An Improved Time- and Labor- Efficient Protocol for Mouse Primary Hepatocyte Isolation

Mingxiao Feng; Sara Divall; Sheng Wu

doi:10.3791/61812

An Improved Time- and Labor- Efficient Protocol for Mouse Primary Hepatocyte Isolation

JoVE Journal
Biology

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JoVE Journal Biology

An Improved Time- and Labor- Efficient Protocol for Mouse Primary Hepatocyte Isolation

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05:42 min

October 25, 2021

DOI: 10.3791/61812-v

Mingxiao Feng¹, Sara Divall³, Sheng Wu^1,2

¹Department of Pediatrics,Johns Hopkins University School of Medicine, ²Department of Physiology,Temple University School of Medicine, ³Department of Pediatrics,Seattle Children’s Hospital

Overview

This study presents a labor- and time-efficient protocol for isolating mouse primary hepatocytes, which are critical for research in liver function and metabolism. The method utilizes commercially available reagents to enhance yield and minimize interference from non-parenchymal cells.

Key Study Components

Research Area

Hepatic metabolism
Glucose metabolism
Drug testing

Background

Mouse primary hepatocytes are essential for studying liver biology.
Traditional isolation methods are often time-consuming and labor-intensive.
Reducing isolation time can facilitate more efficient biological experimentation.

Methods Used

Isolation protocol using peristaltic pump and collagenase dispase medium.
Mouse model: C57BL/6J female mice.
Techniques for cell culture and measurement of gene expression.

Main Results

Increased mRNA levels of hepatocyte markers compared to whole liver.
Decreased presence of immune and endothelial cells post-isolation.
Demonstrated insulin sensitivity and activated glucose production pathways.

Conclusions

This protocol significantly improves the efficiency of isolating primary hepatocytes.
Enhanced understanding of glucose metabolism and drug response in liver studies.

Frequently Asked Questions

What are mouse primary hepatocytes used for?

They are used for studying liver metabolism, glucose processing, and drug testing.

How does this protocol improve hepatocyte isolation?

It streamlines the process, making it both time- and labor-efficient.

What measures the success of the hepatocyte isolation?

Success is measured by the purity of hepatocyte markers and the functionality in insulin sensitivity assays.

Can this protocol be applied to other research areas?

Yes, it can enhance studies related to liver biology and pharmacology.

What happens to the expression of hepatocyte markers over time?

Some markers decrease over time but remain comparable to whole liver levels for a period.

Is the glucose production pathway affected after isolation?

Yes, the pathway was activated following this isolation protocol.

What type of commercial reagents are needed?

The protocol requires commercially available perfusion and wash media, among other reagents.

Primary hepatocytes are a valuable tool to study liver response and metabolism in vitro. Utilizing commercially available reagents, an improved time- and labor-efficient protocol for mouse primary hepatocyte isolation was developed.

Mouse primary hepatocytes are important in liver studies. Especially for glucose metabolism, and hepatic drug testing. Time wise the isolation process can be time consuming and the energy costly.

This protocol can provide an efficient way to isolate mouse primary hepatocytes. It is time and labor friendly. Requiring very few steps with all regions commercially available.

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