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In This Article

  • Summary
  • Abstract
  • Introduction
  • Protocol
  • Representative Results
  • Discussion
  • Acknowledgements
  • Materials
  • References
  • Reprints and Permissions

Summary

An appropriate animal model is needed to understand the pathologic mechanisms underlying aortic valve stenosis (AVS) and to evaluate the efficacy of therapeutic interventions. The present protocol describes a new procedure for developing the AVS rabbit model via a direct balloon injury in vivo.

Abstract

Animal models are emerging as an important tool to understand the pathologic mechanisms underlying aortic valve stenosis (AVS) because of the lack of access to reliable sources of diseased human aortic valves. Among the various animal models, AVS rabbit models are one of the most commonly used in large animal studies. However, traditional AVS rabbit models require a long-term period of dietary supplementation and genetic manipulation to induce significant stenosis in the aortic valve, limiting their use in experimental studies. To address these limitations, a new AVS rabbit model is proposed, in which stenosis is induced by a direct balloon injury to the aortic valve. The present protocol describes a successful technique for inducing AVS in New Zealand white (NZW) rabbits, with step-by-step procedures for the preparation, the surgical procedure, and the post-operative care. This simple and reproducible model offers a promising approach for studying the initiation and progression of AVS and provides a valuable tool for investigating the underlying pathological mechanisms of the disease.

Introduction

It is increasingly recognized that the use of appropriate animal models can contribute to a better understanding of the pathological mechanisms underlying aortic valve stenosis (AVS) due to the lack of access to reliable sources of diseased human aortic valves associated with the progression of aortic stenosis (AS). Among the various animal models for studying AVS, rabbits are one of the most commonly used large-animal AVS models, and the AVS rabbit model is induced either through cholesterol/vitamin D2 supplementation or genetic manipulation1,2,3,

Protocol

All animal research procedures were approved and performed in accordance with the Laboratory Animals Welfare Act, the Guide for the Care and Use of Laboratory Animals, and the Guidelines and Policies for Animal Experiments provided by the Institutional Animal Care and Use Committee (IACUC) at the College of Medicine of The Catholic University of Korea (approval number: CUMC-2021-0176-05). The present study utilized 3 month old male New Zealand white (NZW) rabbits weighing 3.5-4.0 kg, which were maintained under standard .......

Representative Results

Rabbit AVS model induced by aortic valve injury
To induce the rabbit AVS model, male NZW rabbits weighing 3.5-4.0 kg were used for this study. According to the surgical procedures described in step 2 (Figure 2), the AVS model was established by aortic valve injury, which resulted in mechanical aortic valve degeneration and calcification. The control group included rabbits fed with a 0.5% cholesterol-enriched diet (high-cholesterol, HC) and 50,000 U of vitamin D2 (VitD2.......

Discussion

Animal AVS models are commonly used to study the pathological aspects of AVS, including the initiation and progression of AVS. This protocol introduces a new rabbit AVS model induced by a direct balloon injury to the aortic valve. In this study, the aortic valve injury model showed significant leaflet thickening and calcification. Compared to the mild AVS model induced by dietary supplementation, the aortic valve in the direct balloon injury model was selectively injured, leading to thickened cusps and restricted motion,.......

Acknowledgements

This work was supported by a National Research Foundation of Korea (NRF) grant funded by the Korean government (MSIT) (No. 2020R1A4A3079570), the Ministry of Education (No. 2021R1I1A1A01051425), and the Industrial Strategic Technology Development Program (No. 20014873) funded by the Ministry of Trade, Industry & Energy, Republic of Korea.

....

Materials

NameCompanyCatalog NumberComments
3-0 Silk sutureAILEESK312
4% paraformaldehyde(PFA)IntronIBS-BP031-2
Alizarin red SolutionMillporeTMS-008-C
ASAHI SION BLUE ASAHIGuide wire
Back Table CoverYuhan kimberly80101-30
Balloon In-deflation DeviceDemax MedicalDID30s
Bionet Veterinary monitorBIONETBM3 VET
C-ArmSIEMENS Healthcare GmbHCios alpha
Certified Rabbit DietPurina53224.7% Hydrogenated Coconut Oil, 0.5% Cholesterol, & 1% Molasse
Curadle Smart IncubatorAutoelexCS-CV206Intensive Care Unit (ICU)
ErgocalciferolSigma-aldrich E5750Vitamin D2
Fechtner conjunctiva forceps titaniumWORLD PRECISSION InstrumentWP1820
ForcepsHEBUHB203
GentamicinShin Poong
Glycopyrrolate SamChunDang
Greenflex NSDAI HAN PHARMNormal saline 500 mL
Hematoxylin solutionSigma-aldrich HT1079-1 SET
HeparinJW pharmaceutical25,000 U
Infusion set for single useSWOON MEDICAL
IodineGreen pharmaceutical
IodixanolGE HealthcareVisipaqueInflation solution (contrast agent)
IV catheter 22 GBD 382423
IV catheter 24 GBD382412
KetoprofenSamChunDang
Luer-Lok syringe 10 mLBecton Dickinson Medical
Luer-Lok syringe 3 mLBecton Dickinson Medical
MicroscopeOLYMPUSSZ61
MicrotomeThermoFisher ScientificHM 325
MT stain kitSigma-aldrichHT15-1kt
Needel holderSolco009-1304
Needle Holder with Lock and SutureJEUNGDO BIO & PLANTH-1222-18
ParaffinLK LABKOREAH06-660-107
PBSGibco10010-023
Potassium chloride 40Daihan PharmKCl
Prelude Ideal Hydrophilic SheathMERIT MEDICALPID4F11018SSSheath 4F
PTA Balloon Dilatation catheterBoston ScientificH749-3903280208-0Balloon catheter 8.0 mm
RompunElancoXylaxine
sterile GauzeDAE HAN Medical10 cm x 20 cm 
Surgical GlovesAnsellAnsell
Surgical GownYuhan kimberly90002-02
Surgical ScissorsNopa, GermanyAC020/16
Surgical Tape3M micopore1530-1
Syringe 1 mLShin Chang Medical
Syringe 10 mLShin Chang Medical
Tissue cassetteScilav koreaCas3003
Transducer gel SUNGHEUNGSH102
TridolYuhan Corp.Tramadol HCl
Ultrasound systemPhilpsAffiniti 50
Von Kossa stain kitAbcamab105689
Zoletil 50Virbac koreaTiletamine & zolazepam

References

  1. Aliev, G., Burnstock, G. Watanabe rabbits with heritable hypercholesterolaemia: A model of atherosclerosis. Histology and Histopathology. 13 (3), 797-817 (1998).
  2. Cimini, M., Boughner, D. R., Ronald, J. A., Aldington, L., Rogers, K. A.

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