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Article ; Online: Circulating Tumor Cell-Neutrophil Tango along the Metastatic Process.

Saini, Massimo / Szczerba, Barbara M / Aceto, Nicola

2019 Volume 79, Issue 24, Page(s) 6067–6073

Abstract: The crosstalk between cancer cells and the immune system is crucial for disease progression and its therapeutic targeting is providing exciting results, in particular with newly developed immune checkpoint inhibitors. Current approaches primarily focus ... ...

Abstract	The crosstalk between cancer cells and the immune system is crucial for disease progression and its therapeutic targeting is providing exciting results, in particular with newly developed immune checkpoint inhibitors. Current approaches primarily focus on cellular interactions occurring between tumor cells and T lymphocytes; however, recent data highlight a crucial role of neutrophils in support of tumor progression and suggest yet unexplored treatment opportunities. In this review, we summarize the current understanding of those interactions that occur between neutrophils and cancer cells, focusing on both protumor and antitumor activities of neutrophils at different stages of cancer progression. These include infiltration of neutrophils into the primary tumor, their interactions with circulating tumor cells (CTC) within the bloodstream, and their involvement in the establishment of a metastatic niche. Additionally, we discuss how further investigation of CTCs and their interacting immune cell partners may point towards novel immune checkpoint inhibition strategies and provide new insights on the efficacy of already existing immunotherapies.
MeSH term(s)	Animals ; Antineoplastic Agents, Immunological/pharmacology ; Antineoplastic Agents, Immunological/therapeutic use ; Cell Communication/drug effects ; Cell Communication/immunology ; Costimulatory and Inhibitory T-Cell Receptors/antagonists & inhibitors ; Costimulatory and Inhibitory T-Cell Receptors/immunology ; Costimulatory and Inhibitory T-Cell Receptors/metabolism ; Disease Models, Animal ; Disease Progression ; Humans ; Neoplasm Metastasis/immunology ; Neoplasm Metastasis/prevention & control ; Neoplasms/drug therapy ; Neoplasms/immunology ; Neoplasms/pathology ; Neoplastic Cells, Circulating/immunology ; Neoplastic Cells, Circulating/metabolism ; Neutrophil Infiltration ; Neutrophils/drug effects ; Neutrophils/immunology ; Neutrophils/metabolism ; T-Lymphocytes/drug effects ; T-Lymphocytes/immunology ; T-Lymphocytes/metabolism
Chemical Substances	Antineoplastic Agents, Immunological ; Costimulatory and Inhibitory T-Cell Receptors
Language	English
Publishing date	2019-09-16
Publishing country	United States
Document type	Journal Article ; Review
ZDB-ID	1432-1
ISSN	1538-7445 ; 0008-5472
ISSN (online)	1538-7445
ISSN	0008-5472
DOI	10.1158/0008-5472.CAN-19-1972
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Article: Micromanipulation of circulating tumor cells for downstream molecular analysis and metastatic potential assessment

Donato, Cinzia / Szczerba, Barbara M / Scheidmann, Manuel C / Castro-Giner, Francesc / Aceto, Nicola

Journal of visualized experiments. 2019 May 14, , no. 147

2019

Abstract: Blood-borne metastasis accounts for most cancer-related deaths and involves circulating tumor cells (CTCs) that are successful in establishing new tumors at distant sites. CTCs are found in the bloodstream of patients as single cells (single CTCs) or as ... ...

Abstract	Blood-borne metastasis accounts for most cancer-related deaths and involves circulating tumor cells (CTCs) that are successful in establishing new tumors at distant sites. CTCs are found in the bloodstream of patients as single cells (single CTCs) or as multicellular aggregates (CTC clusters and CTC-white blood cell clusters), with the latter displaying a higher metastatic ability. Beyond enumeration, phenotypic and molecular analysis is extraordinarily important to dissect CTC biology and to identify actionable vulnerabilities. Here, we provide a detailed description of a workflow that includes CTC immunostaining and micromanipulation, ex vivo culture to assess proliferative and survival capabilities of individual cells, and in vivo metastasis-formation assays. Additionally, we provide a protocol to achieve the dissociation of CTC clusters into individual cells and the investigation of intra-cluster heterogeneity. With these approaches, for instance, we precisely quantify survival and proliferative potential of single CTCs and individual cells within CTC clusters, leading us to the observation that cells within clusters display better survival and proliferation in ex vivo cultures compared to single CTCs. Overall, our workflow offers a platform to dissect the characteristics of CTCs at the single cell level, aiming towards the identification of metastasis-relevant pathways and a better understanding of CTC biology.
Keywords	blood flow ; metastasis ; neoplasm cells ; neoplasms ; patients ; phenotype
Language	English
Dates of publication	2019-0514
Size	p. e59677.
Publishing place	Journal of Visualized Experiments
Document type	Article
ISSN	1940-087X
DOI	10.3791/59677
Database	NAL-Catalogue (AGRICOLA)

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Article ; Online: Tracing colonic embryonic transcriptional profiles and their reactivation upon intestinal damage.

Fazilaty, Hassan / Brügger, Michael David / Valenta, Tomas / Szczerba, Barbara M / Berkova, Linda / Doumpas, Nikolaos / Hausmann, George / Scharl, Michael / Basler, Konrad

Cell reports

2021 Volume 36, Issue 5, Page(s) 109484

Abstract: We lack a holistic understanding of the genetic programs orchestrating embryonic colon morphogenesis and governing damage response in the adult. A window into these programs is the transcriptomes of the epithelial and mesenchymal cell populations in the ... ...

Abstract	We lack a holistic understanding of the genetic programs orchestrating embryonic colon morphogenesis and governing damage response in the adult. A window into these programs is the transcriptomes of the epithelial and mesenchymal cell populations in the colon. Performing unbiased single-cell transcriptomic analyses of the developing mouse colon at different embryonic stages (embryonic day 14.5 [E14.5], E15.5, and E18.5), we capture cellular and molecular profiles of the stages before, during, and after the appearance of crypt structures, as well as in a model of adult colitis. The data suggest most adult lineages are established by E18.5. We find embryonic-specific gene expression profiles and cell populations that reappear in response to tissue damage. Comparison of the datasets from mice and human colitis suggests the processes are conserved. In this study, we provide a comprehensive single-cell atlas of the developing mouse colon and evidence for the reactivation of embryonic genes in disease.
MeSH term(s)	Animals ; Cell Differentiation ; Colitis/genetics ; Colon/embryology ; Colon/pathology ; Disease Models, Animal ; Embryo, Mammalian/metabolism ; Gene Expression Profiling ; Gene Expression Regulation, Developmental ; Humans ; Inflammatory Bowel Diseases/genetics ; Inflammatory Bowel Diseases/pathology ; Intestinal Mucosa/embryology ; Intestinal Mucosa/metabolism ; Intestinal Mucosa/pathology ; Mesoderm/embryology ; Mice, Inbred C57BL ; Single-Cell Analysis ; Mice
Language	English
Publishing date	2021-08-04
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2649101-1
ISSN	2211-1247 ; 2211-1247
ISSN (online)	2211-1247
ISSN	2211-1247
DOI	10.1016/j.celrep.2021.109484
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Article ; Online: Micromanipulation of Circulating Tumor Cells for Downstream Molecular Analysis and Metastatic Potential Assessment.

Donato, Cinzia / Szczerba, Barbara M / Scheidmann, Manuel C / Castro-Giner, Francesc / Aceto, Nicola

Journal of visualized experiments : JoVE

2019 , Issue 147

Abstract	Blood-borne metastasis accounts for most cancer-related deaths and involves circulating tumor cells (CTCs) that are successful in establishing new tumors at distant sites. CTCs are found in the bloodstream of patients as single cells (single CTCs) or as multicellular aggregates (CTC clusters and CTC-white blood cell clusters), with the latter displaying a higher metastatic ability. Beyond enumeration, phenotypic and molecular analysis is extraordinarily important to dissect CTC biology and to identify actionable vulnerabilities. Here, we provide a detailed description of a workflow that includes CTC immunostaining and micromanipulation, ex vivo culture to assess proliferative and survival capabilities of individual cells, and in vivo metastasis-formation assays. Additionally, we provide a protocol to achieve the dissociation of CTC clusters into individual cells and the investigation of intra-cluster heterogeneity. With these approaches, for instance, we precisely quantify survival and proliferative potential of single CTCs and individual cells within CTC clusters, leading us to the observation that cells within clusters display better survival and proliferation in ex vivo cultures compared to single CTCs. Overall, our workflow offers a platform to dissect the characteristics of CTCs at the single cell level, aiming towards the identification of metastasis-relevant pathways and a better understanding of CTC biology.
MeSH term(s)	Animals ; Humans ; Mice ; Micromanipulation ; Neoplasm Metastasis/diagnosis ; Neoplasm Metastasis/pathology ; Neoplastic Cells, Circulating
Language	English
Publishing date	2019-05-14
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Video-Audio Media
ZDB-ID	2259946-0
ISSN	1940-087X ; 1940-087X
ISSN (online)	1940-087X
ISSN	1940-087X
DOI	10.3791/59677
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: E-cadherin is required for metastasis in multiple models of breast cancer.

Padmanaban, Veena / Krol, Ilona / Suhail, Yasir / Szczerba, Barbara M / Aceto, Nicola / Bader, Joel S / Ewald, Andrew J

Nature

2019 Volume 573, Issue 7774, Page(s) 439–444

Abstract: Metastasis is the major driver of death in patients with cancer. Invasion of surrounding tissues and metastasis have been proposed to initiate following loss of the intercellular adhesion protein, E- ... ...

Abstract	Metastasis is the major driver of death in patients with cancer. Invasion of surrounding tissues and metastasis have been proposed to initiate following loss of the intercellular adhesion protein, E-cadherin
MeSH term(s)	Animals ; Antigens, CD/metabolism ; Breast Neoplasms/metabolism ; Breast Neoplasms/pathology ; Cadherins/metabolism ; Carcinoma, Ductal, Breast/metabolism ; Carcinoma, Ductal, Breast/pathology ; Female ; Humans ; Mice ; Neoplasm Invasiveness ; Neoplasm Metastasis ; Receptors, Transforming Growth Factor beta/metabolism ; Transforming Growth Factor beta/metabolism
Chemical Substances	Antigens, CD ; CDH1 protein, human ; Cadherins ; Receptors, Transforming Growth Factor beta ; Transforming Growth Factor beta
Language	English
Publishing date	2019-09-04
Publishing country	England
Document type	Journal Article
ZDB-ID	120714-3
ISSN	1476-4687 ; 0028-0836
ISSN (online)	1476-4687
ISSN	0028-0836
DOI	10.1038/s41586-019-1526-3
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Article ; Online: T cell epitope mimicry between Sjögren's syndrome Antigen A (SSA)/Ro60 and oral, gut, skin and vaginal bacteria.

Szymula, Agnieszka / Rosenthal, Jacob / Szczerba, Barbara M / Bagavant, Harini / Fu, Shu Man / Deshmukh, Umesh S

Clinical immunology (Orlando, Fla.)

2014 Volume 152, Issue 1-2, Page(s) 1–9

Abstract: This study was undertaken to test the hypothesis that Sjogren's syndrome Antigen A (SSA)/Ro60-reactive T cells are activated by peptides originating from oral and gut bacteria. T cell hybridomas generated from HLA-DR3 transgenic mice recognized 3 regions ...

Abstract	This study was undertaken to test the hypothesis that Sjogren's syndrome Antigen A (SSA)/Ro60-reactive T cells are activated by peptides originating from oral and gut bacteria. T cell hybridomas generated from HLA-DR3 transgenic mice recognized 3 regions on Ro60, with core epitopes mapped to amino acids 228-238, 246-256 and 371-381. BLAST analysis identified several mimicry peptides, originating from human oral, intestinal, skin and vaginal bacteria, as well as environmental bacteria. Amongst these, a peptide from the von Willebrand factor type A domain protein (vWFA) from the oral microbe Capnocytophaga ochracea was the most potent activator. Further, Ro60-reactive T cells were activated by recombinant vWFA protein and whole Escherichia coli expressing this protein. These results demonstrate that peptides derived from normal human microbiota can activate Ro60-reactive T cells. Thus, immune responses to commensal microbiota and opportunistic pathogens should be explored as potential triggers for initiating autoimmunity in SLE and Sjögren's syndrome.
MeSH term(s)	Amino Acid Sequence ; Animals ; Autoimmunity/immunology ; Capnocytophaga/genetics ; Capnocytophaga/immunology ; Cross Reactions/immunology ; Epitopes, T-Lymphocyte/immunology ; Female ; HLA-DR3 Antigen/immunology ; Humans ; Hybridomas/immunology ; Intestines/microbiology ; Lupus Erythematosus, Systemic/immunology ; Lymphocyte Activation/immunology ; Mice ; Molecular Mimicry/immunology ; Mouth/microbiology ; Peptides/genetics ; Peptides/immunology ; Recombinant Proteins/immunology ; Ribonucleoproteins/immunology ; Sjogren's Syndrome/immunology ; Skin/microbiology ; T-Lymphocytes/immunology ; Vagina/microbiology ; von Willebrand Factor/genetics ; von Willebrand Factor/immunology
Chemical Substances	Epitopes, T-Lymphocyte ; HLA-DR3 Antigen ; Peptides ; Recombinant Proteins ; Ribonucleoproteins ; SS-A antigen ; von Willebrand Factor
Language	English
Publishing date	2014-02-19
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	1459903-x
ISSN	1521-7035 ; 1521-6616
ISSN (online)	1521-7035
ISSN	1521-6616
DOI	10.1016/j.clim.2014.02.004
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Article ; Online: An In Vivo CRISPR Screen Identifies Stepwise Genetic Dependencies of Metastatic Progression.

Cancer research

2021 Volume 82, Issue 4, Page(s) 681–694

Abstract: Blood-borne metastasis of breast cancer involves a series of tightly regulated sequential steps, including the growth of a primary tumor lesion, intravasation of circulating tumor cells (CTC), and adaptation in various distant metastatic sites. The genes ...

Abstract	Blood-borne metastasis of breast cancer involves a series of tightly regulated sequential steps, including the growth of a primary tumor lesion, intravasation of circulating tumor cells (CTC), and adaptation in various distant metastatic sites. The genes orchestrating each of these steps are poorly understood in physiologically relevant contexts, owing to the rarity of experimental models that faithfully recapitulate the biology, growth kinetics, and tropism of human breast cancer. Here, we conducted an in vivo loss-of-function CRISPR screen in newly derived CTC xenografts, unique in their ability to spontaneously mirror the human disease, and identified specific genetic dependencies for each step of the metastatic process. Validation experiments revealed sensitivities to inhibitors that are already available, such as PLK1 inhibitors, to prevent CTC intravasation. Together, these findings present a new tool to reclassify driver genes involved in the spread of human cancer, providing insights into the biology of metastasis and paving the way to test targeted treatment approaches. Significance: A loss-of-function CRISPR screen in human CTC-derived xenografts identifies genes critical for individual steps of the metastatic cascade, suggesting novel drivers and treatment opportunities for metastatic breast cancers.
MeSH term(s)	Animals ; Biomarkers, Tumor/genetics ; Biomarkers, Tumor/metabolism ; Breast Neoplasms/blood ; Breast Neoplasms/genetics ; Breast Neoplasms/pathology ; CRISPR-Cas Systems ; Cell Cycle Proteins/genetics ; Cell Cycle Proteins/metabolism ; Cell Line, Tumor ; Clustered Regularly Interspaced Short Palindromic Repeats/genetics ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; Mice, Inbred NOD ; Mice, Knockout ; Mice, SCID ; Neoplasm Metastasis ; Neoplastic Cells, Circulating/metabolism ; Neoplastic Cells, Circulating/pathology ; Protein Serine-Threonine Kinases/genetics ; Protein Serine-Threonine Kinases/metabolism ; Proto-Oncogene Proteins/genetics ; Proto-Oncogene Proteins/metabolism ; RNA, Guide, CRISPR-Cas Systems/genetics ; RNA, Guide, CRISPR-Cas Systems/metabolism ; RNA-Seq/methods ; Survival Analysis ; Xenograft Model Antitumor Assays/methods ; Polo-Like Kinase 1 ; Mice
Chemical Substances	Biomarkers, Tumor ; Cell Cycle Proteins ; Proto-Oncogene Proteins ; RNA, Guide, CRISPR-Cas Systems ; Protein Serine-Threonine Kinases (EC 2.7.11.1)
Language	English
Publishing date	2021-12-13
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1432-1
ISSN	1538-7445 ; 0008-5472
ISSN (online)	1538-7445
ISSN	0008-5472
DOI	10.1158/0008-5472.CAN-21-3908
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Article ; Online: Interaction between innate immunity and Ro52-induced antibody causes Sjögren's syndrome-like disorder in mice.

Szczerba, Barbara M / Kaplonek, Paulina / Wolska, Nina / Podsiadlowska, Anna / Rybakowska, Paulina D / Dey, Paromita / Rasmussen, Astrid / Grundahl, Kiely / Hefner, Kimberly S / Stone, Donald U / Young, Stephen / Lewis, David M / Radfar, Lida / Scofield, R Hal / Sivils, Kathy L / Bagavant, Harini / Deshmukh, Umesh S

Annals of the rheumatic diseases

2016 Volume 75, Issue 3, Page(s) 617–622

Abstract: Objectives: Autoantibodies reactive with Ro52 are often found in sera of patients with Sjögren's syndrome (SS). This study was undertaken to investigate the role of Ro52-induced immune responses in pathogenesis of SS.: Methods: New Zealand Mixed (NZM) ...

Abstract	Objectives: Autoantibodies reactive with Ro52 are often found in sera of patients with Sjögren's syndrome (SS). This study was undertaken to investigate the role of Ro52-induced immune responses in pathogenesis of SS. Methods: New Zealand Mixed (NZM) 2758 mice were immunised with Ro52 in alum adjuvant. Control mice were immunised either with maltose-binding protein or injected with alum alone. Mice were monitored for anti-Ro52 antibody, sialoadenitis and pilocarpine-induced salivation. Antibody binding to salivary gland (SG) cells was analysed in vivo and in vitro by immunofluorescence. Sera from immunised mice were passively transferred into untreated or alum injected NZM2758 mice. Results: By day 30 post-immunisation, Ro52 immunised mice generated immunoprecipitating anti-Ro52 antibodies and they had the maximum drop in saliva production. Both Ro52 immunised and control mice showed evidence of mild sialoadenitis. However, only Ro52 immunised mice had antibody deposition in their SG. Passive transfer of Ro52-immune sera induced SG dysfunction in recipient mice, only if the recipients were primed with alum. In vitro, antibodies from Ro52-immune sera were internalised by a SG cell line and this uptake was inhibited by cytochalasin D treatment. Conclusions: Our data show for the first time that antibodies induced by Ro52 are capable of inducing SG dysfunction, and that this phenomenon is dependent on the activation of innate immunity. The mouse model described in this study implies that autoantibody deposition in the SG might be an important step in the induction of xerostomia and pathogenesis of SS.
MeSH term(s)	Animals ; Autoantibodies/immunology ; Disease Models, Animal ; Humans ; Immunity, Innate/immunology ; Immunization, Passive ; Immunoglobulin G/immunology ; Mice ; Ribonucleoproteins/immunology ; Sialadenitis/immunology ; Sialadenitis/pathology ; Sjogren's Syndrome/immunology ; Sjogren's Syndrome/pathology ; Submandibular Gland/immunology ; Submandibular Gland/pathology
Chemical Substances	Autoantibodies ; Immunoglobulin G ; Ribonucleoproteins ; SS-A antigen
Language	English
Publishing date	2016-03
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	7090-7
ISSN	1468-2060 ; 0003-4967
ISSN (online)	1468-2060
ISSN	0003-4967
DOI	10.1136/annrheumdis-2014-206297
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