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Measurement of Fatty Acid β-Oxidation in a Suspension of Freshly Isolated Mouse Hepatocytes
Published on: September 9, 2021
A novel perfusion culture system for screening mitochondrial toxicity in primary mouse hepatocytes
Chika Yamamoto1, Akinori Takemura1, Sanae Ishii1
1Laboratory of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University.
Abstract:
The liver microphysiological system (MPS) model is an in-vitro culture method that mimics physiological blood flow, which enhances basal cellular functions. However, the liver MPS model has not been tested in the preclinical stage because of its obscure utility. It can overcome the major problem of conventional systems-rapid loss of mitochondrial activity in cultured hepatocytes due to limited oxygen supply-by supplying oxygen to cultured hepatocytes using a perfusion device. In this study, we developed a new perfusion culture system that can detect mitochondrial toxicity. Primary mouse hepatocytes were cultured under perfusion condition for 48 hr. The hepatocytes showed increased oxygen consumption and reduced lactate release. These results indicated that the ATP-production pathway was switched from glycolysis to mitochondrial oxidative phosphorylation in the perfusion culture system. Furthermore, ATP levels were considerably reduced in the perfusion culture system after exposure to phenformin, a mitochondrial complex I inhibitor. To summarize, the perfusion culture system could improve the mitochondrial activity in primary mouse hepatocytes, and thus, has potential implications in the detection of mitochondrial toxicity.
Insights
A new liver microphysiological system (MPS) using perfusion culture enhances hepatocyte mitochondrial function. This system shows promise for detecting mitochondrial toxicity in preclinical drug development.
Area of Science:
- Hepatology
- Mitochondrial Biology
- In Vitro Toxicology
Background:
- Liver microphysiological systems (MPS) offer enhanced cellular function mimicking physiological blood flow.
- Conventional hepatocyte cultures suffer from rapid mitochondrial dysfunction due to limited oxygen.
- The utility of liver MPS in preclinical settings, particularly for mitochondrial toxicity, remains underexplored.
Purpose of the Study:
- To develop and evaluate a novel perfusion culture system for detecting mitochondrial toxicity in primary mouse hepatocytes.
- To assess the impact of perfusion on hepatocyte mitochondrial activity and metabolic pathways.
- To determine the system's capability in identifying mitochondrial complex I inhibition.
Main Methods:
- Primary mouse hepatocytes were cultured in a newly developed perfusion system for 48 hours.
- Oxygen consumption and lactate release were measured to assess metabolic activity.
- Adenosine triphosphate (ATP) levels were measured following exposure to phenformin, a mitochondrial complex I inhibitor.
Main Results:
- Perfusion culture significantly increased hepatocyte oxygen consumption and decreased lactate release.
- Metabolic analysis indicated a shift from glycolysis to mitochondrial oxidative phosphorylation.
- Phenformin exposure in the perfusion system led to a substantial reduction in ATP levels, confirming mitochondrial toxicity detection.
Conclusions:
- The developed perfusion culture system enhances mitochondrial activity in primary mouse hepatocytes.
- This system effectively detects mitochondrial toxicity, demonstrated by reduced ATP levels after inhibitor exposure.
- The liver MPS perfusion model holds significant potential for preclinical assessment of mitochondrial toxicity.
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