Loss of Serpin E2 alters antimicrobial gene expression by microglia but not astrocytes.

Microglia are the brain-resident immune cells responsible for surveilling and protecting the central nervous system. These cells can express a wide array of immune genes, and that expression can become highly dynamic in response to changes in the environment, such as traumatic injury or neurological disease. Though microglial immune responses are well studied, we still do not know many mechanisms and regulators underlying all the varied microglial responses. Serpin E2 is a serine protease inhibitor that acts on a wide variety of serine proteases, with particularly potent affinity for the blood clotting enzyme thrombin. In the brain, Serpin E2 is highly expressed by many cell types, especially glia, and loss of Serpin E2 leads to behavioral changes as well as deficits in synaptic plasticity. To determine whether Serpin E2 is important for maintaining homeostasis in glia, we performed RNA sequencing of microglia and astrocytes from Serpin E2-deficient mice in a healthy state or under immune activation due to lipopolysaccharide (LPS) injection. We found that microglia in Serpin E2-deficient mice had higher expression of antimicrobial genes, while astrocytes did not display any robust changes in transcription. Furthermore, the lack of Serpin E2 did not affect transcriptional responses to LPS in either microglia or astrocytes. Overall, we find that Serpin E2 is a regulator of antimicrobial genes in microglia.

Lymphocyte deficiency alters the transcriptomes of oligodendrocytes, but not astrocytes or microglia.

Though the brain was long characterized as an immune-privileged organ, findings in recent years have shown extensive communications between the brain and peripheral immune cells. We now know that alterations in the peripheral immune system can affect the behavioral outputs of the central nervous system, but we do not know which brain cells are affected by the presence of peripheral immune cells. Glial cells including microglia, astrocytes, oligodendrocytes, and oligodendrocyte precursor cells (OPCs) are critical for the development and function of the central nervous system. In a wide range of neurological and psychiatric diseases, the glial cell state is influenced by infiltrating peripheral lymphocytes. However, it remains largely unclear whether the development of the molecular phenotypes of glial cells in the healthy brain is regulated by lymphocytes. To answer this question, we acutely purified each type of glial cell from immunodeficient Rag2-/- mice. Interestingly, we found that the transcriptomes of microglia, astrocytes, and OPCs developed normally in Rag2-/- mice without reliance on lymphocytes. In contrast, there are modest transcriptome differences between the oligodendrocytes from Rag2-/- and control mice. Furthermore, the subcellular localization of the RNA-binding protein Quaking, is altered in oligodendrocytes. These results demonstrate that the molecular attributes of glial cells develop largely without influence from lymphocytes and highlight potential interactions between lymphocytes and oligodendrocytes.

Activity-dependent translation dynamically alters the proteome of the perisynaptic astrocyte process.

Within eukaryotic cells, translation is regulated independent of transcription, enabling nuanced, localized, and rapid responses to stimuli. Neurons respond transcriptionally and translationally to synaptic activity. Although transcriptional responses are documented in astrocytes, here we test whether astrocytes have programmed translational responses. We show that seizure activity rapidly changes the transcripts on astrocyte ribosomes, some predicted to be downstream of BDNF signaling. In acute slices, we quantify the extent to which cues of neuronal activity activate translation in astrocytes and show that this translational response requires the presence of neurons, indicating that the response is non-cell autonomous. We also show that this induction of new translation extends into the periphery of astrocytes. Finally, synaptic proteomics show that new translation is required for changes that occur in perisynaptic astrocyte protein composition after fear conditioning. Regulation of translation in astrocytes by neuronal activity suggests an additional mechanism by which astrocytes may dynamically modulate nervous system functioning.

Human Astrocytes Exhibit Tumor Microenvironment-, Age-, and Sex-Related Transcriptomic Signatures.

Astrocytes are critical for the development and function of synapses. There are notable species differences between human astrocytes and commonly used animal models. Yet, it is unclear whether astrocytic genes involved in synaptic function are stable or exhibit dynamic changes associated with disease states and age in humans, which is a barrier in understanding human astrocyte biology and its potential involvement in neurologic diseases. To better understand the properties of human astrocytes, we acutely purified astrocytes from the cerebral cortices of over 40 humans across various ages, sexes, and disease states. We performed RNA sequencing to generate transcriptomic profiles of these astrocytes and identified genes associated with these biological variables. We found that human astrocytes in tumor-surrounding regions downregulate genes involved in synaptic function and sensing of signals in the microenvironment, suggesting involvement of peritumor astrocytes in tumor-associated neural circuit dysfunction. In aging, we also found downregulation of synaptic regulators and upregulation of markers of cytokine signaling, while in maturation we identified changes in ionic transport with implications for calcium signaling. In addition, we identified subtle sexual dimorphism in human cortical astrocytes, which has implications for observed sex differences across many neurologic disorders. Overall, genes involved in synaptic function exhibit dynamic changes in the peritumor microenvironment and aging. These data provide powerful new insights into human astrocyte biology in several biologically relevant states that will aid in generating novel testable hypotheses about homeostatic and reactive astrocytes in humans.SIGNIFICANCE STATEMENT Astrocytes are an abundant class of cells playing integral roles at synapses. Astrocyte dysfunction is implicated in a variety of human neurologic diseases. Yet our knowledge of astrocytes is largely based on mouse studies. Direct knowledge of human astrocyte biology remains limited. Here, we present transcriptomic profiles of human cortical astrocytes, and we identified molecular differences associated with age, sex, and disease state. We found that peritumor and aging astrocytes downregulate genes involved in astrocyte-synapse interactions. These data provide necessary insight into human astrocyte biology that will improve our understanding of human disease.

Astrocyte-to-astrocyte contact and a positive feedback loop of growth factor signaling regulate astrocyte maturation.

Astrocytes are critical for the development and function of the central nervous system. In developing brains, immature astrocytes undergo morphological, molecular, cellular, and functional changes as they mature. Although the mechanisms that regulate the maturation of other major cell types in the central nervous system such as neurons and oligodendrocytes have been extensively studied, little is known about the cellular and molecular mechanisms that control astrocyte maturation. Here, we identified molecular markers of astrocyte maturation and established an in vitro assay for studying the mechanisms of astrocyte maturation. Maturing astrocytes in vitro exhibit similar molecular changes and represent multiple molecular subtypes of astrocytes found in vivo. Using this system, we found that astrocyte-to-astrocyte contact strongly promotes astrocyte maturation. In addition, secreted signals from microglia, oligodendrocyte precursor cells, or endothelial cells affect a small subset of astrocyte genes but do not consistently change astrocyte maturation. To identify molecular mechanisms underlying astrocyte maturation, we treated maturing astrocytes with molecules that affect the function of tumor-associated genes. We found that a positive feedback loop of heparin-binding epidermal growth factor-like growth factor (HBEGF) and epidermal growth factor receptor (EGFR) signaling regulates astrocytes maturation. Furthermore, HBEGF, EGFR, and tumor protein 53 (TP53) affect the expression of genes important for cilium development, the circadian clock, and synapse function. These results revealed cellular and molecular mechanisms underlying astrocytes maturation with implications for the understanding of glioblastoma.