Microvascular endothelial model that can predict the inflammatory responses in human microvessels with high reproducibility: Culture model of an hCMEC/D3 cell line fabricated in a collagen vitrigel membrane chamber

In this study, we investigated the production of endothelin 1 (ET-1) by a human microvascular endothelial cell line, HMEC-1, co-cultured with Plasmodium falciparum-parasitized red blood cells (pRBCs). The results indicate that hypoxia increased the basal level of ET-1 production by HMEC-1 cells after 24 or 48h of treatment. However, the co-incubation of HMEC-1 cells with pRBCs, but not with uninfected RBCs, induced a dose-dependent decrease of both constitutive and hypoxia-induced ET-1 production. The inhibition was not due to a decrease in cell viability, as lactate dehydrogenase release remained constant. These results indicate that pRBCs are able to interfere with both the constitutive and stimulated ET-1 release from the microvascular endothelium, thus inducing local modifications of the vascular tone and of the inflammatory response. This could be of relevance in the pathogenesis of the most severe forms of P. falciparum infections, such as cerebral malaria or malaria during pregnancy.

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A Tissue-Engineered Tracheobronchial In Vitro Co-Culture Model for Determining Epithelial Toxicological and Inflammatory Responses

Biomedicines ◽

10.3390/biomedicines9060631 ◽

2021 ◽

Vol 9 (6) ◽

pp. 631

Author(s):

Luis Soriano ◽

Tehreem Khalid ◽

Fergal J. O'Brien ◽

Cian O'Leary ◽

Sally-Ann Cryan

Keyword(s):

Epithelial Cells ◽

Inflammatory Responses ◽

Bronchial Epithelial Cells ◽

Lung Fibroblasts ◽

Drug Induced ◽

Bronchial Epithelial ◽

Culture Model ◽

Epithelial Cultures

Translation of novel inhalable therapies for respiratory diseases is hampered due to the lack of in vitro cell models that reflect the complexity of native tissue, resulting in many novel drugs and formulations failing to progress beyond preclinical assessments. The development of physiologically-representative tracheobronchial tissue analogues has the potential to improve the translation of new treatments by more accurately reflecting in vivo respiratory pharmacological and toxicological responses. Herein, advanced tissue-engineered collagen hyaluronic acid bilayered scaffolds (CHyA-B) previously developed within our group were used to evaluate bacterial and drug-induced toxicity and inflammation for the first time. Calu-3 bronchial epithelial cells and Wi38 lung fibroblasts were grown on either CHyA-B scaffolds (3D) or Transwell® inserts (2D) under air liquid interface (ALI) conditions. Toxicological and inflammatory responses from epithelial monocultures and co-cultures grown in 2D or 3D were compared, using lipopolysaccharide (LPS) and bleomycin challenges to induce bacterial and drug responses in vitro. The 3D in vitro model exhibited significant epithelial barrier formation that was maintained upon introduction of co-culture conditions. Barrier integrity showed differential recovery in CHyA-B and Transwell® epithelial cultures. Basolateral secretion of pro-inflammatory cytokines to bacterial challenge was found to be higher from cells grown in 3D compared to 2D. In addition, higher cytotoxicity and increased basolateral levels of cytokines were detected when epithelial cultures grown in 3D were challenged with bleomycin. CHyA-B scaffolds support the growth and differentiation of bronchial epithelial cells in a 3D co-culture model with different transepithelial resistance in comparison to the same co-cultures grown on Transwell® inserts. Epithelial cultures in an extracellular matrix like environment show distinct responses in cytokine release and metabolic activity compared to 2D polarised models, which better mimic in vivo response to toxic and inflammatory stimuli offering an innovative in vitro platform for respiratory drug development.

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S100A8, S100A9 and the S100A8/A9 heterodimer complex specifically bind to human endothelial cells: identification and characterization of ligands for the myeloid-related proteins S100A9 and S100A8/A9 on human dermal microvascular endothelial cell line-1 cells

International Immunology ◽

10.1093/intimm/14.3.287 ◽

2002 ◽

Vol 14 (3) ◽

pp. 287-297 ◽

Cited By ~ 22

Author(s):

Ines Eue ◽

Simone König ◽

Jolanthe Pior ◽

Clemens Sorg

Keyword(s):

Endothelial Cells ◽

Endothelial Cell ◽

Cell Line ◽

Microvascular Endothelial Cell ◽

Human Endothelial Cells ◽

Endothelial Cell Line ◽

Microvascular Endothelial ◽

Related Proteins ◽

Identification And Characterization

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Epithelial Cell Line Derived from Endometriotic Lesion Mimics Macrophage Nervous Mechanism of Pain Generation on Proteome and Metabolome Levels

Biomolecules ◽

10.3390/biom11081230 ◽

2021 ◽

Vol 11 (8) ◽

pp. 1230

Author(s):

Benjamin Neuditschko ◽

Marlene Leibetseder ◽

Julia Brunmair ◽

Gerhard Hagn ◽

Lukas Skos ◽

...

Keyword(s):

Cell Line ◽

Specific Protein ◽

Reproductive Age ◽

Epithelial Cell Line ◽

Proteomic Profiling ◽

Metabolomic Profiling ◽

Pelvic Area ◽

Culture Model ◽

Neutrophil Aggregation ◽

Cardinal Symptoms

Endometriosis is a benign disease affecting one in ten women of reproductive age worldwide. Although the pain level is not correlated to the extent of the disease, it is still one of the cardinal symptoms strongly affecting the patients’ quality of life. Yet, a molecular mechanism of this pathology, including the formation of pain, remains to be defined. Recent studies have indicated a close interaction between newly generated nerve cells and macrophages, leading to neurogenic inflammation in the pelvic area. In this context, the responsiveness of an endometriotic cell culture model was characterized upon inflammatory stimulation by employing a multi-omics approach, including proteomics, metabolomics and eicosanoid analysis. Differential proteomic profiling of the 12-Z endometriotic cell line treated with TNFα and IL1β unexpectedly showed that the inflammatory stimulation was able to induce a protein signature associated with neuroangiogenesis, specifically including neuropilins (NRP1/2). Untargeted metabolomic profiling in the same setup further revealed that the endometriotic cells were capable of the autonomous production of 7,8-dihydrobiopterin (BH2), 7,8-dihydroneopterin, normetanephrine and epinephrine. These metabolites are related to the development of neuropathic pain and the former three were found up-regulated upon inflammatory stimulation. Additionally, 12-Z cells were found to secrete the mono-oxygenated oxylipin 16-HETE, a known inhibitor of neutrophil aggregation and adhesion. Thus, inflammatory stimulation of endometriotic 12-Z cells led to specific protein and metabolite expression changes suggesting a direct involvement of these epithelial-like cells in endometriosis pain development.

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Isolation of Primary Mouse Pulmonary Microvascular Endothelial Cells and Generation of an Immortalized Cell Line to Obtain Sufficient Extracellular Vesicles

Frontiers in Immunology ◽

10.3389/fimmu.2021.759176 ◽

2021 ◽

Vol 12 ◽

Author(s):

Xu Liu ◽

Feiping Xia ◽

Xiao Wu ◽

Ying Tang ◽

Lu Wang ◽

...

Keyword(s):

Endothelial Cells ◽

Cell Line ◽

Extracellular Vesicles ◽

Phenotypic Expression ◽

Microvascular Endothelial Cells ◽

Pulmonary Microvascular Endothelial Cells ◽

Immortalized Cell Line ◽

Primary Mouse ◽

Microvascular Endothelial ◽

Immortalized Cell

Pulmonary microvascular endothelial cells (PMECs) and the extracellular vesicles (EVs) derived from PMECs participate in maintaining pulmonary homeostasis and mediating the inflammatory response. However, obtaining a high-purity population of PMECs and their EVs from mouse is still notoriously difficult. Herein we provide a method to isolate primary mouse PMECs (pMPMECs) and to transduce SV40 lentivirus into pMPMECs to establish an immortalized cell line (iMPMECs), which provides sufficient quantities of EVs for further studies. pMPMECs and iMPMECs can be identified using morphologic criteria, a phenotypic expression profile (e.g., CD31, CD144, G. simplicifolia lectin binding), and functional properties (e.g., Dil-acetylated low-density protein uptake, Matrigel angiogenesis). Furthermore, pMPMEC–EVs and iMPMEC–EVs can be identified and compared. The characteristics of pMPMEC–EVs and iMPMEC–EVs are ascertained by transmission electron microscopy, nanoparticle tracking analysis, and specific protein markers. iMPMECs produce far more EVs than pMPMECs, while their particle size distribution is similar. Our detailed protocol to isolate and immortalize MPMECs will provide researchers with an in vitro model to investigate the specific roles of EVs in pulmonary physiology and diseases.

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