Brain microvascular pericytes are immunoactive in culture: cytokine, chemokine, nitric oxide, and LRP-1 expression in response to lipopolysaccharide

Abstract

BackgroundBrain microvascular pericytes are important constituents of the neurovascular unit. These cells are physically the closest cells to the microvascular endothelial cells in brain capillaries. They significantly contribute to the induction and maintenance of the barrier functions of the blood-brain barrier. However, very little is known about their immune activities or their roles in neuroinflammation. Here, we focused on the immunological profile of brain pericytes in culture in the quiescent and immune-challenged state by studying their production of immune mediators such as nitric oxide (NO), cytokines, and chemokines. We also examined the effects of immune challenge on pericyte expression of low density lipoprotein receptor-related protein-1 (LRP-1), a protein involved in the processing of amyloid precursor protein and the brain-to-blood efflux of amyloid-β peptide.MethodsSupernatants were collected from primary cultures of mouse brain pericytes. Release of nitric oxide (NO) was measured by the Griess reaction and the level of S-nitrosylation of pericyte proteins measured with a modified "biotin-switch" method. Specific mitogen-activated protein kinase (MAPK) pathway inhibitors were used to determine involvement of these pathways on NO production. Cytokines and chemokines were analyzed by multianalyte technology. The expression of both subunits of LRP-1 was analyzed by western blot.ResultsLipopolysaccharide (LPS) induced release of NO by pericytes in a dose-dependent manner that was mediated through MAPK pathways. Nitrative stress resulted in S-nitrosylation of cellular proteins. Eighteen of twenty-three cytokines measured were released constitutively by pericytes or with stimulation by LPS, including interleukin (IL)-12, IL-13, IL-9, IL-10, granulocyte-colony stimulating factor, granulocyte macrophage-colony stimulating factor, eotaxin, chemokine (C-C motif) ligand (CCL)-3, and CCL-4. Pericyte expressions of both subunits of LRP-1 were upregulated by LPS.ConclusionsOur results show that cultured mouse brain microvascular pericytes secrete cytokines, chemokines, and nitric oxide and respond to the innate immune system stimulator LPS. These immune properties of pericytes are likely important in their communication within the neurovascular unit and provide a mechanism by which they participate in neuroinflammatory processes in brain infections and neurodegenerative diseases.

Lipopolysaccharide Activates Toll-like Receptor 4 (TLR4)-mediated NF-κB Signaling Pathway and Proinflammatory Response in Human Pericytes

“…At the protein level, we have corroborated the secretion of CXCL1, CXCL2, CXCL3, CCL2, IL-6, and IL-8 by LPS and HMGB1-stimulated pericytes. Interestingly, when compared with previous results using LPS-treated mouse brain pericytes (16), the profile of cytokines and chemokines released by HBVPs is considerably different. Stimulation of mouse brain pericytes with LPS at the same concentration used in this study resulted in significant release of IL-1␣, TNF-␣, IL-3, IL-10, IL-12 (p70), IL-13, G-CSF, GM-CSF, and CCL5, which were undetectable in the conditioned medium of stimulated HBVPs.…”

“…In a series of pioneering works by Edelman et al (13)(14)(15), the authors demonstrated the expression of TLR4 by Western blotting in rat lung pericytes (13) as well as increased vessel permeability (14) and production of IL-1␤ (15) in LPS-treated pericytes, suggesting for the first time an active role of pericytes in the inflammatory cascade. More recently, the release of nitric oxide and several cytokines and chemokines by mouse brain vascular pericytes in response to LPS has been reported (16). Other interesting works studied the expression of immunologically relevant cell surface proteins by human pericytes after stimulation with TNF-␣ or IFN-␥ (17, 18) but not with LPS.…”

Role of nucleotide‐binding oligomerization domain 1 (NOD1) in pericyte‐mediated vascular inflammation

“…In contrast to the numerous studies assessing the crosstalk among brain endothelial cells, astrocytes and microglia during inflammation, little is known about the role played by the brain microvascular pericyte. In fact, the response of brain pericytes to pro‐inflammatory stimuli has only been addressed in the last few years 2, 3, 4. In a pioneering work, Kovac et al .…”

Blood–Brain Barrier Dysfunction as a Cause and Consequence of Alzheimer's Disease

“…The BBB endothelial cells respond to inflammatory stimuli such as cytokines, 186 LPS, 187,188 Ab, [189][190][191] and a truncated tau fragment 192 by activating signaling pathways that result in the upregulation of proinflammatory second messengers and reactive oxygen/ nitrogen species, ultimately causing BBB disruption and paracrine activation of surrounding cells that are responsive to such stimuli, such as astrocytes 193 and pericytes 153,194 of the NVU. Inflammation impairs systemic clearance of Ab by the liver and kidneys, as well as bulk flow of CSF.…”

“…123,151 Whether pericytes have a similar Ab clearing function is presently unknown, but they do express LRP-1. 152,153 The Astrocytes Astrocytes are the most abundant cell type in the brain, and have important functions in the BBB. Their endfeet surround the endothelium of capillaries, arterioles, and venules.…”