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Isolation microglia

Lisa K Gouwens, Mudar S Ismail, Victoria A Rogers, Nathan T Zeller, Evan C Garrad, Fatima S Amtashar, Nyasha J Makoni, David C Osborn, Michael R Nichols
Microvesicles (MVs) and exosomes comprise a class of cell-secreted particles termed extracellular vesicles (EVs). These cargo-holding vesicles mediate cell-to-cell communication and have recently been implicated in neurodegenerative diseases such as Alzheimer's disease (AD). The two types of EVs are distinguished by the mechanism of cell release and their size, with the smaller exosomes and the larger MVs ranging from 30 to 100 nm and 100 nm to 1 μm in diameter, respectively. MV numbers are increased in AD and appear to interact with amyloid-β peptide (Aβ), the primary protein component of the neuritic plaques in the AD brain...
March 15, 2018: ACS Chemical Neuroscience
Laura K Fonken, Matthew G Frank, Andrew D Gaudet, Heather M D'Angelo, Rachel A Daut, Emma C Hampson, Monica T Ayala, Linda R Watkins, Steven F Maier
Exposure to stressors can enhance neuroinflammatory responses, and both stress and neuroinflammation are predisposing factors in the development of psychiatric disorders. Females suffer disproportionately more from several psychiatric disorders, yet stress-induced changes in neuroinflammation have primarily been studied in males. Here we tested whether exposure to inescapable tail shock sensitizes or 'primes' neuroinflammatory responses in male and female rats. At 24 h post-stress, male and female rats exposed to a peripheral immune challenge enhanced neuroinflammatory responses and exacerbated anxiety- and depressive-like behaviors...
March 7, 2018: Brain, Behavior, and Immunity
Kaitryn E Ronning, Gabriel Peinado Allina, Eric B Miller, Robert J Zawadzki, Edward N Pugh, Rolf Hermann, Marie E Burns
Rods and cones mediate visual perception over 9 log units of light intensities, with both photoreceptor types contributing to a middle 3-log unit range that comprises most night-time conditions. Rod function in this mesopic range has been difficult to isolate and study in vivo because of the paucity of mutants that abolish cone signaling without causing photoreceptor degeneration. Here we describe a novel Gnat2 knockout mouse line (Gnat2-/- ) ideal for dissecting rod and cone function. In this line, loss of Gnat2 expression abolished cone phototransduction, yet there was no loss of cones, disruption of the photoreceptor mosaic, nor change in general retinal morphology up to at least 9 months of age...
March 5, 2018: Experimental Eye Research
Fa-Cai Li, Qing Liu, Hany M Elsheikha, Wen-Bin Yang, Jun-Ling Hou, Xing-Quan Zhu
Toxoplasma gondii deploys many effector proteins in order to hijack and manipulate host cell signaling pathways, allowing parasite colonization, subversion of immune responses, and disease progression. T. gondii effector protein 14-3-3 (Tg14-3-3) promotes parasite dissemination inside the body, by enhancing the migratory ability of infected microglia and dendritic cells. Understanding both the mechanism of action and the host targets of Tg14-3-3 effector is important because of their importance to the parasite's virulence...
March 3, 2018: Parasitology Research
Mark Ronald Mizee, Marlijn van der Poel, Inge Huitinga
In order to translate the findings obtained from postmortem brain tissue samples to functional biologic mechanisms of central nervous system disease, it will be necessary to understand how these findings affect the different cell populations in the brain. The acute isolation and analysis of pure glial cell populations are common practice in animal models for neurologic diseases, but are not yet regularly applied to human postmortem brain material. The development of novel cell isolation techniques and methods for transcriptomic and proteomic analysis have made it possible to isolate and phenotype primary human cell populations from the central nervous system...
2018: Handbook of Clinical Neurology
Tanja Zöller, Abdelraheim Attaai, Phani Sankar Potru, Tamara Ruß, Björn Spittau
Microglia are the resident immune cells of the central nervous system (CNS) and participate in physiological and pathological processes. Their unique developmental nature suggests age-dependent structural and functional impairments that might contribute to neurodegenerative diseases. In the present study, we addressed the age-dependent changes in cortical microglia gene expression patterns and the expression of M1- and M2-like activation markers. Iba1 immunohistochemistry, isolation of cortical microglia followed by fluorescence-activated cell sorting and RNA isolation to analyze transcriptional changes in aged cortical microglia was performed...
March 1, 2018: International Journal of Molecular Sciences
Yong-Ying Jiang, Lin Li, Wei-Xing Shen, Yi-Hua Qiu, Yu-Ping Peng
Microglia are the main immune cells in the central nervous system. In the present study, the mechanism for acetylcholine (ACh) inhibiting microglial inflammatory response was investigated. Primary culture of microglia was isolated from cerebral cortex of Sprague-Dawley (SD) rats. Lipopolysaccharide (LPS) was used to activate the microglia to induce inflammatory response, and then the microglia were treated with ACh for 24 h. Protein expressions of several inflammatory factors, insulin-like growth factor 1 (IGF-1) and α7 nicotinic acetylcholine receptor (α7nAChR) were detected by Western blot...
February 25, 2018: Sheng Li Xue Bao: [Acta Physiologica Sinica]
Jonathan R Bumstead, Jasmine J Park, Isaac A Rosen, Andrew W Kraft, Patrick W Wright, Matthew D Reisman, Daniel C Côté, Joseph P Culver
Conventional two-photon microscopy (TPM) is capable of imaging neural dynamics with subcellular resolution, but it is limited to a field-of-view (FOV) diameter [Formula: see text]. Although there has been recent progress in extending the FOV in TPM, a principled design approach for developing large FOV TPM (LF-TPM) with off-the-shelf components has yet to be established. Therefore, we present a design strategy that depends on analyzing the optical invariant of commercially available objectives, relay lenses, mirror scanners, and emission collection systems in isolation...
April 2018: Neurophotonics
Guillaume Hoeffel, Florent Ginhoux
Tissue-resident macrophages have pivotal functions for tissue defense and homeostasis. Two main discoveries have changed our current understanding of macrophage development: Their embryonic origin and their ability to self-renew throughout the lifespan. It is now well accepted that most tissue-resident macrophages are long-lived cells derived from a transient hematopoietic wave of erythro-myeloid progenitors (EMPs) emerging in the yolk sac. At least two distinct pathways derived from EMPs have been implicated in macrophage development...
January 12, 2018: Cellular Immunology
Yi-Je Chen, Hai M Nguyen, Izumi Maezawa, Lee-Way Jin, Heike Wulff
Objective: Inhibitors of the voltage-gated K+ channel Kv1.3 are currently in development as immunomodulators for the treatment of autoimmune diseases. As Kv1.3 is also expressed on microglia and has been shown to be specifically up-regulated on "M1-like" microglia, we here tested the therapeutic hypothesis that the brain-penetrant small-molecule Kv1.3-inhibitor PAP-1 reduces secondary inflammatory damage after ischemia/reperfusion. Methods: We studied microglial Kv1...
February 2018: Annals of Clinical and Translational Neurology
Viljami Jokinen, Yulia Sidorova, Hanna Viisanen, Ilida Suleymanova, Henna Tiilikainen, Zhilin Li, Tuomas O Lilius, Kert Mätlik, Jenni E Anttila, Mikko Airavaara, Li Tian, Pekka V Rauhala, Eija A Kalso
Development of tolerance is a well known pharmacological characteristic of opioids and a major clinical problem. In addition to the known neuronal mechanisms of opioid tolerance, activation of glia has emerged as a potentially significant new mechanism. We studied activation of microglia and astrocytes in morphine tolerance and opioid-induced hyperalgesia in rats using immunohistochemistry, flow cytometry and RNA sequencing in spinal- and supraspinal regions. Chronic morphine treatment that induced tolerance and hyperalgesia also increased immunoreactivity of spinal microglia in the dorsal and ventral horns...
February 6, 2018: Neuroscience
Prasant Kumar Jena, Lili Sheng, Jacopo Di Lucente, Lee-Way Jin, Izumi Maezawa, Yu-Jui Yvonne Wan
The goal of this study was to identify the intrinsic links that explain the effect of a Western diet (WD) on cognitive dysfunction. Specific pathogen-free, wild-type mice were fed either a control diet (CD) or a high-fat, high-sucrose WD after weaning and were euthanized at 10 mo of age to study the pathways that affect cognitive health. The results showed that long-term WD intake reduced hippocampal synaptic plasticity and the level of brain-derived neurotrophic factor mRNA in the brain and isolated microglia...
January 10, 2018: FASEB Journal: Official Publication of the Federation of American Societies for Experimental Biology
Diego Mastroeni, Jennifer Nolz, Omar M Khdour, Shobana Sekar, Elaine Delvaux, Lori Cuyugan, Winnie S Liang, Sidney M Hecht, Paul D Coleman
INTRODUCTION: Our laboratories have demonstrated that accumulation of oligomeric amyloid β (OAβ) in neurons is an essential step leading to OAβ-mediated mitochondrial dysfunction. METHODS: Alzheimer's disease (AD) and matching control hippocampal neurons, astrocytes, and microglia were isolated by laser-captured microdissection from the same subjects, followed by whole-transcriptome sequencing. Complementary in vitro work was performed in OAβ-treated differentiated SH-SY5Y, followed by the use of a novel CoQ10 analogue for protection...
January 20, 2018: Alzheimer's & Dementia: the Journal of the Alzheimer's Association
Jing Wei, Wan-Hui Guo, Chen-Yu Cao, Rong-Wei Kou, Yuan-Zhen Xu, Marcin Górecki, Lorenzo Di Bari, Gennaro Pescitelli, Jin-Ming Gao
In a previous study, we reported ten new polyoxygenated cyathane diterpenoids, neocyathins A-J, and their anti-neuroinflammatory effects from the liquid culture of the medicinal Basidiomycete Cyathus africanus. In the present study, eight new highly polyoxygenated cyathane diterpenoids, named neocyathins K-R (1-8), were isolated from the solid culture of C. africanus cultivated on cooked rice, together with three known congeners (9-11). The structures and the absolute configurations of the new compounds were elucidated through comprehensive NMR and HRESIMS spectroscopic data, electronic circular dichroism (ECD) data, and chemical conversion...
February 1, 2018: Scientific Reports
Atsuko Katsumoto, Aline S Miranda, Oleg Butovsky, Antônio L Teixeira, Richard M Ransohoff, Bruce T Lamb
BACKGROUND: Traumatic brain injury (TBI) is a critical public health and socio-economic problem worldwide. A growing body of evidence supports the involvement of inflammatory events in TBI. It has been reported that resident microglia and infiltrating monocytes promote an inflammatory reaction that leads to neuronal death and eventually behavioral and cognitive impairment. Currently, there is no effective treatment for TBI and the development of new therapeutic strategies is a scientific goal of highest priority...
January 30, 2018: Journal of Neuroinflammation
Joanna A Phipps, Kirstan A Vessey, Alice Brandli, Nupur Nag, Mai X Tran, Andrew I Jobling, Erica L Fletcher
Purpose: This study explored whether the proangiogenic factor Angiotensin II (AngII) had a direct effect on the activation state of microglia via the Angiotensin type 1 receptor (AT1-R). Methods: Microglial dynamic activity was investigated in live retinal flatmounts from adult Cx3Cr1+/GFP mice under control, AngII (5 μM) or AngII (5 μM) + candesartan (0.227 μM) conditions. The effects of intravitreal administration of AngII (10 mM) were also investigated at 24 hours, with retinae processed for immunocytochemistry, flow cytometry, or inflammatory quantitative PCR arrays...
January 1, 2018: Investigative Ophthalmology & Visual Science
Julie E Simpson, Stephen B Wharton, Paul R Heath
Laser-capture microdissection (LCM) has revolutionized the isolation of defined regions and specific cell populations from human tissue. This approach used in combination with immunohistochemistry (immuno-LCM) has become a valuable method for isolating individual cell-types from a complex heterogeneous population. Here, we describe the detailed methodology required for the isolation of enriched populations of GFAP+ astrocytes, OSP+ oligodendrocytes, and CD68+ microglia from frozen post-mortem human central nervous system tissue using immuno-LCM...
2018: Methods in Molecular Biology
Seoyeon Bok, Young-Eun Kim, Youngsik Woo, Soeun Kim, Suk-Jo Kang, Yoontae Lee, Sang Ki Park, Irving L Weissman, G-One Ahn
Cells universally adapt to ischemic conditions by turning on a transcription factor hypoxia-inducible factor (HIF), in which its role is known to differ widely across many different types of cells. Given that microglia have been reported as an essential mediator of neuroinflammation in many brain diseases, we examined the role of HIF in microglia in the progression of an acute phase of ischemic stroke by challenging our novel strains of myeloid-specific Hif-1α or Hif-2α knockout (KO) mice created by Cre-loxP system via middle cerebral artery occlusion (MCAO)...
December 19, 2017: Oncotarget
Adriana-Natalia Murgoci, Dasa Cizkova, Petra Majerova, Eva Petrovova, Lubomir Medvecky, Isabelle Fournier, Michel Salzet
The function and integrity of nervous system require interactive exchanges among neurons and glial cells. Exosomes and other extracellular vesicles (EVs) are emerging as a key mediator of intercellular communication, capable to transfer nucleic acids, proteins and lipids, influencing numerous functional and pathological aspects of both donor and recipient cells. The immune response mediated by microglia derived exosomes are most prominently involved in the spread of neuroinflammation, neurodegenerative disorders and brain cancer...
January 12, 2018: Chemphyschem: a European Journal of Chemical Physics and Physical Chemistry
Xue-Tao Yan, Jing-Min Lu, Yu Wang, Xiao-Li Cheng, Xiang-Hu He, Wen-Zhong Zheng, Hu Chen, Yan-Lin Wang
LncRNAs are reported to participate in neuropathic pain development. LncRNA X-inactive specific transcript (XIST) is involved in the progression of various cancers. However, the role of XIST in neuropathic pain remains unclear. In our present study, we established a chronic constriction injury (CCI) rat model and XIST was found to be greatly upregulated both in the spinal cord tissues and in the isolated microglias of CCI rats. Inhibition of XIST inhibited neuropathic pain behaviors including mechanical and thermal hyperalgesia...
January 11, 2018: Journal of Cellular Physiology
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