The development of an extended normothermic ex vivo reperfusion model of the sheep uterus to evaluate organ quality after cold ischemia in relation to uterus transplantation

Abstract

Introduction: Uterus transplantation has recently proved that infertility in women with uterine factor infertility can be cured. It is still an experimental procedure with numerous critical details remaining to be established, including tolerance to warm and cold ischemic insults. In preparation for human uterus transplantation trials, most teams use the sheep as a model system for research and team training, since the vasculature and the uterus is of similar size as in the human. We, therefore, aimed to develop an ex vivo sheep uterus reperfusion platform that mimics the reperfusion situation so that initial assessments and comparisons can be performed without the need for costly and labor-intensive in vivo transplantation experiments. Material and methods: Isolated sheep uteri were perfused with the preservation solution IGL-1 and were then exposed to cold ischemia for either 4 (n = 6) or 48 hours (n = 7). Uteri were then reperfused for 48 hours under normothermic conditions with an oxygenated recirculating perfusate containing growth factors and synthetic oxygen carriers. Histological and biochemical analysis of the perfusate was conducted to assess reperfusion injury. Results: Quantification of cell density indicated no significant edema in the myometrium or in the endometrium of uteri exposed to 4 hours cold ischemia and then a normothermic ex vivo reperfusion for 48 hours. Only the outer serosa layer and the inner columnar luminal epithelial cells were affected by the reperfusion. However, a much faster and severe reperfusion damage of all uterine layers were evident during the reperfusion experiment following 48 hours of cold ischemia. This was indicated by major accumulation of extracellular fluid, presence of apoptotic-labeled glandular epithelial layer and vascular endothelium. A significant accumulation of lactate was measured in the perfusate with a subsequent decrease in pH. Conclusions: We developed a novel ex vivo sheep uterus model for prolonged perfusion. This model proved to be able to distinguish reperfusion injury-related differences associated to organ preservation. The experimental setup is a platform that

Analysis of blood parameters and molecular endometrial markers during early reperfusion in two ovine models of uterus transplantation

“…occur in transplantation models with deceased donor and after long-time of cold ischemia [39]. Neutrophils infiltration of the endometrium-an early marker of inflammation-was observed in all cases, confirming previous data in sheep after auto transplantation [40].…”

“…The time of reperfusion and ischemia being relatively short in our series, this probably explains why no signs of necrosis, stasis, apoptosis or oedema was detected by histological examination nor by stereology. These signs are observed with a longer period of ischemia which is more likely occur in transplantation models with deceased donor and after long-time of cold ischemia [ 39 ]. Neutrophils infiltration of the endometrium—an early marker of inflammation—was observed in all cases, confirming previous data in sheep after auto transplantation [ 40 ].…”

The Key Role of Warm and Cold Ischemia in Uterus Transplantation: A Review

“…In kidney transplantation, hypothermic perfusion machines, providing a continuous supply of metabolic substrates and maintaining microvascularization during CI, have proved to be effective during the storage of organs in CI; reducing the damage caused to the organ [67,68]. An ex vivo uterus reperfusion platform could be an interesting way in UTx to decrease ischemia- and reperfusion injury [17]. Similarly, the use of hemoglobin from the marine worm Arenicola marina, which has an enhanced affinity for O 2 , has also proved to be effective during CI for lung and kidney transplantation [69,70,71].…”

“…However, a much faster and more severe reperfusion injury of all uterine layers was evident during the reperfusion experiment after 48 h of cold ischemia. Significant accumulation of lactate was measured in the perfusate with a subsequent decrease in pH [17].…”

Custodiol-N Is Superior to Custodiol® Solution in Experimental Rat Uterus Preservation

“…In separate experiments, the uterus graft was proven to be resistant to both warm and cold ischemia [11,12]. To date, several different preservation solutions, such as IGL-1 ® [13], Celsior ® [14][15][16], University of Wisconsin [17,18], NaCl [17], Ringer's acetate [18], Perfadex ® [18], and Custodiol ® [19][20][21][22][23][24][25], were used in experimental and clinical practice of UTx. Based on literature, modulation of the solution used for uterus graft static cold storage (SCS) could prevent ROS formation with resulting in reduced cell damage [17].…”