scholarly journals An Orthotopic Model of Uveal Melanoma in Zebrafish Embryo: A Novel Platform for Drug Evaluation

Biomedicines ◽  
2021 ◽  
Vol 9 (12) ◽  
pp. 1873
Author(s):  
Chiara Tobia ◽  
Daniela Coltrini ◽  
Roberto Ronca ◽  
Alessandra Loda ◽  
Jessica Guerra ◽  
...  
Keyword(s):  
Uveal Melanoma ◽  
Zebrafish Embryo ◽  
Drug Evaluation ◽  
Metastatic Tumor ◽  
Melanoma Cells ◽  
Response To Therapy ◽  
Neoplastic Disease ◽  
Zebrafish Embryos ◽  
Orthotopic Model

Uveal melanoma is a highly metastatic tumor, representing the most common primary intraocular malignancy in adults. Tumor cell xenografts in zebrafish embryos may provide the opportunity to study in vivo different aspects of the neoplastic disease and its response to therapy. Here, we established an orthotopic model of uveal melanoma in zebrafish by injecting highly metastatic murine B16-BL6 and B16-LS9 melanoma cells, human A375M melanoma cells, and human 92.1 uveal melanoma cells into the eye of zebrafish embryos in the proximity of the developing choroidal vasculature. Immunohistochemical and immunofluorescence analyses showed that melanoma cells proliferate during the first four days after injection and move towards the eye surface. Moreover, bioluminescence analysis of luciferase-expressing human 92.1 uveal melanoma cells allowed the quantitative assessment of the antitumor activity exerted by the canonical chemotherapeutic drugs paclitaxel, panobinostat, and everolimus after their injection into the grafted eye. Altogether, our data demonstrate that the zebrafish embryo eye is a permissive environment for the growth of invasive cutaneous and uveal melanoma cells. In addition, we have established a new luciferase-based in vivo orthotopic model that allows the quantification of human uveal melanoma cells engrafted in the zebrafish embryo eye, and which may represent a suitable tool for the screening of novel drug candidates for uveal melanoma therapy.

scholarly journals Melanoma-specific bcl-2 promotes a protumoral M2-like phenotype by tumor-associated macrophages

2020 ◽  
Vol 8 (1) ◽  
pp. e000489 ◽  
Author(s):  
Marta Di Martile ◽  
Valentina Farini ◽  
Francesca Maria Consonni ◽  
Daniela Trisciuoglio ◽  
Marianna Desideri ◽  
...  
Keyword(s):  
Tumor Microenvironment ◽  
Tumor Progression ◽  
Tumor Cells ◽  
Macrophage Polarization ◽  
Melanoma Cells ◽  
Response To Therapy ◽  
Major Constituent ◽  
M2 Macrophage ◽  
Tumor Associated Macrophages

BackgroundA bidirectional crosstalk between tumor cells and the surrounding microenvironment contributes to tumor progression and response to therapy. Our previous studies have demonstrated that bcl-2 affects melanoma progression and regulates the tumor microenvironment. The aim of this study was to evaluate whether bcl-2 expression in melanoma cells could influence tumor-promoting functions of tumor-associated macrophages, a major constituent of the tumor microenvironment that affects anticancer immunity favoring tumor progression.MethodsTHP-1 monocytic cells, monocyte-derived macrophages and melanoma cells expressing different levels of bcl-2 protein were used. ELISA, qRT-PCR and Western blot analyses were used to evaluate macrophage polarization markers and protein expression levels. Chromatin immunoprecipitation assay was performed to evaluate transcription factor recruitment at specific promoters. Boyden chamber was used for migration experiments. Cytofluorimetric and immunohistochemical analyses were carried out to evaluate infiltrating macrophages and T cells in melanoma specimens from patients or mice.ResultsHigher production of tumor-promoting and chemotactic factors, and M2-polarized activation was observed when macrophages were exposed to culture media from melanoma cells overexpressing bcl-2, while bcl-2 silencing in melanoma cells inhibited the M2 macrophage polarization. In agreement, the number of melanoma-infiltrating macrophages in vivo was increased, in parallel with a greater expression of bcl-2 in tumor cells. Tumor-derived interleukin-1β has been identified as the effector cytokine of bcl-2-dependent macrophage reprogramming, according to reduced tumor growth, decreased number of M2-polarized tumor-associated macrophages and increased number of infiltrating CD4+IFNγ+and CD8+IFNγ+effector T lymphocytes, which we observed in response to in vivo treatment with the IL-1 receptor antagonist kineret. Finally, in tumor specimens from patients with melanoma, high bcl-2 expression correlated with increased infiltration of M2-polarized CD163+macrophages, hence supporting the clinical relevance of the crosstalk between tumor cells and microenvironment.ConclusionsTaken together, our results show that melanoma-specific bcl-2 controls an IL-1β-driven axis of macrophage diversion that establishes tumor microenvironmental conditions favoring melanoma development. Interfering with this pathway might provide novel therapeutic strategies.

scholarly journals Suppression of long noncoding RNA MALAT1 inhibits the development of uveal melanoma via microRNA-608-mediated inhibition of HOXC4

2020 ◽  
Vol 318 (5) ◽  
pp. C903-C912 ◽  
Author(s):  
Shuai Wu ◽  
Han Chen ◽  
Ling Zuo ◽  
Hai Jiang ◽  
Hongtao Yan
Keyword(s):  
Cell Cycle ◽  
Uveal Melanoma ◽  
Noncoding Rna ◽  
Cell Cycle Progression ◽  
Melanoma Cells ◽  
Cell Cycle Distribution ◽  
Invasion And Migration ◽  
And Migration

This study explored the effects of the metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) on the development of uveal melanoma. Moreover, the role of the MALAT1/microRNA-608 (miR-608)/homeobox C4 (HOXC4) axis was assessed by evaluating the proliferation, invasion, and migration, as well as the cell cycle distribution of uveal melanoma in vitro after knocking down MALAT1 or HOXC4 and/or overexpression of miR-608 in uveal melanoma cells (MUM-2B and C918). Moreover, the effects of the MALAT1/miR-608/HOXC4 axis in uveal melanoma in vivo were further evaluated by injecting the C918 cells into the NOD/SCID mice. HOXC4 was found to be a gene upregulated in uveal melanoma, while knockdown of its expression resulted in suppression of uveal melanoma cell migration, proliferation, and invasion, as well as cell cycle progression. In addition, the upregulation of miR-608 reduced the expression of HOXC4 in the uveal melanoma cells, which was rescued by overexpression of MALAT1. Hence, MALAT1 could upregulate the HOXC4 by binding to miR-608. The suppressed progression of uveal melanoma in vitro by miR-608 was rescued by overexpression of MALAT1. Additionally, in vivo assays demonstrated that downregulation of MALAT1 could suppress tumor growth through downregulation of HOXC4 expression via increasing miR-608 in uveal melanoma. In summary, MALAT1 downregulation functions to restrain the development of uveal melanoma via miR-608-mediated inhibition of HOXC4.

scholarly journals Decreased Expression of HLA Class II Antigens on Human Uveal Melanoma Cells After In Vivo X-ray Irradiation

1988 ◽  
Vol 105 (1) ◽  
pp. 78-86 ◽  
Author(s):  
Martine J. Jager ◽  
Jeroen P. van der Pol ◽  
Didi de Wolff-Rouendaal ◽  
Paul T.V.M. de Jong ◽  
Dirk J. Ruiter
Keyword(s):  
Uveal Melanoma ◽  
Class Ii ◽  
Melanoma Cells ◽  
Hla Class Ii ◽  
X Ray ◽  
Class Ii Antigens

scholarly journals Signaling of Tumor-Derived sEV Impacts Melanoma Progression

2020 ◽  
Vol 21 (14) ◽  
pp. 5066 ◽  
Author(s):  
Aneta Zebrowska ◽  
Piotr Widlak ◽  
Theresa Whiteside ◽  
Monika Pietrowska
Keyword(s):  
Cancer Progression ◽  
Cancer Biology ◽  
Melanoma Cells ◽  
Response To Therapy ◽  
Host Immune System ◽  
Development Activity ◽  
Tumor Biopsy ◽  
Potential Biomarkers

Small extracellular vesicles (sEV or exosomes) are nanovesicles (30–150 nm) released both in vivo and in vitro by most cell types. Tumor cells produce sEV called TEX and disperse them throughout all body fluids. TEX contain a cargo of proteins, lipids, and RNA that is similar but not identical to that of the “parent” producer cell (i.e., the cargo of exosomes released by melanoma cells is similar but not identical to exosomes released by melanocytes), possibly due to selective endosomal packaging. TEX and their role in cancer biology have been intensively investigated largely due to the possibility that TEX might serve as key component of a “liquid tumor biopsy.” TEX are also involved in the crosstalk between cancer and immune cells and play a key role in the suppression of anti-tumor immune responses, thus contributing to the tumor progression. Most of the available information about the TEX molecular composition and functions has been gained using sEV isolated from supernatants of cancer cell lines. However, newer data linking plasma levels of TEX with cancer progression have focused attention on TEX in the patients’ peripheral circulation as potential biomarkers of cancer diagnosis, development, activity, and response to therapy. Here, we consider the molecular cargo and functions of TEX as potential biomarkers of one of the most fatal malignancies—melanoma. Studies of TEX in plasma of patients with melanoma offer the possibility of an in-depth understanding of the melanoma biology and response to immune therapies. This review features melanoma cell-derived exosomes (MTEX) with special emphasis on exosome-mediated signaling between melanoma cells and the host immune system.

Ce(SO4)2-catalysed the highly diastereoselective synthesis of tetrahydroquinolines via an imino Diels Alder ABB′ type reaction and their in vivo toxicity and imaging in zebrafish embryos

RSC Advances ◽  
2016 ◽  
Vol 6 (44) ◽  
pp. 37478-37486 ◽  
Author(s):  
Carlos A. Martínez Bonilla ◽  
Carlos E. Puerto Galvis ◽  
Leonor Y. Vargas Méndez ◽  
Vladimir V. Kouznetsov
Keyword(s):  
Zebrafish Embryo ◽  
Diels Alder ◽  
Zebrafish Embryos ◽  
Diastereoselective Synthesis ◽  
In Vivo Toxicity ◽  
Type Reaction ◽  
Povarov Reaction

The synthesis of tetrahydroquinolines via the Povarov reaction has been developed using Ce(SO2)4 as a catalyst. This efficient protocol allowed the toxicity and phenotypic study of these products using a zebrafish embryo model.

HLA antigen expression on uveal melanoma cells in vivo and in vitro

1995 ◽  
Vol 44 (2) ◽  
pp. 111-117 ◽  
Author(s):  
Itte de Waard-Siebinga ◽  
Jeanette Kool ◽  
Martine J. Jager
Keyword(s):  
Uveal Melanoma ◽  
Antigen Expression ◽  
Melanoma Cells ◽  
Hla Antigen

scholarly journals Quantifying endothelial cell proliferation in the zebrafish embryo

F1000Research ◽  
2021 ◽  
Vol 10 ◽  
pp. 1032
Author(s):  
George Bowley ◽  
Timothy JA Chico ◽  
Jovana Serbanovic-Canic ◽  
Paul C Evans
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cell ◽  
Zebrafish Embryo ◽  
Time Lapse ◽  
Endothelial Cell Proliferation ◽  
Small Scale ◽  
Zebrafish Embryos ◽  
Time Lapse Imaging

Introduction: Endothelial cell (EC) proliferation is a fundamental determinant of vascular development and homeostasis, and contributes to cardiovascular disease by increasing vascular permeability to blood-borne lipoproteins. Rodents have been traditionally used to analyse EC proliferation mechanisms in vascular health and disease; however, alternative models such as the zebrafish embryo allow researchers to conduct small scale screening studies in a physiologically relevant vasculature whilst reducing the use of mammals in biomedical research. In vitro models of EC proliferation are valuable but do not fully recapitulate the complexity of the in vivo situation. Several groups have used zebrafish embryos for vascular biology research because they offer the advantages of an in vivo model in terms of complexity but are also genetically manipulable and optically transparent. Methods: Here we investigated whether zebrafish embryos can provide a suitable model for the study of EC proliferation. We explored the use of antibody, DNA labelling, and time-lapse imaging approaches. Results: Antibody and DNA labelling approaches were of limited use in zebrafish due to the low rate of EC proliferation combined with the relatively narrow window of time in which they can label proliferating nuclei. By contrast, time-lapse imaging of fluorescent proteins localised to endothelial nuclei was a sensitive method to quantify EC proliferation in zebrafish embryos. Discussion: We conclude that time-lapse imaging is suitable for analysis of endothelial cell proliferation in zebrafish, and that this method is capable of capturing more instances of EC proliferation than immunostaining or cell labelling alternatives. This approach is relevant to anyone studying endothelial cell proliferation for screening genes or small molecules involved in EC proliferation. It offers greater biological relevance than existing in vitro models such as HUVECs culture, whilst reducing the overall number of animals used for this type of research.

scholarly journals Extra-Intestinal Effects of C. difficile Toxin A and B: An In Vivo Study Using the Zebrafish Embryo Model

Cells ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 2575
Author(s):  
Federica Tonon ◽  
Stefano Di Bella ◽  
Gabriele Grassi ◽  
Roberto Luzzati ◽  
Paolo Ascenzi ◽  
...  
Keyword(s):  
Natriuretic Peptide ◽  
Zebrafish Embryo ◽  
Vascular System ◽  
Morphological Changes ◽  
Vital Signs ◽  
Zebrafish Embryos ◽  
Toxin A ◽  
Cardiovascular Damage ◽  
Starting Point

C.difficile infection (CDI) is not a merely “gut-confined” disease as toxemia could drive the development of CDI-related extra-intestinal effects. These effects could explain the high CDI-associated mortality, not just justified by diarrhea and dehydration. Here, the extra-intestinal effects of toxin A (TcdA) and B (TcdB) produced by C. difficile have been studied in vivo using the zebrafish embryo model. Noteworthy, protective properties of human serum albumin (HSA) towards toxins-induced extra-intestinal effects were also addressed. Zebrafish embryos were treated with TcdA, TcdB and/or HSA at 24 h post-fertilization. Embryos were analyzed for 48 h after treatment to check vital signs and morphological changes. Markers related to cardio-vascular damage and inflammation were evaluated by Real-Time quantitative PCR and/or western blotting. Both toxins induced cardiovascular damage in zebrafish embryos by different mechanisms: (i) direct toxicity (i.e., pericardial edema, cardiac chambers enlargement, endothelial alteration); (ii) increased hormonal production and release (i.e., atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP)), (iii) alteration of the vascular system through the increase of the vascular endothelial growth factor (VEGF-A) levels, as well as of its receptors, (iv) pro-inflammatory response through high cytokines production (i.e., CXCL8, IL1B, IL6 and TNFα) and (v) cell-mediated damage due to the increase in neutrophils number. In addition to cardiovascular damage, we observe skin alteration and inflammation. Finally, our data indicate a protective effect of HSA toward the toxins induced extra-intestinal effects. Together, our findings can serve as a starting point for humans’ studies to substantiate and understand the extra-intestinal effects observed in CDI patients.

scholarly journals Antimelanogenic activities of piperlongumine derived from Piper longum on murine B16F10 melanoma cells in vitro and zebrafish embryos in vivo: its molecular mode of depigmenting action

2019 ◽  
Vol 62 (1) ◽  
Author(s):  
Hwang-Ju Jeon ◽  
Kyeongnam Kim ◽  
Yong-Deuk Kim ◽  
Sung-Eun Lee
Keyword(s):  
Melanoma Cells ◽  
Positive Control ◽  
Zebrafish Embryos ◽  
B16f10 Melanoma ◽  
Melanin Production ◽  
Melanocyte Stimulating Hormone ◽  
B16f10 Melanoma Cells ◽  
Pharmaceutical Industries

Abstract In this study, the antimelanogenic activity of piperlongumine in murine B16F10 melanoma cells and zebrafish was investigated, and its mode of antimelanogenic action was elucidated using quantitative reverse transcription-polymerase chain reaction. A melanocyte-stimulating hormone (α-MSH, 200 nM) was used to induce melanin production in B16F10 melanoma cells, and kojic acid (200 μM) was used as a positive control. Piperlongumine had no inhibitory effects on cell growth at the treated concentrations (3 and 6 μM), and it significantly reduced total melanin production. Piperlongumine decreased the expression of Mitf, Tyr, Trp-1, and Trp-2 and tyrosinase activity was also dramatically reduced by the piper amide addition under α-MSH treatment. With these findings, zebrafish embryos were used to confirm antimelanogenic activity of piperlongumine, and it showed the potent antimelanogenic activity at the concentration of 1 μM. Altogether, piperlongumine has potent antimelanogenic activity, and these results support it as a candidate for natural depigmentation agent in a cosmetic and pharmaceutical industries.

scholarly journals In vivo biomolecular imaging of zebrafish embryos using confocal Raman spectroscopy

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Håkon Høgset ◽  
Conor C. Horgan ◽  
James P. K. Armstrong ◽  
Mads S. Bergholt ◽  
Vincenzo Torraca ◽  
...  
Keyword(s):  
Zebrafish Embryo ◽  
Spectroscopic Imaging ◽  
Wound Response ◽  
Three Dimensions ◽  
Zebrafish Embryos ◽  
Biological Processes ◽  
Raman Spectroscopic ◽  
Confocal Raman ◽  
Biomolecular Imaging

AbstractZebrafish embryos provide a unique opportunity to visualize complex biological processes, yet conventional imaging modalities are unable to access intricate biomolecular information without compromising the integrity of the embryos. Here, we report the use of confocal Raman spectroscopic imaging for the visualization and multivariate analysis of biomolecular information extracted from unlabeled zebrafish embryos. We outline broad applications of this method in: (i) visualizing the biomolecular distribution of whole embryos in three dimensions, (ii) resolving anatomical features at subcellular spatial resolution, (iii) biomolecular profiling and discrimination of wild type and ΔRD1 mutant Mycobacterium marinum strains in a zebrafish embryo model of tuberculosis and (iv) in vivo temporal monitoring of the wound response in living zebrafish embryos. Overall, this study demonstrates the application of confocal Raman spectroscopic imaging for the comparative bimolecular analysis of fully intact and living zebrafish embryos.

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