Temporal unsnarling of brain’s acute neuroinflammatory transcriptional profiles reveals panendothelitis as the earliest event preceding microgliosis

Abstract

Sepsis-associated encephalopathy (SAE) is an acutely progressing brain dysfunction induced by systemic inflammation. The mechanism of initiation of neuroinflammation during SAE, which ultimately leads to delirium and cognitive dysfunction, remains elusive. We aimed to study the molecular events of SAE to capture its onset and progression into the central nervous system (CNS), and further identify the cellular players involved in mediating acute inflammatory signaling. Gene expression profiling on the cerebral vessels isolated from the brains of the mice treated with peripheral lipopolysaccharide (LPS) revealed that the cerebral vasculature responds within minutes to acute systemic inflammation by upregulating the expression of immediate early response genes, followed by activation of the nuclear factor-κB pathway. To identify the earliest responding cell type, we used fluorescence-activated cell sorting (FACS) to sort the glial and vascular cells from the brains of the mice treated with LPS at different time points, and RNA-seq was performed on microglia and cerebral endothelial cells (CECs). Bioinformatic analysis followed by further validation in all the cell types revealed that panendothelitis. i.e., the activation of CECs is the earliest event in the CNS during the inception of acute neuroinflammation. Microglial activation occurs later than that of CECs, suggesting that CECs are the most likely initial source of proinflammatory mediators, which could further initiate glial cell activation. This is then followed by the activation of apoptotic signaling in the CECs, which is known to lead to the blood–brain barrier disruption and allow peripheral cytokines to leak into the CNS, exacerbate the gliosis, and result in the vicious neuroinflammatory cascade. Together, our results model the earliest sequential events during the advancement of systemic inflammation into the CNS and facilitate to understand the interplay between the vascular and glial cells in initiating and driving acute neuroinflammation during SAE.

Deep parallel characterization of AAV tropism and AAV-mediated transcriptional changes via single-cell RNA sequencing

“…The highest number of differentially expressed genes being in endothelial cells suggests that vascular cells could be the initial responders to viral transduction and expression of the transgene. This is supported by Kodali et al, who have shown that endothelial cells are the first to elicit a response to peripheral inflammatory stimulation by transcribing genes for proinflammatory mediators and cytokines (Kodali et al, 2020). With regards to p53 differentially expressed genes, Ghouzzi, et al have also shown that the genes Phdla3, Aen, and Cdkn1a were upregulated in cells infected with ZIKA virus, signifying genotoxic stress and apoptosis induction (Ghouzzi et al, 2016).…”

“…Immediate early genes such as Ier2 (Kodali et al, 2020) were upregulated across pericytes, endothelial cells, inhibitory neurons, and OPCs at 3 DPI, while Fos, Ier2, Junb, and Arc were prominent in excitatory neurons at 25 DPI.…”

Role of microRNAs As Biomarkers in Sepsis-Associated Encephalopathy

“…Several mechanisms are involved in SAE pathogenesis, such as disturbance of the blood–brain barrier (BBB), neuronal apoptosis, endothelial activation, hyperinflammation produced by inflammatory cytokines release, oxidative stress, neurotransmission disturbances by alteration in neurotransmitter level, altered brain signaling, altered microcirculation, and dysregulated metabolism [ 5 , 46 ]. Recently, Kodali et al demonstrated that cerebral endothelial cells (CECs) are the first activated cells during the earliest stages of acute neuroinflammation, defined as a spinal cord or brain inflammatory response, mediated by cytokine, chemokine, and oxidative stress production [ 47 , 48 ]. Kodali et al [ 47 ] suggested that CECs are the main source of proinflammatory mediators, which in turn can promote glial cell activation, such as microglial activation.…”

“…Recently, Kodali et al demonstrated that cerebral endothelial cells (CECs) are the first activated cells during the earliest stages of acute neuroinflammation, defined as a spinal cord or brain inflammatory response, mediated by cytokine, chemokine, and oxidative stress production [ 47 , 48 ]. Kodali et al [ 47 ] suggested that CECs are the main source of proinflammatory mediators, which in turn can promote glial cell activation, such as microglial activation. Furthermore, Kodali et al [ 47 ] observed that SAE continued by activating apoptotic signaling in CECs, which is known that causes a BBB disruption, allowing the entrance of peripheral cytokines into the CNS and thus causing an exacerbate gliosis.…”

“…Kodali et al [ 47 ] suggested that CECs are the main source of proinflammatory mediators, which in turn can promote glial cell activation, such as microglial activation. Furthermore, Kodali et al [ 47 ] observed that SAE continued by activating apoptotic signaling in CECs, which is known that causes a BBB disruption, allowing the entrance of peripheral cytokines into the CNS and thus causing an exacerbate gliosis. Finally, it causes a vicious neuroinflammatory cascade, which is commonly observed during SAE.…”