Multi-Scale Imaging and Informatics Pipeline for In Situ Pluripotent Stem Cell Analysis
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{"title"=>"Multi-scale imaging and informatics pipeline for in situ pluripotent stem cell analysis", "type"=>"journal", "authors"=>[{"first_name"=>"Bryan R.", "last_name"=>"Gorman", "scopus_author_id"=>"55788834600"}, {"first_name"=>"Junjie", "last_name"=>"Lu", "scopus_author_id"=>"56076235600"}, {"first_name"=>"Anna", "last_name"=>"Baccei", "scopus_author_id"=>"55788087800"}, {"first_name"=>"Nathan C.", "last_name"=>"Lowry", "scopus_author_id"=>"36840063800"}, {"first_name"=>"Jeremy E.", "last_name"=>"Purvis", "scopus_author_id"=>"24400118300"}, {"first_name"=>"Rami S.", "last_name"=>"Mangoubi", "scopus_author_id"=>"6602248344"}, {"first_name"=>"Paul H.", "last_name"=>"Lerou", "scopus_author_id"=>"9039830200"}], "year"=>2014, "source"=>"PLoS ONE", "identifiers"=>{"sgr"=>"84920462707", "doi"=>"10.1371/journal.pone.0116037", "issn"=>"19326203", "pui"=>"601108050", "isbn"=>"1932-6203 (Electronic)\\r1932-6203 (Linking)", "pmid"=>"25551762", "scopus"=>"2-s2.0-84920462707"}, "id"=>"42807949-a408-30da-aed5-753586916ca5", "abstract"=>"Human pluripotent stem (hPS) cells are a potential source of cells for medical therapy and an ideal system to study fate decisions in early development. However, hPS cells cultured in vitro exhibit a high degree of heterogeneity, presenting an obstacle to clinical translation. hPS cells grow in spatially patterned colony structures, necessitating quantitative single-cell image analysis. We offer a tool for analyzing the spatial population context of hPS cells that integrates automated fluorescent microscopy with an analysis pipeline. It enables high-throughput detection of colonies at low resolution, with single-cellular and sub-cellular analysis at high resolutions, generating seamless in situ maps of single-cellular data organized by colony. We demonstrate the tool's utility by analyzing inter- and intra-colony heterogeneity of hPS cell cycle regulation and pluripotency marker expression. We measured the heterogeneity within individual colonies by analyzing cell cycle as a function of distance. Cells loosely associated with the outside of the colony are more likely to be in G1, reflecting a less pluripotent state, while cells within the first pluripotent layer are more likely to be in G2, possibly reflecting a G2/M block. Our multi-scale analysis tool groups colony regions into density classes, and cells belonging to those classes have distinct distributions of pluripotency markers and respond differently to DNA damage induction. Lastly, we demonstrate that our pipeline can robustly handle high-content, high-resolution single molecular mRNA FISH data by using novel image processing techniques. Overall, the imaging informatics pipeline presented offers a novel approach to the analysis of hPS cells that includes not only single cell features but also colony wide, and more generally, multi-scale spatial configuration.", "link"=>"http://www.mendeley.com/research/multiscale-imaging-informatics-pipeline-situ-pluripotent-stem-cell-analysis", "reader_count"=>31, "reader_count_by_academic_status"=>{"Professor > Associate Professor"=>2, "Student > Doctoral Student"=>1, "Researcher"=>7, "Student > Ph. D. Student"=>11, "Other"=>3, "Student > Master"=>4, "Student > Bachelor"=>2, "Professor"=>1}, "reader_count_by_user_role"=>{"Professor > Associate Professor"=>2, "Student > Doctoral Student"=>1, "Researcher"=>7, "Student > Ph. D. Student"=>11, "Other"=>3, "Student > Master"=>4, "Student > Bachelor"=>2, "Professor"=>1}, "reader_count_by_subject_area"=>{"Engineering"=>1, "Unspecified"=>1, "Biochemistry, Genetics and Molecular Biology"=>6, "Medicine and Dentistry"=>2, "Agricultural and Biological Sciences"=>12, "Chemical Engineering"=>1, "Physics and Astronomy"=>1, "Chemistry"=>1, "Psychology"=>1, "Social Sciences"=>1, "Computer Science"=>3, "Immunology and Microbiology"=>1}, "reader_count_by_subdiscipline"=>{"Engineering"=>{"Engineering"=>1}, "Medicine and Dentistry"=>{"Medicine and Dentistry"=>2}, "Chemistry"=>{"Chemistry"=>1}, "Social Sciences"=>{"Social Sciences"=>1}, "Physics and Astronomy"=>{"Physics and Astronomy"=>1}, "Psychology"=>{"Psychology"=>1}, "Immunology and Microbiology"=>{"Immunology and Microbiology"=>1}, "Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>12}, "Computer Science"=>{"Computer Science"=>3}, "Biochemistry, Genetics and Molecular Biology"=>{"Biochemistry, Genetics and Molecular Biology"=>6}, "Unspecified"=>{"Unspecified"=>1}, "Chemical Engineering"=>{"Chemical Engineering"=>1}}, "reader_count_by_country"=>{"United States"=>3}, "group_count"=>2}

Scopus | Further Information

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Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/1859577"], "description"=>"<p>(A) Quantification of Oct4 mRNA level with smFISH in a hES cell. Top: raw image; Bottom: segmented FISH spots. (B) Segmentation of Oct4 spots in hES cells with nuclear boundaries in red, cell boundaries in yellow, and colonies boundaries in green. (<b>C</b>) Method for setting smFISH spot detection thresholds to ensure spot detection consistency across adjacent fields in a large region. (D) Reconstruction of Oct4 mRNA levels across 6 contiguous fields of view using the spot matching procedure detailed in the main text. Each circle represents an individual cell and circle color denotes the expression level of Oct4. Overall, Oct4 levels diminish through differentiation, although small clusters of cells with elevated Oct4 levels can be observed after 2 days of differentiation.</p>", "links"=>[], "tags"=>["hPS cells", "novel image processing techniques", "mRNA FISH data", "imaging informatics pipeline", "pluripotency marker expression", "DNA damage induction", "study fate decisions", "hPS cell cycle regulation", "Situ Pluripotent Stem Cell Analysis Human pluripotent", "analysis"], "article_id"=>1283512, "categories"=>["Biological Sciences"], "users"=>["Bryan R. Gorman", "Junjie Lu", "Anna Baccei", "Nathan C. Lowry", "Jeremy E. Purvis", "Rami S. Mangoubi", "Paul H. Lerou"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0116037.g004", "stats"=>{"downloads"=>2, "page_views"=>49, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Large_scale_mapping_of_single_cell_heterogeneity_in_Oct4_mRNA_expression_/1283512", "title"=>"Large-scale mapping of single-cell heterogeneity in Oct4 mRNA expression.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-12-31 03:25:49"}
  • {"files"=>["https://ndownloader.figshare.com/files/1859627", "https://ndownloader.figshare.com/files/1859628", "https://ndownloader.figshare.com/files/1859629", "https://ndownloader.figshare.com/files/1859630", "https://ndownloader.figshare.com/files/1859633", "https://ndownloader.figshare.com/files/1859634"], "description"=>"<div><p>Human pluripotent stem (hPS) cells are a potential source of cells for medical therapy and an ideal system to study fate decisions in early development. However, hPS cells cultured <i>in vitro</i> exhibit a high degree of heterogeneity, presenting an obstacle to clinical translation. hPS cells grow in spatially patterned colony structures, necessitating quantitative single-cell image analysis. We offer a tool for analyzing the spatial population context of hPS cells that integrates automated fluorescent microscopy with an analysis pipeline. It enables high-throughput detection of colonies at low resolution, with single-cellular and sub-cellular analysis at high resolutions, generating seamless <i>in situ</i> maps of single-cellular data organized by colony. We demonstrate the tool's utility by analyzing inter- and intra-colony heterogeneity of hPS cell cycle regulation and pluripotency marker expression. We measured the heterogeneity within individual colonies by analyzing cell cycle as a function of distance. Cells loosely associated with the outside of the colony are more likely to be in G1, reflecting a less pluripotent state, while cells within the first pluripotent layer are more likely to be in G2, possibly reflecting a G2/M block. Our multi-scale analysis tool groups colony regions into density classes, and cells belonging to those classes have distinct distributions of pluripotency markers and respond differently to DNA damage induction. Lastly, we demonstrate that our pipeline can robustly handle high-content, high-resolution single molecular mRNA FISH data by using novel image processing techniques. Overall, the imaging informatics pipeline presented offers a novel approach to the analysis of hPS cells that includes not only single cell features but also colony wide, and more generally, multi-scale spatial configuration.</p></div>", "links"=>[], "tags"=>["hPS cells", "novel image processing techniques", "mRNA FISH data", "imaging informatics pipeline", "pluripotency marker expression", "DNA damage induction", "study fate decisions", "hPS cell cycle regulation", "Situ Pluripotent Stem Cell Analysis Human pluripotent", "analysis"], "article_id"=>1283550, "categories"=>["Biological Sciences"], "users"=>["Bryan R. Gorman", "Junjie Lu", "Anna Baccei", "Nathan C. Lowry", "Jeremy E. Purvis", "Rami S. Mangoubi", "Paul H. Lerou"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0116037.s001", "https://dx.doi.org/10.1371/journal.pone.0116037.s002", "https://dx.doi.org/10.1371/journal.pone.0116037.s003", "https://dx.doi.org/10.1371/journal.pone.0116037.s004", "https://dx.doi.org/10.1371/journal.pone.0116037.s005", "https://dx.doi.org/10.1371/journal.pone.0116037.s006"], "stats"=>{"downloads"=>4, "page_views"=>22, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Multi_Scale_Imaging_and_Informatics_Pipeline_for_In_Situ_Pluripotent_Stem_Cell_Analysis_/1283550", "title"=>"Multi-Scale Imaging and Informatics Pipeline for In Situ Pluripotent Stem Cell Analysis", "pos_in_sequence"=>0, "defined_type"=>4, "published_date"=>"2014-12-31 03:25:49"}
  • {"files"=>["https://ndownloader.figshare.com/files/1859536"], "description"=>"<p>(A) Illustration of the distance transform applied to cells in all colonies. Outside of the boundaries of the colony, the distance from edge is “0”, and within the border of the colony, the distance ranges between 0 and the maximum radius of the colony. (B) Frequencies of cells as a function of distance from edge for the G1 (red), S (magenta), and G2 (blue) and M (green) subpopulations. Error bars indicate the 95% confidence interval based on 1000 bootstrap samples. Abscissa tick marks indicate the distance intervals from which the population and bootstrap frequencies were calculated. Each distance interval contains approximately 1500 cells. The data points are located halfway in between each interval. The frequencies at each point add up to 1. Of note is the statistically significant peak in the G2/M subpopulation at 25 microns. (C) A cellular map illustrating the periphery (blue), Cell Layer 1 (green), Cell Layer 2 (red), and Cell Layer 3 (blue) subpopulations. (D) Distributions of cell cycle phases for each cell layer across many colonies. There is a significant enrichment of G2 cells in Cell Layer 1 relative to Cell Layers 2 and 3.</p>", "links"=>[], "tags"=>["hPS cells", "novel image processing techniques", "mRNA FISH data", "imaging informatics pipeline", "pluripotency marker expression", "DNA damage induction", "study fate decisions", "hPS cell cycle regulation", "Situ Pluripotent Stem Cell Analysis Human pluripotent", "analysis"], "article_id"=>1283474, "categories"=>["Biological Sciences"], "users"=>["Bryan R. Gorman", "Junjie Lu", "Anna Baccei", "Nathan C. Lowry", "Jeremy E. Purvis", "Rami S. Mangoubi", "Paul H. Lerou"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0116037.g002", "stats"=>{"downloads"=>1, "page_views"=>17, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Spatial_enrichment_of_G1_and_G2_phase_cells_in_colonies_/1283474", "title"=>"Spatial enrichment of G1 and G2 phase cells in colonies.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-12-31 03:25:49"}
  • {"files"=>["https://ndownloader.figshare.com/files/1859520"], "description"=>"<p>(A) Our system enables researchers to analyze intercellular dynamics in hES cells by structuring relationships between cells within a colony; between cells and the colony they belong to; and from one colony to another. (B) The window of the main GUI controlling the automated image acquisition software. (C) A daughter window of the GUI facilitating pre-scanning of the slide and selection of regions. (D) A workflow to obtain image-derived features from single cells, while placing them in the context of the colony they belong to. Thus, the pipeline involves a multi-scale segmentation of the colonies within the sample, and the cells within the colonies. Each cell not only has a physical address within the colony, but is linked with colony-wide properties, such as the size and shape of the colony it is derived from.</p>", "links"=>[], "tags"=>["hPS cells", "novel image processing techniques", "mRNA FISH data", "imaging informatics pipeline", "pluripotency marker expression", "DNA damage induction", "study fate decisions", "hPS cell cycle regulation", "Situ Pluripotent Stem Cell Analysis Human pluripotent", "analysis"], "article_id"=>1283458, "categories"=>["Biological Sciences"], "users"=>["Bryan R. Gorman", "Junjie Lu", "Anna Baccei", "Nathan C. Lowry", "Jeremy E. Purvis", "Rami S. Mangoubi", "Paul H. Lerou"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0116037.g001", "stats"=>{"downloads"=>2, "page_views"=>14, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Overview_of_the_multi_scale_Imaging_and_Informatics_pipeline_/1283458", "title"=>"Overview of the multi-scale Imaging and Informatics pipeline.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-12-31 03:25:49"}
  • {"files"=>["https://ndownloader.figshare.com/files/1859549"], "description"=>"<p>(A–C) In order to investigate regional heterogeneity, colonies of differentiated and NCS-treated cells were computationally divided into windows, which were then classified according to the density of cells contained. 25 colonies were divided into 164 windows, containing a total of 13,133 cells. As demonstrated, high cell density regions (purple) tend to have low Oct4 (B), and reduced induction of cPARP (C) upon DNA damage, than low (blue) and mixed (red) density regions. (D) Oct4 distribution of the cells in the above classified subregions. Black line shows the distribution of all cells. (E) Proportion of Oct4 positive cells in each subpopulation. (F) proportion of cPARP positive cells.</p>", "links"=>[], "tags"=>["hPS cells", "novel image processing techniques", "mRNA FISH data", "imaging informatics pipeline", "pluripotency marker expression", "DNA damage induction", "study fate decisions", "hPS cell cycle regulation", "Situ Pluripotent Stem Cell Analysis Human pluripotent", "analysis"], "article_id"=>1283487, "categories"=>["Biological Sciences"], "users"=>["Bryan R. Gorman", "Junjie Lu", "Anna Baccei", "Nathan C. Lowry", "Jeremy E. Purvis", "Rami S. Mangoubi", "Paul H. Lerou"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0116037.g003", "stats"=>{"downloads"=>1, "page_views"=>18, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Cells_in_different_regions_of_colonies_respond_differently_to_DNA_damage_/1283487", "title"=>"Cells in different regions of colonies respond differently to DNA damage.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-12-31 03:25:49"}

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Relative Metric

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