New perspectives on heterogeneity in astrocytic reactivity in neuroinflammation

Mathys H., Boix C.A., Akay L.A., Xia Z., Davila-Velderrain J., Ng A.P., Jiang X., Abdelhady G., Galani K., Mantero J., Band N., James B.T., Babu S., Galiana-Melendez F., Louderback K., et. al.

AbstractAlzheimer’s disease is the leading cause of dementia worldwide, but the cellular pathways that underlie its pathological progression across brain regions remain poorly understood1–3. Here we report a single-cell transcriptomic atlas of six different brain regions in the aged human brain, covering 1.3 million cells from 283 post-mortem human brain samples across 48 individuals with and without Alzheimer’s disease. We identify 76 cell types, including region-specific subtypes of astrocytes and excitatory neurons and an inhibitory interneuron population unique to the thalamus and distinct from canonical inhibitory subclasses. We identify vulnerable populations of excitatory and inhibitory neurons that are depleted in specific brain regions in Alzheimer’s disease, and provide evidence that the Reelin signalling pathway is involved in modulating the vulnerability of these neurons. We develop a scalable method for discovering gene modules, which we use to identify cell-type-specific and region-specific modules that are altered in Alzheimer’s disease and to annotate transcriptomic differences associated with diverse pathological variables. We identify an astrocyte program that is associated with cognitive resilience to Alzheimer’s disease pathology, tying choline metabolism and polyamine biosynthesis in astrocytes to preserved cognitive function late in life. Together, our study develops a regional atlas of the ageing human brain and provides insights into cellular vulnerability, response and resilience to Alzheimer’s disease pathology.

Potokar M., Zorec R., Jorgačevski J.

Astrocytes are increasingly recognized as important viral host cells in the central nervous system. These cells can produce relatively high quantities of new virions. In part, this can be attributed to the characteristics of astrocyte metabolism and its abundant and dynamic cytoskeleton network. Astrocytes are anatomically localized adjacent to interfaces between blood capillaries and brain parenchyma and between blood capillaries and brain ventricles. Moreover, astrocytes exhibit a larger membrane interface with the extracellular space than neurons. These properties, together with the expression of various and numerous viral entry receptors, a relatively high rate of endocytosis, and morphological plasticity of intracellular organelles, render astrocytes important target cells in neurotropic infections. In this review, we describe factors that mediate the high susceptibility of astrocytes to viral infection and replication, including the anatomic localization of astrocytes, morphology, expression of viral entry receptors, and various forms of autophagy.

da Silva E.V., Fontes-Dantas F.L., Dantas T.V., Dutra A., Nascimento O.J., Alves-Leon S.V.

Zika virus (ZIKV) is an arbovirus of the Flaviviridae genus that has rapidly disseminated from across the Pacific to the Americas. Robust evidence has indicated a crucial role of ZIKV in congenital virus syndrome, including neonatal microcephaly. Moreover, emerging evidence suggests an association between ZIKV infection and the development of an extensive spectrum of central nervous system inflammatory demyelinating diseases (CNS IDD), such as multiple sclerosis–like clinical phenotypes. However, the underlying mechanisms of host-pathogen neuro-immune interactions remain to be elucidated. This study aimed to identify common transcriptional signatures between multiple sclerosis (MS) and ZIKV infection to generate molecular interaction networks, thereby leading to the identification of deregulated processes and pathways, which could give an insight of these underlying molecular mechanisms. Our investigation included publicly available transcriptomic data from MS patients in either relapse or remission (RR-MS) and datasets of subjects acutely infected by ZIKV for both immune peripheral cells and central nervous system cells. The protein-protein interaction (PPI) analysis showed upregulated AP-1 transcription factors (JUN and FOS) among the top hub and bottleneck genes in RR-MS and ZIKV data. Gene enrichment analysis retrieved a remarkable presence of ontologies and pathways linked to oxidative stress responses, immune cell function, inflammation, interleukin signaling, cell division, and transcriptional regulation commonly enriched in both scenarios. Considering the recent findings concerning AP-1 function in immunological tolerance breakdown, regulation of inflammation, and its function as an oxidative stress sensor, we postulate that the ZIKV trigger may contribute as a boost for the activation of such AP-1-regulated mechanisms that could favor the development of MS-like phenotypes following ZIKV infection in a genetically susceptible individual.

Jorgačevski J., Potokar M.

Neuroinfections of the central nervous system (CNS) can be triggered by various pathogens. Viruses are the most widespread and have the potential to induce long-term neurologic symptoms with potentially lethal outcomes. In addition to directly affecting their host cells and inducing immediate changes in a plethora of cellular processes, viral infections of the CNS also trigger an intense immune response. Regulation of the innate immune response in the CNS depends not only on microglia, which are fundamental immune cells of the CNS, but also on astrocytes. These cells align blood vessels and ventricle cavities, and consequently, they are one of the first cell types to become infected after the virus breaches the CNS. Moreover, astrocytes are increasingly recognized as a potential viral reservoir in the CNS; therefore, the immune response initiated by the presence of intracellular virus particles may have a profound effect on cellular and tissue physiology and morphology. These changes should be addressed in terms of persisting infections because they may contribute to recurring neurologic sequelae. To date, infections of astrocytes with different viruses originating from genetically distinct families, including Flaviviridae, Coronaviridae, Retroviridae, Togaviridae, Paramyxoviridae, Picomaviridae, Rhabdoviridae, and Herpesviridae, have been confirmed. Astrocytes express a plethora of receptors that detect viral particles and trigger signaling cascades, leading to an innate immune response. In this review, we summarize the current knowledge on virus receptors that initiate the release of inflammatory cytokines from astrocytes and depict the involvement of astrocytes in immune functions of the CNS.

Gotoh M., Miyamoto Y., Ikeshima-Kataoka H.

Both astrocytic and microglial functions have been extensively investigated in healthy subjects and neurodegenerative diseases. For astrocytes, not only various sub-types were identified but phagocytic activity was also clarified recently and is making dramatic progress. In this review paper, we mostly focus on the functional role of astrocytes in the extracellular matrix and on interactions between reactive astrocytes and reactive microglia in normal states and in neurodegenerative diseases, because the authors feel it is necessary to elucidate the mechanisms among activated glial cells in the pathology of neurological diseases in order to pave the way for drug discovery. Finally, we will review cyclic phosphatidic acid (cPA), a naturally occurring phospholipid mediator that induces a variety of biological activities in the brain both in vivo and in vitro. We propose that cPA may serve as a novel therapeutic molecule for the treatment of brain injury and neuroinflammation.

Wu H., Qiu X., Wang T., Chen K., Liu S., Li Y., He T., Zhao J.

Nassar A., Kodi T., Satarker S., Chowdari Gurram P., Upadhya D., SM F., Mudgal J., Nampoothiri M.

Astrocytes are important for maintaining cholesterol metabolism, glutamate uptake, and neurotransmission. Indeed, inflammatory processes and neurodegeneration contribute to the altered morphology, gene expression, and function of astrocytes. Astrocytes, in collaboration with numerous microRNAs, regulate brain cholesterol levels as well as glutamatergic and inflammatory signaling, all of which contribute to general brain homeostasis. Neural electrical activity, synaptic plasticity processes, learning, and memory are dependent on the astrocyte–neuron crosstalk. Here, we review the involvement of astrocytic microRNAs that potentially regulate cholesterol metabolism, glutamate uptake, and inflammation in Alzheimer’s disease (AD). The interaction between astrocytic microRNAs and long non-coding RNA and transcription factors specific to astrocytes also contributes to the pathogenesis of AD. Thus, astrocytic microRNAs arise as a promising target, as AD conditions are a worldwide public health problem. This review examines novel therapeutic strategies to target astrocyte dysfunction in AD, such as lipid nanodiscs, engineered G protein-coupled receptors, extracellular vesicles, and nanoparticles.

Kong W., Montano M., Corley M.J., Helmy E., Kobayashi H., Kinisu M., Suryawanshi R., Luo X., Royer L.A., Roan N.R., Ott M., Ndhlovu L.C., Greene W.C.

SARS-CoV-2 infection primarily targets the lung but may also damage other organs, including the brain, heart, kidney, and intestine. Central nervous system (CNS) pathologies include loss of smell and taste, headache, delirium, acute psychosis, seizures, and stroke.

Leng K., Rose I.V., Kim H., Xia W., Romero-Fernandez W., Rooney B., Koontz M., Li E., Ao Y., Wang S., Krawczyk M., TCW J., Goate A., Zhang Y., Ullian E.M., et. al.

Astrocytes become reactive in response to insults to the central nervous system by adopting context-specific cellular signatures and outputs, but a systematic understanding of the underlying molecular mechanisms is lacking. In this study, we developed CRISPR interference screening in human induced pluripotent stem cell-derived astrocytes coupled to single-cell transcriptomics to systematically interrogate cytokine-induced inflammatory astrocyte reactivity. We found that autocrine–paracrine IL-6 and interferon signaling downstream of canonical NF-κB activation drove two distinct inflammatory reactive signatures, one promoted by STAT3 and the other inhibited by STAT3. These signatures overlapped with those observed in other experimental contexts, including mouse models, and their markers were upregulated in human brains in Alzheimer’s disease and hypoxic-ischemic encephalopathy. Furthermore, we validated that markers of these signatures were regulated by STAT3 in vivo using a mouse model of neuroinflammation. These results and the platform that we established have the potential to guide the development of therapeutics to selectively modulate different aspects of inflammatory astrocyte reactivity. Leng et al. establish CRISPRi screens in astrocytes to dissect pathways controlling inflammatory reactivity. They uncover two distinct inflammatory reactive signatures that are inversely regulated by STAT3 and validate that these exist in human disease.

Fu Y., Guo Z., Wang Y., Zhang H., Zhang F., Xu Z., Shen X., Roppongi R.T., Mo S., Gu W., Nakajima T., Tsushima Y.

Alzheimer’s disease (AD)-like cognitive impairment, a kind of Neuro-COVID syndrome, is a reported complication of SARS-CoV-2 infection. However, the specific mechanisms remain largely unknown. Here, we integrated single-nucleus RNA-sequencing data to explore the potential shared genes and pathways that may lead to cognitive dysfunction in AD and COVID-19. We also constructed ingenuity AD-high-risk scores based on AD-high-risk genes from transcriptomic, proteomic, and Genome-Wide Association Studies (GWAS) data to identify disease-associated cell subtypes and potential targets in COVID-19 patients. We demonstrated that the primary disturbed cell populations were astrocytes and neurons between the above two dis-eases that exhibit cognitive impairment. We identified significant relationships between COVID-19 and AD involving synaptic dysfunction, neuronal damage, and neuroinflammation. Our findings may provide new insight for future studies to identify novel targets for preventive and therapeutic interventions in COVID-19 patients.

Steardo L., Steardo L., Scuderi C.

COVID-19, initially regarded as specific lung disease, exhibits an extremely broad spectrum of symptoms. Extrapulmonary manifestations of the disease also include important neuropsychiatric symptoms with atypical characteristics. Are these disturbances linked to stress accompanying every systemic infection, or are due to specific neurobiological changes associated with COVID-19? Evidence accumulated so far indicates that the pathophysiology of COVID-19 is characterized by systemic inflammation, hypoxia resulting from respiratory failure, and neuroinflammation (either due to viral neurotropism or in response to cytokine storm), all affecting the brain. It is reasonable to hypothesize that all these events may initiate or worsen psychiatric and cognitive disorders. Damage to the brain triggers a specific type of reactive response mounted by neuroglia cells, in particular by astrocytes which are the homeostatic cell par excellence. Astrocytes undergo complex morphological, biochemical, and functional remodeling aimed at mobilizing the regenerative potential of the central nervous system. If the brain is not directly damaged, resolution of systemic pathology usually results in restoration of the physiological homeostatic status of neuroglial cells. The completeness and dynamics of this process in pathological conditions remain largely unknown. In a subset of patients, glial cells could fail to recover after infection thus promoting the onset and progression of COVID-19-related neuropsychiatric diseases. There is evidence from post-mortem examinations of the brains of COVID-19 patients of alterations in both astrocytes and microglia. In conclusion, COVID-19 activates a huge reactive response of glial cells, that physiologically act as the main controller of the inflammatory, protective and regenerative events. However, in some patients the restoration of glial physiological state does not occur, thus compromising glial function and ultimately resulting in homeostatic failure underlying a set of specific neuropsychiatric symptoms related to COVID-19.

Huang S., Fishell G.

Spurgat M.S., Tang S.

Astrocytes and microglia are non-neuronal cells that maintain homeostasis within the central nervous system via their capacity to regulate neuronal transmission and prune synapses. Both astrocytes and microglia can undergo morphological and transcriptomic changes in response to infection with human immunodeficiency virus (HIV). While both astrocytes and microglia can be infected with HIV, HIV viral proteins in the local environment can interact with and activate these cells. Given that both astrocytes and microglia play critical roles in maintaining neuronal function, it will be critical to have an understanding of their heterogeneity and to identify genes and mechanisms that modulate their responses to HIV. Heterogeneity may include a depletion or increase in one or more astrocyte or microglial subtypes in different regions of the brain or spine as well as the gain or loss of a specific function. Single-cell RNA sequencing (scRNA-seq) has emerged as a powerful tool that can be used to characterise these changes within a given population. The use of this method facilitates the identification of subtypes and changes in cellular transcriptomes that develop in response to activation and various disease processes. In this review, we will examine recent studies that have used scRNA-seq to explore astrocyte and microglial heterogeneity in neurodegenerative diseases including Alzheimer’s disease and amyotrophic lateral sclerosis as well as in response to HIV infection. A careful review of these studies will expand our current understanding of cellular heterogeneity at homeostasis and in response to specific disease states.

Lee J., Kim S.W., Kim K.

Although different regions of the brain are dedicated to specific functions, the intra- and inter-regional heterogeneity of astrocytes and microglia in these regions has not yet been fully understood. Recently, an advancement in various technologies, such as single-cell RNA sequencing, has allowed for the discovery of astrocytes and microglia with distinct molecular fingerprints and varying functions in the brain. In addition, the regional heterogeneity of astrocytes and microglia exhibits different functions in several situations, such as aging and neurodegenerative diseases. Therefore, investigating the region-specific astrocytes and microglia is important in understanding the overall function of the brain. In this review, we summarize up-to-date research on various intra- and inter-regional heterogeneities of astrocytes and microglia, and provide information on how they can be applied to aging and neurodegenerative diseases.

Smajić S., Prada-Medina C.A., Landoulsi Z., Ghelfi J., Delcambre S., Dietrich C., Jarazo J., Henck J., Balachandran S., Pachchek S., Morris C.M., Antony P., Timmermann B., Sauer S., Pereira S.L., et. al.

Abstract Idiopathic Parkinson’s disease is characterized by a progressive loss of dopaminergic neurons, but the exact disease aetiology remains largely unknown. To date, Parkinson’s disease research has mainly focused on nigral dopaminergic neurons, although recent studies suggest disease-related changes also in non-neuronal cells and in midbrain regions beyond the substantia nigra. While there is some evidence for glial involvement in Parkinson’s disease, the molecular mechanisms remain poorly understood. The aim of this study was to characterize the contribution of all cell types of the midbrain to Parkinson’s disease pathology by single-nuclei RNA sequencing and to assess the cell type-specific risk for Parkinson’s disease using the latest genome-wide association study. We profiled >41 000 single-nuclei transcriptomes of post-mortem midbrain from six idiopathic Parkinson’s disease patients and five age-/sex-matched controls. To validate our findings in a spatial context, we utilized immunolabelling of the same tissues. Moreover, we analysed Parkinson’s disease-associated risk enrichment in genes with cell type-specific expression patterns. We discovered a neuronal cell cluster characterized by CADPS2 overexpression and low TH levels, which was exclusively present in idiopathic Parkinson’s disease midbrains. Validation analyses in laser-microdissected neurons suggest that this cluster represents dysfunctional dopaminergic neurons. With regard to glial cells, we observed an increase in nigral microglia in Parkinson’s disease patients. Moreover, nigral idiopathic Parkinson’s disease microglia were more amoeboid, indicating an activated state. We also discovered a reduction in idiopathic Parkinson’s disease oligodendrocyte numbers with the remaining cells being characterized by a stress-induced upregulation of S100B. Parkinson’s disease risk variants were associated with glia- and neuron-specific gene expression patterns in idiopathic Parkinson’s disease cases. Furthermore, astrocytes and microglia presented idiopathic Parkinson’s disease-specific cell proliferation and dysregulation of genes related to unfolded protein response and cytokine signalling. While reactive patient astrocytes showed CD44 overexpression, idiopathic Parkinson’s disease microglia revealed a pro-inflammatory trajectory characterized by elevated levels of IL1B, GPNMB and HSP90AA1. Taken together, we generated the first single-nuclei RNA sequencing dataset from the idiopathic Parkinson’s disease midbrain, which highlights a disease-specific neuronal cell cluster as well as ‘pan-glial’ activation as a central mechanism in the pathology of the movement disorder. This finding warrants further research into inflammatory signalling and immunomodulatory treatments in Parkinson’s disease.