scholarly journals Tumor-Derived Extracellular Vesicles Induce Abnormal Angiogenesis via TRPV4 Downregulation and Subsequent Activation of YAP and VEGFR2

2021 ◽  
Vol 9 ◽  
Author(s):  
Brianna Guarino ◽  
Venkatesh Katari ◽  
Ravi Adapala ◽  
Neha Bhavnani ◽  
Julie Dougherty ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Extracellular Vesicles ◽  
Tumor Angiogenesis ◽  
Lung Tumor ◽  
Rho Kinase ◽  
Paracrine Factors ◽  
Human Lung Tumor ◽  
Subsequent Activation

Tumor angiogenesis is initiated and maintained by the tumor microenvironment through secretion of autocrine and paracrine factors, including extracellular vesicles (EVs). Although tumor-derived EVs (t-EVs) have been implicated in tumor angiogenesis, growth and metastasis, most studies on t-EVs are focused on proangiogenic miRNAs and growth factors. We have recently demonstrated that conditioned media from human lung tumor cells (A549) downregulate TRPV4 channels and transform normal endothelial cells to a tumor endothelial cell-like phenotype and induce abnormal angiogenesis in vitro, via t-EVs. However, the underlying molecular mechanism of t-EVs on endothelial cell phenotypic transition and abnormal angiogenesis in vivo remains unknown. Here, we demonstrate that t-EVs downregulate TRPV4 expression post-translationally and induce abnormal angiogenesis by activating Rho/Rho kinase/YAP/VEGFR2 pathways. Further, we demonstrate that t-EVs induce abnormal vessel formation in subcutaneously implanted Matrigel plugs in vivo (independent of tumors), which are characterized by increased VEGFR2 expression and reduced pericyte coverage. Taken together, our findings demonstrate that t-EVs induce abnormal angiogenesis via TRPV4 downregulation-mediated activation of Rho/Rho kinase/YAP/VEGFR2 pathways and suggest t-EVs and TRPV4 as novel targets for vascular normalization and cancer therapy.

Carcinoembryonic Antigen (CEA) Regulates Tumor-Angiogenesis in Colon Carcinomas.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1897-1897
Author(s):  
Kira Braemswig ◽  
Marina Poettler ◽  
Wazlawa Kalinowska ◽  
Christoph Zielinski ◽  
Gerald W Prager
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Tumor Angiogenesis ◽  
Endothelial Cell Proliferation ◽  
Erk Phosphorylation ◽  
Dose Dependent Increase ◽  
Dose Dependent

Abstract Human carcinoembryonic antigen (CEA) is a cell surface adhesion molecule member of the Immunoglobulin Superfamily (IgSF). Aberrant upregulation and secretion of soluble CEA is a common feature found in a wide variety of human cancers such as colon, breast and lung. Previous in vitro and in vivo results have demonstrated that CEA can affect tumor cell behavior including the inhibition of cell differentiation and apoptosis. However, any functional effects on angiogenic endothelial cell behavior are so far unknown. In the present work we found that in endothelial cells exogenous CEA led to a time and dose dependent increase in ERK phosphorylation, which was inhibited by the specific MEK inhibitor U0126. Thereby, the observed CEA effect was comparable in time and intense with the canonical angiogenic growth factor VEGF. The CEA-induced ERK phosphorylation was not affected by the blockage of VEGFR-2 / flk-1 using a specific inhibiting peptide (CBO-P11), which indicates a VEGF-independent mechanism. Furthermore, co-stimulation of endothelial cells with VEGF and CEA shows synergistic effects on ERK phosphorylation. While in endothelial cells no endogenous expression of CEA is detected, its putative receptor, the CEA receptor (CEAR), is highly expressed as shown by immunohistochemical staining of paraffin-embedded colon carcinoma sections as well as in biochemical analyses. When an activating antibody against CEAR was used, CEA-induced ERK phosphorylation was mimicked, while downregulation of CEAR by siRNA diminished CEA-induced signal transduction, significantly. To test a biological relevance of our findings, we first measured endothelial cell proliferation: CEA led to a dose dependent increase in endothelial cell proliferation in vitro, which again revealed a synergistic effect with VEGF. Thereby, CEA-induced endothelial cell proliferation was again independent of VEGFR-2 / flk-1. A biological role of CEA in tumor-angiogenesis was reflected by an in vivo model using CEA Mimotope immunized BALB/c mice, which were transplanted with MethA/CEA overexpressing tumor cells. Immunohistological analyses of these tumors revealed a significantly reduced vascular density, which was accompanied with diminished tumor growth. Our data provide first evidence of CEA as a novel pro-angiogenic activator of endothelial cells, which results in an increase in endothelial cell proliferation, independent of VEGFR-2. Furthermore, by targeting CEA in an in vivo mouse model, tumor-angiogenesis was markley reduced, indicating a potential therapeutic target in cancer.

scholarly journals Mesenchymal Stromal Cell-Derived Extracellular Vesicles – Silver Linings for Cartilage Regeneration?

2020 ◽  
Vol 8 ◽  
Author(s):  
Andrea De Luna ◽  
Alexander Otahal ◽  
Stefan Nehrer
Keyword(s):  
Extracellular Vesicles ◽  
Treatment Options ◽  
Cartilage Regeneration ◽  
Bioactive Molecules ◽  
Paracrine Factors ◽  
Differentiation Potential ◽  
Vast Number ◽  
The Impact

As the world’s population is aging, the incidence of the degenerative disease Osteoarthritis (OA) is increasing. Current treatment options of OA focus on the alleviation of the symptoms including pain and inflammation rather than on restoration of the articular cartilage. Cell-based therapies including the application of mesenchymal stromal cells (MSCs) have been a promising tool for cartilage regeneration approaches. Due to their immunomodulatory properties, their differentiation potential into cells of the mesodermal lineage as well as the plurality of sources from which they can be isolated, MSCs have been applied in a vast number of studies focusing on the establishment of new treatment options for Osteoarthritis. Despite promising outcomes in vitro and in vivo, applications of MSCs are connected with teratoma formation, limited lifespan of differentiated cells as well as rejection of the cells after transplantation, highlighting the need for new cell free approaches harboring the beneficial properties of MSCs. It has been demonstrated that the regenerative potential of MSCs is mediated by the release of paracrine factors rather than by differentiation into cells of the desired tissue. Besides soluble factors, extracellular vesicles are the major component of a cell’s secretome. They represent novel mechanisms by which (pathogenic) signals can be communicated between cell types as they deliver bioactive molecules (nucleic acids, proteins, lipids) from the cell of origin to the target cell leading to specific biological processes upon uptake. This review will give an overview about extracellular vesicles including general characteristics, isolation methods and characterization approaches. Furthermore, the role of MSC-derived extracellular vesicles in in vitro and in vivo studies for cartilage regeneration will be summarized with special focus on transported miRNA which either favored the progression of OA or protected the cartilage from degradation. In addition, studies will be reviewed investigating the impact of MSC-derived extracellular vesicles on inflammatory arthritis. As extracellular vesicles are present in all body fluids, their application as potential biomarkers for OA will also be discussed in this review. Finally, studies exploring the combination of MSC-derived extracellular vesicles with biomaterials for tissue engineering approaches are summarized.

scholarly journals A Chemically Defined, Xeno- and Blood-Free Culture Medium Sustains Increased Production of Small Extracellular Vesicles From Mesenchymal Stem Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Aliosha I. Figueroa-Valdés ◽  
Catalina de la Fuente ◽  
Yessia Hidalgo ◽  
Ana María Vega-Letter ◽  
Rafael Tapia-Limonchi ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Large Scale ◽  
Culture Medium ◽  
Culture Media ◽  
Cell Phenotype ◽  
Target Cells ◽  
Scale Production ◽  
Paracrine Factors

Cell therapy is witnessing a notable shift toward cell-free treatments based on paracrine factors, in particular, towards small extracellular vesicles (sEV), that mimic the functional effect of the parental cells. While numerous sEV-based applications are currently in advanced preclinical stages, their promised translation depends on overcoming the manufacturing hurdles posed by the large-scale production of purified sEV. Unquestionably, the culture medium used with the parental cells plays a key role in the sEV’s secretion rate and content. An essential requisite is the use of a serum-, xeno-, and blood-free medium to meet the regulatory entity requirements of clinical-grade sEV’s production. Here, we evaluated OxiumTMEXO, a regulatory complying medium, with respect to production capacity and conservation of the EV’s characteristics and functionality and the parental cell’s phenotype and viability. A comparative study was established with standard DMEM and a commercially available culture medium developed specifically for sEV production. Under similar conditions, OxiumTMEXO displayed a three-fold increase of sEV secretion, with an enrichment of particles ranging between 51 and 200 nm. These results were obtained through direct quantification from the conditioned medium to avoid the isolation method’s interference and variability and were compared to the two culture media under evaluation. The higher yield obtained was consistent with several harvest time points (2, 4, and 6 days) and different cell sources, incluiding umbilical cord-, menstrual blood-derived mesenchymal stromal cells and fibroblasts. Additionally, the stem cell phenotype and viability of the parental cell remained unchanged. Furthermore, OxiumTMEXO-sEV showed a similar expression pattern of the vesicular markers CD63, CD9, and CD81, with respect to sEV derived from the other conditions. The in vitro internalization assays in different target cell types and the pharmacokinetic profile of intraperitoneally administered sEV in vivo indicated that the higher EV production rate did not affect the uptake kinetics or the systemic biodistribution in healthy mice. In conclusion, the OxiumTMEXO medium sustains an efficient and robust production of large quantities of sEV, conserving the classic functional properties of internalization into acceptor target cells and biodistribution in vivo, supplying the amount and quality of EVs for the development of cell-free therapies.

scholarly journals Corneal Endothelial Cell Migration and Proliferation Enhanced by Rho Kinase (ROCK) Inhibitors in In Vitro and In Vivo Models

2016 ◽  
Vol 57 (15) ◽  
pp. 6731 ◽  
Author(s):  
Landon C. Meekins ◽  
Noel Rosado-Adames ◽  
Rupalatha Maddala ◽  
Jiagang J. Zhao ◽  
Ponugoti V. Rao ◽  
...  
Keyword(s):  
Cell Migration ◽  
Endothelial Cell ◽  
Rho Kinase ◽  
Corneal Endothelial Cell ◽  
Endothelial Cell Migration ◽  
In Vivo Models ◽  
Cell Migration And Proliferation

scholarly journals A BMP7 Variant Inhibits Tumor Angiogenesis In Vitro and In Vivo through Direct Modulation of Endothelial Cell Biology

PLoS ONE ◽  
2015 ◽  
Vol 10 (4) ◽  
pp. e0125697 ◽  
Author(s):  
Courtney M. Tate ◽  
Jacquelyn Mc Entire ◽  
Roberto Pallini ◽  
Eliza Vakana ◽  
Lisa Wyss ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Tumor Angiogenesis ◽  
Cell Biology ◽  
Direct Modulation

Endogenous hydrogen sulfide sulfhydrates IKKβ at cysteine 179 to control pulmonary artery endothelial cell inflammation

2019 ◽  
Vol 133 (20) ◽  
pp. 2045-2059 ◽  
Author(s):  
Da Zhang ◽  
Xiuli Wang ◽  
Siyao Chen ◽  
Selena Chen ◽  
Wen Yu ◽  
...  
Keyword(s):  
Hydrogen Sulfide ◽  
Endothelial Cell ◽  
Pulmonary Artery ◽  
Cell Model ◽  
Site Directed Mutagenesis ◽  
Critical Event ◽  
Hypertensive Rats ◽  
Pulmonary Artery Endothelial Cell

Abstract Background: Pulmonary artery endothelial cell (PAEC) inflammation is a critical event in the development of pulmonary arterial hypertension (PAH). However, the pathogenesis of PAEC inflammation remains unclear. Methods: Purified recombinant human inhibitor of κB kinase subunit β (IKKβ) protein, human PAECs and monocrotaline-induced pulmonary hypertensive rats were employed in the study. Site-directed mutagenesis, gene knockdown or overexpression were conducted to manipulate the expression or activity of a target protein. Results: We showed that hydrogen sulfide (H2S) inhibited IKKβ activation in the cell model of human PAEC inflammation induced by monocrotaline pyrrole-stimulation or knockdown of cystathionine γ-lyase (CSE), an H2S generating enzyme. Mechanistically, H2S was proved to inhibit IKKβ activity directly via sulfhydrating IKKβ at cysteinyl residue 179 (C179) in purified recombinant IKKβ protein in vitro, whereas thiol reductant dithiothreitol (DTT) reversed H2S-induced IKKβ inactivation. Furthermore, to demonstrate the significance of IKKβ sulfhydration by H2S in the development of PAEC inflammation, we mutated C179 to serine (C179S) in IKKβ. In purified IKKβ protein, C179S mutation of IKKβ abolished H2S-induced IKKβ sulfhydration and the subsequent IKKβ inactivation. In human PAECs, C179S mutation of IKKβ blocked H2S-inhibited IKKβ activation and PAEC inflammatory response. In pulmonary hypertensive rats, C179S mutation of IKKβ abolished the inhibitory effect of H2S on IKKβ activation and pulmonary vascular inflammation and remodeling. Conclusion: Collectively, our in vivo and in vitro findings demonstrated, for the first time, that endogenous H2S directly inactivated IKKβ via sulfhydrating IKKβ at Cys179 to inhibit nuclear factor-κB (NF-κB) pathway activation and thereby control PAEC inflammation in PAH.

Organische Nitrate und Thrombozytenfunktion

1988 ◽  
Vol 08 (02) ◽  
pp. 90-99 ◽  
Author(s):  
H. Schröder ◽  
K. Schrör
Keyword(s):  
Endothelial Cell ◽  
Angina Pectoris ◽  
Ex Vivo

ZusammenfassungOrganische Nitrate unterschiedlicher chemischer Struktur sowie Nitroprussidnatrium und Molsidomin (bzw. ihre biologisch aktiven Metaboliten) können die (primäre) Aggregation und Sekretion von Humanthrombozyten in vitro und ex vivo hemmen. Eine solche Wirkung wird für Molsidomin (SIN-1) und Nitroprussidnatrium in vitro in Konzentrationen beobachtet, die in der gleichen Größenordnung liegen wie die vasodilatierenden Effekte der Substanzen. Dagegen sind für eine direkte Antiplättchenwirkung organischer Nitrate (Glyzeryltrinitrat, Isosorbiddinitr at, Isosorbidmononitrate, Teopranitol) in vitro Konzentrationen erforderlich, die ca. 100- bis 1000fach höher sind als die Plasmaspiegel der Substanzen nach therapeutischer Dosierung bzw. die Konzentrationen, die isolierte Gefäßstreifen relaxieren. Als gemeinsamer Wirkungsmechanismus der direkten thrombozy-tenfunktionshemmenden und gefäßerweiternden Wirkung all dieser Substanzen kann heute eine Stickoxid-(NO)-vermittelte Stimulation der cGMP-Bildung angenommen werden, das aus organischen Nitraten als »Pro-drug« entsteht. Die Freisetzung von NO, eines »endothelial cell-derived relaxing factors« (EDRF) aus Nitroprussidnatrium und SIN-1 erfolgt spontan. Dagegen erfordert die Freisetzung von NO aus organischen Nitraten einen enzymatischen Stoffwechselweg, der in isolierten Thrombozyten nicht vorhanden ist. Eine Antiplättchenwirkung organischer Nitrate in vivo bzw. ex vivo wird daher über die Stimulation eines endothelialen, thrombozyteninhibitorischen Faktors erklärt. Hierbei sind Prostazyklin sowie ein bisher unbekannter Endothel-zellfaktor neben einer synergistischen Wirkung organischer Nitrate mit endogenem Prostazyklin in Diskussion. Eine thrombozytenfunktionshemmen-de Wirkung organischer Nitrate könnte in Kombination mit ihren hämody-namischen Effekten auch für die an-tianginöse Wirkung in der Klinik bedeutsam sein, insbesondere zur Verhinderung vasospastischer Zustände bei der instabilen Angina pectoris.

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