Nonalcoholic steatohepatitis (NASH) is a disease of the liver characterized by inflammation, cell injury and fibrosis. Progression of this disease can lead to the cirrhosis, possible hepatocellular carcinoma and may ultimately require liver transplantation. NASH is a progression of non-alcoholic fatty liver disease (NAFLD), a condition that affects 30-40% of adults in the United States. Early stages of NAFLD can be treated with alterations in a patient’s diet and activity levels, and thus the Food and Drug Administration (FDA) does not recommend therapeutic drug intervention at those stages. However, once the disease progresses to NASH with fibrosis there are much clearer negative clinical outcomes, and thus pharmacologic treatment may have more benefits than risks for this stage. Currently, there are no FDA approved drugs for NASH, but several compounds have shown promise.
Visikol tested two of these clinically relevant compounds, Elafibranor and Pioglitazone, in Visikol’s HepaRG™ NP 3D liver model to assess their effect on markers of fibrosis. The HepaRG™ NP 3D liver model combines hepatocytes and primary non-parenchymal cells (hepatic stellate cells, LSECs, Kupffer cells) to examine end points such as: collagen deposition, hepatic stellate cell activation, lipid deposition etc. Specifically for this study, collagen deposition and stellate cell activation were assessed using immunofluorescent labeling for pan-collagen and alpha-smooth muscle actin, respectively. Using the Visikol® HISTO-M™ clearing reagent we were able to image through these 3D tissues. Clearing allowed for the quantification of immunofluorescent labeling throughout entire spheroids using our custom in-house image analysis pipelines. Fibrosis was induced in the HepaRG™ NP 3D liver model using alone TGFβ1, resulting in both increased collagen deposition and alpha-smooth muscle actin expression. Treatment with TGFβ1 and either Elafibranor or Pioglitazone showed qualitative and quantitative decreases in collagen deposition and alpha-smooth muscle actin expression with a clear dose-dependent response in collagen deposition for pioglitazone.
Figure 1. Representative maximum intensity projections of the HepaRGTM NP 3D liver model spheroids after treatment with 100 ng/mL TGFB1 and either elafibranor or pioglitazone. Immunofluorescent labeling indicates DAPI (blue), pan-collagen (green) and alpha-smooth muscle actin (red).
The results of these experiments demonstrate the clinical relevance of the HepaRG™ NP 3D liver model, and Visikol is thrilled to be able to offer this 3D human model for use in the drug discovery pipeline. Please feel free to reach out if you are interested in learning more about this NASH/fibrosis model or any of the other services Visikol offers. Our Clients’ interests and requirements are paramount to our success and we look forward to working together to develop assays that best suit your needs!
Figure 2. Quantification of collagen deposition and alpha-smooth muscle actin expression. Data represent mean ± SEM of percent expression of pan-collagen or alpha-smooth muscle actin for treatment with 100 ng/mL TGFB1 and either elafibranor or pioglitazone. Treatment with either compound resulted in a decrease in both collagen deposition and alpha-smooth muscle actin expression compared to the TGFB1-only control.