Development of a Computational High-Throughput Tool for the Quantitative Examination of Dose-Dependent Histological Features

Abstract

High-resolution digitalizing of histology slides facilitates the development of computational alternatives to manual quantitation of features of interest. We developed a MATLAB-based quantitative histological analysis tool (QuHAnT) for the high-throughput assessment of distinguishable histological features. QuHAnT validation was demonstrated by comparison with manual quantitation using liver sections from mice orally gavaged with sesame oil vehicle or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; 0.001–30 µg/kg) every 4 days for 28 days, which elicits hepatic steatosis with mild fibrosis. A quality control module of QuHAnT reduced the number of quantifiable Oil Red O (ORO)-stained images from 3,123 to 2,756. Increased ORO staining was measured at 10 and 30 µg/kg TCDD with a high correlation between manual and computational volume densities (Vv), although the dynamic range of QuHAnT was 10-fold greater. Additionally, QuHAnT determined the size of each ORO vacuole, which could not be accurately quantitated by visual examination or manual point counting. PicroSirius Red quantitation demonstrated superior collagen deposition detection due to the ability to consider all images within each section. QuHAnT dramatically reduced analysis time and facilitated the comprehensive assessment of features improving accuracy and sensitivity and represents a complementary tool for tissue/cellular features that are difficult and tedious to assess via subjective or semiquantitative methods.

From the Cover: Coagulation-Driven Hepatic Fibrosis Requires Protease Activated Receptor-1 (PAR-1) in a Mouse Model of TCDD-Elicited Steatohepatitis

“…Development of periportal fibrosis as indicated by excess collagen deposition was determined by qualitative and quantitative assessment of PSR-stained liver sections. In agreement with previous studies (Nault et al, 2015a;Pierre et al, 2014), TCDD treatment significantly increased collagen deposition in the liver (Fig. 6A and B).…”

“…All histological processing and staining were performed at the Michigan State University Investigative Histopathology Laboratory (https://humanpathology.natsci.msu.edu/; last accessed September 14, 2016) (Nault et al, 2015a). Briefly, frozen livers were sectioned at 6 mm and stained with an Oil Red O (ORO) solution and Gill 2 hematoxylin counterstain.…”

“…Slides were digitized using an Olympus Virtual Slide System (VS110) at 20x magnification and images were sampled using the Visiomorph Microimager (Visiopharm, Denmark). Quantitation of histological features was performed using an in-house developed tool (Nault et al, 2015a). Semi-quantitative severity scoring of H&E-stained liver sections was performed by a board certified veterinary pathologist experienced in hepatic toxicologic pathology of laboratory rodents.…”

“…Recent studies also suggest exposure to environmental contaminants including persistent organic pollutants and organochlorines contributes to metabolic disease development (Cave et al, 2010;Lee et al, 2006Lee et al, , 2007Taylor et al, 2013). For example,2,3,7,, the prototypical aryl hydrocarbon receptor (AhR) ligand, has been shown to contribute to the progression of NAFLD spectrum from simple, reversible steatosis, to steatohepatitis and early signs of fibrosis in mice (Kopec et al, 2013;Nault et al, 2015a;Pierre et al, 2014). Notably, all of these pathologies are elements of the NAFLD spectrum in humans.…”

“…Collectively, these studies indicate a central role for coagulation and thrombin signaling across the spectrum of NAFLD/NASH pathologies. Repeated TCDD exposure in mice has been shown to promote the development of steatosis, steatohepatitis, and fibrosis (Fader et al, 2015;Nault et al, 2015a;Pierre et al, 2014). However, no studies have examined whether the progression of liver pathologies elicited by TCDD is coupled to coagulation cascade activation, and the role of PAR-1 in this experimental context is unknown.…”