VIPAR, a quantitative approach to 3D histopathology applied to lymphatic malformations
René Hägerling, Dominik Drees, Aaron Scherzinger, Cathrin Dierkes, Silvia Martin-Almedina, Stefan Butz, Kristiana Gordon, Michael Schäfers, Klaus Hinrichs, Pia Ostergaard, Dietmar Vestweber, Tobias Goerge, Sahar Mansour, Xiaoyi Jiang, Peter S. Mortimer, Friedemann Kiefer
René Hägerling, Dominik Drees, Aaron Scherzinger, Cathrin Dierkes, Silvia Martin-Almedina, Stefan Butz, Kristiana Gordon, Michael Schäfers, Klaus Hinrichs, Pia Ostergaard, Dietmar Vestweber, Tobias Goerge, Sahar Mansour, Xiaoyi Jiang, Peter S. Mortimer, Friedemann Kiefer
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Clinical Research and Public Health Dermatology Vascular biology

VIPAR, a quantitative approach to 3D histopathology applied to lymphatic malformations

Abstract

BACKGROUND. Lack of investigatory and diagnostic tools has been a major contributing factor to the failure to mechanistically understand lymphedema and other lymphatic disorders in order to develop effective drug and surgical therapies. One difficulty has been understanding the true changes in lymph vessel pathology from standard 2D tissue sections. METHODS. VIPAR (volume information-based histopathological analysis by 3D reconstruction and data extraction), a light-sheet microscopy–based approach for the analysis of tissue biopsies, is based on digital reconstruction and visualization of microscopic image stacks. VIPAR allows semiautomated segmentation of the vasculature and subsequent nonbiased extraction of characteristic vessel shape and connectivity parameters. We applied VIPAR to analyze biopsies from healthy lymphedematous and lymphangiomatous skin. RESULTS. Digital 3D reconstruction provided a directly visually interpretable, comprehensive representation of the lymphatic and blood vessels in the analyzed tissue volumes. The most conspicuous features were disrupted lymphatic vessels in lymphedematous skin and a hyperplasia (4.36-fold lymphatic vessel volume increase) in the lymphangiomatous skin. Both abnormalities were detected by the connectivity analysis based on extracted vessel shape and structure data. The quantitative evaluation of extracted data revealed a significant reduction of lymphatic segment length (51.3% and 54.2%) and straightness (89.2% and 83.7%) for lymphedematous and lymphangiomatous skin, respectively. Blood vessel length was significantly increased in the lymphangiomatous sample (239.3%). CONCLUSION. VIPAR is a volume-based tissue reconstruction data extraction and analysis approach that successfully distinguished healthy from lymphedematous and lymphangiomatous skin. Its application is not limited to the vascular systems or skin. FUNDING. Max Planck Society, DFG (SFB 656), and Cells-in-Motion Cluster of Excellence EXC 1003.

Authors

René Hägerling, Dominik Drees, Aaron Scherzinger, Cathrin Dierkes, Silvia Martin-Almedina, Stefan Butz, Kristiana Gordon, Michael Schäfers, Klaus Hinrichs, Pia Ostergaard, Dietmar Vestweber, Tobias Goerge, Sahar Mansour, Xiaoyi Jiang, Peter S. Mortimer, Friedemann Kiefer

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Figure 6

Determination of distinguishing characteristic parameters of the blood vessels in healthy and patient samples.

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Determination of distinguishing characteristic parameters of the blood v...
Following segmentation and skeletonization, the segment properties of the blood vasculature in three different control skin biopsies (n = 3), the lymphedematous skin biopsy (n = 1), and the lymphangiomatous skin biopsy (n = 1) were quantified, according to the definitions provided in Supplemental Figure 3. (A) Distance describes the length of the direct connection between branching points. (B) Segment length corresponds to the length of the center line of a vessel between two branching points. (C) The volume of the corresponding vessel segment is calculated from the total number of voxels associated with the segment. The vessel cross section is calculated as the quotient of volume and length. Data are presented as box-and-whisker plots, with the line depicting the median, the boxes showing upper and lower quartile, and the end of the whiskers representing the 1.5-fold interquartile range. Note the increased segment length and distance in lymphedematous and lymphangiomatous skin biopsies. Mann-Whitney U test with a Bonferroni correction for multiple comparison was used to compare data between groups. *P < 0.05, **P < 0.01, ***P < 0.001.