Automated Biochemical, Morphological, and Organizational Assessment of Precancerous Changes from Endogenous Two-Photon Fluorescence Images
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{"title"=>"Automated biochemical, morphological, and organizational assessment of precancerous changes from endogenous Two-Photon fluorescence images", "type"=>"journal", "authors"=>[{"first_name"=>"Jonathan M.", "last_name"=>"Levitt", "scopus_author_id"=>"7006925040"}, {"first_name"=>"Margaret E.", "last_name"=>"McLaughlin-Drubin", "scopus_author_id"=>"6508142180"}, {"first_name"=>"Karl", "last_name"=>"Münger", "scopus_author_id"=>"7006292295"}, {"first_name"=>"Irene", "last_name"=>"Georgakoudi", "scopus_author_id"=>"6603938579"}], "year"=>2011, "source"=>"PLoS ONE", "identifiers"=>{"sgr"=>"80052587887", "pmid"=>"21931846", "scopus"=>"2-s2.0-80052587887", "pui"=>"362523750", "isbn"=>"1932-6203 (Electronic) 1932-6203 (Linking)", "issn"=>"19326203", "doi"=>"10.1371/journal.pone.0024765"}, "id"=>"1e249f6e-5447-3d25-b4cb-5d11241e8ed3", "abstract"=>"BACKGROUND: Multi-photon fluorescence microscopy techniques allow for non-invasive interrogation of live samples in their native environment. These methods are particularly appealing for identifying pre-cancers because they are sensitive to the early changes that occur on the microscopic scale and can provide additional information not available using conventional screening techniques. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we developed novel automated approaches, which can be employed for the real-time analysis of two-photon fluorescence images, to non-invasively discriminate between normal and pre-cancerous/HPV-immortalized engineered tissues by concurrently assessing metabolic activity, morphology, organization, and keratin localization. Specifically, we found that the metabolic activity was significantly enhanced and more uniform throughout the depths of the HPV-immortalized epithelia, based on our extraction of the NADH and FAD fluorescence contributions. Furthermore, we were able to separate the keratin contribution from metabolic enzymes to improve the redox estimates and to use the keratin localization as a means to discriminate between tissue types. To assess morphology and organization, Fourier-based, power spectral density (PSD) approaches were employed. The nuclear size distribution throughout the epithelial depths was quantified by evaluating the variance of the corresponding spatial frequencies, which was found to be greater in the normal tissue compared to the HPV-immortalized tissues. The PSD was also used to calculate the Hurst parameter to identify the level of organization in the tissues, assuming a fractal model for the fluorescence intensity fluctuations within a field. We found the range of organization was greater in the normal tissue and closely related to the level of differentiation. CONCLUSIONS/SIGNIFICANCE: A wealth of complementary morphological, biochemical and organizational tissue parameters can be extracted from high resolution images that are acquired based entirely on endogenous sources of contrast. They are promising diagnostic parameters for the non-invasive identification of early cancerous changes and could improve significantly diagnosis and treatment for numerous patients.", "link"=>"http://www.mendeley.com/research/automated-biochemical-morphological-organizational-assessment-precancerous-changes-endogenous-twopho", "reader_count"=>14, "reader_count_by_academic_status"=>{"Unspecified"=>1, "Professor > Associate Professor"=>1, "Researcher"=>4, "Student > Ph. D. Student"=>3, "Student > Master"=>3, "Student > Bachelor"=>1, "Librarian"=>1}, "reader_count_by_user_role"=>{"Unspecified"=>1, "Professor > Associate Professor"=>1, "Researcher"=>4, "Student > Ph. D. Student"=>3, "Student > Master"=>3, "Student > Bachelor"=>1, "Librarian"=>1}, "reader_count_by_subject_area"=>{"Engineering"=>3, "Unspecified"=>3, "Agricultural and Biological Sciences"=>4, "Physics and Astronomy"=>2, "Chemistry"=>1, "Computer Science"=>1}, "reader_count_by_subdiscipline"=>{"Engineering"=>{"Engineering"=>3}, "Chemistry"=>{"Chemistry"=>1}, "Physics and Astronomy"=>{"Physics and Astronomy"=>2}, "Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>4}, "Computer Science"=>{"Computer Science"=>1}, "Unspecified"=>{"Unspecified"=>3}}, "group_count"=>2}

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Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/372066"], "description"=>"<div><h3>Background</h3><p>Multi-photon fluorescence microscopy techniques allow for non-invasive interrogation of live samples in their native environment. These methods are particularly appealing for identifying pre-cancers because they are sensitive to the early changes that occur on the microscopic scale and can provide additional information not available using conventional screening techniques.</p> <h3>Methodology/Principal Findings</h3><p>In this study, we developed novel automated approaches, which can be employed for the real-time analysis of two-photon fluorescence images, to non-invasively discriminate between normal and pre-cancerous/HPV-immortalized engineered tissues by concurrently assessing metabolic activity, morphology, organization, and keratin localization. Specifically, we found that the metabolic activity was significantly enhanced and more uniform throughout the depths of the HPV-immortalized epithelia, based on our extraction of the NADH and FAD fluorescence contributions. Furthermore, we were able to separate the keratin contribution from metabolic enzymes to improve the redox estimates and to use the keratin localization as a means to discriminate between tissue types. To assess morphology and organization, Fourier-based, power spectral density (PSD) approaches were employed. The nuclear size distribution throughout the epithelial depths was quantified by evaluating the variance of the corresponding spatial frequencies, which was found to be greater in the normal tissue compared to the HPV-immortalized tissues. The PSD was also used to calculate the Hurst parameter to identify the level of organization in the tissues, assuming a fractal model for the fluorescence intensity fluctuations within a field. We found the range of organization was greater in the normal tissue and closely related to the level of differentiation.</p> <h3>Conclusions/Significance</h3><p>A wealth of complementary morphological, biochemical and organizational tissue parameters can be extracted from high resolution images that are acquired based entirely on endogenous sources of contrast. They are promising diagnostic parameters for the non-invasive identification of early cancerous changes and could improve significantly diagnosis and treatment for numerous patients.</p> </div>", "links"=>[], "tags"=>["automated", "precancerous", "changes", "endogenous", "two-photon", "fluorescence", "images"], "article_id"=>133491, "categories"=>["Physics", "Biotechnology", "Cancer"], "users"=>["Jonathan M. Levitt", "Margaret E. McLaughlin-Drubin", "Karl Münger", "Irene Georgakoudi"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0024765", "stats"=>{"downloads"=>1, "page_views"=>15, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/Automated_Biochemical_Morphological_and_Organizational_Assessment_of_Precancerous_Changes_from_Endogenous_Two_Photon_Fluorescence_Images/133491", "title"=>"Automated Biochemical, Morphological, and Organizational Assessment of Precancerous Changes from Endogenous Two-Photon Fluorescence Images", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-09-09 00:58:11"}
  • {"files"=>["https://ndownloader.figshare.com/files/737494"], "description"=>"<p>Normal (left column) and HPV- immortalized tissues (right column). Histological hematoxylin and eosin staining (<i>a–b</i>). False-color, depth-resolved TPEF and SHG images (<i>c–f</i>) from tissue excited at 755 nm and 800 nm. Green is fluorescence in the 455 nm channel (from NADH and keratin), while red is signal from the 525 nm channel (from FAD and keratin), acquired at 755 nm excitation. Blue represents 400 nm SHG detected at 800 nm excitation. Three-dimensional reconstructions were created from the fluorescence images (<i>c–d</i>). Corresponding depth images are shown in (<i>e–f</i>) with increasing depth from i to iv with zoomed insets to highlight the cell sizes. The scale bar represents 47.5 µm in the images and 10 µm in the zoomed insets. The transverse views of the normal and HPV-immortalized tissues (zx & zy) are shown with scale bars of 100 µm and 82 µm, respectively.</p>", "links"=>[], "tags"=>["fluorescence", "images"], "article_id"=>407846, "categories"=>["Physics", "Biotechnology", "Cancer", "Virology"], "users"=>["Jonathan M. Levitt", "Margaret E. McLaughlin-Drubin", "Karl Münger", "Irene Georgakoudi"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0024765.g001", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Histology_and_fluorescence_based_images_of_tissue_/407846", "title"=>"Histology and fluorescence based images of tissue.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-09-09 02:10:46"}
  • {"files"=>["https://ndownloader.figshare.com/files/737983"], "description"=>"<p>NAD(P)H (blue) and FAD (red) contribution after being converted to a relative concentration value as a function of depth for normal and HPV-immortalized tissues (a) and the corresponding concentration based redox ratio (b). The keratin contribution is reported by the number of pixels that were removed at each depth increment for the normal and HPV tissue (c).</p>", "links"=>[], "tags"=>["keratin", "localization"], "article_id"=>408359, "categories"=>["Physics", "Biotechnology", "Cancer", "Virology"], "users"=>["Jonathan M. Levitt", "Margaret E. McLaughlin-Drubin", "Karl Münger", "Irene Georgakoudi"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0024765.g005", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Redox_ratio_and_keratin_localization_from_a_representative_sample_/408359", "title"=>"Redox ratio and keratin localization from a representative sample.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-09-09 02:19:19"}
  • {"files"=>["https://ndownloader.figshare.com/files/737651"], "description"=>"<p>The (a) variation of the PSD as a function of frequency and the (b) Hurst parameter, as a function of normalized depth for the two tissue types, from the basal (value of 0) to the upper most tissue layer (value of 1).</p>", "links"=>[], "tags"=>["Virology", "biotechnology", "oncology", "physics"], "article_id"=>408008, "categories"=>["Physics", "Biotechnology", "Cancer", "Virology"], "users"=>["Jonathan M. Levitt", "Margaret E. McLaughlin-Drubin", "Karl Münger", "Irene Georgakoudi"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0024765.g002", "stats"=>{"downloads"=>0, "page_views"=>1, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Morphological_and_organizational_assessment_/408008", "title"=>"Morphological and organizational assessment.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-09-09 02:13:28"}
  • {"files"=>["https://ndownloader.figshare.com/files/737757"], "description"=>"<p>Fluorescence emission from 755 nm (A), 800 nm (B), and 860 nm (C) excitation at the cornified (blue), suprabasal (red), and basal (green) layers and the corresponding fields (D) before (i–iii) and after (iv–vi) thresholding to identify the keratin(+) regions (scale bar – 47.5 µm). Spectral decomposition was performed on the keratin(−) (E) and keratin(+) (F) areas to extract the relative contributions of NAD(P)H, FAD and keratin.</p>", "links"=>[], "tags"=>["Virology", "biotechnology", "oncology", "physics"], "article_id"=>408121, "categories"=>["Physics", "Biotechnology", "Cancer", "Virology"], "users"=>["Jonathan M. Levitt", "Margaret E. McLaughlin-Drubin", "Karl Münger", "Irene Georgakoudi"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0024765.g003", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Spectra_analysis_from_normal_tissue_/408121", "title"=>"Spectra analysis from normal tissue.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-09-09 02:15:21"}
  • {"files"=>["https://ndownloader.figshare.com/files/737871"], "description"=>"<p>Fluorescence emission from 755 nm (A), 800 nm (B), and 860 nm (C) excitation at the cornified (blue), suprabasal (red), and basal (green) layers and the corresponding fields (D) before (i–iii) and after (iv–vi) thresholding to identify the keratin(+) regions (scale bar – 47.5 µm). Spectral decomposition was performed on the keratin(−) (E) and keratin(+) (F) intensity areas to extract the relative contributions of NAD(P)H, FAD and keratin.</p>", "links"=>[], "tags"=>["hpv", "immortalized"], "article_id"=>408249, "categories"=>["Physics", "Biotechnology", "Cancer", "Virology"], "users"=>["Jonathan M. Levitt", "Margaret E. McLaughlin-Drubin", "Karl Münger", "Irene Georgakoudi"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0024765.g004", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Spectral_analysis_from_HPV_immortalized_tissues_/408249", "title"=>"Spectral analysis from HPV immortalized tissues.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-09-09 02:17:29"}
  • {"files"=>["https://ndownloader.figshare.com/files/738139"], "description"=>"<p>Possible combinations of three parameters to discriminate between normal (green circles) and HPV-immortalized (blue triangles) tissues.</p>", "links"=>[], "tags"=>["Virology", "biotechnology", "oncology", "physics"], "article_id"=>408510, "categories"=>["Physics", "Biotechnology", "Cancer", "Virology"], "users"=>["Jonathan M. Levitt", "Margaret E. McLaughlin-Drubin", "Karl Münger", "Irene Georgakoudi"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0024765.g006", "stats"=>{"downloads"=>1, "page_views"=>4, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Multi_parametric_analysis_/408510", "title"=>"Multi-parametric analysis.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-09-09 02:21:50"}

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Relative Metric

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