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Article ; Online: High-Throughput, Parallel Flow Cytometry Screening of Hundreds of Cell Surface Antigens Using Fluorescent Barcoding.

Drápela, Stanislav / Fedr, Radek / Vacek, Ondřej / Remšík, Ján / Souček, Karel

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

2022 Volume 2543, Page(s) 99–111

Abstract: Multicolor flow cytometry allows for analysis of tens of cellular parameters in millions of cells at a single-cell resolution within minutes. The lack of technologies that would facilitate feasible and relatively cheap profiling of such a number of cells ...

Abstract	Multicolor flow cytometry allows for analysis of tens of cellular parameters in millions of cells at a single-cell resolution within minutes. The lack of technologies that would facilitate feasible and relatively cheap profiling of such a number of cells with an antibody-based approach led us to the development of a high-throughput cytometry-based platform for surface profiling. We coupled the fluorescent cell barcoding with preexisting, commercially available screening tools to analyze cell surface fingerprint at a large scale. This powerful approach will help to identify novel biomarkers and druggable targets and facilitate the discovery of new concepts in immunology, oncology, and developmental biology.
MeSH term(s)	Antigens, Surface ; Biomarkers/analysis ; Flow Cytometry ; Fluorescent Dyes ; Research
Chemical Substances	Antigens, Surface ; Biomarkers ; Fluorescent Dyes
Language	English
Publishing date	2022-09-10
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ISSN	1940-6029
ISSN (online)	1940-6029
DOI	10.1007/978-1-0716-2553-8_9
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Variability of fluorescence intensity distribution measured by flow cytometry is influenced by cell size and cell cycle progression.

Fedr, Radek / Kahounová, Zuzana / Remšík, Ján / Reiterová, Michaela / Kalina, Tomáš / Souček, Karel

Scientific reports

2023 Volume 13, Issue 1, Page(s) 4889

Abstract: The distribution of fluorescence signals measured with flow cytometry can be influenced by several factors, including qualitative and quantitative properties of the used fluorochromes, optical properties of the detection system, as well as the ... ...

Abstract	The distribution of fluorescence signals measured with flow cytometry can be influenced by several factors, including qualitative and quantitative properties of the used fluorochromes, optical properties of the detection system, as well as the variability within the analyzed cell population itself. Most of the single cell samples prepared from in vitrocultures or clinical specimens contain a variable cell cycle component. Cell cycle, together with changes in the cell size, are two of the factors that alter the functional properties of analyzed cells and thus affect the interpretation of obtained results. Here, we describe the association between cell cycle status and cell size, and the variability in the distribution of fluorescence intensity as determined with flow cytometry, at population scale. We show that variability in the distribution of background and specific fluorescence signals is related to the cell cycle state of the selected population, with the 10% low fluorescence signal fraction enriched mainly in cells in their G0/G1 cell cycle phase, and the 10% high fraction containing cells mostly in the G2/M phase. Therefore we advise using caution and additional experimental validation when comparing populations defined by fractions at both ends of fluorescence signal distribution to avoid biases caused by the effect of cell cycle and cell size.
MeSH term(s)	Flow Cytometry/methods ; Cell Division ; Cell Cycle/physiology ; G2 Phase ; Cell Size
Language	English
Publishing date	2023-03-25
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	2615211-3
ISSN	2045-2322 ; 2045-2322
ISSN (online)	2045-2322
ISSN	2045-2322
DOI	10.1038/s41598-023-31990-1
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Article: Phenotypic Heterogeneity of Triple-Negative Breast Cancer Mediated by Epithelial-Mesenchymal Plasticity.

Kvokačková, Barbora / Remšík, Ján / Jolly, Mohit Kumar / Souček, Karel

Cancers

2021 Volume 13, Issue 9

Abstract: Triple-negative breast cancer (TNBC) is a subtype of breast carcinoma known for its unusually aggressive behavior and poor clinical outcome. Besides the lack of molecular targets for therapy and profound intratumoral heterogeneity, the relatively quick ... ...

Abstract	Triple-negative breast cancer (TNBC) is a subtype of breast carcinoma known for its unusually aggressive behavior and poor clinical outcome. Besides the lack of molecular targets for therapy and profound intratumoral heterogeneity, the relatively quick overt metastatic spread remains a major obstacle in effective clinical management. The metastatic colonization of distant sites by primary tumor cells is affected by the microenvironment, epigenetic state of particular subclones, and numerous other factors. One of the most prominent processes contributing to the intratumoral heterogeneity is an epithelial-mesenchymal transition (EMT), an evolutionarily conserved developmental program frequently hijacked by tumor cells, strengthening their motile and invasive features. In response to various intrinsic and extrinsic stimuli, malignant cells can revert the EMT state through the mesenchymal-epithelial transition (MET), a process that is believed to be critical for the establishment of macrometastasis at secondary sites. Notably, cancer cells rarely undergo complete EMT and rather exist in a continuum of E/M intermediate states, preserving high levels of plasticity, as demonstrated in primary tumors and, ultimately, in circulating tumor cells, representing a simplified element of the metastatic cascade. In this review, we focus on cellular drivers underlying EMT/MET phenotypic plasticity and its detrimental consequences in the context of TNBC cancer.
Language	English
Publishing date	2021-05-02
Publishing country	Switzerland
Document type	Journal Article ; Review
ZDB-ID	2527080-1
ISSN	2072-6694
ISSN	2072-6694
DOI	10.3390/cancers13092188
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Article ; Online: Variability of fluorescence intensity distribution measured by flow cytometry is influenced by cell size and cell cycle progression

Radek Fedr / Zuzana Kahounová / Ján Remšík / Michaela Reiterová / Tomáš Kalina / Karel Souček

Scientific Reports, Vol 13, Iss 1, Pp 1-

2023 Volume 13

Abstract: Abstract The distribution of fluorescence signals measured with flow cytometry can be influenced by several factors, including qualitative and quantitative properties of the used fluorochromes, optical properties of the detection system, as well as the ... ...

Abstract	Abstract The distribution of fluorescence signals measured with flow cytometry can be influenced by several factors, including qualitative and quantitative properties of the used fluorochromes, optical properties of the detection system, as well as the variability within the analyzed cell population itself. Most of the single cell samples prepared from in vitrocultures or clinical specimens contain a variable cell cycle component. Cell cycle, together with changes in the cell size, are two of the factors that alter the functional properties of analyzed cells and thus affect the interpretation of obtained results. Here, we describe the association between cell cycle status and cell size, and the variability in the distribution of fluorescence intensity as determined with flow cytometry, at population scale. We show that variability in the distribution of background and specific fluorescence signals is related to the cell cycle state of the selected population, with the 10% low fluorescence signal fraction enriched mainly in cells in their G0/G1 cell cycle phase, and the 10% high fraction containing cells mostly in the G2/M phase. Therefore we advise using caution and additional experimental validation when comparing populations defined by fractions at both ends of fluorescence signal distribution to avoid biases caused by the effect of cell cycle and cell size.
Keywords	Medicine ; R ; Science ; Q
Subject code	612
Language	English
Publishing date	2023-03-01T00:00:00Z
Publisher	Nature Portfolio
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: The covariance environment defines cellular niches for spatial inference.

Haviv, Doron / Remšík, Ján / Gatie, Mohamed / Snopkowski, Catherine / Takizawa, Meril / Pereira, Nathan / Bashkin, John / Jovanovich, Stevan / Nawy, Tal / Chaligne, Ronan / Boire, Adrienne / Hadjantonakis, Anna-Katerina / Pe'er, Dana

Nature biotechnology

2024

Abstract: A key challenge of analyzing data from high-resolution spatial profiling technologies is to suitably represent the features of cellular neighborhoods or niches. Here we introduce the covariance environment (COVET), a representation that leverages the ... ...

Abstract	A key challenge of analyzing data from high-resolution spatial profiling technologies is to suitably represent the features of cellular neighborhoods or niches. Here we introduce the covariance environment (COVET), a representation that leverages the gene-gene covariate structure across cells in the niche to capture the multivariate nature of cellular interactions within it. We define a principled optimal transport-based distance metric between COVET niches that scales to millions of cells. Using COVET to encode spatial context, we developed environmental variational inference (ENVI), a conditional variational autoencoder that jointly embeds spatial and single-cell RNA sequencing data into a latent space. ENVI includes two decoders: one to impute gene expression across the spatial modality and a second to project spatial information onto single-cell data. ENVI can confer spatial context to genomics data from single dissociated cells and outperforms alternatives for imputing gene expression on diverse spatial datasets.
Language	English
Publishing date	2024-04-02
Publishing country	United States
Document type	Journal Article
ZDB-ID	1311932-1
ISSN	1546-1696 ; 1087-0156
ISSN (online)	1546-1696
ISSN	1087-0156
DOI	10.1038/s41587-024-02193-4
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Zs.A 2167: Show issues			Location: Je nach Verfügbarkeit (siehe Angabe bei Bestand) bis Jg. 1994: Bestellungen von Artikeln über das Online-Bestellformular Jg. 1995 - 2021: Lesesall (1.OG) ab Jg. 2022: Lesesaal (EG)
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Article ; Online: Single-cell protein profiling defines cell populations associated with triple-negative breast cancer aggressiveness.

Kvokačková, Barbora / Fedr, Radek / Kužílková, Daniela / Stuchlý, Jan / Vávrová, Adéla / Navrátil, Jiří / Fabian, Pavel / Ondruššek, Róbert / Ovesná, Petra / Remšík, Ján / Bouchal, Jan / Kalina, Tomáš / Souček, Karel

Molecular oncology

2023 Volume 17, Issue 6, Page(s) 1024–1040

Abstract: Triple-negative breast cancer (TNBC) is an aggressive and complex subtype of breast cancer that lacks targeted therapy. TNBC manifests characteristic, extensive intratumoral heterogeneity that promotes disease progression and influences drug response. ... ...

Abstract	Triple-negative breast cancer (TNBC) is an aggressive and complex subtype of breast cancer that lacks targeted therapy. TNBC manifests characteristic, extensive intratumoral heterogeneity that promotes disease progression and influences drug response. Single-cell techniques in combination with next-generation computation provide an unprecedented opportunity to identify molecular events with therapeutic potential. Here, we describe the generation of a comprehensive mass cytometry panel for multiparametric detection of 23 phenotypic markers and 13 signaling molecules. This single-cell proteomic approach allowed us to explore the landscape of TNBC heterogeneity, with particular emphasis on the tumor microenvironment. We prospectively profiled freshly resected tumors from 26 TNBC patients. These tumors contained phenotypically distinct subpopulations of cancer and stromal cells that were associated with the patient's clinical status at the time of surgery. We further classified the epithelial-mesenchymal plasticity of tumor cells, and molecularly defined phenotypically diverse populations of tumor-associated stroma. Furthermore, in a retrospective tissue-microarray TNBC cohort, we showed that the level of CD97 at the time of surgery has prognostic potential.
MeSH term(s)	Humans ; Triple Negative Breast Neoplasms/metabolism ; Proteomics ; Retrospective Studies ; Signal Transduction ; Stromal Cells/metabolism ; Cell Line, Tumor ; Tumor Microenvironment
Language	English
Publishing date	2023-01-25
Publishing country	United States
Document type	Journal Article
ZDB-ID	2415106-3
ISSN	1878-0261 ; 1574-7891
ISSN (online)	1878-0261
ISSN	1574-7891
DOI	10.1002/1878-0261.13365
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Zs.A 6637: Show issues

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Article ; Online: Correction Alternative mechanisms of miR-34a regulation in cancer.

Slabáková, Eva / Culig, Zoran / Remšík, Ján / Souček, Karel

Cell death & disease

2018 Volume 9, Issue 8, Page(s) 783

Abstract: The PDF and HTML versions of the article have been updated to include the Creative Commons Attribution 4.0 International License information. ...

Abstract	The PDF and HTML versions of the article have been updated to include the Creative Commons Attribution 4.0 International License information.
Language	English
Publishing date	2018-07-16
Publishing country	England
Document type	Published Erratum
ZDB-ID	2541626-1
ISSN	2041-4889 ; 2041-4889
ISSN (online)	2041-4889
ISSN	2041-4889
DOI	10.1038/s41419-018-0833-1
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Article: Trop2: Jack of All Trades, Master of None.

Lenárt, Sára / Lenárt, Peter / Šmarda, Jan / Remšík, Ján / Souček, Karel / Beneš, Petr

Cancers

2020 Volume 12, Issue 11

Abstract: Trophoblast cell surface antigen 2 (Trop2) is a widely expressed glycoprotein and an epithelial cell adhesion molecule (EpCAM) family member. Although initially identified as a transmembrane protein, other subcellular localizations and processed forms ... ...

Abstract	Trophoblast cell surface antigen 2 (Trop2) is a widely expressed glycoprotein and an epithelial cell adhesion molecule (EpCAM) family member. Although initially identified as a transmembrane protein, other subcellular localizations and processed forms were described. Its congenital mutations cause a gelatinous drop-like corneal dystrophy, a disease characterized by loss of barrier function in corneal epithelial cells. Trop2 is considered a stem cell marker and its expression associates with regenerative capacity in various tissues. Trop2 overexpression was described in tumors of different origins; however, functional studies revealed both oncogenic and tumor suppressor roles. Nevertheless, therapeutic potential of Trop2 was recognized and clinical studies with drug-antibody conjugates have been initiated in various cancer types. One of these agents, sacituzumab govitecan, has been recently granted an accelerated approval for therapy of metastatic triple-negative breast cancer. In this article, we review the current knowledge about the yet controversial function of Trop2 in homeostasis and pathology.
Language	English
Publishing date	2020-11-11
Publishing country	Switzerland
Document type	Journal Article ; Review
ZDB-ID	2527080-1
ISSN	2072-6694
ISSN	2072-6694
DOI	10.3390/cancers12113328
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Article ; Online: TGF-β regulates Sca-1 expression and plasticity of pre-neoplastic mammary epithelial stem cells.

Remšík, Ján / Pícková, Markéta / Vacek, Ondřej / Fedr, Radek / Binó, Lucia / Hampl, Aleš / Souček, Karel

Scientific reports

2020 Volume 10, Issue 1, Page(s) 11396

Abstract: The epithelial-mesenchymal plasticity, in tight association with stemness, contributes to the mammary gland homeostasis, evolution of early neoplastic lesions and cancer dissemination. Focused on cell surfaceome, we used mouse models of pre-neoplastic ... ...

Abstract	The epithelial-mesenchymal plasticity, in tight association with stemness, contributes to the mammary gland homeostasis, evolution of early neoplastic lesions and cancer dissemination. Focused on cell surfaceome, we used mouse models of pre-neoplastic mammary epithelial and cancer stem cells to reveal the connection between cell surface markers and distinct cell phenotypes. We mechanistically dissected the TGF-β family-driven regulation of Sca-1, one of the most commonly used adult stem cell markers. We further provided evidence that TGF-β disrupts the lineage commitment and promotes the accumulation of tumor-initiating cells in pre-neoplastic cells.
MeSH term(s)	Animals ; Ataxin-1/metabolism ; Breast Neoplasms/genetics ; Breast Neoplasms/pathology ; Cell Line, Tumor/transplantation ; Cell Plasticity/genetics ; Epithelial Cells/pathology ; Epithelial-Mesenchymal Transition/genetics ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; Mammary Glands, Animal/pathology ; Mammary Neoplasms, Experimental/genetics ; Mammary Neoplasms, Experimental/pathology ; Mice ; Neoplastic Stem Cells/pathology ; Receptor, ErbB-2/genetics ; Recombinant Proteins/genetics ; Recombinant Proteins/metabolism ; Signal Transduction/genetics ; Transforming Growth Factor beta/genetics ; Transforming Growth Factor beta/metabolism
Chemical Substances	Ataxin-1 ; Atxn1 protein, mouse ; Recombinant Proteins ; Transforming Growth Factor beta ; Erbb2 protein, mouse (EC 2.7.10.1) ; Receptor, ErbB-2 (EC 2.7.10.1)
Language	English
Publishing date	2020-07-09
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2615211-3
ISSN	2045-2322 ; 2045-2322
ISSN (online)	2045-2322
ISSN	2045-2322
DOI	10.1038/s41598-020-67827-4
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Article ; Online: Alternative mechanisms of miR-34a regulation in cancer.

Slabáková, Eva / Culig, Zoran / Remšík, Ján / Souček, Karel

Cell death & disease

2017 Volume 8, Issue 10, Page(s) e3100

Abstract: MicroRNA miR-34a is recognized as a master regulator of tumor suppression. The strategy of miR-34a replacement has been investigated in clinical trials as the first attempt of miRNA application in cancer treatment. However, emerging outcomes promote the ... ...

Abstract	MicroRNA miR-34a is recognized as a master regulator of tumor suppression. The strategy of miR-34a replacement has been investigated in clinical trials as the first attempt of miRNA application in cancer treatment. However, emerging outcomes promote the re-evaluation of existing knowledge and urge the need for better understanding the complex biological role of miR-34a. The targets of miR-34a encompass numerous regulators of cancer cell proliferation, survival and resistance to therapy. MiR-34a expression is transcriptionally controlled by p53, a crucial tumor suppressor pathway, often disrupted in cancer. Moreover, miR-34a abundance is fine-tuned by context-dependent feedback loops. The function and effects of exogenously delivered or re-expressed miR-34a on the background of defective p53 therefore remain prominent issues in miR-34a based therapy. In this work, we review p53-independent mechanisms regulating the expression of miR-34a. Aside from molecules directly interacting with MIR34A promoter, processes affecting epigenetic regulation and miRNA maturation are discussed. Multiple mechanisms operate in the context of cancer-associated phenomena, such as aberrant oncogene signaling, EMT or inflammation. Since p53-dependent tumor-suppressive mechanisms are disturbed in a substantial proportion of malignancies, we summarize the effects of miR-34a modulation in cell and animal models in the clinically relevant context of disrupted or insufficient p53 function.
MeSH term(s)	Animals ; Epigenesis, Genetic/genetics ; Epithelial-Mesenchymal Transition/genetics ; Gene Expression Regulation, Neoplastic/genetics ; Genes, Tumor Suppressor ; Humans ; MicroRNAs/genetics ; Neoplasms/genetics ; Neoplasms/pathology ; Promoter Regions, Genetic/genetics ; Tumor Suppressor Protein p53/genetics
Chemical Substances	MIRN34 microRNA, human ; MicroRNAs ; TP53 protein, human ; Tumor Suppressor Protein p53
Language	English
Publishing date	2017-10-12
Publishing country	England
Document type	Journal Article ; Review ; Research Support, Non-U.S. Gov't
ZDB-ID	2541626-1
ISSN	2041-4889 ; 2041-4889
ISSN (online)	2041-4889
ISSN	2041-4889
DOI	10.1038/cddis.2017.495
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