PUBLICATION

Generation and Application of the Zebrafish heg1 Mutant as a Cardiovascular Disease Model

Authors

Lu, S., Hu, M., Wang, Z., Liu, H., Kou, Y., Lyu, Z., Tian, J.

ID

ZDB-PUB-201120-147

Date

2020

Source

Biomolecules 10(11): (Journal)

Registered Authors

Tian, Jing

Keywords

CRISPR/Cas9, Traditional Chinese Medicine (TCM), cardiovascular disease (CVD), dilated cardiomyopathy (DCM), drug screening, heg1, thrombosis, zebrafish model

MeSH Terms

Female
Zebrafish Proteins/genetics
Zebrafish Proteins/metabolism*
Membrane Glycoproteins/genetics
Membrane Glycoproteins/metabolism*
Blood Circulation
Gene Knockout Techniques
Mutation
Zebrafish/genetics*
Zebrafish/metabolism
Humans
Cardiovascular Diseases/genetics
Cardiovascular Diseases/metabolism*
Cardiovascular Diseases/physiopathology
Disease Models, Animal*
Heart Rate
Animals
Male

(all 18)

PubMed

33198188 Full text @ Biomolecules

Abstract

Cardiovascular disease (CVD) is the leading cause of global mortality, which has caused a huge burden on the quality of human life. Therefore, experimental animal models of CVD have become essential tools for analyzing the pathogenesis, developing drug screening, and testing potential therapeutic strategies. In recent decades, zebrafish has entered the field of CVD as an important model organism. HEG1, a heart development protein with EGF like domains 1, plays important roles in the development of vertebrate cardiovascular system. Loss of HEG1 will affect the stabilization of vascular endothelial cell connection and eventually lead to dilated cardiomyopathy (DCM). Here, we generated a heg1-specific knockout zebrafish line using CRISPR/Cas9 technology. Zebrafish heg1 mutant demonstrated severe cardiovascular malformations, including atrial ventricular enlargement, heart rate slowing, venous thrombosis and slow blood flow, which were similar to human heart failure and thrombosis phenotype. In addition, the expression of zebrafish cardiac and vascular markers was abnormal in heg1 mutants. In order to apply zebrafish heg1 mutant in cardiovascular drug screening, four Traditional Chinese Medicine (TCM) herbs and three Chinese herbal monomers were used to treat heg1 mutant. The pericardial area, the distance between sinus venosus and bulbus arteriosus (SV-BA), heart rate, red blood cells (RBCs) accumulation in posterior cardinal vein (PCV), and blood circulation in the tail vein were measured to evaluate the therapeutic effects of those drugs on DCM and thrombosis. Here, a new zebrafish model of DCM and thrombosis was established, which was verified to be suitable for drug screening of cardiovascular diseases. It provided an alternative method for traditional in vitro screening, and produced potential clinical related drugs in a rapid and cost-effective way.

Genes / Markers

Figures

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Expression

Gene	Fish	Conditions	Stage	Anatomy	Assay	Figure
fabp2	twu34Tg	standard conditions	Protruding-mouth	heart	RTPCR	Figure 1
fabp2	twu34Tg	standard conditions	Protruding-mouth	whole organism	RTPCR	Figure 1
flt4	heg1^nwc1/nwc1	standard conditions	Long-pec	whole organism	RTPCR	Figure 4
flt4	WT	standard conditions	Long-pec	whole organism	RTPCR	Figure 4
heg1	heg1^nwc1/nwc1	standard conditions	Protruding-mouth	whole organism	RTPCR	Figure 2
heg1	twu34Tg	standard conditions	Protruding-mouth	heart	RTPCR	Figure 1
heg1	WT	standard conditions	1k-cell	whole organism	RTPCR	Figure 1
heg1	WT	standard conditions	30%-epiboly	whole organism	RTPCR	Figure 1
heg1	WT	standard conditions	Germ-ring	whole organism	RTPCR	Figure 1
heg1	WT	standard conditions	5-9 somites	whole organism	RTPCR	Figure 1

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Phenotype

Fish	Conditions	Stage	Phenotype	Figure
heg1^nwc1/nwc1	standard conditions	Long-pec	heart increased size, abnormal	Figure 3
heg1^nwc1/nwc1	standard conditions	Long-pec	whole organism flt4 expression increased amount, abnormal	Figure 4
heg1^nwc1/nwc1	standard conditions	Long-pec	whole organism kdrl expression increased amount, abnormal	Figure 4
heg1^nwc1/nwc1	standard conditions	Long-pec	whole organism myb expression increased amount, abnormal	Figure 4
heg1^nwc1/nwc1	standard conditions	Long-pec	whole organism myh6 expression increased amount, abnormal	Figure 4
heg1^nwc1/nwc1	standard conditions	Long-pec	whole organism myl7 expression increased amount, abnormal	Figure 4
heg1^nwc1/nwc1	standard conditions	Long-pec	whole organism sox7 expression increased amount, abnormal	Figure 4
heg1^nwc1/nwc1	standard conditions	Protruding-mouth	eye edematous, abnormal	Figure 3
heg1^nwc1/nwc1	standard conditions	Protruding-mouth	heart increased size, abnormal	Figure 3
heg1^nwc1/nwc1	standard conditions	Protruding-mouth	pericardium edematous, abnormal	Figure 3

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Mutations / Transgenics

Allele	Construct	Type	Affected Genomic Region
nwc1		Small Deletion	heg1
s843Tg	Tg(kdrl:EGFP)	Transgenic Insertion
twu34Tg	Tg(myl7:EGFP)	Transgenic Insertion

Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
heg1	CRISPR1-heg1	CRISPR

Fish

Fish
heg1^nwc1/nwc1
twu34Tg
WT

Antibodies

Orthology

Engineered Foreign Genes

Marker	Marker Type	Name
EGFP	EFG	EGFP

Mapping

Lu et al., 2020 ZDB-PUB-201120-147 PMID:33198188

Generation and Application of the Zebrafish heg1 Mutant as a Cardiovascular Disease Model

Lu et al., 2020

ZDB-PUB-201120-147
PMID:33198188