Allo- and Xeno-Reassembly of Human and Rat Myometrium from Cells and Scaffolds

Abstract

Neo-myometrium was created by culturing isolated myocytes into decellularized rat and human myometrial scaffolds. The dual purpose of the uterus is to accommodate the growing fetus, and then expel the fetus at term by phasically contracting it. The first function requires physical robustness as well as the ability to expand and remodel. Congenital anomalies or previous surgeries can mechanically compromise the uterus and lead to major complications in pregnancy. The second function utilizes multiple interactions of complex physiological mechanisms that have yet to be fully elucidated, and this knowledge gap contributes to the continuation of serious complications of pregnancy. To address both problems, we reconstructed myometrium from isolated myocytes and scaffold. From both rat and human myometrium, myocytes were isolated using collagenase digestion, and scaffolds were isolated using ethanol/ trypsin protocols. The number of myocytes was amplified using monolayer culture, and then, the myocytes were cultured back into the scaffolds. We called this engineered tissue neo-myometrium, with allo-neo-myometrium being made from components of the same species, and xeno-neo-myometrium from across species. By artificially creating defects in rat scaffold, alloneo-myometrium was created that demonstrated rapid scaffold remodeling. Xeno-neo myometrium (human myocytes/rat scaffold) was created and demonstrated myocytes occurring in bundles 500 mm deep in the scaffold. These experiments provide proof of principle that modest numbers of myocytes can be amplified and used to create a larger volume of engineered tissue, which may be useful for semi-autologous transplantation to repair structural defects of the human uterus. In isometric contractility experiments, coordinated contractions were observed in xeno-neo-myometrium (human myocytes, rat scaffold), but not allo-neo-myometrium (human myocytes, human scaffold).

Uterine Tissue Engineering and the Future of Uterus Transplantation

“…These constructs were cultured for up to 51 days in vitro which gave the cells enough time to form multicellular layers on the scaffold surface and clusters of cells within the depths of the rat scaffold. These constructs showed some contractility in an organ bath, indicating elementary uterus phenotype-like functionality 113. However, no repopulation of the vascular conduits was observed which may have affected the cell homeostasis deeper in the tissue layers.…”

Towards the development of a bioengineered uterus: Comparison of different protocols for rat uterus decellularization

“…Concerning the female internal genital tract, small patches of myometrial tissue were recently decellularized using a modified protocol that had been optimized for decellularization of blood vessels [22]. Other tissue engineering approaches for uterine tissue include the use of scaffolds of collagen/matrigel [23][24][25] or silk/collagen [26], as well as scaffolds made from blood clots [27].…”

Methods and Model Systems Used to Study Pregnant Human Uterine Smooth Muscle

“…This naturally occurring 3D biologic scaffold can be then recellularized with primary SMCs that will differentiate into the original type of tissue (Figure 3). This approach has been successfully used in animal studies as well as in human clinical applications [53][54][55][56][57][58][59], and more recently, in myometrium [60]. Young et al [60] used tissue-engineering techniques to reconstruct allo-and xeno-neo-myometrium from isolated rat and human myocytes and Figure 3.…”