Surgical Approach, Challenges, and Resolutions for Uterus Transplantation in Rats

Summary

The present protocol describes all the essential steps for successful uterine transplantation (UTx) in rats. The rat model has proven suitable to promote the clinical implementation of UTx; however, rat UTx is a highly complex procedure requiring careful instructions.

Abstract

Uterine transplantation (UTx) is a new approach for treating women with absolute uterine factor infertility (AUFI). An estimated 3%-5% of women suffer from AUFI. These women were deprived of the option to have children until the advent of UTx. The clinical application of UTx was driven by experimental studies in animals, and the first successful UTx was achieved in rats. Given their physiological, immunological, genetic, and reproductive characteristics, rats are a suitable model system for such transplants. In particular, their short gestation period is a clear advantage, as the usual endpoint of experimental UTx is successful pregnancy with live birth. The biggest challenge for rat models remains the small anatomy, which requires advanced microsurgical skills and experience. Although UTx has led to pregnancy in the clinic, the procedure is not established and requires continuous experimental optimization. Here, a detailed protocol is presented, including essential troubleshooting for rat UTx, which is expected to make the entire procedure easier to grasp for those without experience in this type of microsurgery.

Introduction

Uterine transplantation (UTx) is a novel treatment for absolute uterine factor infertility (AUFI). AUFI results from an absence (congenital or acquired) or malformation of the uterus and affects 3%-5% of women worldwide¹. Ethical, legal, or religious reasons rule out adoption or surrogacy for many women who have a desire for motherhood but suffer from AUFI². For these women, UTx remains the only option to start their own family. UTx has been applied in the clinic, albeit with mixed success; the procedure is technically challenging and requires steady improvement for its clinical establishment.

Protocol

All animal experiments were performed following Swiss Federal Animal Regulations and approved by the Veterinary Office of Zurich (n° 225/2019), assuring human care. Female virgin Lewis rats (body weight of 170-200 g) and female virgin Brown Norway rats (170-200 g) were used as uterus donors/recipients, while male Lewis rats (300-320 g) were used for mating. The rats were aged from 12-15 months. The animals were obtained from commercial sources (see Table of Materials) and were housed in controlled conditions and an enriched environment with free access to water and standard food.

1. Uterus retrieval

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Results

Results from two groups of rats are presented. UTx was carried out before (group 1, n = 8) and after (group 2, n = 8) protocol adjustment (Table 1) to demonstrate the effects of our modifications (please see the Discussion for an explanation of our modifications)^12,15,21.

The outcome of rat UTx is associated with three key phases. The first phase is successful recover.......

Discussion

The protocol presented here offers detailed instructions for the surgical approach behind uterus transplantation in rats. The protocol has been optimized to increase the odds of live births following UTx and subsequent mating. The original protocol has been taken over from the Brännström group^12,13, inspired by the mouse work of Akouri et al.¹⁰, and modified based on the authors' experiences over the past years. As such, the .......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Angled to Side Scissor 5 mm	F.S.T	15008-08
Big Paper Clip	No specific		Used as retractor
Blunt Bend Needle G30	Unimed S.A.
Bupivacain 0.5%	Sintetica
Buprenorphine 0.3 mg/mL	Temgesic
Dosiernadel G25	H.SIGRIST& PARTNER AG
Dumont #5SF Forceps	F.S.T	11252-00
Ethilon 10/0	Ethicon	2810G
Ethilon 6/0	Ethicon	667H
Ethilon 7/0	Ethicon	EH7446H
Ethilon 8/0	Ethicon	2808G
Femal Brown Norway Rats (150-170 g)	Janvier
Femal Lewis Rats (150-170 g)	Charles River Deutschland
Fine Scissors - Sharp	F.S.T	14060-09	Any other small scissor works too
Halsey Micro Needle Holder	F.S.T	12500-12	Any other small needholder works too
Heparin Natrium 25000 I.E./ 5 mL	B. Braun
Institute Georges Lopez Perfusion Solution (IGL)	Institute Georges Lopez		Organ preservation solution
Male Lewis Rats (300-320 g)	Charles River Deutschland
Micro Serrefines 13 mm	F.S.T	18055-04
Micro Serrefines 16 mm gebogen	F.S.T	18055-06
Micro-Serrefine Clamp Applicator with Lock	F.S.T	18056-14
Mölnlyncke Op Towel	Mölnlyncke	800300	Sterile drape
NaCl 0.9%	B.Braun
Octenisept	Schülke
Paper Tape	Tesa		For fixing the animal
Philips Avent Schneller Flaschenwärmer SCF358/02	Philips	12824216
Ringerfundin	B.Braun
Rompun 2%	Bayer		Xylazine
Round Handled Needle Holders	F.S.T	12075-12
Round Handled Needle Holders	F.S.T	12075-12
S&T Vessel Dilating Forceps - Angled 45°	F.S.T	00276-13
Sacryl Naht	KRUUSE	152575
Scapel No 10	Swann Morton	201
Small Histo-Container			Any small histo-container works fine-for coldstorage of the graft
Small Plastik Bags			Any transparant plastic bags are fine
Steril Cotton swab	Lohmann-Rauscher		Any steril cotton swab is fine
Sterile Gauze	Lohmann-Rauscher		Any steril gauze is fine
Straight Scissor 8mm	F.S.T	15024-10
Surgical microscope – SZX9	Olympus	OLY-SZX9-B
Sutter Non Stick GLISS 0.4 mm	Sutter	78 01 69 SLS
Suture Tying Forceps	F.S.T	00272-13
ThermoLux warming mat	ThermoLux
Tissue Forceps for Skin			Any tissue forceps are fine
Vesseldilatator Forceps	F.S.T	00125-11
Vicryl  plus 4/0	Ethicon	VCP292H