Stereological analysis of size and density of hepatocytes in the porcine liver

Abstract

The porcine liver is frequently used as a large animal model for verification of surgical techniques, as well as experimental therapies. Often, a histological evaluation is required that include measurements of the size, nuclearity or density of hepatocytes. Our aims were to assess the mean number-weighted volume of hepatocytes, the numerical density of hepatocytes, and the fraction of binuclear hepatocytes (BnHEP) in the porcine liver, and compare the distribution of these parameters among hepatic lobes and macroscopic regions of interest (ROIs) with different positions related to the liver vasculature. Using disector and nucleator as design-based stereological methods, the morphometry of hepatocytes was quantified in seven healthy piglets. The samples were obtained from all six hepatic lobes and three ROIs (peripheral, paracaval and paraportal) within each lobe. Histological sections (thickness 16 μm) of formalin-fixed paraffin-embedded material were stained with the periodic acid-Schiff reaction to indicate the cell outlines and were assessed in a series of 3-μm-thick optical sections. The mean number-weighted volume of mononuclear hepatocytes (MnHEP) in all samples was 3670 ± 805 μm (mean ± SD). The mean number-weighted volume of BnHEP was 7050 ± 2550 μm . The fraction of BnHEP was 4 ± 2%. The numerical density of all hepatocytes was 146 997 ± 15 738 cells mm of liver parenchyma. The porcine hepatic lobes contained hepatocytes of a comparable size, nuclearity and density. No significant differences were identified between the lobes. The peripheral ROIs of the hepatic lobes contained the largest MnHEP with the smallest numerical density. The distribution of a larger MnHEP was correlated with a larger volume of BnHEP and a smaller numerical density of all hepatocytes. Practical recommendations for designing studies that involve stereological evaluations of the size, nuclearity and density of hepatocytes in porcine liver are provided.

Evaluation of the Cholesterol-Lowering Mechanism of Enterococcus faecium Strain 132 and Lactobacillus paracasei Strain 201 in Hypercholesterolemia Rats

“…The epididymal fat and liver were rinsed with PBS and fixed in 10% buffered formalin for 24 h. After fixation, the tissues were dehydrated using an alcohol gradient, paraffinembedded and sliced with a cut angle of 10 • . The thickness of liver slices were 3 µm and that of fat slices were 5 µm; tissues were then dewaxed with xylene and stained with haematoxylin, and the ScopeImage 9.0 system was used at a magnification of 200× for observation [30]. Steatosis and hepatocyte inflammation were scored based on non-alcoholic fatty liver disease (NAFS) scores [31].…”

Mesenchymal Stromal Cell Therapy in Novel Porcine Model of Diffuse Liver Damage Induced by Repeated Biliary Obstruction

“…The distribution of volume, nuclearity, and density within a healthy porcine liver was recently mapped [ 38 ]. Despite extensive sampling, no differences were identified between porcine liver lobes.…”

Estimating volumes from common carp hepatocytes using design‐based stereology and examining correlations with profile areas: Revisiting a nutritional assay and unveiling guidelines to microscopists

“…Indeed, based on their 3D morphology and orientation, hepatocytes (both cell body and nuclei) have been regarded as isotropic for all practical stereological purposes (Mandarim‐de‐Lacerda, ; Marcos et al, ). In a recent study on porcine liver, where the nucleator was used, the authors generated isotropic uniform random sections not to cope with anisotropy of the hepatocytes, but because of prospective analysis of microvessels (Junatas et al, ). Additionally, and particularly in fish species—in which hepatocytes are spatially organized and oriented in so many diverse ways in 3D, making intricately anastomosed cell tubules and/or plates, (Hardman, Volz, Kullman, & Hinton, ; Rocha, Monteiro, & Pereira, ) – any section of the liver will offer vast numbers of hepatocytes that are sectioned in all possible directions in 3D, with no preferential orientation, therefore granting isotropy of particle–probe intersections with insignificant possibilities of biases (Rocha et al, ).…”