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  1. Article ; Online: Fluorescence-Activated Cell Sorting and Quantitative Real-Time PCR to Reveal VEGF-Expressing Macrophage Populations in the Zebrafish Larvae.

    Herman, Andrew / Greenhough, Alexander / Gurevich, David B

    Methods in molecular biology (Clifton, N.J.)

    2022  Volume 2475, Page(s) 325–337

    Abstract: The transparent, genetically tractable zebrafish is increasingly recognized as a useful model to both live image and uncover mechanistic insight into cell interactions governing tissue homeostasis, pathology, and regeneration. Here, we describe a ... ...

    Abstract The transparent, genetically tractable zebrafish is increasingly recognized as a useful model to both live image and uncover mechanistic insight into cell interactions governing tissue homeostasis, pathology, and regeneration. Here, we describe a protocol for the isolation of macrophages from zebrafish wounds using fluorescence-activated cell sorting (FACS), and the identification of specific pro-angiogenic macrophage populations that express high levels of vascular endothelial growth factor (vegf) using quantitative real-time PCR (qPCR). The cell dissociation and FACS sorting techniques have been optimized for immune cells and successfully used to isolate other fluorescently marked populations within the wound such as neutrophils and endothelial cells. More broadly, this protocol can be easily adapted to other contexts where identification of pro-angiogenic immune cells is transformative for understanding, from development to pathologies such as infection, cancer, and diabetes.
    MeSH term(s) Animals ; Endothelial Cells ; Flow Cytometry/methods ; Larva/genetics ; Macrophages ; Real-Time Polymerase Chain Reaction ; Vascular Endothelial Growth Factor A/genetics ; Zebrafish/genetics
    Chemical Substances Vascular Endothelial Growth Factor A
    Language English
    Publishing date 2022-04-22
    Publishing country United States
    Document type Journal Article
    ISSN 1940-6029
    ISSN (online) 1940-6029
    DOI 10.1007/978-1-0716-2217-9_24
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  2. Article: Interacting Proteins, Polymorphisms and the Susceptibility of Animals to SARS-CoV-2.

    Hancock, John T / Rouse, Ros C / Stone, Emma / Greenhough, Alexander

    Animals : an open access journal from MDPI

    2021  Volume 11, Issue 3

    Abstract: COVID-19, caused by SARS-CoV-2, is a world-wide problem for the human population. It is known that some animal species, such as mink, can become infected and transmit the virus. However, the susceptibility of most animals is not known. Here, we review ... ...

    Abstract COVID-19, caused by SARS-CoV-2, is a world-wide problem for the human population. It is known that some animal species, such as mink, can become infected and transmit the virus. However, the susceptibility of most animals is not known. Here, we review the use of sequence analysis of the proteins which are known to interact with SARS-CoV-2 as a way to estimate an animal's susceptibility. Although most such work concentrates on the angiotensin-converting enzyme 2 receptor (ACE2), here TMPRSS2 (Transmembrane Serine Protease 2), neuropilin-1 and furin are also considered. Polymorphisms, especially ones which are known to alter viral/host interactions are also discussed. Analysis of ACE2 and TMPRSS2 protein sequences across species suggests this approach may be of some utility in predicting susceptibility; however, this analysis fails to highlight some susceptible animals such as mink. However, combined with observational data which emerges over time about which animals actually become infected, this may, in the future, be a useful tool to assist the management of risks associated with human/animal contact and support conservation and animal welfare measures.
    Language English
    Publishing date 2021-03-12
    Publishing country Switzerland
    Document type Journal Article ; Review
    ZDB-ID 2606558-7
    ISSN 2076-2615
    ISSN 2076-2615
    DOI 10.3390/ani11030797
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  3. Article: Wnt Signaling Is a Major Determinant of Neuroblastoma Cell Lineages.

    Szemes, Marianna / Greenhough, Alexander / Malik, Karim

    Frontiers in molecular neuroscience

    2019  Volume 12, Page(s) 90

    Abstract: The neural crest (NC), which has been referred to as the fourth germ layer, comprises a multipotent cell population which will specify diverse cells and tissues, including craniofacial cartilage and bones, melanocytes, the adrenal medulla and the ... ...

    Abstract The neural crest (NC), which has been referred to as the fourth germ layer, comprises a multipotent cell population which will specify diverse cells and tissues, including craniofacial cartilage and bones, melanocytes, the adrenal medulla and the peripheral nervous system. These cell fates are known to be determined by gene regulatory networks (GRNs) acting at various stages of NC development, such as induction, specification, and migration. Although transcription factor hierarchies and some of their interplay with morphogenetic signaling pathways have been characterized, the full complexity of activities required for regulated development remains uncharted. Deregulation of these pathways may contribute to tumorigenesis, as in the case of neuroblastoma, a frequently lethal embryonic cancer thought to arise from the sympathoadrenal lineage of the NC. In this "Hypothesis and Theory" article, we utilize the next generation sequencing data from neuroblastoma cells and tumors to evaluate the possible influences of Wnt signaling on NC GRNs and on neuroblastoma cell lineages. We propose that Wnt signaling is a major determinant of regulatory networks that underlie mesenchymal/neural crest cell (NCC)-like cell identities through PRRX1 and YAP/TAZ transcription factors. Furthermore, Wnt may also co-operate with Hedgehog signaling in driving proneural differentiation programmes along the adrenergic (ADRN) lineage. Elucidation of Signaling Regulatory Networks can augment and complement GRNs in characterizing cell identities, which may in turn contribute to the design of improved therapeutics tailored to primary and relapsing neuroblastoma.
    Language English
    Publishing date 2019-04-16
    Publishing country Switzerland
    Document type Journal Article
    ZDB-ID 2452967-9
    ISSN 1662-5099
    ISSN 1662-5099
    DOI 10.3389/fnmol.2019.00090
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  4. Article ; Online: Interacting Proteins, Polymorphisms and the Susceptibility of Animals to SARS-CoV-2

    John T. Hancock / Ros C. Rouse / Emma Stone / Alexander Greenhough

    Animals, Vol 11, Iss 797, p

    2021  Volume 797

    Abstract: COVID-19, caused by SARS-CoV-2, is a world-wide problem for the human population. It is known that some animal species, such as mink, can become infected and transmit the virus. However, the susceptibility of most animals is not known. Here, we review ... ...

    Abstract COVID-19, caused by SARS-CoV-2, is a world-wide problem for the human population. It is known that some animal species, such as mink, can become infected and transmit the virus. However, the susceptibility of most animals is not known. Here, we review the use of sequence analysis of the proteins which are known to interact with SARS-CoV-2 as a way to estimate an animal’s susceptibility. Although most such work concentrates on the angiotensin-converting enzyme 2 receptor (ACE2), here TMPRSS2 (Transmembrane Serine Protease 2), neuropilin-1 and furin are also considered. Polymorphisms, especially ones which are known to alter viral/host interactions are also discussed. Analysis of ACE2 and TMPRSS2 protein sequences across species suggests this approach may be of some utility in predicting susceptibility; however, this analysis fails to highlight some susceptible animals such as mink. However, combined with observational data which emerges over time about which animals actually become infected, this may, in the future, be a useful tool to assist the management of risks associated with human/animal contact and support conservation and animal welfare measures.
    Keywords ACE2 ; COVID-19 ; furin ; neuropilin-1 ; polymorphisms ; SARS-CoV-2 ; Veterinary medicine ; SF600-1100 ; Zoology ; QL1-991
    Subject code 630
    Language English
    Publishing date 2021-03-01T00:00:00Z
    Publisher MDPI AG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  5. Article: Transcriptomic Analyses of MYCN-Regulated Genes in Anaplastic Wilms' Tumour Cell Lines Reveals Oncogenic Pathways and Potential Therapeutic Vulnerabilities.

    Szemes, Marianna / Melegh, Zsombor / Bellamy, Jacob / Park, Ji Hyun / Chen, Biyao / Greenhough, Alexander / Catchpoole, Daniel / Malik, Karim

    Cancers

    2021  Volume 13, Issue 4

    Abstract: ... ...

    Abstract The
    Language English
    Publishing date 2021-02-06
    Publishing country Switzerland
    Document type Journal Article
    ZDB-ID 2527080-1
    ISSN 2072-6694
    ISSN 2072-6694
    DOI 10.3390/cancers13040656
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  6. Article ; Online: A Wnt-BMP4 Signaling Axis Induces MSX and NOTCH Proteins and Promotes Growth Suppression and Differentiation in Neuroblastoma.

    Szemes, Marianna / Melegh, Zsombor / Bellamy, Jacob / Greenhough, Alexander / Kollareddy, Madhu / Catchpoole, Daniel / Malik, Karim

    Cells

    2020  Volume 9, Issue 3

    Abstract: The Wnt and bone morphogenetic protein (BMP) signaling pathways are known to be crucial in the development of neural crest lineages, including the sympathetic nervous system. Surprisingly, their role in paediatric neuroblastoma, the prototypic tumor ... ...

    Abstract The Wnt and bone morphogenetic protein (BMP) signaling pathways are known to be crucial in the development of neural crest lineages, including the sympathetic nervous system. Surprisingly, their role in paediatric neuroblastoma, the prototypic tumor arising from this lineage, remains relatively uncharacterised. We previously demonstrated that Wnt/b-catenin signaling can have cell-type-specific effects on neuroblastoma phenotypes, including growth inhibition and differentiation, and that BMP4 mRNA and protein were induced by Wnt3a/Rspo2. In this study, we characterised the phenotypic effects of BMP4 on neuroblastoma cells, demonstrating convergent induction of MSX homeobox transcription factors by Wnt and BMP4 signaling and BMP4-induced growth suppression and differentiation. An immunohistochemical analysis of BMP4 expression in primary neuroblastomas confirms a striking absence of BMP4 in poorly differentiated tumors, in contrast to a high expression in ganglion cells. These results are consistent with a tumor suppressive role for BMP4 in neuroblastoma. RNA sequencing following BMP4 treatment revealed induction of Notch signaling, verified by increases of Notch3 and Hes1 proteins. Together, our data demonstrate, for the first time, Wnt-BMP-Notch signaling crosstalk associated with growth suppression of neuroblastoma.
    MeSH term(s) Bone Morphogenetic Protein 4/metabolism ; Bone Morphogenetic Protein 4/pharmacology ; Cell Differentiation/drug effects ; Cell Line, Tumor ; Cell Proliferation/drug effects ; Gene Expression Regulation, Neoplastic/drug effects ; Homeodomain Proteins/metabolism ; Humans ; MSX1 Transcription Factor/metabolism ; Models, Biological ; Neuroblastoma/genetics ; Neuroblastoma/metabolism ; Neuroblastoma/pathology ; Prognosis ; Receptor, Notch3/metabolism ; Signal Transduction ; Transcriptome/genetics ; Wnt Proteins/metabolism
    Chemical Substances Bone Morphogenetic Protein 4 ; Homeodomain Proteins ; MSX1 Transcription Factor ; MSX1 protein, human ; MSX2 protein ; NOTCH3 protein, human ; Receptor, Notch3 ; Wnt Proteins
    Language English
    Publishing date 2020-03-23
    Publishing country Switzerland
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 2661518-6
    ISSN 2073-4409 ; 2073-4409
    ISSN (online) 2073-4409
    ISSN 2073-4409
    DOI 10.3390/cells9030783
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  7. Article ; Online: BCL‑3 promotes cyclooxygenase‑2/prostaglandin E2 signalling in colorectal cancer.

    Collard, Tracey Jane / Fallatah, Hafsah Mohammed / Greenhough, Alexander / Paraskeva, Christos / Williams, Ann Caroline

    International journal of oncology

    2020  Volume 56, Issue 5, Page(s) 1304–1313

    Abstract: First discovered as an oncogene in leukaemia, recent reports highlight an emerging role for the proto‑oncogene BCL‑3 in solid tumours. Importantly, BCL‑3 expression is upregulated in >30% of colorectal cancer cases and is reported to be associated with a ...

    Abstract First discovered as an oncogene in leukaemia, recent reports highlight an emerging role for the proto‑oncogene BCL‑3 in solid tumours. Importantly, BCL‑3 expression is upregulated in >30% of colorectal cancer cases and is reported to be associated with a poor prognosis. However, the mechanism by which BCL‑3 regulates tumorigenesis in the large intestine is yet to be fully elucidated. In the present study, it was shown for the first time that knocking down BCL‑3 expression suppressed cyclooxygenase‑2 (COX‑2)/prostaglandin E2 (PGE2) signalling in colorectal cancer cells, a pathway known to drive several of the hallmarks of cancer. RNAi‑mediated suppression of BCL‑3 expression decreased COX‑2 expression in colorectal cancer cells both at the mRNA and protein level. This reduction in COX‑2 expression resulted in a significant and functional reduction (30‑50%) in the quantity of pro‑tumorigenic PGE2 produced by the cancer cells, as shown by enzyme linked immunoassays and medium exchange experiments. In addition, inhibition of BCL‑3 expression also significantly suppressed cytokine‑induced (TNF‑α or IL‑1β) COX‑2 expression. Taken together, the results of the present study identified a novel role for BCL‑3 in colorectal cancer and suggested that expression of BCL‑3 may be a key determinant in the COX‑2‑meditated response to inflammatory cytokines in colorectal tumour cells. These results suggest that targeting BCL‑3 to suppress PGE2 synthesis may represent an alternative or complementary approach to using non‑steroidal anti‑inflammatory drugs [(NSAIDs), which inhibit cyclooxygenase activity and suppress the conversion of arachidonic acid to prostaglandin], for prevention and/or recurrence in PGE2‑driven tumorigenesis.
    MeSH term(s) B-Cell Lymphoma 3 Protein/genetics ; B-Cell Lymphoma 3 Protein/metabolism ; Cell Line, Tumor ; Colorectal Neoplasms/genetics ; Colorectal Neoplasms/metabolism ; Cyclooxygenase 2/genetics ; Cyclooxygenase 2/metabolism ; Dinoprostone/metabolism ; Gene Expression Regulation, Neoplastic/drug effects ; Gene Knockdown Techniques ; Humans ; Interleukin-1beta/pharmacology ; Signal Transduction ; Tumor Necrosis Factor-alpha/pharmacology ; Up-Regulation/drug effects
    Chemical Substances B-Cell Lymphoma 3 Protein ; BCL3 protein, human ; Interleukin-1beta ; Tumor Necrosis Factor-alpha ; Cyclooxygenase 2 (EC 1.14.99.1) ; PTGS2 protein, human (EC 1.14.99.1) ; Dinoprostone (K7Q1JQR04M)
    Language English
    Publishing date 2020-03-16
    Publishing country Greece
    Document type Journal Article
    ZDB-ID 1154403-x
    ISSN 1791-2423 ; 1019-6439
    ISSN (online) 1791-2423
    ISSN 1019-6439
    DOI 10.3892/ijo.2020.5013
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  8. Article ; Online: 5-Aminosalicylic acid inhibits stem cell function in human adenoma-derived cells: implications for chemoprophylaxis in colorectal tumorigenesis.

    Dixon, Steven William / Collard, Tracey Jane / Mortensson, Eleanor May Harrisdotter / Legge, Danny Nigel / Chambers, Adam Christian / Greenhough, Alexander / Creed, Tom Julian / Williams, Ann Caroline

    British journal of cancer

    2021  Volume 124, Issue 12, Page(s) 1959–1969

    Abstract: Background: Most colorectal cancers (CRC) arise sporadically from precursor lesions: colonic polyps. Polyp resection prevents progression to CRC. Risk of future polyps is proportional to the number and size of polyps detected at screening, allowing ... ...

    Abstract Background: Most colorectal cancers (CRC) arise sporadically from precursor lesions: colonic polyps. Polyp resection prevents progression to CRC. Risk of future polyps is proportional to the number and size of polyps detected at screening, allowing identification of high-risk individuals who may benefit from effective chemoprophylaxis. We aimed to investigate the potential of 5-aminosalicylic acid (5-ASA), a medication used in the treatment of ulcerative colitis, as a possible preventative agent for sporadic CRC.
    Methods: Human colorectal adenoma (PC/AA/C1, S/AN/C1 and S/RG/C2), transformed adenoma PC/AA/C1/SB10 and carcinoma cell lines (LS174T and SW620) were treated with 5-ASA. The effect on growth in two- and three-dimensional (3D) culture, β-catenin transcriptional activity and on cancer stemness properties of the cells were investigated.
    Results: 5-ASA was shown, in vitro, to inhibit the growth of adenoma cells and suppress β-catenin transcriptional activity. Downregulation of β-catenin was found to repress expression of stem cell marker LGR5 (leucine-rich G protein-coupled receptor-5) and functionally suppress stemness in human adenoma and carcinoma cells using 3D models of tumorigenesis.
    Conclusions: 5-ASA can suppress the cancer stem phenotype in adenoma-derived cells. Affordable and well-tolerated, 5-ASA is an outstanding candidate as a chemoprophylactic medication to reduce the risk of colorectal polyps and CRC in those at high risk.
    MeSH term(s) Adenoma/drug therapy ; Adenoma/genetics ; Adenoma/pathology ; Adenoma/prevention & control ; Carcinogenesis/drug effects ; Carcinogenesis/genetics ; Carcinogenesis/pathology ; Carcinoma/genetics ; Carcinoma/pathology ; Carcinoma/prevention & control ; Cell Line, Tumor ; Cell Transformation, Neoplastic/drug effects ; Cell Transformation, Neoplastic/genetics ; Chemoprevention/methods ; Colitis, Ulcerative/drug therapy ; Colitis, Ulcerative/genetics ; Colitis, Ulcerative/pathology ; Colorectal Neoplasms/drug therapy ; Colorectal Neoplasms/genetics ; Colorectal Neoplasms/pathology ; Colorectal Neoplasms/prevention & control ; Gene Expression Regulation, Neoplastic/drug effects ; Humans ; Mesalamine/pharmacology ; Mesalamine/therapeutic use ; Neoplastic Stem Cells/drug effects ; Neoplastic Stem Cells/physiology ; Wnt Signaling Pathway/drug effects ; Wnt Signaling Pathway/genetics
    Chemical Substances Mesalamine (4Q81I59GXC)
    Language English
    Publishing date 2021-03-30
    Publishing country England
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 80075-2
    ISSN 1532-1827 ; 0007-0920
    ISSN (online) 1532-1827
    ISSN 0007-0920
    DOI 10.1038/s41416-021-01354-5
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    Uh III Zs.142: Show issues Location:
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  9. Article ; Online: Circulating prostaglandins as biomarkers for colorectal cancer?

    Greenhough, Alexander / Paraskeva, Chris / Williams, Ann C

    EBioMedicine

    2015  Volume 2, Issue 2, Page(s) 105–106

    MeSH term(s) Biomarkers, Tumor ; Colorectal Neoplasms ; Humans ; Prostaglandins
    Chemical Substances Biomarkers, Tumor ; Prostaglandins
    Language English
    Publishing date 2015-01-28
    Publishing country Netherlands
    Document type Journal Article ; Comment
    ZDB-ID 2851331-9
    ISSN 2352-3964
    ISSN (online) 2352-3964
    DOI 10.1016/j.ebiom.2015.01.016
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    In stock of ZB MED Cologne/Königswinter

    Zs.A 7090: Show issues Location:
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  10. Article: Interacting Proteins, Polymorphisms and the Susceptibility of Animals to SARS-CoV-2

    Hancock, John T / Rouse, Ros C / Stone, Emma / Greenhough, Alexander

    Animals. 2021 Mar. 12, v. 11, no. 3

    2021  

    Abstract: COVID-19, caused by SARS-CoV-2, is a world-wide problem for the human population. It is known that some animal species, such as mink, can become infected and transmit the virus. However, the susceptibility of most animals is not known. Here, we review ... ...

    Abstract COVID-19, caused by SARS-CoV-2, is a world-wide problem for the human population. It is known that some animal species, such as mink, can become infected and transmit the virus. However, the susceptibility of most animals is not known. Here, we review the use of sequence analysis of the proteins which are known to interact with SARS-CoV-2 as a way to estimate an animal’s susceptibility. Although most such work concentrates on the angiotensin-converting enzyme 2 receptor (ACE2), here TMPRSS2 (Transmembrane Serine Protease 2), neuropilin-1 and furin are also considered. Polymorphisms, especially ones which are known to alter viral/host interactions are also discussed. Analysis of ACE2 and TMPRSS2 protein sequences across species suggests this approach may be of some utility in predicting susceptibility; however, this analysis fails to highlight some susceptible animals such as mink. However, combined with observational data which emerges over time about which animals actually become infected, this may, in the future, be a useful tool to assist the management of risks associated with human/animal contact and support conservation and animal welfare measures.
    Keywords COVID-19 infection ; Severe acute respiratory syndrome coronavirus 2 ; animal welfare ; human population ; humans ; mink ; observational studies ; peptidyl-dipeptidase A ; sequence analysis ; serine proteinases ; viruses
    Language English
    Dates of publication 2021-0312
    Publishing place Multidisciplinary Digital Publishing Institute
    Document type Article
    Note NAL-light
    ISSN 2076-2615
    DOI 10.3390/ani11030797
    Database NAL-Catalogue (AGRICOLA)

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