Left Atrial Stenosis Induced Pulmonary Venous Arterialization and Group 2 Pulmonary Hypertension in Rat

Summary

Left atrial stenosis (LAS) is a novel surgical technique used for studying group 2 pulmonary hypertension (PH) and mechanisms underlying pulmonary venous arterialization. Here, we present a protocol to constrict the left atrium using a titanium clip to cause pulmonary venous arterialization and moderate PH in a rat.

Abstract

The mechanism of mitral stenosis-induced pulmonary venous arterialization and group 2 pulmonary hypertension (PH) is unclear. There is no rodent model of group 2 PH, due to mitral stenosis (MS), to facilitate the investigation of disease mechanisms and potential therapeutic strategies. We present a novel rat model of pulmonary venous congestion-induced pulmonary venous arterialization and group 2 PH caused by left atrial stenosis (LAS). LAS is achieved by constricting the left atrium using a half-closed titanium clip. After the LAS surgery, a rat model with a transmitral inflow velocity greater than or equal to 2.0 m/s on echocardiography gradually develops pulmonary venous arterialization and group 2 PH over an 8- to 10-week period. In this protocol, we provide the step-by-step procedure of how to perform the LAS surgery. The presented LAS rat model mimics MS in humans and is useful for studying the underlying molecular mechanism of pulmonary venous arterialization and for the preclinical evaluation of therapies for group 2 PH.

Introduction

The purpose of this article is to demonstrate the step-by-step procedure of how to perform the LAS surgery in rats. Surgically induced LAS closely mimics MS and cor triatriatum in humans, which involve the creation of a mechanical obstruction in the left atrium¹. Obstruction of the left ventricular (LV) inflow often causes a congestion of the pulmonary venous circulation, and patients gradually develop PH. The World Health Organization classifies PH due to left heart diseases as group 2, which is the most prevalent group of PH^2,3,4. The diagnosis of PH....

Protocol

The LAS experimental protocol has been approved by the Jikei University School of Medicine Animal Care Committee and the University Research and Ethics Committee (protocol #2015-118).

1. Pre-operative Preparation

1. After arriving at the animal facility, provide 5-week-old male Sprague Dawley rats between 150 to 200 g with 1 week to acclimate to their new home prior to the operation.
2. Prepare the following equipment before the surgery by autoclaving: 1) a small animal respirator, 2) an anesthetic machine, 3) an intubation kit (composed of a pair of hemostat forceps, a tongue depressor, and an 18 G angiocatheter), 4) surgic....

Results

The effectiveness of the LAS is confirmed using echocardiography, 2 weeks postoperative. Rats with an LV inflow velocity greater than 2.0 m/s, measured with a four-chamber view, are considered to have developed significant stenosis (Figure 1) and reliably develop moderate PH and pulmonary venous arterialization 8 - 10 weeks post-LAS surgery.

Ten weeks post-LAS surgery, the rats in the LAS group show.......

Discussion

The LAS rat is a novel group 2 PH model that has already received substantial interest from researchers in the field^12,13. Comparing to the two existing group 2 models, namely the pulmonary vein stenosis (PVS) model¹⁴, using piglets, and the supracoronary aortic banding (SAB) rat model^8,9,10, the LAS rat model has several advantages. Compared to t.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
5-0 Prolene Suture	Johnson & Johnson - Ethicon	8725H	Polypropylene suture with HEMO-SEAL Technology
Anaesthesia Machine	Wakenyaku Co., Ltd.	BRTK-100A	Air pump and anaethesia machine
Angiocatheter guidewire	Self-made		10 cm guidewire glued to a 1 cc syringe
Chest retractor	Natsume Seisakusho Co., Ltd.	F-2
Chest tube 23G	Self-made		10 cc syringe attached to a 23G needle plus plastic tube
Curved forceps	Natsume Seisakusho Co., Ltd.	A-14
Heating pad	Vivaria	MP-916-NV	Keep body temperature at 37 degree celsius
Horizon Ligating Clips	Teleflex	REF 003200	Size Medium-Large
Horizon Manual-Load Ligating Clip Applier For Medium-Large Size Horizon	Teleflex	REF 337085	Ligation Clips Angled Jaw, (20cm)
Needle holder	Natsume Seisakusho Co., Ltd.	MC-40
Rodent Respirator	CWE Inc	SAR-830/P	Small animal ventilator
Scissors	Natsume Seisakusho Co., Ltd.	B-12	Straight scissors ideally with round tips
Straight forceps	Natsume Seisakusho Co., Ltd.	A-7
Tongue depressor	Uchida Yoko Co., Ltd.	8-615-2417	Use the wide end