TY - JOUR
T1 - Multispectral Imaging Enables Characterization of Intrahepatic Macrophages in Patients With Chronic Liver Disease
AU - Saldarriaga, Omar A.
AU - Freiberg, Benjamin
AU - Krishnan, Santhoshi
AU - Rao, Arvind
AU - Burks, Jared
AU - Booth, Adam L.
AU - Dye, Bradley
AU - Utay, Netanya
AU - Ferguson, Monique
AU - Akil, Abdellah
AU - Yi, Minkyung
AU - Beretta, Laura
AU - Stevenson, Heather L.
N1 - Funding Information:
We would like to thank Judy Pham for her assistance with fibrosis quantification of liver biopsies. We also thank Kevin Mottershead and Grady Carlson from Akoya Biosciences for their support and guidance in using the Vectra Platform. We extend sincere gratitude to Jeffrey East, physician assistant, and Shana White, study coordinator, for their assistance in acquiring and organizing the clinical samples. Sam Diaz de Leon and Blanca Flores provided secretarial support and assistance with obtaining the archived liver biopsy tissue blocks. Drs. David Walker and Cornelius Elferink provided a critical review of the manuscript.
Publisher Copyright:
© 2020 The Authors. Hepatology Communications published by Wiley Periodicals, Inc., on behalf of the American Association for the Study of Liver Diseases.
PY - 2020/5/1
Y1 - 2020/5/1
N2 - Intrahepatic macrophages influence the composition of the microenvironment, host immune response to liver injury, and development of fibrosis. Compared with stellate cells, the role of macrophages in the development of fibrosis remains unclear. Multispectral imaging allows detection of multiple markers in situ in human formalin-fixed, paraffin-embedded tissue. This cutting-edge technology is ideal for analyzing human liver tissues, as it allows spectral unmixing of fluorophore signals, subtraction of auto-fluorescence, and preservation of hepatic architecture. We analyzed five different antibodies commonly observed on macrophage populations (CD68, MAC387, CD163, CD14, and CD16). After optimization of the monoplex stains and development of a Spectral Library, we combined all of the antibodies into a multiplex protocol and used them to stain biopsies collected from representative patients with chronic liver diseases, including chronic hepatitis C, nonalcoholic steatohepatitis, and autoimmune hepatitis. Various imaging modalities were tested, including cell phenotyping, tissue segmentation, t-distributed stochastic neighbor embedding plots, and phenotype matrices that facilitated comparison and visualization of the identified macrophage and other cellular profiles. We then tested the feasibility of this platform to analyze numerous regions of interest from liver biopsies with multiple patients per group, using batch analysis algorithms. Five populations showed significant differences between patients positive for hepatitis C virus with advanced fibrosis when compared with controls. Three of these were significantly increased in patients with advanced fibrosis when compared to controls, and these included CD163+CD16+, CD68+, and CD68+MAC387+. Conclusion: Spectral imaging microscopy is a powerful tool that enables in situ analysis of macrophages and other cells in human liver biopsies and may lead to more personalized therapeutic approaches in the future.
AB - Intrahepatic macrophages influence the composition of the microenvironment, host immune response to liver injury, and development of fibrosis. Compared with stellate cells, the role of macrophages in the development of fibrosis remains unclear. Multispectral imaging allows detection of multiple markers in situ in human formalin-fixed, paraffin-embedded tissue. This cutting-edge technology is ideal for analyzing human liver tissues, as it allows spectral unmixing of fluorophore signals, subtraction of auto-fluorescence, and preservation of hepatic architecture. We analyzed five different antibodies commonly observed on macrophage populations (CD68, MAC387, CD163, CD14, and CD16). After optimization of the monoplex stains and development of a Spectral Library, we combined all of the antibodies into a multiplex protocol and used them to stain biopsies collected from representative patients with chronic liver diseases, including chronic hepatitis C, nonalcoholic steatohepatitis, and autoimmune hepatitis. Various imaging modalities were tested, including cell phenotyping, tissue segmentation, t-distributed stochastic neighbor embedding plots, and phenotype matrices that facilitated comparison and visualization of the identified macrophage and other cellular profiles. We then tested the feasibility of this platform to analyze numerous regions of interest from liver biopsies with multiple patients per group, using batch analysis algorithms. Five populations showed significant differences between patients positive for hepatitis C virus with advanced fibrosis when compared with controls. Three of these were significantly increased in patients with advanced fibrosis when compared to controls, and these included CD163+CD16+, CD68+, and CD68+MAC387+. Conclusion: Spectral imaging microscopy is a powerful tool that enables in situ analysis of macrophages and other cells in human liver biopsies and may lead to more personalized therapeutic approaches in the future.
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U2 - 10.1002/hep4.1494
DO - 10.1002/hep4.1494
M3 - Article
C2 - 32363321
AN - SCOPUS:85094166310
SN - 2471-254X
VL - 4
SP - 708
EP - 723
JO - Hepatology Communications
JF - Hepatology Communications
IS - 5
ER -