Repeated ethanol exposure and withdrawal alters angiotensin-converting enzyme 2 expression in discrete brain regions: Implications for SARS-CoV-2 neuroinvasion.

BACKGROUND: People with alcohol use disorder (AUD) may be at higher risk for COVID-19. Angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) are required for cellular entry by SARS-CoV-2, but information on their expression in specific brain regions after alcohol exposure is limited. We sought to clarify how chronic alcohol exposure affects ACE2 expression in monoaminergic brainstem circuits and other putative SARS-CoV-2 entry points. METHODS: Brains were examined for ACE2 using immunofluorescence after 4 weeks of chronic intermittent ethanol (CIE) vapor inhalation. We also examined TMPRSS2, Cathepsin L, and ADAM17 by Western blot and RAS pathway mediators and pro-inflammatory markers via RT-qPCR. RESULTS: ACE2 was increased in most brain regions following CIE including the olfactory bulb (OB), hypothalamus (HT), raphe magnus (RMG), raphe obscurus (ROB), locus coeruleus (LC), and periaqueductal gray (PAG). We also observed increased colocalization of ACE2 with monoaminergic neurons in brainstem nuclei. Moreover, soluble ACE2 (sACE2) was elevated in OB, HT, and LC. The increase in sACE2 in OB and HT was accompanied by upregulation of ADAM17, an ACE2 sheddase, while TMPRSS2 increased in HT and LC. Cathepsin L, an endosomal receptor involved in viral entry, was also increased in OB. Alcohol can increase Angiotensin II, which triggers a pro-inflammatory response that may upregulate ACE2 via activation of RAS pathway receptors AT1R/AT2R. ACE2 then metabolizes Angiotensin II to Angiotensin (1-7) and provokes an anti-inflammatory response via MAS1. Accordingly, we report that AT1R/AT2R mRNA decreased in OB and increased in the LC, while MAS1 mRNA increased in both OB and LC. Other mRNAs for pro-inflammatory markers were also dysregulated in OB, HT, raphe, and LC. CONCLUSIONS: Our results suggest that alcohol triggers a compensatory upregulation of ACE2 in the brain due to disturbed RAS and may increase the risk or severity of SARS-CoV-2 infection.

Repeated ethanol exposure and withdrawal alters ACE2 expression in discrete brain regions: Implications for SARS-CoV-2 infection (preprint)

Emerging evidence suggests that people with alcohol use disorders are at higher risk for SARS-CoV-2. SARS-CoV-2 engages angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) receptors for cellular entry. While ACE2 and TMPRSS2 genes are upregulated in the cortex of alcohol-dependent individuals, information on expression in specific brain regions and neural populations implicated in SARS-CoV-2 neuroinvasion, particularly monoaminergic neurons, is limited. We sought to clarify how chronic alcohol exposure affects ACE2 and TMPRSS2 expression in monoaminergic brainstem circuits and other putative SARS-CoV-2 entry points. C57BL/6J mice were exposed to chronic intermittent ethanol (CIE) vapor for 4 weeks and brains were examined using immunofluorescence. We observed increased ACE2 levels in the olfactory bulb and hypothalamus following CIE, which are known to mediate SARS-CoV-2 neuroinvasion. Total ACE2 immunoreactivity was also elevated in the raphe magnus (RMG), raphe obscurus (ROB), and locus coeruleus (LC), while in the dorsal raphe nucleus (DRN), ROB, and LC we observed increased colocalization of ACE2 with monoaminergic neurons. ACE2 also increased in the periaqueductal gray (PAG) and decreased in the amygdala. Whereas ACE2 was detected in most brain regions, TMPRSS2 was only detected in the olfactory bulb and DRN but was not significantly altered after CIE. Our results suggest that previous alcohol exposure may increase the risk of SARS-CoV-2 neuroinvasion and render brain circuits involved in cardiovascular and respiratory function as well as emotional processing more vulnerable to infection, making adverse outcomes more likely. Additional studies are needed to define a direct link between alcohol use and COVID-19 infection.

Neuroprotective effects of flavone luteolin in neuroinflammation and neurotrauma.

Neuroinflammation leads to neurodegeneration, cognitive defects, and neurodegenerative disorders. Neurotrauma/traumatic brain injury (TBI) can cause activation of glial cells, neurons, and neuroimmune cells in the brain to release neuroinflammatory mediators. Neurotrauma leads to immediate primary brain damage (direct damage), neuroinflammatory responses, neuroinflammation, and late secondary brain damage (indirect) through neuroinflammatory mechanism. Secondary brain damage leads to chronic inflammation and the onset and progression of neurodegenerative diseases. Currently, there are no effective and specific therapeutic options to treat these brain damages or neurodegenerative diseases. Flavone luteolin is an important natural polyphenol present in several plants that show anti-inflammatory, antioxidant, anticancer, cytoprotective, and macrophage polarization effects. In this short review article, we have reviewed the neuroprotective effects of luteolin in neurotrauma and neurodegenerative disorders and pathways involved in this mechanism. We have collected data for this study from publications in the PubMed using the keywords luteolin and mast cells, neuroinflammation, neurodegenerative diseases, and TBI. Recent reports suggest that luteolin suppresses systemic and neuroinflammatory responses in Coronavirus disease 2019 (COVID-19). Studies have shown that luteolin exhibits neuroprotective effects through various mechanisms, including suppressing immune cell activation, such as mast cells, and inflammatory mediators released from these cells. In addition, luteolin can suppress neuroinflammatory response, activation of microglia and astrocytes, oxidative stress, neuroinflammation, and the severity of neuroinflammatory diseases such as Alzheimer's disease, Parkinson's disease, multiple sclerosis, and TBI pathogenesis. In conclusion, luteolin can improve cognitive decline and enhance neuroprotection in neurodegenerative diseases, TBI, and stroke.

Acute Traumatic Brain Injury-Induced Neuroinflammatory Response and Neurovascular Disorders in the Brain.

Acute traumatic brain injury (TBI) leads to neuroinflammation, neurodegeneration, cognitive decline, psychological disorders, increased blood-brain barrier (BBB) permeability, and microvascular damage in the brain. Inflammatory mediators secreted from activated glial cells, neurons, and mast cells are implicated in the pathogenesis of TBI through secondary brain damage. Abnormalities or damage to the neurovascular unit is the indication of secondary injuries in the brain after TBI. However, the precise mechanisms of molecular and ultrastructural neurovascular alterations involved in the pathogenesis of acute TBI are not yet clearly understood. Moreover, currently, there are no precision-targeted effective treatment options to prevent the sequelae of TBI. In this study, mice were subjected to closed head weight-drop-induced acute TBI and evaluated neuroinflammatory and neurovascular alterations in the brain by immunofluorescence staining or quantitation by enzyme-linked immunosorbent assay (ELISA) procedure. Mast cell stabilizer drug cromolyn was administered to inhibit the neuroinflammatory response of TBI. Results indicate decreased level of pericyte marker platelet-derived growth factor receptor-beta (PDGFR-ß) and BBB-associated tight junction proteins junctional adhesion molecule-A (JAM-A) and zonula occludens-1 (ZO-1) in the brains 7 days after weight-drop-induced acute TBI as compared with the brains from sham control mice indicating acute TBI-associated BBB/tight junction protein disruption. Further, the administration of cromolyn drug significantly inhibited acute TBI-associated decrease of PDGFR-ß, JAM-A, and ZO-1 in the brain. These findings suggest that acute TBI causes BBB/tight junction damage and that cromolyn administration could protect this acute TBI-induced brain damage as well as its long-time consequences.

COVID-19, Mast Cells, Cytokine Storm, Psychological Stress, and Neuroinflammation.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a new pandemic infectious disease that originated in China. COVID-19 is a global public health emergency of international concern. COVID-19 causes mild to severe illness with high morbidity and mortality, especially in preexisting risk groups. Therapeutic options are now limited to COVID-19. The hallmark of COVID-19 pathogenesis is the cytokine storm with elevated levels of interleukin-6 (IL-6), IL-1ß, tumor necrosis factor-alpha (TNF-α), chemokine (C-C-motif) ligand 2 (CCL2), and granulocyte-macrophage colony-stimulating factor (GM-CSF). COVID-19 can cause severe pneumonia, and neurological disorders, including stroke, the damage to the neurovascular unit, blood-brain barrier disruption, high intracranial proinflammatory cytokines, and endothelial cell damage in the brain. Mast cells are innate immune cells and also implicated in adaptive immune response, systemic inflammatory diseases, neuroinflammatory diseases, traumatic brain injury and stroke, and stress disorders. SARS-CoV-2 can activate monocytes/macrophages, dendritic cells, T cells, mast cells, neutrophils, and induce cytokine storm in the lung. COVID-19 can activate mast cells, neurons, glial cells, and endothelial cells. SARS-CoV-2 infection can cause psychological stress and neuroinflammation. In conclusion, COVID-19 can induce mast cell activation, psychological stress, cytokine storm, and neuroinflammation.