Aging and sex: Impact on microglia phagocytosis

Microglia dysfunction and activation are important hallmarks of the aging brain and are concomitant with age‐related neurodegeneration and cognitive decline. Age‐associated changes in microglia migration and phagocytic capacity result in maladaptive responses, chronic neuroinflammation, and worsened outcomes in neurodegenerative disorders. Given the sex bias in the incidence, prevalence, and therapy response of most neurological disorders, we have here examined whether the phagocytic activity of aged microglia is different in males and females. With this aim, the phagocytosis activity of male and female cells was compared in an in vitro aged microglia model and in microglia isolated from adult (5‐month‐old) or aged (18‐month‐old) mice. In both models, the phagocytosis of neural debris increased with aging in male and female cells and was higher in aged female microglia than in aged male cells. However, female aged microglia lost its ability to adapt its phagocytic activity to inflammatory conditions. These findings suggest that microglia phagocytosis of neural debris may represent a previously unexplored neuroprotective characteristic of aged microglia that may contribute to the generation of sex differences in the manifestation of neurodegenerative diseases.     

Ccr2 deficiency impairs microglial accumulation and accelerates progression of Alzheimer-like disease

Microglia are the principal immune cells of the brain. In Alzheimer disease, these brain mononuclear phagocytes are recruited from the blood and accumulate in senile plaques. However, the role of microglia in Alzheimer disease has not been resolved. Microglia may be neuroprotective by phagocytosing amyloid-beta (Abeta), but their activation and the secretion of neurotoxins may also cause neurodegeneration. Ccr2 is a chemokine receptor expressed on microglia, which mediates the accumulation of mononuclear phagocytes at sites of inflammation. Here we show that Ccr2 deficiency accelerates early disease progression and markedly impairs microglial accumulation in a transgenic mouse model of Alzheimer disease (Tg2576). Alzheimer disease mice deficient in Ccr2 accumulated Abeta earlier and died prematurely, in a manner that correlated with Ccr2 gene dosage, indicating that absence of early microglial accumulation leads to decreased Abeta clearance and increased mortality. Thus, Ccr2-dependent microglial accumulation plays a protective role in the early stages of Alzheimer disease by promoting Abeta clearance.     

小胶质细胞的神经递质受体在阿尔茨海默病中的作用

阿尔茨海默病(Alzheimer's disease,AD)是一种因蛋白错误折叠、聚积影响神经细胞功能,从而导致认知功能下降、行为异常的神经退行性疾病.小胶质细胞是中枢神经系统(central nervous system,CNS)中重要的免疫细胞之一,在AD病理过程中,根据其激活状态的不同小胶质细胞发挥神经保护或神经...     

小胶质细胞在帕金森病病理进展中的作用

帕金森病是中老年人常见的中枢神经系统退行性疾病,研究表明小胶质细胞的活化及其介导的神经炎症在帕金森病的病程进展中发挥重要作用,适度干预小胶质细胞的活化有望延缓帕金森病的进程。小胶质细胞是中枢神经系统固有的巨噬细胞,Notch信号途径可以调控小鼠外周巨噬细胞的分化及功能。Notch通路也参与调控小胶质细胞的激活、细胞因子的表达、吞噬活性的变化等,而这与活化的小胶质细胞介导的帕金森病等神经退行性疾病的病情进展相关。因此,本文将综述Notch信号途径与小胶质细胞介导的相关疾病的研究进展。     

Cathepsin regulation on microglial function

Microglia, the resident mononuclear phagocyte population in the brain, have long been implicated in the pathology of neurodegenerative age-associated disorders. However, activated microglia have now been identified as homeostatic keepers in the brain, because they are involved in the initiation and resolution of neuropathology. The complex roles of activated microglia appear to be linked to change from inflammatory and neurotoxic to anti-inflammatory and neuroprotective phenotypes. Increased expression and secretion of various cathepsins support roles of activated microglia in chronic neuroinflammation, the neurotoxic M1-like polarization and neuronal death. Moreover, changes in expression and localization of microglial cathepsin B play a critical role in the acceleration of the brain aging. Beyond the role as brain-resident macrophages, many lines of evidence have shown that microglia have essential roles in the maturation and maintenance of neuronal circuits in the developing and adult brain. Cathepsin S secreted from microglia induces the diurnal variation of spine density of cortical neurons though proteolytic modification of peri-synaptic extracellular matrix molecules. In this review, I highlight the emerging roles of cathepsins that support the roles of microglia in both normal healthy and pathological brains. In addition, I discuss cathepsin inhibitors as potential therapeutic targets for brain disorders.     

Microglia-Specific Metabolic Changes in Neurodegeneration

The high energetic demand of the brain deems this organ rather sensitive to changes in energy supply. Therefore, even minor alterations in energy metabolism may underlie detrimental disturbances in brain function, contributing to the generation and progression of neurodegenerative diseases. Considerable evidence supports the key role of deficits in cerebral energy metabolism, particularly hypometabolism of glucose and mitochondrial dysfunction, in the pathophysiology of brain disorders. Major breakthroughs in the field of bioenergetics and neurodegeneration have been achieved through the use of in vitro and in vivo models of disease as well as sophisticated neuroimaging techniques in patients, yet these have been mainly focused on neuron and astrocyte function. Remarkably, the subcellular metabolic mechanisms linked to neurodegeneration that operate in other crucial brain cell types such as microglia have remain obscured, although they are beginning to be unraveled. Microglia, the brain-resident immune sentinels, perform a diverse range of functions that require a high-energy expenditure, namely, their role in brain development, maintenance of the neural environment, response to injury and infection, and activation of repair programs. Interestingly, another key mechanism underlying several neurodegenerative diseases is neuroinflammation, which can be associated with chronic microglia activation. Considering that many brain disorders are accompanied by changes in brain energy metabolism and sustained inflammation, and that energy metabolism has a strong influence on the inflammatory responses of microglia, the emerging significance of microglial energy metabolism in neurodegeneration is highlighted in this review.     

Homeostatic and injury‐induced microglia behavior in the aging brain

Microglia cells are essential for brain homeostasis and have essential roles in neurodegenerative diseases. Aging is the main risk factor for most neurodegenerative diseases, and age‐related changes in microglia may contribute to the susceptibility of the aging brain to dysfunction and neurodegeneration. We have analyzed morphology and dynamic behavior of neocortical microglia in their physiological environment in young adult (3‐month‐old), adult (11‐ to 12‐month‐old), and aged (26‐ to 27‐month‐old) C57BL/6J‐Iba1‐eGFP mice using in vivo 2‐photon microscopy. Results show that surveying microglial cells in the neocortex exhibit age‐related soma volume increase, shortening of processes, and loss of homogeneous tissue distribution. Furthermore, microglial process speed significantly decreased with age. While only a small population of microglia showed soma movement in adult mice, the microglia population with soma movement was increased in aged mice. However, in response to tissue injury, the dynamic microglial response was age‐dependently diminished. These results provide novel insights into microglial behavior and indicate that microglial dysfunction in the aging brain may contribute to age‐related cognitive decline and neurodegenerative diseases.     

Suppressed Retinal Degeneration in Aged Wild Type and APPswe/PS1ΔE9 Mice by Bone Marrow Transplantation

Alzheimer''s disease (AD) is an age-related condition characterized by accumulation of neurotoxic amyloid β peptides (Aβ) in brain and retina. Because bone marrow transplantation (BMT) results in decreased cerebral Aβ in experimental AD, we hypothesized that BMT would mitigate retinal neurotoxicity through decreased retinal Aβ. To test this, we performed BMT in APPswe/PS1ΔE9 double transgenic mice using green fluorescent protein expressing wild type (wt) mice as marrow donors. We first examined retinas from control, non-transplanted, aged AD mice and found a two-fold increase in microglia compared with wt mice, prominent inner retinal Aβ and paired helical filament-tau, and decreased retinal ganglion cell layer neurons. BMT resulted in near complete replacement of host retinal microglia with BMT-derived cells and normalized total AD retinal microglia to non-transplanted wt levels. Aβ and paired helical filament-tau were reduced (61.0% and 44.1% respectively) in BMT-recipient AD mice, which had 20.8% more retinal ganglion cell layer neurons than non-transplanted AD controls. Interestingly, aged wt BMT recipients also had significantly more neurons (25.4%) compared with non-transplanted aged wt controls. Quantitation of retinal ganglion cell layer neurons in young mice confirmed age-related retinal degeneration was mitigated by BMT. We found increased MHC class II expression in BMT-derived microglia and decreased oxidative damage in retinal ganglion cell layer neurons. Thus, BMT is neuroprotective in age-related as well as AD-related retinal degeneration, and may be a result of alterations in innate immune function and oxidative stress in BMT recipient mice.     

Microglia in cerebral ischemia: molecular actions and interactions

The precise role of microglia in stroke and cerebral ischemia has been the subject of debate for a number of years. Microglia are capable of synthesizing numerous soluble and membrane-bound biomolecules, some known to be neuroprotective, some neurotoxic, whereas others have less definitive bioactivities. The molecular mechanisms through which microglia activate these molecules have thus become an important area of ischemia research. Here we provide a survey review that summarizes the key actions of microglial factors in cerebral ischemia including complement proteins, chemokines, pro-inflammatory cytokines, neurotrophic factors, hormones, and proteinases, as well several important messenger molecules that play a part in how these factors respond to extracellular signals during ischemic injuries. We also provide some new perspectives on how microglial intracellular signaling may contribute to the seemingly contradictory roles of several microglial effector molecules.     

Microglia in Alzheimer's Disease: Exploring How Genetics and Phenotype Influence Risk

Research into the function of microglia has dramatically accelerated during the last few years, largely due to recent genetic findings implicating microglia in virtually every neurodegenerative disorder. In Alzheimer's disease (AD), a majority of risk loci discovered through genome-wide association studies were found in or near genes expressed most highly in microglia leading to the hypothesis that microglia play a much larger role in disease progression than previously thought. From this body of work produced in the last several years, we find that almost every function of microglia has been proposed to influence the progression of AD from altered phagocytosis and synaptic pruning to cytokine secretion and changes in trophic support. By studying key Alzheimer's risk genes such as TREM2, CD33, ABCA7, and MS4A6A, we will be able to distinguish true disease-modulatory pathways from the full range of microglial-related functions. To successfully carry out these experiments, more advanced microglial models are needed. Microglia are quite sensitive to their local environment, suggesting the need to more fully recapitulate an in vivo environment to study this highly plastic cell type. Likely only by combining the above approaches will the field fully elucidate the molecular pathways that regulate microglia and influence neurodegeneration, in turn uncovering potential new targets for future therapeutic development.     

Chronic stress and Alzheimer's disease: the interplay between the hypothalamic–pituitary–adrenal axis,genetics and microglia

Chronic psychosocial stress is increasingly being recognised as a risk factor for sporadic Alzheimer's disease (AD). The hypothalamic–pituitary–adrenal axis (HPA axis) is the major stress response pathway in the body and tightly regulates the production of cortisol, a glucocorticoid hormone. Dysregulation of the HPA axis and increased levels of cortisol are commonly found in AD patients and make a major contribution to the disease process. The underlying mechanisms remain poorly understood. In addition, within the general population there are interindividual differences in sensitivities to glucocorticoid and stress responses, which are thought to be due to a combination of genetic and environmental factors. These differences could ultimately impact an individuals’ risk of AD. The purpose of this review is first to summarise the literature describing environmental and genetic factors that can impact an individual's HPA axis reactivity and function and ultimately AD risk. Secondly, we propose a mechanism by which genetic factors that influence HPA axis reactivity may also impact inflammation, a key driver of neurodegeneration. We hypothesize that these factors can mediate glucocorticoid priming of the immune cells of the brain, microglia, to become pro-inflammatory and promote a neurotoxic environment resulting in neurodegeneration. Understanding the underlying molecular mechanisms and identifying these genetic factors has implications for evaluating stress-related risk/progression to neurodegeneration, informing the success of interventions based on stress management and potential risks associated with the common use of glucocorticoids.     

Microglial neurotransmitter receptors trigger superoxide production in microglia; consequences for microglial-neuronal interactions

Microglia express three isoforms of the NADPH oxidase, Nox1, Nox2 and Nox4, with the potential to produce superoxide (O(2) ˙(-) ). Microglia also express neurotransmitter receptors, which can modulate microglial responses. In this study, microglial activity of Nox1, Nox2 and Nox4 in primary rat cultured microglia or the rodent BV2 cell line were altered by microglial neurotransmitter receptor modulation. Glutamate, GABA or ATP triggered microglial O(2) ˙(-) production via Nox activation. Nox activation was elicited by agonists of metabotropic mGlu3 receptors and by group III receptors, by GABA(A) but not GABA(B) receptors, and by purinergic P2X(7) or P2Y(2/4) receptors but not P2Y(1) receptors, and inhibited by metabotropic glutamate receptor 5 antagonists. The neurotransmitters also modulated Nox mRNA expression and NADPH activity. The activation of Nox by BzATP or GABA promoted a neuroprotective phenotype whilst the activation of Nox by glutamate promoted a neurotoxic phenotype. Taken together, these data indicate that microglial neurotransmitter receptors can signal via Nox to promote neuroprotection or neurotoxicity. This has implications for the subsequent neurotoxic profile of microglia when neurotransmitter levels may become skewed in neurodegeneration.     

Friend or Foe? Resident Microglia vs Bone Marrow-Derived Microglia and Their Roles in the Retinal Degeneration

Microglia are immune cells in the central nervous system (CNS) that originate from the yolk sac in an embryo. The renewal of the microglia pool in the adult eye consists of two components. In addition to the self-proliferation of resident cells, microglia in the CNS also derive from the bone marrow (BM). BM-derived cells pass through the blood–brain barrier (BBB) or blood–retina barrier (BRB) and differentiate into microglia under specific conditions which involves a complex mechanism. Recent studies have widely investigated the role of resident microglia and BM-derived microglia in the retinal degenerative disease. Both two cell types play dual roles and share many similar functions. However, resident microglia tend to polarize to the M1 phenotype which is pro-inflammatory and neurotoxic, whereas BM-derived microglia mainly polarize to the neuroprotective M2 phenotype in retinal degeneration. The molecular mechanism that underlines the invasion of peripheral cells has led to extensive discussions. In addition to the BBB and BRB disruption, many signaling pathways are involved in this process. Based on these studies, we discuss the roles of these two types of microglia in retinal degeneration disease and the potential clinical application of BM-derived microglia, which may benefit future therapies.     

Neuron–glia crosstalk in health and disease: fractalkine and CX3CR1 take centre stage

An essential aspect of normal brain function is the bidirectional interaction and communication between neurons and neighbouring glial cells. To this end, the brain has evolved ligand–receptor partnerships that facilitate crosstalk between different cell types. The chemokine, fractalkine (FKN), is expressed on neuronal cells, and its receptor, CX₃CR1, is predominantly expressed on microglia. This review focuses on several important functional roles for FKN/CX₃CR1 in both health and disease of the central nervous system. It has been posited that FKN is involved in microglial infiltration of the brain during development. Microglia, in turn, are implicated in the developmental synaptic pruning that occurs during brain maturation. The abundance of FKN on mature hippocampal neurons suggests a homeostatic non-inflammatory role in mechanisms of learning and memory. There is substantial evidence describing a role for FKN in hippocampal synaptic plasticity. FKN, on the one hand, appears to prevent excess microglial activation in the absence of injury while promoting activation of microglia and astrocytes during inflammatory episodes. Thus, FKN appears to be neuroprotective in some settings, whereas it contributes to neuronal damage in others. Many progressive neuroinflammatory disorders that are associated with increased microglial activation, such as Alzheimer''s disease, show disruption of the FKN/CX₃CR1 communication system. Thus, targeting CX₃CR1 receptor hyperactivation with specific antagonists in such neuroinflammatory conditions may eventually lead to novel neurotherapeutics.     

Accelerated microglial pathology is associated with Aβ plaques in mouse models of Alzheimer's disease

Microglia integrate within the neural tissue with a distinct ramified morphology through which they scan the surrounding neuronal network. Here, we used a digital tool for the quantitative morphometric characterization of fine cortical microglial structures in mice, and the changes they undergo with aging and in Alzheimer's‐like disease. We show that, compared with microglia in young mice, microglia in old mice are less ramified and possess fewer branches and fine processes along with a slightly increased proinflammatory cytokine expression. A similar microglial pathology appeared 6–12 months earlier in mouse models of Alzheimer's disease (AD), along with a significant increase in brain parenchyma lacking coverage by microglial processes. We further demonstrate that microglia near amyloid plaques acquire unique activated phenotypes with impaired process complexity. We thus show that along with a chronic proinflammatory reaction in the brain, aging causes a significant reduction in the capacity of microglia to scan their environment. This type of pathology is markedly accelerated in mouse models of AD, resulting in a severe microglial process deficiency, and possibly contributing to enhanced cognitive decline.     

Brain microglia express steroid-converting enzymes in the mouse

In the CNS, steroid hormones play a major role in the maintenance of brain homeostasis and it's response to injury. Since activated microglia are the pivotal immune cell involved in neurodegeneration, we investigated the possibility that microglia provide a discrete source for the metabolism of active steroid hormones. Using RT-PCR, our results showed that mouse microglia expressed mRNA for 17β-hydroxysteroid dehydrogenase type 1 and steroid 5-reductase type 1, which are involved in the metabolism of androgens and estrogens. Microglia also expressed the peripheral benzodiazepine receptor and steroid acute regulatory protein; however, the enzymes required for de novo formation of progesterone and DHEA from cholesterol were not expressed. To test the function of these enzymes, primary microglia cultures were incubated with steroid precursors, DHEA and AD. Microglia preferentially produced delta-5 androgens (Adiol) from DHEA and 5-reduced androgens from AD. Adiol behaved as an effective estrogen receptor agonist in neuronal cells. Activation of microglia with pro-inflammatory factors, LPS and INFγ did not affect the enzymatic properties of these proteins. However, PBR ligands reduced TNF production signifying an immunomodulatory role for PBR. Collectively, our results suggest that microglia utilize steroid-converting enzymes and related proteins to influence inflammation and neurodegeneration within microenvironments of the brain.     

Primary Microglia Isolation from Mixed Glial Cell Cultures of Neonatal Rat Brain Tissue

Microglia account for approximately 12% of the total cellular population in the mammalian brain. While neurons and astrocytes are considered the major cell types of the nervous system, microglia play a significant role in normal brain physiology by monitoring tissue for debris and pathogens and maintaining homeostasis in the parenchyma via phagocytic activity ^1,2. Microglia are activated during a number of injury and disease conditions, including neurodegenerative disease, traumatic brain injury, and nervous system infection ³. Under these activating conditions, microglia increase their phagocytic activity, undergo morpohological and proliferative change, and actively secrete reactive oxygen and nitrogen species, pro-inflammatory chemokines and cytokines, often activating a paracrine or autocrine loop ^4-6. As these microglial responses contribute to disease pathogenesis in neurological conditions, research focused on microglia is warranted.Due to the cellular heterogeneity of the brain, it is technically difficult to obtain sufficient microglial sample material with high purity during in vivo experiments. Current research on the neuroprotective and neurotoxic functions of microglia require a routine technical method to consistently generate pure and healthy microglia with sufficient yield for study. We present, in text and video, a protocol to isolate pure primary microglia from mixed glia cultures for a variety of downstream applications. Briefly, this technique utilizes dissociated brain tissue from neonatal rat pups to produce mixed glial cell cultures. After the mixed glial cultures reach confluency, primary microglia are mechanically isolated from the culture by a brief duration of shaking. The microglia are then plated at high purity for experimental study.The principle and protocol of this methodology have been described in the literature ^7,8. Additionally, alternate methodologies to isolate primary microglia are well described ^9-12. Homogenized brain tissue may be separated by density gradient centrifugation to yield primary microglia ¹². However, the centrifugation is of moderate length (45 min) and may cause cellular damage and activation, as well as, cause enriched microglia and other cellular populations. Another protocol has been utilized to isolate primary microglia in a variety of organisms by prolonged (16 hr) shaking while in culture ^9-11. After shaking, the media supernatant is centrifuged to isolate microglia. This longer two-step isolation method may also perturb microglial function and activation. We chiefly utilize the following microglia isolation protocol in our laboratory for a number of reasons: (1) primary microglia simulate in vivo biology more faithfully than immortalized rodent microglia cell lines, (2) nominal mechanical disruption minimizes potential cellular dysfunction or activation, and (3) sufficient yield can be obtained without passage of the mixed glial cell cultures.It is important to note that this protocol uses brain tissue from neonatal rat pups to isolate microglia and that using older rats to isolate microglia can significantly impact the yield, activation status, and functional properties of isolated microglia. There is evidence that aging is linked with microglia dysfunction, increased neuroinflammation and neurodegenerative pathologies, so previous studies have used ex vivo adult microglia to better understand the role of microglia in neurodegenerative diseases where aging is important parameter. However, ex vivo microglia cannot be kept in culture for prolonged periods of time. Therefore, while this protocol extends the life of primary microglia in culture, it should be noted that the microglia behave differently from adult microglia and in vitro studies should be carefully considered when translated to an in vivo setting.     

The 21st century epidemic: infections as inductors of neuro-degeneration associated with Alzheimer’s Disease

Alzheimer’s disease (AD) is a complex disease resulting in neurodegeneration and cognitive impairment. Investigations on environmental factors implicated in AD are scarce and the etiology of the disease remains up to now obscure. The disease’s pathogenesis may be multi-factorial and different etiological factors may converge during aging and induce an activation of brain microglia and macrophages. This microglia priming will result in chronic neuro-inflammation under chronic antigen activation. Infective agents may prime and drive iper-activation of microglia and be partially responsible of the induction of brain inflammation and decline of cognitive performances. Age-associated immune dis-functions induced by chronic sub-clinical infections appear to substantially contribute to the appearance of neuro-inflammation in the elderly. Individual predisposition to less efficient immune responses is another relevant factor contributing to impaired regulation of inflammatory responses and accelerated cognitive decline.Life-long virus infection may play a pivotal role in activating peripheral and central inflammatory responses and in turn contributing to increased cognitive impairment in preclinical and clinical AD.