Abstract 17016: The Role of Polymerase Delta-Interacting Protein 2 in Pulmonary Vascular Inflammation

Circulation ◽  
2018 ◽  
Vol 138 (Suppl_1) ◽  
Author(s):  
Steven J Forrester ◽  
Qian Xu ◽  
Daniel S Kikuchi ◽  
Derick Okwan-Duodu ◽  
Bernard Lassegue ◽  
...  
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
Evans Blue ◽  
Inflammatory Marker ◽  
Vascular Inflammation ◽  
Blue Staining ◽  
Leukocyte Infiltration ◽  
Leukocyte Adherence ◽  
Interacting Protein ◽  
Marker Expression

Introduction: Vascular inflammation is an underlying causative factor in the progression of various ailments including acute lung injury. In the inflamed lung, chronic endothelial cell (EC) activation facilitates leukocyte recruitment and extravasation into tissue beds, leading to end organ damage. Delineating the mechanisms of EC activation may uncover novel areas of therapeutic opportunity. Our laboratory has identified polymerase delta-interacting protein 2 (Poldip2) as a novel regulator of endothelial activation and permeability. Methods: Poldip2 +/- and +/+ mice were injected with either saline or lipopolysaccharide (LPS, 18 mg/kg) for either 6 or 18 hours to induce acute lung injury. Bronchoalveolar lavage (BAL) fluid was collected to assess leukocyte infiltration via flow cytometry and cytokine expression via ELISA. Evans blue staining was used to evaluate lung permeability, and tissue lysates were analyzed for inflammatory marker expression. For in vitro analysis, inflammatory marker expression, leukocyte adherence and cell permeability were tested in human pulmonary microvascular endothelial cells (HPMVECs) treated with saline or LPS (1 μg/mL). To test the efficacy of Poldip2 silencing in HPMVECs, siRNA was used at 100 nM (siControl vs siPoldip2). Comparisons between groups were analyzed with a one-way ANOVA followed by Tukey’s post-hoc test. Results: Compared to Poldip2 +/+, Poldip2 +/- mice treated with LPS exhibited significantly reduced mortality (80% vs. 20% survival), Evans blue staining, and leukocyte infiltration. FACS analysis of BAL fluid revealed a decrease in LPS-induced monocyte and neutrophil infiltration in Poldip2 +/- mice compared to Poldip2 +/+ mice. RT-qPCR analysis indicated a reduction in inflammatory gene induction in lung tissue. In HPMVECs, LPS induced increases in VCAM-1 protein expression, THP-1 leukocyte adherence and endothelial permeability, which were attenuated with siRNA-mediated knockdown of Poldip2 ( P<0.05 for all). Conclusion: Poldip2 is an important regulator of EC activation, leukocyte adherence and EC permeability. In scenarios of persistent vascular inflammation and endothelial barrier dysfunction, such as in acute lung injury, inhibition of Poldip2 may provide clinical benefit.

scholarly journals Long non-coding RNA OIP5-AS1 aggravates acute lung injury by promoting inflammation and cell apoptosis via regulating the miR-26a-5p/TLR4 axis

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Qingsong Sun ◽  
Man Luo ◽  
Zhiwei Gao ◽  
Xiang Han ◽  
Weiqin Wu ◽  
...  
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
Inflammatory Response ◽  
Cell Apoptosis ◽  
Quantitative Polymerase Chain Reaction ◽  
Luciferase Reporter ◽  
Enzyme Linked Immunosorbent Assay ◽  
Interacting Protein ◽  
Wide Range

Abstract Background Acute lung injury (ALI) is a pulmonary disorder that leads to acute respiration failure and thereby results in a high mortality worldwide. Increasing studies have indicated that toll-like receptor 4 (TLR4) is a promoter in ALI, and we aimed to explore the underlying upstream mechanism of TLR4 in ALI. Methods We used lipopolysaccharide (LPS) to induce an acute inflammatory response in vitro model and a murine mouse model. A wide range of experiments including reverse transcription quantitative polymerase chain reaction, western blot, enzyme linked immunosorbent assay, flow cytometry, hematoxylin–eosin staining, RNA immunoprecipitation, luciferase activity and caspase-3 activity detection assays were conducted to figure out the expression status, specific role and potential upstream mechanism of TLR4 in ALI. Result TLR4 expression was upregulated in ALI mice and LPS-treated primary bronchial/tracheal epithelial cells. Moreover, miR-26a-5p was confirmed to target TLR4 according to results of luciferase reporter assay. In addition, miR-26a-5p overexpression decreased the contents of proinflammatory factors and inhibited cell apoptosis, while upregulation of TLR4 reversed these effects of miR-26a-5p mimics, implying that miR-26a-5p alleviated ALI by regulating TLR4. Afterwards, OPA interacting protein 5 antisense RNA 1 (OIP5-AS1) was identified to bind with miR-26a-5p. Functionally, OIP5-AS1 upregulation promoted the inflammation and miR-26a-5p overexpression counteracted the influence of OIP5-AS1 upregulation on cell inflammatory response and apoptosis. Conclusion OIP5-AS1 promotes ALI by regulating the miR-26a-5p/TLR4 axis in ALI mice and LPS-treated cells, which indicates a promising insight into diagnostics and therapeutics in ALI.

scholarly journals Gene engineered mesenchymal stem cells: greater transgene expression and efficacy with minicircle vs. plasmid DNA vectors in a mouse model of acute lung injury

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Maria Florian ◽  
Jia-Pey Wang ◽  
Yupu Deng ◽  
Luciana Souza-Moreira ◽  
Duncan J. Stewart ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Transgene Expression ◽  
Vascular Inflammation ◽  
Pulmonary Inflammation ◽  
Necrosis Factor Alpha ◽  
Monocyte Chemoattractant Protein 1 ◽  
Therapeutic Benefits

Abstract Background Acute lung injury (ALI) and in its severe form, acute respiratory distress syndrome (ARDS), results in increased pulmonary vascular inflammation and permeability and is a major cause of mortality in many critically ill patients. Although cell-based therapies have shown promise in experimental ALI, strategies are needed to enhance the potency of mesenchymal stem cells (MSCs) to develop more effective treatments. Genetic modification of MSCs has been demonstrated to significantly improve the therapeutic benefits of these cells; however, the optimal vector for gene transfer is not clear. Given the acute nature of ARDS, transient transfection is desirable to avoid off-target effects of long-term transgene expression, as well as the potential adverse consequences of genomic integration. Methods Here, we explored whether a minicircle DNA (MC) vector containing human angiopoietin 1 (MC-ANGPT1) can provide a more effective platform for gene-enhanced MSC therapy of ALI/ARDS. Results At 24 h after transfection, nuclear-targeted electroporation using an MC-ANGPT1 vector resulted in a 3.7-fold greater increase in human ANGPT1 protein in MSC conditioned media compared to the use of a plasmid ANGPT1 (pANGPT1) vector (2048 ± 567 pg/mL vs. 552.1 ± 33.5 pg/mL). In the lipopolysaccharide (LPS)-induced ALI model, administration of pANGPT1 transfected MSCs significantly reduced bronchoalveolar lavage (BAL) neutrophil counts by 57%, while MC-ANGPT1 transfected MSCs reduced it by 71% (p < 0.001) by Holm-Sidak’s multiple comparison test. Moreover, compared to pANGPT1, the MC-ANGPT1 transfected MSCs significantly reduced pulmonary inflammation, as observed in decreased levels of proinflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α), interferon-gamma (IFN-γ), interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1), and macrophage inflammatory protein-2 (MIP-2). pANGPT1-transfected MSCs significantly reduced BAL albumin levels by 71%, while MC-ANGPT1-transfected MSCs reduced it by 85%. Conclusions Overall, using a minicircle vector, we demonstrated an efficient and sustained expression of the ANGPT1 transgene in MSCs and enhanced the therapeutic effect on the ALI model compared to plasmid. These results support the potential benefits of MC-ANGPT1 gene enhancement of MSC therapy to treat ARDS.

Gene silencing of receptor-interacting protein 2 protects against cigarette smoke-induced acute lung injury

2019 ◽  
Vol 139 ◽  
pp. 560-568 ◽  
Author(s):  
Jinrui Dong ◽  
Wupeng Liao ◽  
Lay Hong Tan ◽  
Amy Yong ◽  
Wen Yan Peh ◽  
...  
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
Gene Silencing ◽  
Cigarette Smoke ◽  
Interacting Protein

scholarly journals Gene Engineered Mesenchymal Stem Cells: Greater Transgene Expression and Efficacy With Minicircle Vs. Plasmid DNA Vectors in a Mouse Model of Acute Lung Injury

2020 ◽  
Author(s):  
Maria Florian ◽  
Jia-Pey Wang ◽  
Yupu Deng ◽  
Luciana Souza-Moreira ◽  
Duncan John Stewart ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Transgene Expression ◽  
Vascular Inflammation ◽  
Pulmonary Inflammation ◽  
Necrosis Factor Alpha ◽  
Monocyte Chemoattractant Protein 1 ◽  
Therapeutic Benefits

Abstract Background: Acute lung injury (ALI), and in its severe form the Acute respiratory distress syndrome (ARDS), results in increased pulmonary vascular inflammation and permeability, and is a major cause of mortality in many critically ill patients. Although cell-based therapies have shown promise in experimental ALI, strategies are needed to enhance potency of mesenchymal stem cells (MSC) to develop more effective treatments. Genetic modification of MSC has been demonstrated to significantly improve therapeutic benefits of these cells; however, the optimal vector for gene transfer is not clear. Given the acute nature of ARDS, transient transfection is desirable to avoid off target effects of long-term transgene expression, as well as the potential adverse consequences of genomic integration. Methods: Here, we explored whether a minicircle DNA (MC) DNA vector containing human angiopoietin 1 (MC-ANGPT-1) can provide more effective platform for gene-enhanced MSC therapy of ALI/ARDS. Results: At 24 hours after transfection, nuclear-targeted electroporation using a MC-ANGPT1 vector resulted in a 3,7 fold greater increase in human ANGPT1 protein in MSC conditioned media compared to the use of a plasmid ANGPT1 (pANGPT1) vector (2048±567pg/mL vs. 552.1±33.5pg/mL). In the lipopolysaccharide (LPS)-induced ALI model, administration of pANGPT1 transfected MSC significantly reduced bronchoalveolar lavage (BAL) neutrophil counts by 57%, while MC-ANGPT1 transfected MSC reduced it by 71% (p<0.001) by Holm-Sidak’s multiple comparison test. Moreover, compared to pANGPT1, the MC-ANGPT1 transfected MSC significantly reduced pulmonary inflammation, as observed in decreased levels of proinflammatory cytokines tumor necrosis factor alpha (TNF-α), interferron gamma (IFN-γ), interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-2 (MIP-2). pANGPT1 transfected MSC significantly reduced BAL albumin levels by 71% , while MC-ANGPT1 transfected MSC reduced it by 85%. Coclusions: Overall, using a minicircle vector, we demonstrated an efficient and sustained expression of the ANGPT1 transgene in MSC and enhanced therapeutic effect on ALI model compared to plasmid. These results support the potential benefits of the MC-ANGPT1 gene enhancement of MSC therapy to treat ARDS.

scholarly journals Acetylharpagide Protects Mice from Staphylococcus Aureus-Induced Acute Lung Injury by Inhibiting NF-κB Signaling Pathway

Molecules ◽  
2020 ◽  
Vol 25 (23) ◽  
pp. 5523
Author(s):  
Zhaoxin Zhang ◽  
Yun Wang ◽  
Yating Shan ◽  
Wu Yin
Keyword(s):  
Staphylococcus Aureus ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Signaling Pathway ◽  
Inflammatory Factors ◽  
Therapeutic Effects ◽  
Gas Barrier ◽  
Interacting Protein ◽  
Risk Of Death ◽  
Serious Disease

Staphylococcus aureus (S. aureus)-induced acute lung injury (ALI) is a serious disease that has a high risk of death among infants and teenagers. Acetylharpagide, a natural compound of Ajuga decumbens Thunb. (family Labiatae), has been found to have anti-tumor, anti-inflammatory and anti-viral effects. This study investigates the therapeutic effects of acetylharpagide on S. aureus-induced ALI in mice. Here, we found that acetylharpagide alleviated S. aureus-induced lung pathological morphology damage, protected the pulmonary blood-gas barrier and improved the survival of S. aureus-infected mice. Furthermore, S. aureus-induced myeloperoxidase (MPO) activity of lung homogenate and pro-inflammatory factors in bronchoalveolar lavage (BAL) fluid were suppressed by acetylharpagide. Mechanically, acetylharpagide inhibited the interaction between polyubiquitinated receptor interacting protein 1 (RIP1) and NF-κB essential modulator (NEMO), thereby suppressing NF-κB activity. In summary, these results show that acetylharpagide protects mice from S. aureus-induced ALI by suppressing the NF-κB signaling pathway. Acetylharpagide is expected to become a potential treatment for S. aureus-induced ALI.

scholarly journals Glucagon-like peptide-1 receptor mediates the beneficial effect of liraglutide in an acute lung injury mouse model involving the thioredoxin-interacting protein

2020 ◽  
Vol 319 (3) ◽  
pp. E568-E578 ◽  
Author(s):  
Wenyong Zhou ◽  
Weijuan Shao ◽  
Yu Zhang ◽  
Dinghui Liu ◽  
Mingyao Liu ◽  
...  
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
Weight Ratio ◽  
Lung Injury Score ◽  
Dependent Manner ◽  
Interacting Protein ◽  
Thioredoxin Interacting Protein ◽  
Beneficial Effects ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1

Repurposing clinically used drugs is among the important strategies in drug discovery. Glucagon-like peptide-1 (GLP-1) and its diabetes-based drugs, such as liraglutide, possess a spectrum of extra-pancreatic functions, while GLP-1 receptor (GLP-1R) is most abundantly expressed in the lung. Recent studies have suggested that GLP-1-based drugs exert beneficial effects in chronic, as well as acute, lung injury rodent models. Here, we show that liraglutide pretreatment reduced LPS induced acute lung injury in mice. It significantly reduced lung injury score, wet/dry lung weight ratio, bronchoalveolar lavage fluid immune cell count and protein concentration, and cell apoptosis in the lung, and it was associated with reduced lung inflammatory cytokine and chemokine gene expression. Importantly, these effects were virtually absent in GLP-1R−/− mice. A well-known function of GLP-1 and GLP-based drugs in pancreatic β-cells is the attenuation of high-glucose stimulated expression of thioredoxin-interacting protein (TxNIP), a key component of inflammasome. LPS-challenged lungs showed elevated TxNIP mRNA and protein expression, which was attenuated by liraglutide treatment in a GLP-1R-dependent manner. Hence, our observations suggest that GLP-1R is essential in mediating beneficial effects of liraglutide in acute lung injury, with the inflammasome component TxNIP as a potential target.

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