The Chick Chorioallantoic Membrane (CAM) Model as a Tool to Study Ovarian Tissue Transplantation.

Ovarian tissue cryopreservation and transplantation is an effective strategy for preserving fertility but has one major drawback, namely massive follicle loss occurring shortly after reimplantation due to abnormal follicle activation and death. Rodents are benchmark models for investigating follicle activation, but the cost, time, and ethical considerations are becoming increasingly prohibitive, thus driving the development of alternatives. The chick chorioallantoic membrane (CAM) model is particularly attractive, being inexpensive and maintaining natural immunodeficiency up to day 17 postfertilization, making it ideal to study short-term xenografting of human ovarian tissue. The CAM is also highly vascularized and has been widely used as a model to explore angiogenesis. This gives it a remarkable advantage over in vitro models and allows the investigation of mechanisms affecting the early post-grafting follicle loss process. The protocol outlined herein aims to describe the development of a CAM xenografting model for human ovarian tissue, with specific insights into the effectiveness of the technique, the graft revascularization time frame, and the tissue viability across a 6 day grafting period.

Proteome-wide and matrisome-specific atlas of the human ovary computes fertility biomarker candidates and open the way for precision oncofertility.

Our modern era is witnessing an increasing infertility rate worldwide. Although some of the causes can be attributed to our modern lifestyle (e.g., persistent organic pollutants, late pregnancy), our knowledge of the human ovarian tissue has remained limited and insufficient to reverse the infertility statistics. Indeed, all efforts have been focused on the endocrine and cellular function in support of the cell theory that dates back to the 18th century, while the human ovarian matrisome is still under-described. Hereby, we unveil the extracellular side of the story during different periods of the ovary life, demonstrating that follicle survival and development, and ultimately fertility, would not be possible without its involvement. We examined the human ovarian matrisome and described its remodeling from prepuberty until menopause, creating the first ovarian proteomic codex. Here, we confidently identified and quantified 98 matrisome proteins present in the three ovary groups. Among them, 26 were expressed differently among age groups, delineating a peculiar matrisomal fingerprint at each stage. Such proteins could be potential biomarkers phenotyping ovarian ECM at each age phase of female reproductive life. Beyond proteomics, our study presents a unique approach to understanding the data and depicting the spatiotemporal ECM-intracellular signaling networks and remodeling with age through imaging, advanced text-mining based on natural language processing technology, machine learning, and data sonification. Our findings provide essential context for healthy ovarian physiology, identifying and characterizing disease states, and recapitulating physiological tissues or development in vitro. This comprehensive proteomics analysis represents the ovarian proteomic codex and contributes to an improved understanding of the critical roles that ECM plays throughout the ovarian life span.

A blueprint of the topology and mechanics of the human ovary for next-generation bioengineering and diagnosis.

Although the first dissection of the human ovary dates back to the 17th century, the biophysical characteristics of the ovarian cell microenvironment are still poorly understood. However, this information is vital to deciphering cellular processes such as proliferation, morphology and differentiation, as well as pathologies like tumor progression, as demonstrated in other biological tissues. Here, we provide the first readout of human ovarian fiber morphology, interstitial and perifollicular fiber orientation, pore geometry, topography and surface roughness, and elastic and viscoelastic properties. By determining differences between healthy prepubertal, reproductive-age, and menopausal ovarian tissue, we unravel and elucidate a unique biophysical phenotype of reproductive-age tissue, bridging biophysics and female fertility. While these data enable to design of more biomimetic scaffolds for the tissue-engineered ovary, our analysis pipeline is applicable for the characterization of other organs in physiological or pathological states to reveal their biophysical markers or design their bioinspired analogs.

Important role of collective cell migration and nerve fiber density in the development of deep nodular endometriosis.

OBJECTIVE: To evaluate deep nodular endometriotic lesions induced in baboons over 12 months and analyze collective cell migration and nerve fiber density. DESIGN: Morphologic and immunohistochemical analysis of endometriotic lesions induced in baboons over the course of 1 year. SETTING: Academic research unit. ANIMAL(S): Three female baboons (Papio anubis). INTERVENTION(S): Recovery of induced deep nodular endometriotic nodules from baboons. MAIN OUTCOME MEASURE(S): Evaluation of the morphology of glands by analysis of the center of lesions and the invasion front; immunohistochemical staining with Ki67, E-cadherin, and ß-catenin for investigation of mitotic activity and cell-cell junctions, and with protein gene product 9.5 and nerve growth factor (NGF) for study of nerve fiber density (NFD). RESULT(S): All (100%) of the lesions were invasive 1 year after induction, compared with 42.29% after 6 months. Glands from the invasion front showed significantly reduced thickness but significantly higher mitotic activity. E-Cadherin and ß-catenin expression were similar between the center and front. NFD was significantly higher in lesions induced after 1 year than after 6 months, and NGF expression was significantly lower in 1-year lesions than in 6-month lesions. CONCLUSION(S): Nodular endometriotic lesions induced in the baboon model were found to be significantly more invasive and innervated after 12 months than after 6 months. The invasive phenotype was highly expressed in glands at the invasion front, and our study suggests that nerve fibers play a role in the development of lesions as observed in women.

Invasion process of induced deep nodular endometriosis in an experimental baboon model: similarities with collective cell migration?

OBJECTIVE: To determine the implications of collective cell migration in the invasion phenomenon observed in deep endometriotic lesions induced in a baboon model. DESIGN: Study of morphology and collective cell migration markers in invasive and noninvasive deep endometriotic lesions induced in a baboon model. Invasive lesions were defined as the presence of endometrial glands and stroma in surrounding organs, and a distinction was made between the center of the lesion (glands present in the main lesion) and the invasion front (glands present in surrounding organs). SETTING: Academic research unit. ANIMAL(S): Ten female baboons (Papio anubis). INTERVENTION(S): Recovery of induced deep nodular endometriotic nodules. MAIN OUTCOME MEASURE(S): Evaluation of the morphology of glands by analysis of noninvasive and invasive lesions (center of the lesion and invasion front); staining with specific antibodies (Ki67, E-cadherin, ß-catenin) for immunohistochemical study of mitotic activity and cell-cell junctions. RESULT(S): Glands from invasive lesions, particularly from the invasion front, showed a significantly lower thickness coefficient, higher mitotic activity, and lower expression of E-cadherin and ß-catenin than glands from noninvasive lesions and the center of invasive lesions. CONCLUSION(S): We report altered morphology, increased mitotic activity, and fewer adhesion molecules in invasive glands present in induced nodular endometriosis, particularly along the invasion front, suggesting that collective cell migration is involved in the invasion process of deep endometriotic lesions induced in a baboon model.

Differential expression of steroidogenic enzymes according to endometriosis type.

OBJECTIVE: To evaluate, in peritoneal, ovarian, and rectovaginal endometriotic lesions, expression of steroidogenic enzymes involved in the activation and inactivation of estrogens: 17ß-hydroxysteroid dehydrogenase type 1 (HSD17B1) and 2 (HSD17B2), estrone sulfotransferase (EST), and steroid sulfatase (STS). SETTING: Academic gynecology research unit. DESIGN: Retrospective study. PATIENT(S): Disease-free (n = 41) patients and patients with endometriosis (n = 79) were included for quantitative polymerase chain reaction (q-PCR) (15 disease-free, 33 endometriosis) and immunohistochemistry (26 disease-free, 46 endometriosis) studies. INTERVENTION(S): Q-PCR and immunohistochemistry. MAIN OUTCOME MEASURE(S): Evaluation of mRNA and protein expression. RESULT(S): Glandular HSD17B1, HSD17B2, and STS protein expression were demonstrated. HSD17B2 mRNA values were higher in the secretory phase of the menstrual cycle in the endometrium of disease-free women, but not in the eutopic endometrium of patients with endometriosis. HSD17B1 mRNA was equally expressed in the various tissues investigated, and EST mRNA was expressed at low levels in the different lesion types. HSD17B2 mRNA expression was decreased in ovarian and rectovaginal endometriosis compared with eutopic endometrium, while STS mRNA was increased in rectovaginal lesions compared with ovarian lesions. Ratios between pro- and antiestrogenic enzymes (STS/EST and HSD17B1/HSD17B2) were more in favor of estrogens in ovarian and rectovaginal endometriosis. CONCLUSION(S): In endometriosis development, local activation of estrogens appears to be important. STS and HSD17B1 inhibitors may therefore prove useful to treat the disease.

Induction of endometriotic nodules in an experimental baboon model mimicking human deep nodular lesions.

OBJECTIVE: To establish an experimental model for the study of deep nodular endometriosis. DESIGN: Induction of nodular endometriosis in baboons by grafting different uterine specimens to the peritoneal cavity. SETTING: Research and university facilities. ANIMAL(S): Ten baboons, to develop a model of induced deep nodular endometriosis. INTERVENTION(S): Biopsies of endometrium, and endometrium plus the junctional zone (JZ), full uterine thickness, and myometrium grafted to the peritoneum. MAIN OUTCOME MEASURE(S): Macroscopic descriptions recorded for observed induced lesions; staining with hematoxylin and eosin for histological evaluation and specific antibodies (CK22, CD10) for immunohistochemical studies; and analysis of surface area and volume of lesions, glandular density, and invasion of surrounding organs. RESULT(S): The incidence of induced nodular endometriosis was 100%, but the extent depended on the tissue grafted. Lesions induced after grafting specimens containing the JZ were statistically significantly larger than those not containing the JZ. Surrounding organ invasion was reported in more than 40% of lesions after grafting specimens containing the JZ. CONCLUSION(S): The first experimental model of nodular endometriosis allows investigation of deeper nodular lesions as well as invasion phenomena associated with nodular lesions.