scholarly journals Induced Pluripotent Stem Cell-Derived Conditioned Medium Promotes Endogenous Leukemia Inhibitory Factor to Attenuate Endotoxin-Induced Acute Lung Injury

2021 ◽  
Vol 22 (11) ◽  
pp. 5554
Author(s):  
Vincent Yi-Fong Su ◽  
Shih-Hwa Chiou ◽  
Wei-Chih Chen ◽  
Wen-Kuang Yu ◽  
Huai-Hsuan Wu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Conditioned Medium ◽  
Adhesion Molecules ◽  
Leukemia Inhibitory Factor ◽  
In Vitro Models ◽  
Inhibitory Factor ◽  
Induced Pluripotent

The conditioned medium of induced pluripotent stem cells (iPSC-CM) can attenuate neutrophil recruitment and endothelial leakage of lipopolysaccharide (LPS)-induced acute lung injury (ALI). Therefore, we investigated the mechanisms by which iPSC-CM regulate the interaction between neutrophils and the endothelium in ALI. Murine iPSCs (miPSCs) were delivered intravenously to male C57BL/6 mice (8–12 weeks old) 4 h after intratracheal LPS injection. A miPSC-derived conditioned medium (miPSC-CM) was delivered intravenously to mice after intratracheal LPS injection. DMSO-induced HL-60 cells (D-HL-60, neutrophil-like cells) and human umbilical vein endothelial cells (HUVECs) were used as in vitro models to assess the interaction of neutrophils and endothelial cells. miPSC-CM diminished the histopathological changes in the lungs and the neutrophil count in bronchoalveolar lavage fluids of ALI mice. miPSC-CM attenuated the expression of adhesion molecules in the lungs of ALI mice. Human iPSC conditioned medium (hiPSC-CM) reduced the expression of adhesion molecules in a HUVEC and D-HL-60 co-culture after LPS stimulation, which decreased the transendothelial migration (TEM) of D-HL-60. A human angiogenesis factors protein array revealed that leukemia inhibitory factor (LIF) was not detected in the absence of D-HL-60 and hiPSC-CM groups. hiPSC-CM significantly promoted the production of endogenous LIF in in vitro models. Administration of an anti-LIF antibody not only reversed the effect of iPSC-CM in ALI mice, but also blocked the effect of iPSC-CM on neutrophils TEM in in vitro models. However, a controlled IgG had no such effect. Our study demonstrated that iPSC-CM promoted endogenous LIF to inhibit neutrophils TEM and attenuate the severity of sepsis-induced ALI.

scholarly journals Metformin-stimulated AMPK-α1 promotes microvascular repair in acute lung injury

2013 ◽  
Vol 305 (11) ◽  
pp. L844-L855 ◽  
Author(s):  
Ming-Yuan Jian ◽  
Mikhail F. Alexeyev ◽  
Paul E. Wolkowicz ◽  
Jaroslaw W. Zmijewski ◽  
Judy R. Creighton
Keyword(s):  
Endothelial Cells ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Ex Vivo ◽  
Endothelial Permeability ◽  
Beneficial Effects ◽  
Isolated Lung

Acute lung injury secondary to sepsis is a leading cause of mortality in sepsis-related death. Present therapies are not effective in reversing endothelial cell dysfunction, which plays a key role in increased vascular permeability and compromised lung function. AMP-activated protein kinase (AMPK) is a molecular sensor important for detection and mediation of cellular adaptations to vascular disruptive stimuli. In this study, we sought to determine the role of AMPK in resolving increased endothelial permeability in the sepsis-injured lung. AMPK function was determined in vivo using a rat model of endotoxin-induced lung injury, ex vivo using the isolated lung, and in vitro using cultured rat pulmonary microvascular endothelial cells (PMVECs). AMPK stimulation using N1-(α-d-ribofuranosyl)-5-aminoimidizole-4-carboxamide or metformin decreased the LPS-induced increase in permeability, as determined by filtration coefficient ( Kf) measurements, and resolved edema as indicated by decreased wet-to-dry ratios. The role of AMPK in the endothelial response to LPS was determined by shRNA designed to decrease expression of the AMPK-α1 isoform in capillary endothelial cells. Permeability, wounding, and barrier resistance assays using PMVECs identified AMPK-α1 as the molecule responsible for the beneficial effects of AMPK in the lung. Our findings provide novel evidence for AMPK-α1 as a vascular repair mechanism important in the pulmonary response to sepsis and identify a role for metformin treatment in the management of capillary injury.

scholarly journals Characterization of the iPSC-derived conditioned medium that promotes the growth of bovine corneal endothelial cells

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e6734
Author(s):  
Qing Liu ◽  
Yonglong Guo ◽  
Shiwei Liu ◽  
Peiyuan Wang ◽  
Yunxia Xue ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Conditioned Medium ◽  
Cell Morphology ◽  
Cell Cycle Progression ◽  
Cellular Proliferation ◽  
Activin A ◽  
Corneal Endothelial Cells ◽  
Induced Pluripotent

Corneal endothelial cells (CECs) maintain corneal transparency and visual acuity. However, the limited proliferative capability of these cells in vitro has prompted researchers to find efficient culturing techniques for them. The aim of our study was to evaluate the use of conditioned medium (CM) obtained from induced pluripotent stem cells (iPSCs) as a source for the effective proliferation of bovine CECs (B-CECs). In our study, the proliferative ability of B-CECs was moderately enhanced when the cells were grown in 25% iPSC conditioned medium (iPSC-CM). Additionally, hexagonal cell morphology was maintained until passage 4, as opposed to the irregular and enlarged shape observed in control corneal endothelial medium (CEM). B-CECs in both the 25% iPSC-CM and CEM groups expressed and Na+-K+-ATPase. The gene expression levels of NIFK, Na+-K+-ATPase, Col4A and Col8A and the percentage of cells entering S and G2 phases were higher in the iPSC-CM group. The number of apoptotic cells also decreased in the iPSC-CM group. In comparison to the control cultures, iPSC-CM facilitated cell migration, and these cells showed better barrier functions after several passages. The mechanism of cell proliferation mediated by iPSC-CM was also investigated, and phosphorylation of Akt was observed in B-CECs after exposure to iPSC-CM and showed sustained phosphorylation induced for up to 180 min in iPSC-CM. Our findings indicate that iPSC-CM may employ PI3-kinase signaling in regulating cell cycle progression, which can lead to enhanced cellular proliferation. Effective component analysis of the CM showed that in the iPSC-CM group, the expression of activin-A was significantly increased. If activin-A is added as a supplement, it could help to maintain the morphology of the cells, similar to that of CM. Hence, we conclude that activin-A is one of the effective components of CM in promoting cell proliferation and maintaining cell morphology.

Endostatin Inhibits VEGF-Induced Lung Edema by Reducing Vascular Permeability.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2614-2614
Author(s):  
Gunter Schuch ◽  
Suleyman Erguen ◽  
Shay Soker ◽  
Dieter K. Hossfeld ◽  
Walter Fiedler
Keyword(s):  
Endothelial Cells ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Vascular Permeability ◽  
Pulmonary Hemorrhage ◽  
Inflammatory Cells ◽  
Endothelial Cell Proliferation ◽  
Lung Edema ◽  
Vegf Receptor

Abstract INTRODUCTION: Acute lung injury is a severe condition developing in patients with acute sepsis characterized by lung edema with extravasation of plasma proteins, infiltration by inflammatory cells and pulmonary hemorrhage. Vascular endothelial growth factor (VEGF), also known as vascular permeability factor, is known to not only promote angiogenesis but also increase vascular permeability and has been linked to pathological conditions like retinopathy and acute lung injury. Patients with sepsis and acute lung injury have increased VEGF levels in their plasma. As shown recently, mice treated with VEGF develop histological changes comparable to acute lung injury in septic patients. Endostatin (ES), a endogenous inhibitor of angiogenesis, has been shown to inhibit tumor growth in various models. The mechanisms by which ES inhibits endothelial cell proliferation and function are not clear, yet. It was proposed that one mechanism is the inhibition of VEGF-induced activation of VEGF receptor 2 (VEGFR-2). This study was performed in order to examine whether ES may antagonize VEGF-induced effects leading to increased permeability and lung injury. METHODS: We established an in vitro permeability model using transwell chambers with human dermal microvascular endothelial cells (HDMEC) seeded in the upper chamber on a porous membrane. VEGF-induced permeability was determined by measuring methylene blue to the lower chamber. In order to test the effect of ES on VEGFR-2 activation porcine aortic endothelial cells expressing human VEGFR-2 were incubated with 50 ng/ml VEGF with or without 5 ug/ml ES for 30 min ice. Cell lysate was precipitated with conacavalin A, separated by SDS-PAGE, blotted and analyzed for KDR phosphorylation using phosphotyrosine specific and KDR antibodies. We generated cells with strong expresssion of either VEGF or mouse ES. The cells were encapsulated in alginate beads and injected s.c. to SCID mice divided in the following groups: A) control, B) VEGF, C) ES, D) VEGF + ES with 5 animals per group. After 5 days lungs were harvested and analyzed by H&E staining of tissue sections. RESULTS: In an in vitro permeability assay VEGF enhanced permeation of dye through a monolayer of endothelial cells. ES significantly inhibited VEGF induced permeability (fig.1). ES alone had no effect compared to controls. On the molecular level VEGF causes phosphorylation of its receptor VEGFR-2 as seen in VEGFR-2 expressing cells (fig.2). This effect was abolished by coincubation with ES showing a direct antagonism of VEGF signalling by ES. In a SCID mouse model animals were treated with VEGF, ES or the combination of both (fig.3). Animals in the VEGF group developed general edema and lung injury resembling acute lung injury with extravasation, accumulation of inflammatory cells and hemorrhage. The animals treated with a combination of VEGF and ES had less generalized edema. The lung sections showed alterations less pronounced than in the VEGF group. ES alone had no effect. CONCLUSIONS: Our results demonstrate that ES inhibits VEGF-induced permeability by blocking VEGFR-2 activation. ES treatment partially restored VEGF-induced lung injury in vivo. The incomplete inhibition may be due to excess VEGF protein levels. Taken together, VEGF blocking with ES may serve as a useful new treatment option for conditions with increased vascular permeability like acute lung injury.

scholarly journals In vitro biosynthesis of leukemia inhibitory factor/human interleukin for DA cells by human endothelial cells: differential regulation by interleukin-1 alpha and glucocorticoids

Blood ◽  
1995 ◽  
Vol 86 (10) ◽  
pp. 3763-3770 ◽  
Author(s):  
C Grosset ◽  
B Jazwiec ◽  
JL Taupin ◽  
H Liu ◽  
S Richard ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Endothelial Cells ◽  
Leukemia Inhibitory Factor ◽  
Umbilical Vein ◽  
Interleukin 1 ◽  
Suppressive Effect ◽  
Inhibitory Factor ◽  
Basal Secretion ◽  
In Vitro Biosynthesis

Endothelial cell (EC) may represent a major source of cytokines in the bone marrow. In this study we have examined the production and the regulation of the production of leukemia inhibitory factor/human interleukin for DA cells (LIF/HILDA) by EC. Human umbilical vein endothelial cells (HUVEC) were chosen as a working model as they are a well known source of cytokines. These cells secrete LIF/HILDA (90 pg/mL/10(6) cells/48 h) in basal conditions. This secretion is profoundly altered by interleukin-1 alpha (IL-1 alpha). Secretion of LIF/HILDA is increased threefold on stimulation with IL-1 alpha at a concentration of 100 IU/mL. The secreted protein is bioactive as demonstrated by its proliferative effects on DA1a cells. Modulation of the production of LIF/HILDA by glucocorticoids (GC) was also examined. In striking contrast to what was observed for IL-1 alpha, the synthetic GC dexamethasone (DXM) at a concentration of 10(-6) mol/L consistently inhibited the basal secretion of LIF/HILDA by an average of threefold and suppressed the IL-1 alpha-induced increase of the secretion of this cytokine by HUVEC. In an effort to extend results obtained with HUVEC to the bone marrow endothelium, we have also examined the production of LIF/HILDA by human bone marrow endothelial cells (HBMEC). Our study shows that HBMEC are quantitatively a very important source of this cytokine (above 7.25 ng/mL/10(6) cells/48 h) suggesting that they are a major source of LIF/HILDA in the bone marrow. Again, IL-1 alpha proved to be a very potent stimulus for the secretion of LIF/HILDA and synthetic GC such as DXM when used at a concentration of 10(-6) mol/L inhibited by an average of threefold the basal secretion of LIF/HILDA and had suppressive effect on the IL-1 alpha-induced increase of this secretion. The downregulation of LIF/HILDA production in the bone marrow by GC may be important to understand the effects of GC on hematopoiesis.

Modelling Of Influenza A-Induced Acute Lung Injury (ALI) And Repair: The Development Of Novel Ex Vivo And In Vitro Models

2012 ◽  
Author(s):  
Mark H. Almond ◽  
Alastair G. Proudfoot ◽  
Neeltje Van Doremalen ◽  
Mark J. Griffiths ◽  
Wendy S. Barclay
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
Influenza A ◽  
Ex Vivo ◽  
In Vitro Models

scholarly journals Contribution of Connexin Hemichannels to the Pathogenesis of Acute Lung Injury

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Shuaiwei Wang ◽  
Yafang Sun ◽  
Yu Bai ◽  
Nannan Zhou ◽  
Na Chen ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Vascular Endothelial Cells ◽  
High Mobility ◽  
Lavage Fluid ◽  
Alveolar Cells ◽  
Blocking Property

Connexin (Cx) family members form hemichannels (HCs) and gap junctions (GJs). Biological functions of Cx HCs have not been adequately characterized due to the inability to selectively target HCs or GJs. Recently, we developed a 6-mer peptide mimetic (P5) of the first extracellular loop of Cx43 and showed that it can block the permeability of HCs but not GJs formed by Cx43. In this study, we further characterized the HC blocking property of P5 and investigated the role of Cx HCs in acute lung injury (ALI). We found that P5 administration decreased HC permeability, in pulmonary microvascular endothelial cells, HepG2 cells, and even Cx43-deficient astrocytes, which express different sets of Cxs, suggesting that P5 is a broad spectrum Cx HC blocker. In addition, P5 reduced HC permeability of alveolar cells in vivo. Moreover, P5 decreased endotoxin-induced release, by vascular endothelial cells in vitro, of high mobility group box protein 1 (HMGB1), a critical mediator of acute lung injury (ALI), and reduced HMGB1 accumulation in bronchoalveolar lavage fluid (BALF) of mice subjected to intratracheal endotoxin instillation. Furthermore, P5 administration resulted in a significant decrease in the concentrations of ALT, AST, and LDH in the BALF, the accumulation of leukocytes in alveoli, and the mortality rate of mice subjected to ALI. Wright-Giemsa staining showed that P5 caused similar reductions of both neutrophils and monocytes in BALF of ALI mice. Together, these results suggest that Cx HCs mediate HMGB1 release, augment leukocyte recruitment, and contribute to ALI pathology.

Matrix Metalloproteinases And Their Tissue Inhibitors In In-Vitro Models Of Acute Lung Injury

2010 ◽  
Author(s):  
Alexander J.P. Edwards ◽  
Murali Shyamsundar ◽  
Joseph S. Elborn ◽  
Cecilia M. O'Kane ◽  
Danny F. McAuley
Keyword(s):  
Acute Lung Injury ◽  
Matrix Metalloproteinases ◽  
Lung Injury ◽  
In Vitro Models ◽  
Tissue Inhibitors

scholarly journals Mesenchymal Stem Cell-Conditioned Medium Induces Neutrophil Apoptosis Associated with Inhibition of the NF-κB Pathway in Endotoxin-Induced Acute Lung Injury

2019 ◽  
Vol 20 (9) ◽  
pp. 2208 ◽  
Author(s):  
Vincent Yi-Fong Su ◽  
Chi-Shiuan Lin ◽  
Shih-Chieh Hung ◽  
Kuang-Yao Yang
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
Conditioned Medium ◽  
Mouse Lung ◽  
Human Neutrophils ◽  
Neutrophil Apoptosis ◽  
Neutrophil Accumulation ◽  
Mobility Shift

The immunomodulatory effects of mesenchymal stem cells (MSCs) are established. However, the effects of MSCs on neutrophil survival in acute lung injury (ALI) remain unclear. The goal of this study was to investigate the effect of an MSC-conditioned medium (MSC-CM) on neutrophil apoptosis in endotoxin-induced ALI. In this study, an MSC-CM was delivered via tail vein injection to wild-type male C57BL/6 mice 4 h after an intratracheal injection of lipopolysaccharide (LPS). Twenty-four hours later, bronchoalveolar lavage fluid (BALF) and lung tissue were collected to perform histology, immunohistochemistry, apoptosis assay of neutrophil, enzyme-linked immunosorbent assays, and an electrophoretic mobility shift assay. Human neutrophils were also collected from patients with sepsis-induced acute respiratory distress syndrome (ARDS). Human neutrophils were treated in vitro with LPS, with or without subsequent MSC-CM co-treatment, and were then analyzed. Administration of the MSC-CM resulted in a significant attenuation of histopathological changes, the levels of interleukin-6 and macrophage inflammatory protein 2, and neutrophil accumulation in mouse lung tissues of LPS-induced ALI. Additionally, MSC-CM therapy enhanced the apoptosis of BALF neutrophils and reduced the expression of the anti-apoptotic molecules, Bcl-xL and Mcl-1, both in vivo and in vitro experiments. Furthermore, phosphorylated and total levels of nuclear factor (NF)-κB p65 were reduced in lung tissues from LPS + MSC-CM mice. Human MSC-CM also reduced the activity levels of NF-κB and matrix metalloproteinase-9 in the human neutrophils from ARDS patients. Thus, the results of this study suggest that the MSC-CM attenuated LPS-induced ALI by inducing neutrophil apoptosis, associated with inhibition of the NF-κB pathway.

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