Increased cellular senescence in doxorubicin-induced murine ovarian injury: effect of senolytics.

Ovarian injury caused by chemotherapy can lead to early menopause, infertility, and even premature senility in female cancer patients, impairing the quality of life and overall health of the cancer survivors seriously. However, there is still a lack of effective protection strategies against such injury. Cellular senescence can be induced by chemotherapeutic agents in multiple organs and may corrode the structure and function of normal tissues. We hypothesized that the widely used first-line chemotherapy drug, doxorubicin, could increase senescent cell burden in normal ovarian tissue during the therapeutic process and that elimination of senescent cells with senolytics would ameliorate doxorubicin-induced ovarian injury. Here, we demonstrated an accumulation of cellular senescence in doxorubicin-treated ovaries through detecting p16 and p21 expression levels and senescence-associated ß-galactosidase (SA-ß-gal) activity as well as senescence-associated secretory phenotype (SASP) factors. Short-term intervention with the classic senolytic combination dasatinib and quercetin (DQ) or fisetin significantly reduced the load of senescent cells in ovaries after doxorubicin treatment. However, neither DQ nor fisetin alleviated doxorubicin-related ovarian dysfunction. Further experiments showed that ovarian apoptosis and fibrosis following doxorubicin exposure could not be improved by senolytics. Collectively, our study shows that senolytic treatment can eliminate accumulated senescent cells, but cannot reverse the massive follicle loss and ovarian stromal fibrosis caused by doxorubicin, suggesting that cellular senescence may not be one of the key mechanisms in doxorubicin-induced ovarian injury.

Construction of a competing endogenous RNA network to identify drug targets against polycystic ovary syndrome.

STUDY QUESTION: Would the construction of a competing endogenous RNA (ceRNA) network help identify new drug targets for the development of potential therapies for polycystic ovary syndrome (PCOS)? SUMMARY ANSWER: Both Food and Drug Administartion (FDA)-approved and candidate drugs could be identified by combining bioinformatics approaches with clinical sample analysis based on our established ceRNA network. WHAT IS KNOWN ALREADY: Thus far, no effective drugs are available for treating PCOS. ceRNAs play crucial roles in multiple diseases, and some of them are in current use as prognostic biomarkers as well as for chemo-response and drug prediction. STUDY DESIGN, SIZE, DURATION: For the bioinformatics part, five microarrays of human granulosa cells were considered eligible after applying strict screening criteria and were used to construct the ceRNA network for target identification. For population-based validation, samples from 24 women with and without PCOS were collected from January 2021 to July 2021. PARTICIPANTS/MATERIALS, SETTING, METHODS: The public data included 27 unaffected women and 25 women with PCOS, according to the Rotterdam criteria proposed in 2003. The limma and RobustRankAggreg R packages were used to identify differentially expressed messenger RNAs and noncoding RNAs. Gene Ontology, Reactome and Kyoto Encyclopedia of Genes and Gemomes (KEGG) enrichment analyses were performed. A ceRNA network was constructed by integrating the differentially expressed genes and target genes. The population-based validation included human luteinized granulosa cell samples from 12 unaffected women and 12 women with PCOS. Quantitative real-time polymerase chain reaction was conducted to detect the levels of mRNAs and microRNAs (miRNAs). Connectivity map and computational model algorithms were implemented to predict therapeutic drugs from the ceRNA network. Additionally, we compared the predicted drugs with known clinical medications in DrugBank. MAIN RESULTS AND THE ROLE OF CHANCE: A set of 10 mRNAs, 11 miRNAs and 53 long non-coding RNAs (lncRNAs) were differentially expressed. Functional enrichment analysis revealed the highest relevance to immune system-related biological processes and signalling pathways, such as cytokine secretion and leucocyte chemotaxis. A ceRNA consisting of two lncRNAs, two miRNAs and five mRNAs was constructed. Through network construction via bioinformatic analysis, we identified some already approved drugs (such as metformin) that could target some molecules in the network as potential drug candidates for PCOS. LARGE SCALE DATA: Public sequencing data were obtained from GSE34526, GSE84376, GSE102293, GSE106724 and GSE114419, which have been deposited in the Gene Expression Omnibus database. LIMITATIONS, REASONS FOR CAUTION: Experiments, such as immunoprecipitation, luciferase reporter assays and animal model studies, are needed to validate the potential targets in the ceRNA network before the identified drug candidates can be tested using cellular and animal model systems. WIDER IMPLICATIONS OF THE FINDINGS: Our findings provide new bioinformatic insight into the possible pathogenesis of PCOS from ceRNA network analysis, which has not been previously studied in the human reproductive field. Our study also reveals some potential drug candidates for the future development of possible therapies against PCOS. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by grants from the National Key Research and Development Program of China (2021YFC2700400) and the National Natural Science Foundation of China (82001498). The authors have no conflicts of interest to disclose.

Construction of Artificial Ovaries with Decellularized Porcine Scaffold and Its Elicited Immune Response after Xenotransplantation in Mice.

Substitution by artificial ovary is a promising approach to restore ovarian function, and a decellularized extracellular matrix can be used as a supporting scaffold. However, biomimetic ovary fabrication and immunogenicity requires more investigation. In this study, we proposed an effective decellularization protocol to prepare ovarian scaffolds, which were characterized by few nuclear substances and which retained the extracellular matrix proteins. The ovarian tissue shape and 3-dimensional structure were well-preserved after decellularization. Electron micrographs demonstrated that the extracellular matrix fibers in the decellularized group had similar porosity and structure to those of native ovaries. Semi-quantification analysis confirmed that the amount of extracellular matrix proteins was reduced, but the collagen fiber length, width, and straightness did not change significantly. Granulosa cells were attached and penetrated into the decellularized scaffold and exhibited high proliferative activity with no visible apoptotic cells on day 15. Follicle growth was compromised on day 7. The implanted artificial ovaries did not restore endocrine function in ovariectomized mice. The grafts were infiltrated with immune cells within 3 days, which damaged the artificial ovary morphology. The findings suggest that immune rejection plays an important role when using artificial ovaries.

Senotherapy Protects against Cisplatin-Induced Ovarian Injury by Removing Senescent Cells and Alleviating DNA Damage.

Ovarian damage induced by platinum-based chemotherapy seriously affects young women with cancer, manifesting as infertility, early menopause, and premature ovarian insufficiency. However, effective prevention strategies for such damage are lacking. Senescent cells may be induced by chemotherapeutic agents. We hypothesized that cisplatin can lead to senescence in ovarian cells during the therapeutic process, and senolytic drugs can protect animals against cisplatin-induced ovarian injury. Here, we demonstrated the existence of senescent cells in cisplatin-treated ovaries, identified the senescence-associated secretory phenotype, and observed significant improvement of ovarian function by treatment with metformin or dasatinib and quercetin (DQ) independently or in combination. These senotherapies improved both oocyte quality and fertility, increased the ovarian reserve, and enhanced hormone secretion in cisplatin-exposed mice. Additionally, attenuated fibrosis, reorganized subcellular structure, and mitigated DNA damage were observed in the ovaries of senotherapeutic mice. Moreover, RNA sequencing analysis revealed upregulation of the proliferation-related genes Ki, Prrx2, Sfrp4, and Megfl0; and the antioxidative gene H2-Q10 after metformin plus DQ treatment. Gene ontology analysis further revealed that combining senotherapies enhanced ovarian cell differentiation, development, and communication. In this study, we demonstrated that metformin plus DQ recovered ovarian function to a greater extent compared to metformin or DQ independently, with more follicular reserve, increased pups per litter, and reduced DNA damage. Collectively, our work indicates that senotherapies might prevent cisplatin-induced ovarian injury by removing senescent cells and reducing DNA damage, which represent a promising therapeutic avenue to prevent chemotherapy-induced ovarian damage.

Fertility outcomes of IVF/ICSI after Caesarean section: a cohort study.

RESEARCH QUESTION: The study objective was to evaluate the impact of a previous Caesarean section on fertility outcomes in women undergoing IVF/intracytoplasmic sperm injection (ICSI). DESIGN: A retrospective cohort study was designed that included 1793 women undergoing IVF/ICSI who had had a previous delivery from January 2015 to December 2016. The primary outcome was live birth. Secondary outcomes were implantation, clinical pregnancy, miscarriage, ectopic pregnancy, multiple pregnancy and perinatal complications. RESULTS: Of the 1793 women included, 796 had had a previous Caesarean section and 997 a previous vaginal delivery. Propensity score matching in a 1:1 ratio resulted in 538 women per group. Compared with women with a previous vaginal delivery, women with a previous Caesarean section had a lower live birth rate (30.1% versus 38.1%, odds ratio [OR] 0.70, 95% confidence interval [CI] 0.54-0.90) and a higher miscarriage rate (25.9% versus 17.5%, OR 1.65, 95% CI 1.06-2.56). Among other secondary outcomes, implantation rates were 32.9% and 37.1% (OR 0.83, 95% CI 0.69-1.01), and clinical pregnancy rates were 42.4% and 46.8% (OR 0.84, 95% CI 0.66-1.06), in the Caesarean section group and vaginal delivery group, respectively. There were no statistically significant differences in terms of ectopic pregnancy, multiple pregnancy or perinatal outcomes between the groups. Further adjustment for confounders did not change the result of the primary outcome (OR 0.64, 95% CI 0.49-0.84). CONCLUSIONS: Women undergoing IVF/ICSI who have had a previous Caesarean section have a lower live birth rate and a higher miscarriage rate than those with a previous vaginal delivery.

Influence of Isoflurane Exposure for 15 Consecutive Days on Ovarian Function in Adult Female Mice.

Female infertility after occupational exposure to inhaled anesthetic agents has attracted critical attention, but systematic studies focusing on the impact of inhaled anesthetics on the female reproductive system have not been well-established. We used a murine model to study the effect of isoflurane exposure on infertility in female adult mice and investigated the potential underlying mechanism. One hundred adult female C57 mice were randomly allocated into 5 groups exposed in air containing 0, 2500, 5000, 10 000 or 20 000 ppm isoflurane for 15 consecutive days. Estrous cycle length was measured based on vaginal smear examination, ovarian histopathologic enumeration of follicles, and serum estradiol (E2), anti-Mullerian hormone (AMH), follicle-stimulating hormone (FSH), and luteinizing hormone (LH) levels to assess the effect of isoflurane on ovarian reserve. Compared to the control group, significant prolongation of the estrous cycle of the adult female mice was observed in the 20 000 ppm isoflurane exposure group. Serum AMH was significantly decreased, and FSH and LH levels profoundly increased in the 5000, 10 000, and 20 000 ppm isoflurane exposure groups compared to the control group. The histopathologic examination revealed a reduced number of developing follicles and an increased number of atretic follicles after isoflurane exposure, but the difference was not statistically significant. Thus, exposure to a higher concentration of isoflurane might have an adverse effect on ovarian reserve in sexually-mature female mice.