Microglial cell dysregulation in brain aging and neurodegeneration

Abstract

Aging is the main risk factor for neurodegenerative diseases. In aging, microglia undergoes phenotypic changes compatible with their activation. Glial activation can lead to neuroinflammation, which is increasingly accepted as part of the pathogenesis of neurodegenerative diseases, including Alzheimer's disease (AD). We hypothesize that in aging, aberrant microglia activation leads to a deleterious environment and neurodegeneration. In aged mice, microglia exhibit an increased expression of cytokines and an exacerbated inflammatory response to pathological changes. Whereas LPS increases nitric oxide (NO) secretion in microglia from young mice, induction of reactive oxygen species (ROS) predominates in older mice. Furthermore, there is accumulation of DNA oxidative damage in mitochondria of microglia during aging, and also an increased intracellular ROS production. Increased ROS activates the redox-sensitive nuclear factor kappa B, which promotes more neuroinflammation, and can be translated in functional deficits, such as cognitive impairment. Mitochondria-derived ROS and cathepsin B, are also necessary for the microglial cell production of interleukin-1β, a key inflammatory cytokine. Interestingly, whereas the regulatory cytokine TGFβ1 is also increased in the aged brain, neuroinflammation persists. Assessing this apparent contradiction, we have reported that TGFβ1 induction and activation of Smad3 signaling after inflammatory stimulation are reduced in adult mice. Other protective functions, such as phagocytosis, although observed in aged animals, become not inducible by inflammatory stimuli and TGFβ1. Here, we discuss data suggesting that mitochondrial and endolysosomal dysfunction could at least partially mediate age-associated microglial cell changes, and, together with the impairment of the TGFβ1-Smad3 pathway, could result in the reduction of protective activation and the facilitation of cytotoxic activation of microglia, resulting in the promotion of neurodegenerative diseases.

Keywords: Alzheimer’s disease; glia; mitochondria; neurodegenerative diseases; neuroinflammation; oxidative stress; reactive oxygen species; transforming growth factor-β.

Figure 1

Reactivespecies participate in normal cellular function orin pathological mechanisms depending on their overproduction. Reactive oxygen species (ROS) and reactive nitrogen species (RNS), are produced through several mechanisms by the cell: the electron transport chain in mitochondria, various cytosolic and membrane enzymes (i.e., xanthine oxidase (XO), nitric oxide synthase (NOS), NADPH oxidase complex, etc.), as well as exogenously provided by the environment. At the same time, cells have several antioxidant defense mechanisms for detoxifying ROS and RNS, including enzymes (i.e., superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR) and nonenzymatic antioxidants (i.e., reduced glutathione (GSH), vitamins E and C. The main generation pathways of ROS and RNS are also shown: the reduction of O₂ occurs by diverse mechanisms (i.e., mitochondria, XO, NADPH-oxidase complex) leading to formation of superoxide anion (O₂^•-); which is easily transformed to hydrogen peroxide (H₂O₂) either nonenzymatically or by SOD. H₂O₂ is converted to H₂O by CAT, or by GPx, which together with the GR regenerate GSH. In addition, under stress conditions and high concentration of transition metal (i.e., iron ions—Fe), O₂ •- can generate hydroxyl radical (OH•), which in turn can react with polyunsaturated fatty acids (PUFAs) and generate peroxyl radical (ROO•). Finally, O₂ •- can react with nitric oxide (NO; depending on NOS), producing the highly reactive peroxinitrite (ONOO•) anion, whereas H₂O₂ is converted to hypochlorous acid (HOCl) by myeloperoxidase (MPO). The balance between oxidants compounds and antioxidant defense determines the end result. Optimal physiologic levels leads to beneficial effects, with ROS and RNS acting as second messengers in intracellular signaling cascades (modulation of gene regulation and signal transduction pathways, mainly by activation of NFκB), regulating several physiological functions (i.e., cognitive and immune functions). However, when overproduction of ROS/RNS is higher than the antioxidant system, the equilibrium status favors oxidant vs. antioxidant reactions, leading to oxidative stress, in which ROS/RNS have harmful effects, because of their reaction with various macromolecules (lipids, proteins and nucleic acids), contributing to cellular and tissue oxidative damage, and the development of age-related impairments. Oxidation products: 3-NT, 3-nitrotyrosine; 8-OHdG, 8-hydroxy-2-deoxyguanosine; malondialdehyde (MDA); alkoxyl radical (RO•).

Figure 2

Reactive oxygen species and inflammation in the aged microglia. Representation of the participation of mitochondria and lysosomes in the increased production of ROS and inflammatory cytokines by aged microglia. Increased intracellular ROS activate redox-sensitive NFκB through a pathway mediated by mitochondrial ROS (associated with decreased energetic production and increased release of ROS by the electron transport chain) and a ROS-independent pathway, potentiating neuroinflammation. The activation of NFκB induces production of pro-CatB and pro-IL1β, and the activation of inflammasome in the cytoplasm. Pro-CatB is processed into CatB in the lysosome, which in turn, mediates the activation of pro-caspase-1 to caspase 1 and increases the processing of pro-IL1β, releasing increased amounts of IL1β both in the phagolysosome and the cytoplasm, as well as potentially potentiate apoptosis. Changes on the expression of pattern recognition receptors, like TLR4 CD14 and SRA, result in changes on neuroinflammatory activation and oxidative stress by activating NFκB and the release of ROS.

Figure 3

Aging-related morphological changes of microglia. Microglial cell morphology changes with aging. Immunohistochemistry for Iba-1 (a constitutive identity marker for monocyte-macrophage cells) and counterstaining with hematoxylin of hippocampal sections from animals of different ages (1- to 18-month old). Microglia obtained from young mice have a small cell body and very long and slender ramifications. As mice age, microglia gradually show bigger cell bodies and progressively shorter and thicker cell processes.

Figure 4

Age-related changes of microglial cell function. In aged brains, there is an increased number, size and activation of microglia. This is affected by additional systemic pathophysiological changes associated with other age related changes, environmental factors and disease processes, such as cardiovascular risk factors and metabolic syndrome or injuries. Deleterious processes further promote an inflammatory environment, increasing cytotoxic microglial cell activation, whereas risk factor management and pharmacological interventions can promote a healthy aging. Aged microglia changes depend both on gained and lost functions. They have increased basal phagocytic activity, although a reduced capacity to induce phagocytosis when stimulated, together with reduced lysosomal activity, resulting in a decreased clearance activity. Microglia also shows an increased production of inflammatory cytokines and reactive species. Those changes result in a shift of balance towards decreased protective functions and an increased neurotoxicity.

Figure 5

Aging of the nervous and immune system and the neuroimmune crosstalk. Healthy aging of the nervous and immune systems depend both on genetic and environmental (lifestyle) factors. Aging is associated with a state of low grade chronic oxidative stress and inflammation (with production of reactive mediators and inflammatory compounds and a decreased antioxidant and anti-inflammatory capacity), which appear to be the cause of an important part of age-related deterioration of the nervous and the immune systems, as well as of the neuroimmune communication. Because of their complex functions, the central nervous system (CNS) and the immune system are especially vulnerable to oxidative damage (i.e., lipid peroxidation, protein oxidation, DNA damage), which contributes to oxidative stress and inflammation. Age-related changes in the immune function, known as immunosenescence, results in increased susceptibility to infections and cancer, inflammation and autoimmune diseases. In the CNS, oxidative stress has a negative impact on function, leading to mitochondrial dysfunction and impaired energetic metabolism, altered neuronal and glial signaling. There may be disruption of the cycle glutamate-glutamine and increased levels of neuronal calcium, which are involved in mechanisms of neuronal damage leading to loss of function, excitotoxicity and apoptosis. In addition, dysfunction of the neuron-glia crosstalk leads to a chronic neuroinflammation, which promotes a prolonged activation of microglia and further induction of dysfunction and degenerative changes. All these alterations contribute to functional decline and the development of neurodegenerative diseases. NO, nitric oxide; NOS, nitric oxide synthase; RNS, reactive nitrogen species; ROS, reactive oxygen species.

Figure 6

The “Glial Cell Dysregulation Hypothesis” of Alzheimer’s disease (AD). The glial cell dysregulation hypothesis proposes that AD has its cause on changes on the activation of microglia and on impaired regulation, which become increasingly cytotoxic decreasing their defensive functions. Impaired activation results in oxidative stress, persistent neuroinflammation and neuronal dysfunction, all of which can also induce production and aggregation of Aβ, and additional neuronal dysfunction. Inflammatory activation, secondary to aging and to various forms of stimuli or injury through life, can result in glial cell dysregulation. Dysregulated activation of glia, through the abnormal release of cytokines, reactive species, and other mediators, contributes to the increased expression of Aβ as well to functional and degenerative changes of neurons, perpetuating abnormal activation of glia, synaptic dysfunction and cell damage.

Figure 7

Life style changes as a strategy for aging well. Cognitive activity, dietary caloric restriction and moderate physical exercise induce mild stress responses which results in a decreased production of stress proteins and reduction of oxidative stress. In additions, there is an increased production of neurotrophic factors, among which brain-derived neurotrophic factor (BDNF) appears to be one of the most important, but also participate growth hormone (GH) and insulin growth factor 1 (IGF1). Decreased stress signal and increased trophic signal acts on mitochondrial function, improving energetic metabolism and reducing oxidative stress to a protective level. Stress signals and ROS, below a certain threshold concentration, induce survival signals capable of restoring cellular homeostasis but, at higher or continued levels, can contribute to aging and degenerative changes.