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Browsing by Subject "microglia"

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  • Pasculli, Maria Samuela (2024)
    The S209F variant of the Abelson Interactor family member 3 (ABI3) gene has emerged as a risk factor for late-onset Alzheimer’s Disease (LOAD). The ABI3 protein is functionally related to the WAVE Regulatory Complex (WRC) participating in the control of cytoskeletal processes favoring either filopodia for chemotaxis or pseudopodia for phagocytosis. The S209F coding variant is thought to impair phosphorylation of the ABI3 protein leading to dysfunctional association with WRC. In the brain, the ABI3 gene is mainly expressed by microglia, macrophages representing the resident immune cells of the brain. Despite some research about the variant based on rodent models and reporting sometimes contrasting results, the role of the ABI3 S209F variant in AD remains poorly understood. Here, human-induced pluripotent stem cells (h-iPSCs) reprogrammed from fibroblasts of controls and variant carriers are sequenced to ensure retention of the original phenotype upon reprogramming. H-iPSCs are differentiated into microglia (iPSC-derived microglia, iMGL) following an established protocol. Morphological changes and microglia-specific gene expression partially show that iMGL between days 31 and 38 of differentiation in vitro can be considered mature. To assess the functional properties of microglia, cytokines/chemokines production, cathepsin gene expression, lysosomal activity, and Apolipoprotein E (ApoE) protein levels are measured. It is found that S209F microglia downregulate CCL5/RANTES and upregulate cathepsins B and L (CTSB and CTSL) upon LPS+IFNg stimulation which may lead to motility, migratory and endo-lysosomal dysfunctions. Lysosomal activity is found to positively correlate with CD163, but not with either CTSB or CTSL expression. ApoE protein levels show an upregulation trend in S209F microglia which may indicate modifications in lipid metabolism. Metabolic assessment based on mitochondrial respiration and glycolysis does not show any difference between S209F and control microglia, but ABI3 knock-out (KO) shows glycolysis dysfunctions. Overall, this study offers some hints into the mechanisms that make the ABI3 S209F variant a risk factor for AD pointing at the need to investigate microglia motility and migration focusing on pathologically relevant protein aggregates and their clearance and with particular attention to phagocytosis and endo-lysosomal pathway.
  • Mäkinen, Hilla (2023)
    Morphological features are considered as markers of microglial functionality, and they show regional heterogeneity in the brain. Recently the sleep-wake cycle was shown to affect microglial morphology in mice and correlate with cortical sleep slow wave activity (SWA). Microglial sizes and ramification increased during the dark period and decreased during the light period in cerebral areas associated with SWA, suggesting that neuronal activation could be affecting microglial morphology through SWA. I studied microglia in the hindbrain areas with and without functional connection to SWA to further investigate the association between SWA and alterations in morphology, and to investigate if there are differences in microglial morphology and their diurnal alterations in brain regions other than those commonly investigated. I examined three hindbrain areas (cerebellar cortex (CC), deep cerebellar nucleus (DCN) and medial vestibular nucleus (MVN)) and somatosensory cortex (SC) of mice (n=15) at two timepoints: 6 hours after the light onset (high SWA) and offset (low SWA). My aims were to answer if there are morphological differences in microglia between 1) the four brain areas at both timepoints and 2) between the two timepoints in each brain area. My hypotheses were that CC and DCN which have functional connections to cortical SWA, would show similar diurnal morphology alterations as demonstrated in the cerebral areas, and MVN that has no known cortical SWA connection, would lack significant alterations. As microglia are heterogenous throughout brain, I expected microglia to differ between different brain areas, especially the hindbrain and the SC. I found that microglial morphologies significantly differed between the hindbrain and the cortex, while the hindbrain areas were more similar in morphology. Moreover, the brain areas demonstrated diurnal morphology alterations of microglia with varying extent: CC and DCN microglial morphology did not correlate with SWA as clearly as SC did, and interestingly, morphological features of MVN microglia showed a pattern opposite to other areas, microglia being larger during the light period than the dark period. These results highlight the importance of the diurnal time to microglial morphology and the heterogeneity of microglia between different brain regions.
  • Pihl, Enni-Eveliina (2023)
    Microglia, the resident macrophage-like glial cells of the central nervous system (CNS), form the first line of defense against pathogens in the brain, and regulate both innate and adaptive immunity. Any abnormalities in their microenvironment lead to microglial activation, characterized by alterations in their gene expression, morphology, and functional behavior. Once activated, microglia respond to CNS injury and inflammation by, e.g., migrating to the site of damage, releasing pro-inflammatory cytokines, as well as phagocyting cell debris and pathogens. Prolonged activation of microglia expressing pro-inflammatory phenotypes can lead to exacerbated CNS damage. Hence, limiting CNS inflammation by stimulating microglial polarization towards their pro-resolving phenotypes would be of great clinical relevance. The research of our laboratory focuses on CNS injury and repair, as well as finding novel therapies for ischemic stroke. Specialized pro-resolving mediators (SPMs) derived from essential fatty acids have been proposed to offer a potential therapeutic approach for ischemic stroke via promoting resolution of post-stroke inflammation. Previous studies have revealed the ability of SPMs to induce a transformation of macrophages, the immune cells strongly resembling microglia, towards their anti-inflammatory phenotypes. The aim of this study was therefore to assess whether SPMs have similar effects on BV2 microglia, specifically on their lipopolysaccharide (LPS)-induced production of pro-inflammatory cytokines, tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6). In addition to assessing the cytokine levels, our aim was to determine the optimal conditions for studying the effects of SPMs on microglial migration. In the present study, the levels of TNF-α and IL-6 were determined by specific ELISAs, and the transwell assay was used to measure microglial migration. Resolvins E1 (RvE1) and D1 (RvD1), as well as protectin D1 (PD1) and 15-epimer of lipoxin A4 (15-epi-LXA4) were all associated with decreased levels of TNF-α and IL-6, with RvE1 having the most potential as a resolving agent. In addition, we observed that serum starvation notably decreases the release of IL-6 and affects microglial migration. Overall, our results support the idea that SPMs could provide a novel therapeutic strategy for stroke therapy as they contribute to the resolution of CNS inflammation.
  • Lahtinen, Lilja (2022)
    Microglia, the resident immune cells of the central nervous system, react to inflammatory stimuli in the brain in a variety of ways. These include migrating to the site of damage and releasing pro- and anti-inflammatory factors. Previous research indicates that these microglial functions require extensive intracellular calcium signaling. Microglial overactivation can exacerbate neuronal damage, especially in cases of chronic inflammation. The ability to modulate the microglial response to damage would therefore be of great clinical relevance. The endoplasmic reticulum (ER) acts as the cell’s main calcium store and regulates cellular calcium levels primarily through the activity of ryanodine receptors (RYR), inositol-triphosphate receptors (IP3R), and the sarco-endoplasmic reticulum calcium ATPase (SERCA) pump. Calcium depletion from the ER is associated with cellular stress and microglial reactivity and therefore the ER may be an important target for modulating the microglial reactive response. The aim of this study is to show whether ER calcium depletion in a microglial cell line causes changes in protein expression, cellular infiltration, and the release of key pro-inflammatory factors. Drugs that block the pumping of calcium from the cytosol via the SERCA pump, such as thapsigargin, effectively induce a state of calcium depletion in the ER. In the present study, treatment with the SERCA pump inhibitor thapsigargin was found to increase SERCA2 expression in BV2, but not SV40, microglial cell lines. Treatment of microglia with thapsigargin was associated with large increases in the release of pro-inflammatory factors IL-6 and TNF-alpha but had no effect on microglial migration.
  • Vasques Ojeda, Ariel Olivia (2024)
    Faculty: Faculty of Biological and Environmental Sciences Degree programme: Master’s Programme in Neuroscience Study track: Neuroscience track Author: Ariel Olivia Vasques Ojeda Title: The effects of sleep disruption on sleep architecture and microglial morphology Level: Master’s thesis Month and year: May 2024 Number of pages: 50 pages Keywords: Sleep disruption, microglia, frontal cortex, adolescents, older mice, EEG, microglial morphology, hippocampus Supervisor or supervisors: Birgitte Rahbek Kornum, Christine Egebjerg Jensen Where deposited: University of Helsinki library Additional information: Abstract: Although sleep is an essential biological need for all beings, we have yet to understand why exactly it is a crucial aspect of our lives. The loss of sleep is seen as a natural occurrence that increases as we begin to age. The consequences of sleep deprivation are not yet fully understood but have been associated with a range of detrimental effects on comorbid conditions, including reduced quality of life, cognitive impairments, immune suppression, and various other adverse outcomes. The role of microglia in response to sleep deprivation is a discussion that is also yet to be understood, but that can be a pivotal point for future understanding. This master's thesis investigates the impact of sleep deprivation on sleep architecture in aged mice and microglial activation in adolescents. The study aims to understand how sleep disruption affects these age groups, focusing on microglial morphology and overall sleep patterns. Using EEG/EMG recordings, sleep disruption was induced by introducing novel objects for four hours daily at ZT 2-6 over seven days. The study found that older mice experienced a shift in their sleep patterns, with significant changes in NREM and REM sleep occurring during the dark phase, highlighting the influence of the circadian rhythm. In adolescent mice, sleep disruption led to increased morphological changes, suggesting a reduction in microglial activity or an intermediary state of activation. The results underscore the importance of sleep in maintaining neural homeostasis and highlight age-dependent differences in the response to sleep loss. The study discusses the implications of these findings for understanding the neurobiological mechanisms underlying sleep and its disruption, particularly in relation to microglial function and brain health. 
  • Tallberg, Robert Georg Michael (2021)
    The immune system is crucial in the central nervous system (CNS), protecting sensitive tissues, promoting regeneration, and maintaining homeostasis. It is involved in CNS-disorders, such as neurodegenerative diseases and neurological insults related to stroke. Critical myeloid leukocytes in the CNS are microglia, divided into pro-inflammatory M1 and anti-inflammatory M2 phenotypes. This polarization achieves modulation of the inflammatory response by amplifying or dampening it. Therefore, microglia are widely investigated in CNS-disorders. β2-integrins are adhesion proteins that mediate inflammation. They are expressed explicitly on leukocytes, including microglia. Important processes, such as phagocytosis and cell motility, are regulated by β2-integrins. They also relay downstream signals, altering inflammation in many settings, although their effects on microglial properties and stroke are currently poorly understood. We here aimed to investigate the role of β2-integrins in stroke-related injury and microglia polarization in vivo using knock-in (KI) mice, which lack functional β2-integrins. Our results show that in a mouse model of haemorrhagic stroke, the functional outcome was less severe in β2-integrin KI versus wild-type (WT) mice (P = 0.0147), suggesting that β2-integrins are involved in stroke pathophysiology. Furthermore, by using flow cytometry we observed significantly lower frequencies of M1 microglia in the KI mouse brain (P = 0.0096). Therefore, our findings reveal neuroprotective aspects by inhibiting β2-integrins in neuroinflammation. Investigating microglial properties mediated by β2-integrins could contribute to the understanding of neuroinflammatory events, leading to the development of therapies for poorly treated CNS-disorders. Our results suggest that β2-integrins should be further explored as molecular targets for novel stroke treatments.
  • Jasikova, Sara (2024)
    Schizophrenia (SCZ) is a chronic neuropsychiatric disorder believed to arise from the intricate interplay between genetic predisposition and environmental factors. Though the aetiology of SCZ is unknown many findings support an excessive synaptic pruning hypothesis. Maternal immune activation (MIA), encompassing prenatal infection and systemic inflammation, constitutes a significant environmental risk factor implicated in SCZ onset (Patterson, 2009; Brown, 2012). MIA induces persistent alterations in the microglia of offspring termed microglial priming, characterized by heightened reactivity to inflammatory stimuli (Choudhury and Lennox, 2021). Notably, studies have reported increased sensitivity to activation, elevated expression of inflammatory markers, and an increase in the total number of microglia (Perry and Holmes, 2014; Choudhury and Lennox, 2021). Primed microglia may contribute to excessive synaptic pruning, thereby compromising neuronal connectivity and potentially leading to the onset of SCZ. This thesis investigated the impact of microglia on neurons and explored the microglial tendency for hyperactivation in the context of SCZ predisposition. It utilized induced pluripotent stem cell (iPSC) technology to create a rat astrocyte/unaffected control human iPSC-derived neuron/induced microglia-like cell (iMGL) tri-culture model. Uniquely, iMGLs were differentiated from a library of monozygotic twin lines discordant for SCZ, and unaffected controls. This allows for exploration of the differences between iMGLs from unaffected twins with a genetic predisposition for SCZ, affected twins with clinical manifestation of SCZ, and unaffected controls without a known genetic predisposition for SCZ. The tri-culture system was subjected to lipopolysaccharide (LPS) and polyinosinic:polycytidylic acid (poly(I:C)) treatments to activate iMGLs, and differences in cytokine release, synapse pruning, and neuronal activity were assessed. The principal outcomes of our investigation revealed enhanced cytokine release from SCZ-derived iMGLs when exposed to inflammatory stimuli, alongside increased network connectivity among samples containing genetically predisposed iMGLs. While most of the results did not reach significance, they suggest a potential link between SCZ pathophysiology and hyperactive microglia. Future research will focus on enlarging the study cohort, establishing tri-culture models featuring neurons and iMGLs derived from the iPSCs of the same patient, conducting CBA analysis to confirm the elevated cytokines finding, and scrutinizing iMGL morphology.