Rat multicellular 3D liver microtissues to explore TGF-β1 induced effects

Area of Science:

• Cell Biology
• Toxicology
• Hepatology

Background:

• Chronic liver damage progresses to fibrosis through hepatocellular injury, Kupffer cell (KC) activation, and hepatic stellate cell (HSC) activation.
• Inflammation and TGF-β1 are key mediators in the liver fibrosis adverse outcome pathway (AOP).

Purpose of the Study:

• To develop a rodent cell culture model for investigating liver fibrosis development.
• To study responses to pathophysiological stimuli like TGF-β1 and LPS-induced inflammation.

Main Methods:

• Optimized a protocol for purifying rat primary hepatocytes (Hep), HSC, and KC cells.
• Generated 3D co-cultures using the hanging drop method.
• Utilized 3D-monocultures and co-cultures for comparative analysis.

Main Results:

• The 3D co-culture model showed hepatocellular damage, inflammation, and HSC activation (increased αSMA) upon TGF-β1 stimulation.
• LPS induced an inflammatory response with increased cytokine expression.
• 3D-monocultures of Hep cells showed different responses, highlighting the necessity of multiple cell types.
• Pre-activated HSC may revert to a quiescent state in 3D culture, suggesting physiological relevance.

Conclusions:

• The developed 3D co-culture system effectively models key events in liver fibrosis.
• This model is valuable for studying the effects of stimuli like TGF-β1 and LPS on liver fibrosis.
• The presence of both parenchymal and non-parenchymal cells is essential for recapitulating fibrosis in vitro.