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The Alternative Complement System Mediates Cell Death in Retinal Ischemia Reperfusion Injury.

ABSTRACT: Ischemia reperfusion (IR) injury induces retinal cell death and contributes to visual impairment. Previous studies suggest that the complement cascade plays a key role in IR injury in several systemic diseases. However, the role of the complement pathway in the ischemic retina has not been investigated. The aim of this study is to determine if the alternative complement cascade plays a role in retinal IR injury, and identify which components of the pathway mediate retinal degeneration in response to IR injury. To accomplish this, we utilized the mouse model of retinal IR injury, wherein the intraocular pressure (IOP) is elevated for 45 min, collapsing the retinal blood vessels and inducing retinal ischemia, followed by IOP normalization and subsequent reperfusion. We found that mRNA expression of complement inhibitors complement receptor 1-related gene/protein-y (Crry), Cd55 and Cd59a was down-regulated after IR. Moreover, genetic deletion of complement component 3 (C3^-/-) and complement factor b (Fb^-/-) decreased IR-induced retinal apoptosis. Because vascular dysfunction is central to IR injury, we also assessed the role of complement in a model of shear stress. In human retinal endothelial cells (HRECs), shear stress up-regulated complement inhibitors Cd46, Cd55, and Cd59, and suppressed complement-mediated cell death, indicating that a lack of vascular flow, commonly observed in IR injury, allows for complement mediated attack of the retinal vasculature. These results suggested that in retinal IR injury, the alternative complement system is activated by suppression of complement inhibitors, leading to vascular dysfunction and neuronal cell death.

SUBMITTER: Inafuku S

PROVIDER: S-EPMC6107794 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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The Alternative Complement System Mediates Cell Death in Retinal Ischemia Reperfusion Injury.

Inafuku Saori S Klokman Garrett G Connor Kip M KM

Frontiers in molecular neuroscience 20180817

Ischemia reperfusion (IR) injury induces retinal cell death and contributes to visual impairment. Previous studies suggest that the complement cascade plays a key role in IR injury in several systemic diseases. However, the role of the complement pathway in the ischemic retina has not been investigated. The aim of this study is to determine if the alternative complement cascade plays a role in retinal IR injury, and identify which components of the pathway mediate retinal degeneration in respons ...[more]

PMID: 30174588

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Project description:ObjectiveRetinal ischemic disease is a major cause of vision loss. Current treatment options are limited to late-stage diseases, and the molecular mechanisms of the initial insult are not fully understood. We have previously shown that the deletion of the mitochondrial arginase isoform, arginase 2 (A2), limits neurovascular injury in models of ischemic retinopathy. Here, we investigated the involvement of A2-mediated alterations in mitochondrial dynamics and function in the pathology.MethodsWe used wild-type (WT), global A2 knockout (A2KO-) mice, cell-specific A2 knockout mice subjected to retinal ischemia/reperfusion (I/R), and bovine retinal endothelial cells (BRECs) subjected to an oxygen-glucose deprivation/reperfusion (OGD/R) insult. We used western blotting to measure levels of cell stress and death markers and the mitochondrial fragmentation protein, dynamin related protein 1 (Drp1). We also used live cell mitochondrial labeling and Seahorse XF analysis to evaluate mitochondrial fragmentation and function, respectively.ResultsWe found that the global deletion of A2 limited the I/R-induced disruption of retinal layers, fundus abnormalities, and albumin extravasation. The specific deletion of A2 in endothelial cells was protective against I/R-induced neurodegeneration. The OGD/R insult in BRECs increased A2 expression and induced cell stress and cell death, along with decreased mitochondrial respiration, increased Drp1 expression, and mitochondrial fragmentation. The overexpression of A2 in BREC also decreased mitochondrial respiration, promoted increases in the expression of Drp1, mitochondrial fragmentation, and cell stress and resulted in decreased cell survival. In contrast, the overexpression of the cytosolic isoform, arginase 1 (A1), did not affect these parameters.ConclusionsThis study is the first to show that A2 in endothelial cells mediates retinal ischemic injury through a mechanism involving alterations in mitochondrial dynamics and function.

Project description:BackgroundRetinal function is ordered by interactions between transcriptional and posttranscriptional regulators at the molecular level. These regulators include transcription factors (TFs) and posttranscriptional factors such as microRNAs (miRs). Some studies propose that miRs predominantly target the TFs rather than other types of protein coding genes and such studies suggest a possible interconnection of these two regulators in co-regulatory networks.ResultsOur lab has generated mRNA and miRNA microarray expression data to investigate time-dependent changes in gene expression, following induction of ischemia-reperfusion (IR) injury in the rat retina. Data from different reperfusion time points following retinal IR-injury were analyzed. Paired expression data for miRNA-target gene (TG), TF-TG, miRNA-TF were used to identify regulatory loop motifs whose expressions were altered by the IR injury paradigm. These loops were subsequently integrated into larger regulatory networks and biological functions were assayed. Systematic analyses of the networks have provided new insights into retinal gene regulation in the early and late periods of IR. We found both overlapping and unique patterns of molecular expression at the two time points. These patterns can be defined by their characteristic molecular motifs as well as their associated biological processes. We highlighted the regulatory elements of miRs and TFs associated with biological processes in the early and late phases of ischemia-reperfusion injury.ConclusionsThe etiology of retinal ischemia-reperfusion injury is orchestrated by complex and still not well understood gene networks. This work represents the first large network analysis to integrate miRNA and mRNA expression profiles in context of retinal ischemia. It is likely that an appreciation of such regulatory networks will have prognostic potential. In addition, the computational framework described in this study can be used to construct miRNA-TF interactive systems networks for various diseases/disorders of the retina and other tissues.

Project description:RationaleWe recently implicated a role for CD4(+) T cells and demonstrated elevated IL-17A expression in lung ischemia-reperfusion (IR) injury. However, identification of the specific subset of CD4(+) T cells and their mechanistic role in IR injury remains unknown.ObjectivesWe tested the hypothesis that invariant natural killer T (iNKT) cells mediate lung IR injury via IL-17A signaling.MethodsMice underwent lung IR via left hilar ligation. Pulmonary function was measured using an isolated lung system. Lung injury was assessed by measuring edema (wet/dry weight) and vascular permeability (Evans blue dye). Inflammation was assessed by measuring proinflammatory cytokines in lungs, and neutrophil infiltration was measured by immunohistochemistry and myeloperoxidase levels.Measurements and main resultsPulmonary dysfunction (increased airway resistance and pulmonary artery pressure and decreased pulmonary compliance), injury (edema, vascular permeability), and inflammation (elevated IL-17A; IL-6; tumor necrosis factor-?; monocyte chemotactic protein-1; keratinocyte-derived chemokine; regulated upon activation, normal T-cell expressed and secreted; and neutrophil infiltration) after IR were attenuated in IL-17A(-/-) and Rag-1(-/-) mice. Anti-IL-17A antibody attenuated lung dysfunction in wild-type mice after IR. Reconstitution of Rag-1(-/-) mice with wild-type, but not IL-17A(-/-), CD4(+) T cells restored lung dysfunction, injury, and inflammation after IR. Lung dysfunction, injury, IL-17A expression, and neutrophil infiltration were attenuated in J?18(-/-) mice after IR, all of which were restored by reconstitution with wild-type, but not IL-17A(-/-), iNKT cells. Flow cytometry and enzyme-linked immunosorbent spot assay confirmed IL-17A production by iNKT cells after IR.ConclusionsThese results demonstrate that CD4(+) iNKT cells play a pivotal role in initiating lung injury, inflammation, and neutrophil recruitment after IR via an IL-17A-dependent mechanism.

Dataset Information

The Alternative Complement System Mediates Cell Death in Retinal Ischemia Reperfusion Injury.

Publications

The Alternative Complement System Mediates Cell Death in Retinal Ischemia Reperfusion Injury.

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