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Talking to the Brain at the Blood-Brain Barrier through Inflammation-Induced Prostaglandin E2
Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Center for Diagnostics, Department of Clinical Immunology and Transfusion Medicine.
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The immune-to-brain signaling is a critical survival factor when the body is confronted by pathogens, and in particular by microorganisms. During infections, the ability of the immune system to engage the central nervous system (CNS) in the management of the inflammatory response is just as important as its ability to mount a specific immune response against the pathogen, since the CNS can provide a systemic negative feed-back to the immune activation by release of stress hormones and also can prioritize the usage of the energy resources by the vital organs. Prostaglandin E2 (PGE2) and proinflammatory cytokines were among the first mediators to be identified to participate in the immuneto-brain signaling, a process that is clinically recognized by the development of manifestations of common illness such as fever, anorexia, decreased social interactions, lethargy, sleepiness, and hyperalgesia.

In this thesis the contribution of PGE2 to the immune-to-brain signaling was further characterized at the blood-brain-barrier (BBB) and in the anterior preoptic area (POA) of the hypothalamus (i.e. the thermoregulatory region or, in sickness, the fever generating region).

BBB is the major interface region between peripheral circulating cytokines and the neuronal parenchyma and a critical source of PGE2. Using chimeric mice lacking the inducible enzyme for PGE2 synthesis, microsomal PGE synthase-1 (mPGES-1), in either hematopoietic or non-hematopoietic cells, we demonstrate in paper I that brain endothelial cells are the critical source of PGE2 in BBB during peripheral inflammation. For the demonstration of the mPGES-1 expression in the BBB cells we developed in paper I a method for enzymatic dissociation of these cells, followed by fluorescence activated cell sorting (FACS). Using the same method, we show in paper II that the BBB response to immune stimuli is towards an increased production of PGE2 in endothelial cells and an increased sensitivity of these cells for pro-inflammatory cytokines. These changes are supported by decreased PGE2 degradation and decreased synthesis of other prostanoids in perivascular macrophages, which hence respond in concordance with the endothelial cells in enhancing PGE2 signaling.

Once released in the neuronal tissue, PGE2 has been shown to be critical for the fever response by acting on the type 3 PGE2 receptors (EP3) within POA. By laser capture microdissection (LCM) we extracted the EP3 receptor expressing region in POA, defined by in situ hybridization histochemistry, from mouse brain sections. We demonstrate in paper III that the predominant subtypes of the EP3 receptor in POA are EP3α and EP3γ. In paper IV we further analyze the effect of PGE2 on the LCM dissected EP-rich POA using gene expression microarrays. We demonstrate that PGE2 has a limited effect on the gene expression changes within POA, suggesting that the neuronal activity is modulated by PGE2 in a transcription-independent manner and that the profound gene expression changes that are seen in the CNS during inflammation are accordingly PGE2-independent.

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2015. , 99 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 1435
National Category
Cell and Molecular Biology Cell Biology
Identifiers
URN: urn:nbn:se:liu:diva-114378DOI: 10.3384/diss.diva-114378ISBN: 978-91-7519-155-3 (print)OAI: oai:DiVA.org:liu-114378DiVA: diva2:789634
Public defence
2015-03-20, Berzeliussalen, Campus US, Linköpings universitet, Linköping, 13:15 (Swedish)
Opponent
Supervisors
Available from: 2015-02-19 Created: 2015-02-19 Last updated: 2016-05-04Bibliographically approved
List of papers
1. Lipopolysaccharide-Induced Fever Depends on Prostaglandin E2 Production Specifically in Brain Endothelial Cells
Open this publication in new window or tab >>Lipopolysaccharide-Induced Fever Depends on Prostaglandin E2 Production Specifically in Brain Endothelial Cells
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2012 (English)In: Endocrinology, ISSN 0013-7227, E-ISSN 1945-7170, Vol. 153, no 10, 4849-4861 p.Article in journal (Refereed) Published
Abstract [en]

Immune-induced prostaglandin E2 (PGE2) synthesis is critical for fever and other centrally elicited disease symptoms. The production of PGE2 depends on cyclooxygenase-2 and microsomal prostaglandin E synthase-1 (mPGES-1), but the identity of the cells involved has been a matter of controversy. We generated mice expressing mPGES-1 either in cells of hematopoietic or nonhematopoietic origin. Mice lacking mPGES-1 in hematopoietic cells displayed an intact febrile response to lipopolysaccharide, associated with elevated levels of PGE2 in the cerebrospinal fluid. In contrast, mice that expressed mPGES-1 only in hematopoietic cells, although displaying elevated PGE2 levels in plasma but not in the cerebrospinal fluid, showed no febrile response to lipopolysaccharide, thus pointing to the critical role of brain-derived PGE2 for fever. Immunohistochemical stainings showed that induced cyclooxygenase-2 expression in the brain exclusively occurred in endothelial cells, and quantitative PCR analysis on brain cells isolated by flow cytometry demonstrated that mPGES-1 is induced in endothelial cells and not in vascular wall macrophages. Similar analysis on liver cells showed induced expression in macrophages and not in endothelial cells, pointing at the distinct role for brain endothelial cells in PGE2 synthesis. These results identify the brain endothelial cells as the PGE2-producing cells critical for immune-induced fever.

Place, publisher, year, edition, pages
Endocrine Society, 2012
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-84885 (URN)10.1210/en.2012-1375 (DOI)000309210200027 ()
Note

Funding Agencies|Swedish Research Council|7879|Swedish Cancer Foundation|100533|Swedish Brain Foundation||Gustav V:s 80-ars Fond||

Available from: 2012-11-01 Created: 2012-10-26 Last updated: 2017-12-07
2. Immune challenge by intraperitoneal administration of lipopolysaccharide directs gene expression in distinct blood-brain barrier cells toward enhanced prostaglandin E2 signaling
Open this publication in new window or tab >>Immune challenge by intraperitoneal administration of lipopolysaccharide directs gene expression in distinct blood-brain barrier cells toward enhanced prostaglandin E2 signaling
2015 (English)In: Brain, behavior, and immunity, ISSN 0889-1591, E-ISSN 1090-2139, Vol. 48, 31-41 p.Article in journal (Refereed) Published
Abstract [en]

The cells constituting the blood-brain barrier are critical for the transduction of peripheral immune signals to the brain, but hitherto no comprehensive analysis of the signaling events that occur in these cells in response to a peripheral inflammatory stimulus has been performed. Here, we examined the inflammatory transcriptome in blood-brain barrier cells, including endothelial cells, pericytes, and perivascular macrophages, which were isolated by fluorescent-activated cell sorting, from non-immune-challenged mice and from mice stimulated by bacterial wall lipopolysaccharide. We show that endothelial cells and perivascular macrophages display distinct transcription profiles for inflammatory signaling and respond in distinct and often opposing ways to the immune stimulus. Thus, endothelial cells show induced PGE2 synthesis and transport with attenuation of PGE2 catabolism, increased expression of cytokine receptors and down-stream signaling molecules, and downregulation of adhesion molecules. In contrast, perivascular macrophages show downregulation of the synthesis of prostanoids other than PGE2 and of prostaglandin catabolism, but upregulation of interleukin-6 synthesis. Pericytes were largely unresponsive to the immune stimulation, with the exception of downregulation of proteins involved in pericyte-endothelial cell communication. While the endothelial cells account for most of the immune-induced gene expression changes in the blood-brain barrier, the response of the endothelial cells occurs in a concerted manner with that of the perivascular cells to elevate intracerebral levels of PGE2, hence emphasizing the critical role of PGE2 in immune-induced signal transduction across the blood-brain barrier.

Place, publisher, year, edition, pages
Academic Press, 2015
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:liu:diva-114377 (URN)10.1016/j.bbi.2015.02.003 (DOI)000358460700005 ()25678162 (PubMedID)
Note

This work was supported by Grants from the Swedish Research Council (07879 to AB, and 22241 to HQ), the Swedish Cancer Foundation (13 0295 to AB), the Swedish Brain Foundation (to AB), the County Council of Ostergotland (to AMV), and Knut och Alice Wallenberg Foundation (WIRM to HQ).

Available from: 2015-02-19 Created: 2015-02-19 Last updated: 2017-12-04Bibliographically approved
3. Expression of PGE2 EP3 receptor subtypes in the mouse preoptic region
Open this publication in new window or tab >>Expression of PGE2 EP3 receptor subtypes in the mouse preoptic region
2007 (English)In: Neuroscience Letters, ISSN 0304-3940, Vol. 423, no 3, 179-183 p.Article in journal (Refereed) Published
Abstract [en]

Inflammatory-induced fever is dependent on prostaglandin E2 (PGE2) binding to its EP3 receptor in the thermoregulatory region of the hypothalamus, but it is not known which EP3 receptor isoform(s) that is/are involved. We identified the EP3 receptor expression in the mouse preoptic region by in situ hybridization and isolated the corresponding area by laser capture microdissection. Real-time RT-PCR analysis of microdissected tissue revealed a predominant expression of the EP3α isoform, but there was also considerable expression of EP3γ, corresponding to approximately 15% of total EP3 receptor expression, whereas EP3β was sparsely expressed. This distribution was not changed by immune challenge induced by peripheral administration of LPS, indicating that EP3 receptor splicing and distribution is not activity dependent. Considering that EP3α and EP3γ are associated with inhibitory and stimulatory G-proteins, respectively, the present data demonstrate that the PGE2 response of the target neurons is intricately regulated. © 2007 Elsevier Ireland Ltd. All rights reserved.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-39503 (URN)10.1016/j.neulet.2007.06.048 (DOI)49060 (Local ID)49060 (Archive number)49060 (OAI)
Available from: 2009-10-10 Created: 2009-10-10 Last updated: 2016-05-04
4. Minor Changes in Gene Expression in the Mouse Preoptic Hypothalamic Region by Inflammation-Induced Prostaglandin E2
Open this publication in new window or tab >>Minor Changes in Gene Expression in the Mouse Preoptic Hypothalamic Region by Inflammation-Induced Prostaglandin E2
2013 (English)In: Journal of neuroendocrinology (Print), ISSN 0953-8194, E-ISSN 1365-2826, Vol. 25, no 7, 635-643 p.Article in journal (Refereed) Published
Abstract [en]

We investigated to what extent inflammation-induced prostaglandin E2 (PGE2) regulates gene expression in the central nervous system. Wild-type mice and mice with deletion of the gene encoding microsomal prostaglandin E synthase-1 (mPGES-1), which cannot produce inflammation-induced PGE2, were subjected to peripheral injection of bacterial wall lipopolysaccharide (LPS) and killed after 5 h. The median and medial preoptic nuclei, which are rich in prostaglandin E receptors, were isolated by laser capture microdissection (LCM), and subjected to whole genome microarray analysis. Although the immune stimulus induced robust transcriptional changes in the brain, as seen by a quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) on selected genes, only small PGE2-dependent gene expression changes were observed in the gene array analysis and, for only two genes, a pronounced differential expression between LPS-treated wild-type and mPGES-1 knockout mice could be verified by qRT-PCR. These were Hspa1a and Hspa1b, encoding heat shock proteins, which showed a two- to three-fold higher expression in wild-type mice than in knockout mice after immune challenge. However, the induced expression of these genes was found to be secondary to increased body temperature because they were induced also by cage exchange stress, which did not elicit PGE2 synthesis, and thus were not induced per se by PGE2-elicited transcriptional events. Our findings suggest that inflammation-induced PGE2 has little effect on gene expression in the preoptic region, and that centrally elicited disease symptoms, although PGE2-dependent, occur as a result of regulation of neuronal excitability that is a consequence of intracellular, transcriptional-independent signalling cascades. Our findings also imply that the profound changes in gene expression in the brain that are elicited by peripheral inflammation occur independently of PGE2 via a yet unidentified mechanism.

Place, publisher, year, edition, pages
Wiley-Blackwell, 2013
Keyword
microsomal prostaglandin E synthase-1; prostaglandin E2; fever; preoptic region; laser capture microdissection; whole genome microarray; heat-shock proteins
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-96460 (URN)10.1111/jne.12044 (DOI)000320402900005 ()
Available from: 2013-08-23 Created: 2013-08-20 Last updated: 2017-12-06Bibliographically approved

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Vasilache, Ana Maria

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