A Protocol for Constructing a Mouse Model of Hypertensive Myocardial Fibrosis Using Angiotensin II

Summary

The continuous infusion of angiotensin II (Ang II) via osmotic pumps is one of the classic methods for constructing animal models of hypertensive myocardial fibrosis. This protocol systematically optimized the process of constructing this model and elaborated the key points of osmotic pump implantation, positioning, and Ang II concentration setting.

Abstract

The continuous infusion of Angiotensin II (Ang II) via osmotic pumps is a widely utilized approach for establishing animal models of hypertensive myocardial fibrosis. Nevertheless, this technique exhibits certain limitations in current reports, such as the instability of Ang II, inaccurate pump placement, and the lack of a standardized procedure. Current reports on the methodology for establishment remain relatively limited. This protocol has systematically optimized the establishment of Ang II osmotic pump-induced hypertensive myocardial fibrosis models in mice, including the standardized preparation of Ang II solution, calibration of dosage, precise subcutaneous implantation of osmotic pumps, and quantitative standards for model evaluation indicators. Experimental results demonstrated that Ang II osmotic pump-induced hypertensive myocardial fibrosis in mice with high survival rates. Systolic blood pressure (SBP) levels stabilized at 160 mmHg after seven days of subcutaneous implantation. The model mice had significantly reduced left ventricular ejection fractions and increased end-diastolic dimensions of the left ventricle. Compared with the sham surgery group, collagen deposition within hypertensive myocardial interstitial tissue and perivascular regions increased. This study, based on a small sample size (n = 6), preliminarily validated the feasibility of establishing a mouse model of hypertensive myocardial fibrosis using Ang II combined with an osmotic pump, and established a quantitative phenotype verification system.

Introduction

Myocardial fibrosis (MF), a major pathological outcome of hypertension, is characterized by excessive collagen fiber deposition within the damaged or compressed myocardium. It represents one of the most important forms of hypertension-mediated target organ damage and contributes to progressive cardiac dysfunction^1,2. Establishing appropriate animal models is essential for advancing mechanistic research and developing novel therapeutic strategies for hypertensive myocardial fibrosis. Angiotensin II (Ang II), a central effector of the renin-angiotensin system, plays a pivotal role in hypertension pathogenesis

Protocol

All experimental procedures were carried out according to the Guidelines for the Care and Use of Laboratory Animals of the National Institutes of Health, and the experimental protocols were approved by the Animal Research Committee of Southwest Medical University (approval number SWMU2020317).

1. Preparation of Ang II solution and filling of the osmotic pump

1. Select 12 SPF-grade male C57BL/6J mice (see Table of Materials) weighing 20 ± 1 g by selecting 8-week-old animals for their developmental uniformity to control for age-related confounders. Divide the mice into a sham surgery group and a....

Results

Twelve mice were implanted subcutaneously with osmotic pumps and survived. Model group all developed hypertension (n = 6). Seven days post-implantation, the mean SBP in the model group exceeded 160 mmHg (Figure 1A). On days 14, 21, and 28, the SBP in the model group consistently remained above 160 mmHg. In contrast, the sham surgery group maintained normal SBP levels (approximately 90-110 mmHg) throughout the experiment without significant elevation.

Twenty-eight .......

Discussion

This protocol optimized the key steps involved in the continuous infusion of Ang II to induce hypertensive myocardial fibrosis using an osmotic pump and detailed the critical aspects, such as the implantation positioning of the osmotic pump and the setting of Ang II concentration. The model 2004 pump was chosen for its 4-week duration, and the Ang II dose was calculated based on the nominal infusion rate, accounting for minimal inter-batch variability. The efficacy of this model was confirmed through SBP monitoring, card.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Ang II	Sigma-Aldrich Corporation	Sigma, A9525
Animal anesthesia machine	Shenzhen Reward Life Science Co., Ltd	R530
Animal ventilator	Shenzhen Reward Life Science Co., Ltd	R415
GraphPad Prism	GraphPad Software Inc.	Version 9.0
Intelligent non-invasive blood pressure monitor	Nippon Soft Dragon Co., Ltd	Softron, BP-98A
Light microscope	Yijingtong Optical Technology (Shanghai) Co., Ltd	OLYMPUS, SZ61TR
Masson Tricolor Staining Solution and Kit	Beijing Soleibao Technology Co., Ltd	Solarbio, G1340
Mice	Chengdu Dossy Exprimental Animals CO., Ltd	C57BL/6J
Osmotic pumps	ALZA Corporation	Alzet,Model 2004
Ultrasound imaging systems	FUJIFILM VisualSonics Corporation, Canada	FUJIFILM, Vevo 3100