Decellularization techniques support the creation of biocompatible extracellular matrix hydrogels, providing tissue-specific environments for both in vitro cell culture and in vivo tissue regeneration. We obtained endometrium derived from porcine decellularized uteri to create endometrial extracellular matrix (EndoECM) hydrogels. After decellularization and detergent removal, we investigated the physicochemical features of the EndoECM, including gelation kinetics, ultrastructure, and proteomic profile. The matrisome showed conservation of structural and tissue-specific components with low amounts of immunoreactive molecules. EndoECM supported in vitro culture of human endometrial cells in two- and three-dimensional conditions and improved proliferation of endometrial stem cells with respect to collagen and Matrigel. Further, we developed a three-dimensional endometrium-like co-culture system of epithelial and stromal cells from different origins. Endometrial co-cultures remained viable and showed significant remodeling. Finally, EndoECM was injected subcutaneously in immunocompetent mice in a preliminary study to test a possible hypoimmunogenic reaction. Biomimetic endometrial milieus offer new strategies in reproductive techniques and endometrial repair and our findings demonstrate that EndoECM has potential for in vitro endometrial culture and as treatment for endometrial pathologies.

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Adult stem cells (ASCs) were long suspected to exist in the endometrium. Indeed, several types of endometrial ASCs were identified in rodents and humans through diverse isolation and characterization techniques. Putative stromal and epithelial stem cell niches were identified in murine models using label-retention techniques. In humans, functional methods (clonogenicity, long-term culture, and multi-lineage differentiation assays) and stem cell markers (CD146, SUSD2/W5C5, LGR5, NTPDase2, SSEA-1, or N-cadherin) facilitated the identification of three main types of endogenous endometrial ASCs: stromal, epithelial progenitor, and endothelial stem cells. Further, exogenous populations of stem cells derived from bone marrow may act as key effectors of the endometrial ASC niche. These findings are promoting the development of stem cell therapies for endometrial pathologies, with an evolution towards paracrine approaches. At the same time, promising therapeutic alternatives based on bioengineering have been proposed.

Objective. To study the effect of human plasma from different sources such as 2 umbilical cord blood and adult blood platelet-rich plasma (PRP) on the 3 regeneration of endometrial damage.

BACKGROUND To provide the optimal milieu for implantation and fetal development, the female reproductive system must orchestrate uterine dynamics with the appropriate hormones produced by the ovaries. Mature oocytes may be fertilized in the fallopian tubes, and the resulting zygote is transported toward the uterus, where it can implant and continue developing. The cervix acts as a physical barrier to protect the fetus throughout pregnancy, and the vagina acts as a birth canal (involving uterine and cervix mechanisms) and facilitates copulation. Fertility can be compromised by pathologies that affect any of these organs or processes, and therefore, being able to accurately model them or restore their function is of paramount importance in applied and translational research. However, innate differences in human and animal model reproductive tracts, and the static nature of 2D cell/tissue culture techniques, necessitate continued research and development of dynamic and more complex in vitro platforms, ex vivo approaches and in vivo therapies to study and support reproductive biology. To meet this need, bioengineering is propelling the research on female reproduction into a new dimension through a wide range of potential applications and preclinical models, and the burgeoning number and variety of studies makes for a rapidly changing state of the field. OBJECTIVE AND RATIONALE This review aims to summarize the mounting evidence on bioengineering strategies, platforms and therapies currently available and under development in the context of female reproductive medicine, in order to further understand female reproductive biology and provide new options for fertility restoration. Specifically, techniques used in, or for, the uterus (endometrium and myometrium), ovary, fallopian tubes, cervix and vagina will be discussed. SEARCH METHODS A systematic search of full-text articles available in PubMed and Embase databases was conducted to identify relevant studies published between January 2000 and September 2021. The search terms included: bioengineering, reproduction, artificial, biomaterial, microfluidic, bioprinting, organoid, hydrogel, scaffold, uterus, endometrium, ovary, fallopian tubes, oviduct, cervix, vagina, endometriosis, adenomyosis, uterine fibroids, chlamydia, Asherman’s syndrome, intrauterine adhesions, uterine polyps, polycystic ovary syndrome and primary ovarian insufficiency. Additional studies were identified by manually searching the references of the selected articles and of complementary reviews. Eligibility criteria included original, rigorous and accessible peer-reviewed work, published in English, on female reproductive bioengineering techniques in preclinical (in vitro/in vivo/ex vivo) and/or clinical testing phases. OUTCOMES Out of the 10 390 records identified, 312 studies were included for systematic review. Owing to inconsistencies in the study measurements and designs, the findings were assessed qualitatively rather than by meta-analysis. Hydrogels and scaffolds were commonly applied in various bioengineering-related studies of the female reproductive tract. Emerging technologies, such as organoids and bioprinting, offered personalized diagnoses and alternative treatment options, respectively. Promising microfluidic systems combining various bioengineering approaches have also shown translational value. WIDER IMPLICATIONS The complexity of the molecular, endocrine and tissue-level interactions regulating female reproduction present challenges for bioengineering approaches to replace female reproductive organs. However, interdisciplinary work is providing valuable insight into the physicochemical properties necessary for reproductive biological processes to occur. Defining the landscape of reproductive bioengineering technologies currently available and under development for women can provide alternative models for toxicology/drug testing, ex vivo fertility options, clinical therapies and a basis for future organ regeneration studies.

Organoids are three-dimensional (3D) multicellular tissue models that mimic their corresponding in vivo tissue. Successful efforts have derived organoids from primary tissues such as intestine, liver, and pancreas. For human uterine endometrium, the recent generation of 3D structures from primary endometrial cells is inspiring new studies of this important tissue using precise preclinical models. To improve on these 3D models, we decellularized pig endometrium containing tissue-specific extracellular matrix and generated a hydrogel (EndoECM). Next, we derived three lines of human endometrial organoids and cultured them in optimal and suboptimal culture expansion media with or without EndoECM (0.01 mg/mL) as a soluble additive. We characterized the resultant organoids to verify their epithelial origin, long-term chromosomal stability, and stemness properties. Lastly, we determined their proliferation potential under different culture conditions using proliferation rates and immunohistochemical methods. Our results demonstrate the importance of a bioactive environment for the maintenance and proliferation of human endometrial organoids.

The oviducts (fallopian tubes in mammals) function as the site of fertilization and provide necessary support for early embryonic development, mainly via embryonic exposure to the tubal microenvironment. The main objective of this study was to create an oviduct-specific extracellular matrix (oviECM) hydrogel rich in bioactive components that mimics the native environment, thus optimizing the developmental trajectories of cultured embryos. Rabbit oviducts were decellularized through SDS treatment and enzymatic digestion, and the acellular tissue was converted into oviductal pre-gel extracellular matrix (ECM) solutions. Incubation of these solutions at 37 °C resulted in stable hydrogels with a fibrous structure based on scanning electron microscopy. Histological staining, DNA quantification and colorimetric assays confirmed that the decellularized tissue and hydrogels contained no cellular or nuclear components but retained important components of the ECM, e.g. hyaluronic acid, glycoproteins and collagens. To evaluate the ability of oviECM hydrogels to maintain early embryonic development, two-cell rabbit embryos were cultured on oviECMcoated surfaces and compared to those cultured with standard techniques. Embryo development was similar in both conditions, with 95.9% and 98% of the embryos reaching the late morula/early blastocyst stage by 48 h under standard culture and oviECM conditions, respectively. Metabolomic analysis of culture media in the presence or absence of embryos, however, revealed that the oviECM coating may include signalling molecules and release compounds beneficial to embryo metabolism.

Endometrial function is essential for embryo implantation and pregnancy, but managing endometrial thickness that is too thin to support pregnancy or an endometrium of compromised functionality due to intrauterine adhesions is an ongoing challenge in reproductive medicine. Here, we review current and emerging therapeutic and experimental options for endometrial regeneration with a focus on animal models used to study solutions for Asherman's syndrome and endometrial atrophy, which both involve a damaged endometrium. A review of existing literature was performed that confirmed the lack of consensus on endometrial therapeutic options, though promising new alternatives have emerged in recent years (platelet-rich plasma, exosomes derived from stem cells, bioengineering-based techniques, endometrial organoids, among others). In the future, basic research using established experimental models of endometrial pathologies (combined with new high-tech solutions) and human clinical trials with large population sizes are needed to evaluate these emerging and new endometrial therapies.