The association between the number of oocytes retrieved and cumulative live birth rate in different female age strata.

To evaluate the association between the number of oocytes retrieved and cumulative live birth rate (CLBR) in different female age strata. 17,931 women undergoing their first IVF/ICSI-ET cycle in the Sir Run Run Shaw Hospital of Zhejiang University were grouped by age (A: ≤ 35 years; B: ≥ 36 years) as well as the number of oocytes retrieved (a: ≤ 5; b:6-9; c:10-14; d: ≥ 15). Multivariate regression analysis was performed to assess the OR of CLBR for the variable 'age' and 'number of oocytes retrieved'. The group ≥ 36 years exhibited lower cumulative pregnancy rates (CPRs) and cumulative live birth rates (CLBRs), which are proportional to the number of oocytes retrieved but opposite to increasing age. Multivariate logistic regression analysis revealed that the age and number of oocytes retrieved remain significant independent predictive factors (P < 0.001). Age and number of oocytes retrieved are two independent factors affecting the CLBR. The discrepancy of the minimum number of oocytes retrieved for patients with different ages to achieve ideal CLBR is instructive for clinical practice. The practice of controlling the stimulation dose is feasible for patients ≤ 35 years who can achieve over 60% CLBR once the number of oocytes obtained is more than 6. However, additional stimulation cycles and accumulation of embryos are necessary for elderly group especially those ≥ 38 years old who need more than 14 oocytes to obtain higher live birth rate.

Dysregulation of Histone Deacetylases Inhibits Trophoblast Growth during Early Placental Development Partially through TFEB-Dependent Autophagy-Lysosomal Pathway.

Dysregulated biological behaviors of trophoblast cells can result in recurrent spontaneous abortion (RSA)-whose underlying etiology still remains insufficient. Autophagy, a conserved intracellular physiological process, is precisely monitored throughout whole pregnancy. Although the exact mechanism or role remains elusive, epigenetic modification has emerged as an important process. Herein, we found that a proportion of RSA patients exhibited higher levels of autophagy in villus tissues compared to controls, accompanied with impaired histone deacetylase (HDAC) expression. The purpose of this study is to explore the connection between HDACs and autophagy in the pathological course of RSA. Mechanistically, using human trophoblast cell models, treatment with HDAC inhibitor (HDACI)-trichostatin A (TSA) can induce autophagy by promoting nuclear translocation and transcriptional activity of the central autophagic regulator transcription factor EB (TFEB). Specifically, overactivated autophagy is involved in the TSA-driven growth inhibition of trophoblast, which can be partially reversed by the autophagy inhibitor chloroquine (CQ) or RNA interference of TFEB. In summary, our results reveal that abnormal acetylation and autophagy levels during early gestation may be associated with RSA and suggest the potential novel molecular target TFEB for RSA treatment.

Regulators involved in trophoblast syncytialization in the placenta of intrauterine growth restriction.

Placental dysfunction refers to the insufficiency of placental perfusion and chronic hypoxia during early pregnancy, which impairs placental function and causes inadequate supply of oxygen and nutrients to the fetus, affecting fetal development and health. Fetal intrauterine growth restriction, one of the most common outcomes of pregnancy-induced hypertensions, can be caused by placental dysfunction, resulting from deficient trophoblast syncytialization, inadequate trophoblast invasion and impaired vascular remodeling. During placental development, cytotrophoblasts fuse to form a multinucleated syncytia barrier, which supplies oxygen and nutrients to meet the metabolic demands for fetal growth. A reduction in the cell fusion index and the number of nuclei in the syncytiotrophoblast are found in the placentas of pregnancies complicated by IUGR, suggesting that the occurrence of IUGR may be related to inadequate trophoblast syncytialization. During the multiple processes of trophoblasts syncytialization, specific proteins and several signaling pathways are involved in coordinating these events and regulating placental function. In addition, epigenetic modifications, cell metabolism, senescence, and autophagy are also involved. Study findings have indicated several abnormally expressed syncytialization-related proteins and signaling pathways in the placentas of pregnancies complicated by IUGR, suggesting that these elements may play a crucial role in the occurrence of IUGR. In this review, we discuss the regulators of trophoblast syncytialization and their abnormal expression in the placentas of pregnancies complicated by IUGR.

S100P promotes trophoblast syncytialization during early placenta development by regulating YAP1.

Recurrent pregnancy loss (RPL) is a severe complication of pregnancy that is caused by genetic abnormalities, immune dysfunction, aberrant cell biology, and tissue structure destruction. Among which, placental dysfunction is crucial in the pathogenetic progression of RPL. Although some regulatory factors associated with RPL have been reported, the placental changes correlated with RPL still need to be elucidated. Here, we found that a portion of RPL patients presented with low serum and placental S100P expression. Using a human trophoblast stem cell model, we demonstrated that S100P was exclusively expressed in syncytiotrophoblast (ST)-like syncytia (ST(2D)-TSCT) and that loss of S100P expression in ST(2D)-TSCT cells impaired ß-hCG secretion, leading to syncytialization failure during early placental development. Moreover, we found that S100P is involved in regulating trophoblast syncytialization by downregulating the protein level of Yes-associated protein 1 (YAP1), which plays a pivotal role in maintaining trophoblast stemness. Together, our findings suggest that S100P plays an essential role in regulating trophoblast syncytialization during early placental development in humans via YAP1. Additionally, lower serum S100P levels may predict poor pregnancy outcomes and represent a potentially useful marker for evaluating placental biological function during early pregnancy.

Toxicity from a single injection of human umbilical cord mesenchymal stem cells into rat ovaries.

Intraovarian injection of human umbilical cord mesenchymal stem cells (hUC-MSCs) has been applied and with promising therapeutic effects, but its toxicity and safety remain uncertain. This study evaluated the toxic effects and the affected target organs after a single injection of hUC-MSCs into bilateral rat ovaries. Sixty Sprague-Dawley rats were randomly divided into four groups and intraovarian injected with three different doses of hUC-MSC suspension. Toxicity-related manifestations occurred over the following 14 days postinjection. On day (D)5 and D15, we assessed the clinical pathology; immunotoxicity, including the cytokine IFN-γ, TNF-α, IL-4, and IL-6 levels; the immune organs, and the organ weights. On D5, inflammatory cells mainly infiltrated the ovaries of the low- and medium-dose groups, whereas inflammatory cells infiltrated the oviduct in the medium- and high-dose groups. On D15, inflammatory cells infiltrated the corpus luteal cysts, ovarian sacs and oviducts in each group. Body weights; organ weights; immunotoxicity; clinical pathology and histopathological examinations of the immune organs did not significantly differ among the groups. No obvious hUC-MSC-related clinical symptoms were observed except in the rats that died. The high-dose group exhibited significantly higher mortality than did the control and low-dose groups. Deaths in the high-dose group, who received approximately 50 times the standard clinical dose, were related to the intraovarian hUC-MSC injection. The maximum tolerated dose was approximately ten times the standard clinical dose. The ovary and oviduct may be the target organs for this toxicity. This report provides dosage references and guidance for clinical applications of intraovarian hUC-MSC injections.

Dual Trigger for Final Follicular Maturation Improves Cumulative Live-Birth Rate in Ovarian Stimulation for Freeze-All In Vitro Fertilization/Intracytoplasmic Sperm Injection Cycles.

Study Question: Does dual trigger in freeze-all in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) cycles improve the cumulative live-birth outcome compared with human chorionic gonadotropin (hCG) trigger? Summary Answer: Dual trigger for final follicular maturation improves the cumulative pregnancy and live-birth rates compared with hCG trigger in freeze-all IVF/ICSI cycles. What Is Known Already: Dual trigger could increase the numbers of oocytes and mature oocytes and improve pregnancy rates. Study Design Size Duration: This retrospective cohort analysis included data from 4438 freeze-all IVF/ICSI cycles between January 2012 and December 2017. Participants/Materials Setting Methods: Women aged 20-49 years who underwent ovarian stimulation and oocyte retrieval for autologous IVF/ICSI with a freeze-all policy in our centre were enrolled. Data on number of oocytes retrieved, number of mature oocytes, clinical pregnancy rate, live-birth rate, cumulative pregnancy rate, and cumulative live-birth rate (CLBR) were assessed and compared between patients who underwent a dual trigger and hCG trigger. Multivariate logistic regression was performed to identify and adjust for factors known to independently affect the CLBR. Main Results and the Role of Chance: A total of 4438 IVF/ICSI cycles were analyzed, including 1445 cycles with single hCG trigger and 2993 cycles with dual trigger. The cumulative biochemical pregnancy rate (60.8% vs. 68.1%, P<0.001; odds ratio (OR): 0.727; 95% confidence interval (CI): 0.638-0.828), cumulative clinical pregnancy rate (52.9% vs. 58.5%, P<0.001; OR: 0.796; 95%CI: 0.701-0.903), and CLBR (44.3% vs. 50.5%, P<0.001; OR: 0.781; 95%CI: 0.688-10.886) were all significantly lower in the hCG-trigger group compared with the dual-trigger group. The clinical pregnancy rate (48.2% vs. 58.2%, P=0.002; OR: 0.829; 95%CI: 0.737-0.934) and embryo implantation rate (34.4% vs. 38.9%, P<0.001; OR: 0.823; 95%CI: 0.750-0.903) in each transfer cycle were also significantly lower in the hCG-trigger group compared with the dual-trigger group. After controlling for all potential confounding variables, the trigger method was identified as an independent factor affecting the CLBR. The OR and 95%CI for hCG trigger were 0.780 and 0.641-0.949 (P=0.013). Limitations Reasons for Caution: The data used to analyse the effect of dual trigger on cumulative pregnancy and live-birth outcomes were retrospective, and the results may thus have been subject to inherent biases. Further prospective randomized controlled trials are required to verify the beneficial effects of dual trigger. Wider Implications of the Findings: Dual trigger had a positive effect on CLBRs, suggesting that it could be used as a routine trigger method in freeze-all cycles. Study Funding/Competing Interests: This study was supported by grants from National Key Research and Development Program of China (2018YFC1004800), the Natural Science Program of Zhejiang (LY19H040009), the National Natural Science Foundation of China (No. 81601236). No authors have competing interests to declare.

Predicting the Likelihood of Live Birth in Assisted Reproductive Technology According to the Number of Oocytes Retrieved and Female Age Using a Generalized Additive Model: A Retrospective Cohort Analysis of 17,948 Cycles.

Capsule: We designed a predictive reference model to evaluate how many stimulation cycles are needed for a patient to achieve an ideal live birth rate using assisted reproductive technology. Objective: To develop a counseling tool for women who wish to undergo assisted reproductive technology (ART) treatment to predict the likelihood of live birth based on age and number of oocytes retrieved. Methods: This was a 6-year population-based retrospective cohort analysis using individual patient ART data. Between 2012 and 2017, 17,948 women were analyzed from their single ovarian stimulation cycle until they had a live birth or had used all their embryos. All consecutive women between 20 and 49 years old undergoing their ovarian stimulation cycles for ART in our center were enrolled. The cumulative live birth rate (CLBR) was defined as the delivery of a live neonate born during fresh or subsequent frozen-thawed embryo transfer cycles. Only the first delivery was considered in the analysis. Binary logistic regression was performed to identify and adjust for factors known to affect the CLBR independently. A generalized additive model was used to build a predictive model of CLBR according to the woman's age and the number of oocytes retrieved. Results: An evidenced-based counseling tool was created to predict the probability of an individual woman having a live birth, based on her age and the number of oocytes retrieved in ART cycles. The model was verified by 10 times 10-fold cross-validation using the preprocessed data, and 100 area under the curve (AUC) values for receiver operating characteristic (ROC) curves were obtained on the test set. The mean AUC value was 0.7394. Our model predicts different CLBRs ranging from nearly 90% to less than 20% for women aged 20-49 years with at least 22 oocytes retrieved. The CLBRs of women aged 20-28 years were very similar, nearly on one trend line with a certain number of oocytes retrieved. Differences in the CLBR began to appear by the age of 29 years; these increased gradually in women aged >35 years. Conclusion: A predictive model of the CLBR was designed to serve as a guide for physicians and for patients considering ART treatment. The number of oocytes needed to be retrieved to achieve a live birth depends on the woman's age.

The biological characteristics of sheep umbilical cord mesenchymal stem cells.

Although mesenchymal stem cells (MSCs) are now regarded as a promising cell resource for tissue repair and regeneration, the optimal source of MSCs has not yet been determined. The objective of this study was to provide a theoretical basis for the clinical application of umbilical cord mesenchymal stem cells (UCMSCs) in the future. Umbilical cord is an easily obtainable tissue resource, which is one reason that it has become a candidate resource for mesenchymal stem cells. In this study, we analyzed the biological characteristics of UCMSCs, such as their multiple differentiation and clone-forming ability, through morphological observation, reverse transcription polymerase chain reaction (RT-PCR), growth curve, positive rate test, and immunophenotype. Umbilical cord MSCs were successfully isolated and passaged to 29 generations. The results from RT-PCR showed that UCMSCs were positive for CD29, CD44, CD73, but negative for CD34. The expression of the stem cell marker nucleostemin and tenocyte-related markers showed similar positive results with CD44, CD73, and CD90. In addition, UCMSCs can be induced to differentiate into osteoblasts, adipocytes, or chondrocytes. Our study showed that UCMSCs not only have the ability to self-renew, but also have the potential to differentiate into multiple lineages. In general, we concluded that UCMSCs are a reliable source for use in cell therapy.

Bien que les cellules souches mésenchymateuses (CSMs) soient maintenant considérées comme une ressource prometteuse de cellules pour la réparation tissulaire et la régénération, la source optimale de CSMs n'a pas encore été déterminée. L'objectif de la présente étude était de fournir une base théorique pour l'application clinique de cellules souches mésenchymateuses de cordon ombilical (CSMCO) dans le futur. Le cordon ombilical est une ressource tissulaire pouvant être obtenue facilement, une des raisons pour laquelle il est devenu un candidat pour les CSMs. Dans cette étude nous avons analysé les caractéristiques biologiques des CSMCO, telles que leur différenciation multiple et la capacité à former des clones, par des observations morphologiques, par réaction d'amplification en chaîne par la polymérase avec la transcriptase réverse (ACP-TR), courbe de croissance, test de ratio positif, et immunophénotype. Les CSMCO ont été isolées avec succès et des passages obtenus jusqu'à la 29e génération. Les résultats d'ACP-TR ont montré que les CSMCO étaient positives pour CD29, CD44, CD73, mais négative pour CD34. L'expression de nucléostémine, un marqueur de cellule souche, et de marqueurs apparentés aux ténocytes ont montré des résultats positifs similaires à ceux de CD44, CD73, et CD90. De plus, les CSMCO peuvent être induites à se différencier en ostéoblastes, adipocytes, ou chondrocytes. Notre étude a démontré que les CSMCO ont non seulement la capacité de s'auto-renouveler, mais ont également le potentiel de se différencier en des lignées multiples. En général, nous avons conclu que les CSMCO sont une source fiable pour utilisation en thérapie cellulaire.(Traduit par Docteur Serge Messier).

Characteristics and multi­lineage differentiation of bone marrow mesenchymal stem cells derived from the Tibetan mastiff.

Bone marrow mesenchymal stem cells (BM­MSCs) are pluripotent stem cells that are regarded as ideal resources for the reconstruction of tissues and organs. The Tibetan mastiff is a breed of domesticated Chinese native dog that is well­adjusted to the high­altitude environments of Tibet. To the best of our knowledge, the biological characterization and multi­lineage differentiation of Tibetan mastiff BM­MSCs have not been reported previously. Therefore, the present study aimed to investigate the biological characteristics and therapeutic potential of Tibetan mastiff BM­MSCs. A cell culture system was constructed and cells were cultured to 23 passages in vitro. Growth curves and colony formation studies suggested that BM­MSCs had a high self­renewal capacity and that their proliferation rate declined with age. Karyotype analysis demonstrated that BM­MSCs were diploid and genetically stable. Semi­quantitative polymerase chain reaction analysis revealed that BM­MSCs positively expressed cluster of differentiation (CD)73, CD90, CD105, CD166 and vimentin, although they were negative for the endothelial cell marker CD31. Additionally, immunofluorescence staining revealed that the cells expressed CD29, CD44, CD90, CD105 and vimentin. Flow cytometric analysis revealed that the rates of positive expression of vimentin, CD44, CD90 and CD105 were all >97%. BM­MSCs were able to differentiate into adipocytes, osteoblasts, cartilage cells, hepatocytes and functional insulin­secreting cells. In conclusion, Tibetan mastiff BM­MSCs may be purified successfully using a whole bone marrow culture method. The findings of the current study suggested important potential applications of BM­MSCs as a source for regenerative therapies.

Therapeutic potential of Bama miniature pig adipose stem cells induced hepatocytes in a mouse model with acute liver failure.

The role of mesenchymal stem cells (MSCs) in cellular therapy is well recognized in this work. MSCs have advantages of high proliferation, clone formation, multi-lineage differentiation and immunosuppression. Furthermore, adipose-resident MSCs (ADSCs) are extensively employed due to its advantages of abundant source, low cost and simple operation. Many researchers have emphasized the role of adipose-resident MSCs in the development of therapies for liver injury, but few attentions were paid on the use of induced functional hepatocytes. Therefore, in this work the role of adipose-resident MSCs induced functional hepatocytes was mainly investigated. The function of induced hepatocytes by ELISA and the induction rate was confirmed by flow cytometry and evaluated by experimental observations. The induced hepatocytes were firstly transplanted into CCl4-caused liver damage ICR mice by tail vein. After transplantation, both liver fibrosis and function could be improved by hepatocytes, which were examined through histology, immunofluorescence staining, serum profile and biochemical parameters levels. The production of cytokines was then compared with normal mice and injury mice to explore the therapeutic mechanisms of hepatocytes. Finally, the secretions of TGF-ß1, IL-6 and IL-10 in hepatocytes transplanted mice were determined and found to be higher than that of the normal and injury mice. The hepatocytes derived from ADSCs were proven to have a great significance in the therapeutic efficacy and clinical settings of liver disease animal models.

Establishment and biological characterization of a dermal mesenchymal stem cells line from bovine.

The DMSCs (dermal mesenchymal stem cells) are multipotent stem cells, which can differentiate in vitro into many cell types. Much work has been done on DMSCs from humans, mice, rabbits and other mammals, but the related literature has not been published about these cells in cattle. In this study, we isolated and established the DMSC lines from cattle, thereby initiating further research on these cells, such as growth kinetics, detection of special surface antigen and RT-PCR (reverse transcription-PCR) assays to identify the biological characterization of the cell line. Furthermore, the DMSCs are induced to differentiate into adipocytes, osteoblasts and neural cells in vitro Our results suggest that DMSCs isolated from cattle possess similar biological characteristics with those from other species. Their multi-lineage differentiation capabilities herald a probable application model in tissue engineering and induced pluripotent stem cells.