Establishing a Swine Model to Study Uterus Dynamic Preservation and Transplantation

Summary

A detailed and reproducible swine uterus model is described, from surgical procurement to the initiation of machine perfusion, allowing for the study of uterus preservation in transplantation.

Abstract

To date, uterus transplantation is the only option for women with absolute uterine infertility, such as those with Rokitansky syndrome, to experience pregnancy and give birth. Despite the growing interest in uterus transplantation in recent years, several issues still require further research, including ischemia-reperfusion injury and its impact on graft quality and rejection. Recent literature has highlighted a thrombotic complication rate of up to 20% following uterus transplantation. This type of complication may result from hypoxia-induced endothelial cell damage, often leading to uterine graft rejection. Hypoxia is induced during static cold storage, which remains the gold standard for graft preservation in solid organ transplantation. Recently, dynamic preservation using machine perfusion has been shown to improve the long-term storage of conventional and marginal organs by reducing ischemic and hypoxic injury. In this protocol, we aim to describe every surgical step involved in porcine uterus procurement and dynamic preservation, based on both uterine pedicles, to enable the connection and initiation of the machine perfusion protocol.

Introduction

Uterus transplantation (UTx) has significantly developed over the last ten years, with several teams starting clinical research programs. To date, the main indication of UTx is absolute uterine infertility due to uterine agenesis, including the Mayer-Rokitansky-Küster-Hauser (MRKH) syndrome. MRKH syndrome is a congenital disorder with a prevalence of one in 5,000 female live births¹. UTx could potentially address additional causes of infertility, including those resulting from hysterectomy due to malignant disease, postpartum hemorrhage, uterine fibroids, infectious sequelae, and various congenital malformations. This suggests that approxi....

Protocol

All animals received humane care following the National Institute of Health Guide for the Care and Use of Laboratory Animals, and the protocols were approved by the Massachusetts General Hospital Institutional Animal Care and Use Committee (IACUC). Overall, 6 female Yucatan minipigs weighing 30-40 kg were used for uterus procurement, with four uteri undergoing SNMP. All animals were heparinized with one full dose (100 IU/kg) before euthanasia. Organ procurement occurred post-mortem with less than 60 min of warm ischemia. Other organs could have been harvested from the same donor for different studies, according to the "3Rs" principles²⁵....

Results

During perfusion, the system was connected to a pressure sensor that recorded the pressure during the experiment. The pressure was initially recorded for a uterus-free system, which was subtracted from pressure recordings during uterus perfusion to obtain the real organ pressure. The flow rate was adapted to maintain the pressure within the desired range and was controlled by the roller pump. The resistance was calculated using the formula R = P / Q (R: resistance (mmHg.mL.min^-1); P: pressure (mmHg); Q: flow r.......

Discussion

Uterus transplantation, often considered part of VCA, has rapidly developed in the last few years. In parallel, machine perfusion started to be explored in VCA since it demonstrated robust evidence in improving solid organ preservation. Hypothermic and subnormothermic machine perfusion has allowed up to 24 h preservation in swine models of myocutaneous and bone-containing VCA^26,27,28. Since the uterus presents challenges compara.......

Materials

Name	Company	Catalog Number	Comments
Affinity Pixie Oxygenation System	Medtronic	BBP241	Oxygenator
Bovin serum albumin	Sigma-Aldrich	A9647	Perfusate component
Calcium chloride dihydrate	Sigma-Aldrich	223506	Perfusate component
Carbon Dioxide Oxygen	Airgas	UN3156	Carbon Dioxide Oxygen mix gas
D-(+)-Glucose monohydrate	Sigma-Aldrich	49159	Perfusate component
Dexamethasone	Sigma-Aldrich	D2915	Perfusate component
Dextran	Thermo scientific	406271000	Perfusate component
Heparin sodium injection	Eugia Pharma	63739-953-25	Perfusate component
Humulin Regular Insulin human	Lilly	0002-8215-01	Perfusate component
Hydrocortisone sodium succinate	Pfizer	0009-0011-03	Perfusate component
Magnesium chloride hexa-hydrate	Sigma-Aldrich	M9272	Perfusate component
MasterFlex L/S	Cole-Parmer	77200-32	Roller pump
Polyethylene glycol 35000	Sigma-Aldrich	25322-68-3	Perfusate component
Potassium chloride	Sigma-Aldrich	7447-40-7	Perfusate component
Pressure Monitor, Portable, PM-P-1	Living Systems Instrumentation	PM-P-1	Pressure sensor
Radnoti Bubble Trap Compliance Chamber	Radnoti	130149	Bubble trap
RAPIDPoint500	Siemens	500	Blood Gas System
Sodium bicarbonate	Sigma-Aldrich	S5761	Perfusate component
Sodium chloride	Sigma-Aldrich	S9888	Perfusate component
Sodium hydroxide	Sigma-Aldrich	72068	Perfusate component
Sodium phosphate monobasique dihydrate	Sigma-Aldrich	71505	Perfusate component
Syringe 1 mL	BD	309659	Sample procurement
Vancomycine hydrochloride	Slate run pharmaceuticals	70436-021-82	Perfusate component