Partial Hepatectomy in Adult Zebrafish

Summary

This protocol describes the procedure for removing the ventral lobe of the liver in adult zebrafish to enable the study of liver regeneration.

Abstract

Liver failure is one of the leading causes of death worldwide, and mortality from chronic liver disease is rising sharply in the United States. Healthy livers are capable of regenerating from toxic damage, but in advanced liver disease, the natural ability of the liver to regenerate is impaired. Zebrafish have emerged as a powerful experimental system for studying regeneration. They are an ideal model for studying liver regeneration from partial hepatectomy, a procedure with direct clinical relevance in which part of the liver is surgically removed, leaving the rest intact. There is no standard protocol for partial hepatectomy; previous studies using this model have used slightly different protocols and reported disparate results. Described here is an efficient, reproducible protocol for performing a partial hepatectomy in adult zebrafish. We use this technique to demonstrate that zebrafish are capable of epimorphic regeneration of the resected lobe. This protocol can be used to further interrogate the mechanisms required for liver regeneration in zebrafish.

Introduction

Among the solid organs in humans, the liver is the only organ capable of regeneration¹. This is critical, as the liver is an essential organ, responsible for key metabolic functions, energy storage, blood detoxification, secretion of plasma proteins, and bile production². Hepatocytes lost due to toxic or inflammatory damage are replaced primarily via division of the remaining hepatocytes¹. One classical experimental model for studying liver regeneration is partial hepatectomy, where individual lobes of the liver are removed, leaving the remaining lobes intact³. This procedu....

Protocol

Zebrafish were raised and bred according to standard procedures. Experiments were approved by the Brigham and Women's Hospital's Institutional Animal Care and Use Committee (2016N000405). Adult zebrafish were fasted for 24 h prior to the start of the protocol. System water refers to the water in zebrafish housing tanks in the aquatic facility.

1. Preparation and anesthetization

1. Prepare 0.016% Tricaine solution in system water.
   CAUTION: Tricaine is an irritant if it c.......

Discussion

The anatomical differences between zebrafish and mammalian models for liver regeneration present unique challenges to liver resection. The liver in zebrafish is in close proximity to the heart and the intestine; inadvertently damaging either organ results in increased mortality. The zebrafish liver is not encapsulated, making it more difficult to separate from the intestine. The liver receives nutrient-rich blood from the intestine through portal veins. In mammals, veins leaving the intestine converge on a primary portal.......

Materials

Name	Company	Catalog Number	Comments
16% Paraformaldehyde Aqueous Solution, EM Grade	Electron Microscopy Sciences	15700
50 mL Falcon Centrifuge Tubes, Polypropylene, Sterile	Corning	352098
AS 82/220.R2 PLUS Analytical Balance	Bay State Scale & Systems, INC.	WL-104-1051
Dumont #55 Forceps	Fine Science Tools	11295-51
EMS Kuehne Coverglass/Specimen Forceps	Electron Microscopy Sciences	72997-07
Epifluorescence microscope	Zeiss	Discovery.V8
Mastertop Cellulose Cleaning Scrub Sponge	Amazon	B07CBSM53Z
PBS10X Liquid Conc 4L	EMD Millipore	6505-4L
Super Fine Micro Scissors, 3 1/4" straight	Biomedical Research Instruments	11-1020
Tricaine methanesulfonate	Syndel	TRIC-M-GR-0010
Tween 20, Fisher BioReagents	Fischer Scientific	BP337-500