Michela Calanni-Pileri

Preeclampsia is a leading cause of morbidity and mortality in pregnant women, affecting 5–8% of gestations worldwide. Its development is influenced by maternal immune abnormalities, metabolic disorders, and gut dysbiosis. In this study, we show that gut dysbiosis in pregnant C57BL/6J dams leads to increased fetal resorption, impaired placental development and altered vascularization. These adverse outcomes are associated with key pathological features of preeclampsia, including hypoxia, endoplasmic reticulum (ER) stress and reduction in uterine natural killer (NK) cell numbers. Furthermore, gut dysbiosis significantly perturbs placental carbohydrate metabolism, which impairs NK cell IFN-γ secretion. Notably, glucose supplementation restores placental NK cell function and reduces fetal resorption, suggesting that the observed impairment is reversible and dependent on a lower glycolytic rate. These findings highlight maternal gut microbiota as a key player in carbohydrate metabolism, with a pivotal role in modulating placental immunity and pregnancy outcome. The results provide valuable insights into potential metabolic biomarkers and suggest that targeting the gut microbiota may offer a strategy for preventing preeclampsia.

Declining human fertility worldwide is an attractive research target for the search for “high fertility” genes and pathways to counteract this problem. To study these genes and pathways for high fertility, the superfertile Dummerstorf mouse lines FL1 and FL2 are two unique model organisms representing an improved fertility phenotype. A direct reason for this remarkable characteristic of increased litter size, which reaches >20 pups/litter in both FLs, is the raised ovulation rate by approximately 100%, representing an impressive record in this field. Dummerstorf high-fertility lines incarnate extraordinary and singular models of high-fertility for other species, mostly farm animals, with the aim of improving production and reducing costs. Our main goal is to describe the genetic and molecular pathways to reach their phenotypical excellence, and to reproduce them using the control population. The large litter size and ovulation rate in Dummerstorf lines are mostly due to an increase in the quality of their oocytes, which receive a different intake of fat and are composed of different types and concentrations of fatty acids. As the follicular microenvironment plays a fundamental role during the oocytes development, in the present manuscript, we tried to improve the in vitro maturation technique by mimicking the fatty acid profile of FLs oocytes during the IVM of control oocytes. Currently, the optimization of the IVM system is fundamental mostly for prepubertal girls and oncological patients whose main source of gametes to restore fertility may be their maturation in vitro. Our data suggest that the specific fatty acid composition of FLs COCs can contribute to their high-fertility phenotype. Indeed, COCs from the control line matured in IVM-medium supplemented with C14:0 (high in FL2 COCs) or with C20:0, C21:0, C22:0, and C23:0 (high in FL1 COCs), but also control oocytes without cumulus, whose concentration in long-chain FAs are “naturally” higher, showing a slightly higher maturation rate. These findings represent an important starting point for the optimization of the IVM system using FA supplementation.

In recent decades, fertility traits in humans as well as in farm animals have decreased worldwide. As such, it is imperative to know more about the genetics and physiology of increased or high fertility. However, most of the current animal models with reproductive phenotypes describe lower fertility or even infertility (around 99%). The “Dummerstorf high-fertility lines” (FL1 and FL2) are two unique mouse lines selected for higher reproductive performances, more specifically for higher number of pups per litter. We recently described how those superfertile mice managed to increase their reproductive phenotype by doubling the ovulation rate and consequently the litter size compared to the unselected mice of the same founder population. FLs show an unusual estrous cycle length and atypical levels of hormones that link reproduction and metabolism, such as insulin in FL1 and leptin in FL2. Moreover, we described that their higher ovulation rate is mostly due to a higher quality of their oocytes rather than their sheer quantity, as they are characterized by a higher quantity of high-quality oocytes in antral follicles, but the quantity of follicles per ovary is not dissimilar compared to the control. In the present study, we aimed to analyze the lipid composition of the fertility lines from plasma to the gonads, as they can connect the higher reproductive performances with their metabolic atypicalities. As such, we analyzed the fat content of FLs and fatty acid composition in plasma, liver, fat, oocytes of different quality, and granulosa cells. We demonstrated that those mice show higher body weight and increased body fat content, but at the same time, they manage to decrease the lipid content in the ovarian fat compared to the abdominal fat, which could contribute to explaining their ovarian quality. In addition, we illustrate the differences in fatty acid composition in those tissues, especially a lower level of saturated fatty acids in plasma and a different lipid microenvironment of the ovary. Our ongoing and future research may be informative for farm animal biology as well as human reproductive medicine, mostly with cases that present characteristics of lower fertility that could be reversed following the way-of-managing of Dummerstorf high-fertility lines.

Dummerstorf fertility lines FL1 and FL2 represent two models of enhanced fertility characterized by the doubling of the litter size compared with an unselected control population (ctrl line, Dummerstorf FztDU). Both biodiverse FLs managed to reach this goal by increasing the ovulation rate per cycle, even showing decreased pregnancy rate and irregular oestrous cycle and metabolic hormone levels, compared with ctrl. The aim of the present study was to analyse oocytes in terms of quality and quantity by comparing the entire pool of oocytes per ovary, with those from the antral follicles within the same animal. We performed Brilliant Cresyl Blue staining as a non‐invasive marker of oocyte quality in combination with an analysis of additional morphological indicators, e.g. cytoplasm clarity, cumulus cell layers, nuclear anatomy, size and shape. We compared our fertility lines with the unselected control population and with another independent line selected from the same founder population, showing lower litter size (DU6P). Our results suggest that fertility lines show decreased number of oocytes per ovary compared with DU6P but increased number of high‐quality oocytes before ovulation. Hence, the raise in the ovulation rate and litter size of those super fertile mouse lines are not associated with an increased number of oocytes per ovary but rather with an increased number of higher quality fertilizable oocytes per cycle. In addition, the most conspicuous method to acquire oocytes with the highest quality in our lines is to assess their morphology, rather than their status after staining. All these discoveries together may be of fundamental importance for further studies in livestock farm animals showing some similar characteristics, e.g. irregular cycle or hormonal misbalances, to improve production while lowering costs, and in humans to increase the possibilities of successful pregnancies for couples undergoing in vitro fertilization (IVF).

Mouse models of decreased fertility mainly support scientific knowledge in the field of reproductive biology. In this study, we changed the perspective, using Dummerstorf high‐fertility mouse lines FL1 and FL2 selected for increased reproductive performances that doubled the amount of ovulated oocytes per cycle and the number of offspring per litter compared to an unselected control line (founder population, FZTDU, ctrl). After recent observations, both fertility lines seem to show a lower pregnancy rate compared to ctrl together with an atypical reproductive cycle. We analysed the estrous cycle of those mice, but also plasma levels of insulin, glucagon, leptin and ghrelin that, when irregular, may have an impact on the reproductive cycle length by modifying the GnRH release. We included females of another independent line (DU6P), selected from the same founder population but independently of fertility traits, as an additional model of decreased pregnancy rate. We aim to evaluate if do they use a similar mechanism associated in the regulation of the estrous cycle or implicated in altered pregnancy mechanism compared to control, because they show a similar situation as FL2 line does, even without an increase in fertility parameters. We speculate that FLs’ estrous cycle undergoes changes during the selection period and aim to demonstrate that some hormonal dysfunctions link with altered reproductive cycle, dampened pregnancy rates and reduced first delivery rates mostly in FL2, but also with higher‐fertility phenotype rather than lower in both FLs.