Scientists in the U.K. have succeeded in deriving endometrial organoids using cells extracted from menstrual blood, opening the way to more relevant high-throughput screens for drug discovery and to personalized treatments for gynecological conditions.

The organoids also will make it possible to explore the factors influencing implantation and early pregnancy, research which ethical and practical issues make more or less impossible, and where animal models are of limited utility.

The menstrual flow organoids can be frozen and biobanked, allowing for future large-scale retrospective studies. They also can be seeded onto 3D collagen scaffolds, paving the way for co-culturing with recently derived blastocyst-type organoids that are models of the earliest, preimplantation stages of human development.

The noninvasive method for generating the menstrual flow organoids builds on earlier work in which the researchers at the universities of Cambridge and Manchester generated endometrial organoids from tissue biopsies taken from the wall of the uterus. These can be cultured long-term and have been shown to recapitulate the molecular signature of the endometrial glands, which secrete proteins necessary for the establishment of pregnancy and its maintenance in the first trimester.

To confirm the menstrual flow organoids accurately reflect the in vivo state of the endometrium, the researchers compared them with those derived from uterine tissue donated by women undergoing in vitro fertilization (IVF) procedures.

The results demonstrate that menstrual flow contains viable endometrial gland progenitor cells, and can be used to culture physiologically relevant organoids in a noninvasive manner, said Tereza Cindrova-Davies of the department of physiology, development and neuroscience at the University of Cambridge, who is lead author of a paper appearing in the June 17, 2021, issue of Communications Biology describing the research.

"We confirmed that the transcriptome of the resultant organoids is identical to that of organoids generated from an endometrial biopsy taken earlier in the same cycle," Cindrova-Davies told BioWorld Science.

Cindrova-Davies and colleagues, and other researchers elsewhere, previously have shown endometrial organoids are a useful model for investigating maternal/fetal interactions in early pregnancy and for studying the pathophysiology of endometriosis and endometrial cancer.

The noninvasive protocol will increase the potential uses of this model, Cindrova-Davies says. "I am confident that the technique can be used for high throughput screening of endometrial function, in a similar fashion to the use of cancer organoids for high throughput drug screening in other systems," she said.

In a pilot study, the researchers used donations from volunteers with normal menstrual cycles, succeeding in generating organoids from six samples. They repeated this with further donations from three of these volunteers.

To validate the protocol, the researchers then derived organoids from uterine tissue from IVF patients who had an endometrial scratch procedure, and compared them to organoids generated from menstrual flow.

Endometrial scratch is a procedure in which the wall of the uterus is deliberately damaged with a catheter. That is thought to promote an immune response which prompts blood vessel growth and the release of signaling molecules that promote implantation.

"We confirmed that these organoids faithfully reflect the in vivo state by comparing organoids derived from paired endometrial scratches and ensuing menstrual flow from the same cycle in patients undergoing IVF," Cindrova-Davies said.

Organoids obtained from menstrual blood or from scratch biopsies had the same morphology. While the yield of tissue fragments from menstrual flow was lower, the speed of organoid development was comparable with those cultured from scratch biopsies.

Menstrual flow organoids begin to form after 2 to 4 days in culture if grown from fresh samples, or from 5 days if the samples have been frozen.

RNA sequencing showed all the organoids had similar transcriptomes and the organoids derived from the same patient were matched pairs.

The transcriptomes were all rich in markers of progenitor cells; proliferation; epithelial cell lineage; epithelial gland development and function; epithelial secretory activity; receptors for pregnancy hormones and cilia formation.

Response to hormones

As a measure of the physiological relevance of the menstrual flow organoids, the researchers tested how they reacted to treatment with early pregnancy hormones, including progesterone, estrogen, prolactin, placental lactogen and chorionic gonadotrophin. The menstrual flow organoids reacted in "identical fashion" to their scratch biopsy counterparts.

The hormones induced an epithelial morphology analogous to the phenotype of early pregnancy, prompting the generation of hypersecretory tissue that produced 'uterine milk', a mix of enzymes, growth factors, hormones and other protein collectively termed the histotroph. This mix of proteins provides nutrition in the first trimester of pregnancy, before the placenta is established.

The importance of the histotroph in early pregnancy has been demonstrated in animal models. But while it is known that low levels of certain components of the histotroph occur in women who experience implantation failure or early pregnancy loss, details of how factors such as age and body mass index impact the histotroph are sketchy. Generating organoids noninvasively from a broad population of women will provide essential data to fill these knowledge gaps, the researchers say.

"This technique has wide-ranging impact for noninvasive investigation and personalized approaches to treatment of common gynecological conditions, such as endometriosis, and reproductive disorders, including failed implantation after IVF, and recurrent miscarriage," said Cindrova-Davies.

Secretions from the endometrial glands play a key role in determining receptiveness of the endometrium and the implantation competence of the blastocyst, as well as stimulating the development of the placenta during the first months of pregnancy, she noted. Deficient functioning of the glands therefore has a profound impact on pregnancy success and outcome.

"Our technique will, for example, enable patients undergoing fertility treatments to have their endometrial function assessed noninvasively prior to initiating a treatment cycle," Cindrova-Davies said. That will allow the hormonal regimen and timing of treatment to be tailored accordingly.

Cindova-Davies is now planning to explore the use of menstrual flow organoids in patients having IVF and, potentially, to test drugs and other therapeutic approaches that might improve conception and pregnancy outcomes.