blood-brain barrier disruption: roles of cyclooxygenase, oxidative stress, neuroinflammation, and elements of the neurovascular unit

Background: Disruption of the blood-brain barrier (BBB) occurs in many diseases and is often mediated by inflammatory and neuroimmune mechanisms. Inflammation is well established as a cause of BBB disruption, but many mechanistic questions remain. Methods: We used lipopolysaccharide (LPS) to induce inflammation and BBB disruption in mice. BBB disruption was measured using 14C-sucrose and radioactively labeled albumin. Brain cytokine responses were measured using multiplex technology and dependence on cyclooxygenase (COX) and oxidative stress determined by treatments with indomethacin and N-acetylcysteine. Astrocyte and microglia/macrophage responses were measured using brain immunohistochemistry. In vitro studies used Transwell cultures of primary brain endothelial cells co- or tri-cultured with astrocytes and pericytes to measure effects of LPS on transendothelial electrical resistance (TEER), cellular distribution of tight junction proteins, and permeability to 14C-sucrose and radioactive albumin. Results: In comparison to LPS-induced weight loss, the BBB was relatively resistant to LPS-induced disruption. Disruption occurred only with the highest dose of LPS and was most evident in the frontal cortex, thalamus, pons-medulla, and cerebellum with no disruption in the hypothalamus. The in vitro and in vivo patterns of LPS-induced disruption as measured with 14C-sucrose, radioactive albumin, and TEER suggested involvement of both paracellular and transcytotic pathways. Disruption as measured with albumin and 14C-sucrose, but not TEER, was blocked by indomethacin. N-acetylcysteine did not affect disruption. In vivo, the measures of neuroinflammation induced by LPS were mainly not reversed by indomethacin. In vitro, the effects on LPS and indomethacin were not altered when brain endothelial cells (BECs) were cultured with astrocytes or pericytes. Conclusions: The BBB is relatively resistant to LPS-induced disruption with some brain regions more vulnerable than others. LPS-induced disruption appears is to be dependent on COX but not on oxidative stress. Based on in vivo and in vitro measures of neuroinflammation, it appears that astrocytes, microglia/macrophages, and pericytes play little role in the LPS-mediated disruption of the BBB

Brain Microvessels from Dementia and No Dementia Subjects: A Model for Micro to Macro Protein Analysis?

The brain microvessel (MV) is vasculature \u3c100um, which includes capillaries and the adjacent portions of venules and arterioles. It is a key component of the blood brain barrier (BBB), which regulates nutrient delivery, waste removal, and movement of substances from the circulation in the brain parenchyma. Studies in this system have focused primarily on changes in the presence of diseases, such as dementia/Alzheimer\u27s Disease (AD), ischemia and traumatic injuries. Dementia is a disorder characterized by neurodegeneration involving one or more cognitive domains (Götz, 2008; APA, 2013; Taylor, 2018). The most common form of dementia that affects older adults is AD (Ballard, 2011; Soria Lopez, 2019). Though the precise sequence of events is debated, the neuropathologic factors of AD are well studied. Previous studies have indicated increased accumulation of amyloid-beta plaques, neuritic plaques, neurofibrillary tangles, and phospho-tau in AD samples (Wenk, 2003; Drummond, 2016; DeTure & Dickson, 2019; Fyfe 2022). However, less is known about the microvascular changes with AD (Pardridge, 2020). There have been some studies showing increased MV heterogeneity and tortuosity as well as decreased MV number and density in the context of aging (Sonntag, Lynch et al. 1997; Li et al., 2018, Steinman et al., 2021). In addition, other studies have indicated similar overall density of MVs in AD and no AD brains but decreased tight junction proteins, including claudin-5, occludin, and Zo-1 in dementia (Yamazaki et al., 2019; Greene et al., 2019; Lochhead et al., 2020). We wished to examine brain MVs isolated from the superior parietal lobe of male and female subjects with and without dementia to assess whether the MV acts as a good model for evaluation of functional protein changes at multiple levels of analysis using proteomics/mass spectrometry, western blot (WB), immunofluorescence (IF), and immunohistochemistry (IHC). Because the MV samples were not transcriptionally active, acquisition of RNA was omitted among the possible techniques (Lee YK et al., 2019)