Tubal endometriosis (tubal EM) is a subtype of endometriosis (EM) associated with fallopian tube impairments and infertility. Since the molecular mechanism underlying tubal EM is not clear, we assume that an aberrant transcriptome of fallopian tube epithelium and microenvironment changes caused by cytokines in tubal fluid are possible causes. The aim of this study was to identify potential hub mRNAs/proteins of tubal EM through integrated transcriptomic and proteomic analyses and to elucidate significant pathways, cellular functions, and interaction networks during the initiation and progression of tubal EM. We obtained human fallopian tube epithelium and tubal fluid samples from patients with and without tubal EM. Tubal epithelia were analyzed using microarray, and tubal fluid was analyzed using quantitative label-free LC-MS/MS. We identified differentially expressed genes (DEGs) and differentially expressed proteins (DEPs) and determined common mRNAs/protein. We observed 35 commonly deregulated mRNAs/proteins, and IPA indicated that cellular movement, inflammatory response, and immune cell trafficking were significantly activated during the pathogenesis of tubal EM. We also identified acute phase response signaling pathway activation as a unique pathogenesis signature of tubal EM. Our results demonstrate that an integrated analysis of the transcriptome and proteome has the potential to reveal novel disease mechanisms at a molecular level.
Hang Qi, Huiyu Zhang, Xiaoya Zhao, Ya Qin, Guiling Liang, Xiaoqing He and Jian Zhang
Hannah L Morgan, Isaac Ampong, Nader Eid, Charlène Rouillon, Helen R Griffiths and Adam J Watkins
The link between male diet and sperm quality has received significant investigation. However, the impact diet and dietary supplements have on the testicular environment has been examined to a lesser extent. Here, we establish the impact of a sub-optimal low protein diet (LPD) on testicular morphology, apoptosis and serum fatty acid profiles. Furthermore, we define whether supplementing a LPD with specific methyl donors abrogates any detrimental effects of the LPD. Male C57BL6 mice were fed either a control normal protein diet (NPD; 18% protein; n = 8), an isocaloric LPD (LPD; 9% protein; n = 8) or an LPD supplemented with methyl donors (MD-LPD; choline chloride, betaine, methionine, folic acid, vitamin B12; n = 8) for a minimum of 7 weeks. Analysis of male serum fatty acid profiles by gas chromatography revealed elevated levels of saturated fatty acids and lower levels of mono- and polyunsaturated fatty acids in MD-LPD males when compared to NPD and/or LPD males. Testes of LPD males displayed larger seminiferous tubule cross section area when compared to NPD and MD-LPD males, while MD-LPD tubules displayed a larger luminal area. Furthermore, TUNNEL staining revealed LPD males possessed a reduced number of tubules positive for apoptosis, while gene expression analysis showed MD-LPD testes displayed decreased expression of the pro-apoptotic genes Bax, Csap1 and Fas when compared to NPD males. Finally, testes from MD-LPD males displayed a reduced telomere length but increased telomerase activity. These data reveal the significance of sub-optimal nutrition for paternal metabolic and reproductive physiology.
Katerina Menelaou, Malwina Prater, Simon J Tunster, Georgina E T Blake, Colleen Geary Joo, James C Cross, Russell S Hamilton and Erica D Watson
Assisted reproduction technologies (ARTs) are becoming increasingly common. Therefore, how these procedures influence gene regulation and foeto-placental development are important to explore. Here, we assess the effects of blastocyst transfer on mouse placental growth and transcriptome. C57Bl/6 blastocysts were transferred into uteri of B6D2F1 pseudopregnant females and dissected at embryonic day 10.5 for analysis. Compared to non-transferred controls, placentas from transferred conceptuses weighed less even though the embryos were larger on average. This suggested a compensatory increase in placental efficiency. RNA sequencing of whole male placentas revealed 543 differentially expressed genes (DEGs) after blastocyst transfer: 188 and 355 genes were downregulated and upregulated, respectively. DEGs were independently validated in male and female placentas. Bioinformatic analyses revealed that DEGs represented expression in all major placental cell types and included genes that are critical for placenta development and/or function. Furthermore, the direction of transcriptional change in response to blastocyst transfer implied an adaptive response to improve placental function to maintain foetal growth. Our analysis revealed that CpG methylation at regulatory regions of two DEGs was unchanged in female transferred placentas and that DEGs had fewer gene-associated CpG islands (within ~20 kb region) compared to the larger genome. These data suggested that altered methylation at proximal promoter regions might not lead to transcriptional disruption in transferred placentas. Genomic clustering of some DEGs warrants further investigation of long-range, cis-acting epigenetic mechanisms including histone modifications together with DNA methylation. We conclude that embryo transfer, a protocol required for ART, significantly impacts the placental transcriptome and growth.
Federica Lopes, Jin Liu, Stephanie Morgan, Rebecca Matthews, Lucy Nevin, Richard A Anderson and Norah Spears
Chemotherapy drugs are administered to patients using combination regimens, and as such the possibility of multiplicative effects between drugs need to be investigated. This study examines the individual and combined effects of the chemotherapy drugs cisplatin and doxorubicin on the human ovary. Although cisplatin and doxorubicin are known to affect female fertility, there is limited information about their direct effects on the human ovary, and none examining the possibility of combined, multiplicative effects of co-exposure to these drugs. Here, human ovarian biopsies were obtained from 14 women at the time of caesarean section, with 38 mouse ovaries also obtained from neonatal C57Bl/6J mice. Tissue was cultured for 6 days prior to analyses, with chemotherapy drugs added to culture medium on the second day of culture only. Treatment groups of a single (5 μg/mL human; 0.5 μg/mL mouse) or double (10 μg/mL human; 1.0 μg/mL mouse) dose of cisplatin, a single (1 μg/mL human; 0.05 μg/mL mouse) or double (2 μg/mL human; 0.01 μg/mL mouse) dose of doxorubicin or a combination of a single dose of both drugs together were compared to controls without drug exposure. Exposure to cisplatin or doxorubicin significantly decreased follicle health in human and mouse, supporting the suitability of the mouse as a model for the human ovary. There was also a significant reduction of mouse follicle number. Human ovarian stromal tissue exhibited increased apoptosis and decreased cell proliferation. Crucially, there was no evidence indicating the occurrence of multiplicative effects between cisplatin and doxorubicin.
Shelby L Oke, Gurjeev Sohi and Daniel B Hardy
Epidemiological data suggest an inverse relationship between birth weight and long-term metabolic deficits, which is exacerbated by postnatal catch-up growth. We have previously demonstrated that rat offspring subject to maternal protein restriction (MPR) followed by catch-up growth exhibit impaired hepatic function and ER stress. Given that mitochondrial dysfunction is associated with various metabolic pathologies, we hypothesized that altered expression of p66Shc, a gatekeeper of oxidative stress and mitochondrial function, contributes to the hepatic defects observed in MPR offspring. To test this hypothesis, pregnant Wistar rats were fed a control (20% protein) diet or an isocaloric low protein (8%; LP) diet throughout gestation. Offspring born to control dams received a control diet in postnatal life, while MPR offspring remained on a LP diet (LP1) or received a control diet post weaning (LP2) or at birth (LP3). At four months, LP2 offspring exhibited increased protein abundance of both p66Shc and the cis-trans isomerase PIN1. This was further associated with aberrant markers of oxidative stress (i.e. elevated 4-HNE, SOD1 and SOD2, decreased catalase) and aerobic metabolism (i.e., increased phospho-PDH and LDHa, decreased complex II, citrate synthase and TFAM). We further demonstrated that tunicamycin-induced ER stress in HepG2 cells led to increased p66Shc protein abundance, suggesting that ER stress may underlie the programmed effects observed in vivo. In summary, because these defects are exclusive to adult LP2 offspring, it is possible that a low protein diet during perinatal life, a period of liver plasticity, followed by catch-up growth is detrimental to long-term mitochondrial function.
Lixin Wen, Rongfang Li, Ji Wang and Jine Yi
In this paper, we propose the reproductive stress hypothesis that describes the pregnant females response to reproductive events based upon the activation of the hypothalamic–pituitary–adrenal axis and sympathetic adrenomedullary system. The main components of the reproductive stress hypothesis can be summarized as follows: (1) events unique to reproduction including empathema, pregnancy, parturition and lactation cause non-specific responses in females, called active reproductive stress; (2) the fetus is a special stressor for pregnant females where endocrine hormones, including corticotropin-releasing hormones and fetal glucocorticoids secreted by the fetus and placenta, enter the maternal circulatory system, leading to another stress response referred to as passive reproductive stress and (3) response to uterine tension and intrauterine infection is the third type of stress, called fetal intrauterine stress. Appropriate reproductive stress is a crucial prerequisite in normal reproductive processes. By contrast, excessive or inappropriate reproductive stress may result in dysfunctions of the reproductive system, such as compromised immune function, leading to susceptibility to disease. The novel insights of the reproductive stress hypothesis have important implications for deciphering the pathogenesis of certain diseases in pregnant animals, including humans, which in turn may be applied to preventing and treating their occurrence.
Guangdong Li, Xiuzhi Tian, Dongying Lv, Lu Zhang, Zhenzhen Zhang, Jing Wang, Minghui Yang, Jingli Tao, Teng Ma, Hao Wu, Pengyun Ji, Yingjie Wu, Zhengxing Lian, Wei Cui and Guoshi Liu
NLRP (NACHT, LRR and PYD domain-containing proteins) family plays pivotal roles in mammalian reproduction. Mutation of NLRP7 is often associated with human recurrent hydatidiform moles. Few studies regarding the functions of NLRP7 have been performed in other mammalian species rather than humans. In the current study, for the first time, the function of NLRP7 has been explored in ovine ovary. NLRP7 protein was mainly located in ovarian follicles and in in vitro pre-implantation embryos. To identify its origin, 763 bp partial CDS of NLRP7 deriving from sheep cumulus oocyte complexes (COCs) was cloned, it showed a great homology with Homo sapiens. The high levels of mRNA and protein of NLRP7 were steadily expressed in oocytes, parthenogenetic embryos or IVF embryos. NLRP7 knockdown by the combination of siRNA and shRNA jeopardized both the parthenogenetic and IVF embryo development. These results strongly suggest that NLRP7 plays an important role in ovine reproduction. The potential mechanisms of NLRP7 will be fully investigated in the future.
Renjie Wang, Wei Pan, Lei Jin, Yuehan Li, Yudi Geng, Chun Gao, Gang Chen, Hui Wang, Ding Ma and Shujie Liao
Artificial intelligence (AI) has experienced rapid growth over the past few years, moving from the experimental to the implementation phase in various fields, including medicine. Advances in learning algorithms and theories, the availability of large datasets and improvements in computing power have contributed to breakthroughs in current AI applications. Machine learning (ML), a subset of AI, allows computers to detect patterns from large complex datasets automatically and uses these patterns to make predictions. AI is proving to be increasingly applicable to healthcare, and multiple machine learning techniques have been used to improve the performance of assisted reproductive technology (ART). Despite various challenges, the integration of AI and reproductive medicine is bound to give an essential direction to medical development in the future. In this review, we discuss the basic aspects of AI and machine learning, and we address the applications, potential limitations and challenges of AI. We also highlight the prospects and future directions in the context of reproductive medicine.
Neil A Youngson, G Mezbah Uddin, Abhirup Das, Carl Martinez, Haley S Connaughton, Sara Whiting, Josephine Yu, David A Sinclair, R John Aitken and Margaret J Morris
Male fertility and sperm quality are negatively impacted by obesity. Furthermore, recent evidence has shown that male offspring from obese rat mothers also have reduced sperm quality and fertility. Here, we extend work in this area by comparing the effects of both maternal obesity and offspring post-weaning diet-induced obesity, as well as their combination, on sperm quality in mice. We additionally tested whether administration of the NAD+-booster nicotinamide mononucleotide (NMN) can ameliorate the negative effects of obesity and maternal obesity on sperm quality. We previously showed that intraperitoneal (i.p.) injection of NMN can reduce the metabolic deficits induced by maternal obesity or post-weaning dietary obesity in mice. In this study, female mice were fed a high-fat diet (HFD) for 6 weeks until they were 18% heavier than a control diet group. Thereafter, HFD and control female mice were mated with control diet males, and male offspring were weaned into groups receiving control or HFD. At 30 weeks of age, mice received 500 mg/kg body weight NMN or vehicle PBS i.p. for 21 days. As expected, adiposity was increased by both maternal and post-weaning HFD but reduced by NMN supplementation. Post-weaning HFD reduced sperm count and motility, while maternal HFD increased offspring sperm DNA fragmentation and levels of aberrant sperm chromatin. There was no evidence that the combination of post-weaning and maternal HFD exacerbated the impacts in sperm quality suggesting that they impact spermatogenesis through different mechanisms. Surprisingly NMN reduced sperm count, vitality and increased sperm oxidative DNA damage, which was associated with increased NAD+ in testes. A subsequent experiment using oral NMN at 400 mg/kg body weight was not associated with reduced sperm viability, oxidative stress, mitochondrial dysfunction or increased NAD+ in testes, suggesting that the negative impacts on sperm could be dependent on dose or mode of administration.
V A van der Weijden, J T Bick, S Bauersachs, G J Arnold, T Fröhlich, B Drews and S E Ulbrich
The uterine microenvironment during pre-implantation presents a pro-survival milieu and is essential for embryo elongation in ruminants. The European roe deer (Careolus capreolus) pre-implantation embryo development is characterised by a 4-month period of reduced development, embryonic diapause, after which the embryo rapidly elongates and implants. We investigated the uterine fluid proteome by label-free liquid chromatography tandem mass spectrometry at four defined stages covering the phase of reduced developmental pace (early diapause, mid-diapause and late diapause) and embryo elongation. We hypothesised that embryo development during diapause is halted by the lack of signals that support progression past the blastocyst stage. Three clusters of differentially abundant proteins were identified by a self-organising tree algorithm: (1) gradual reduction over development; (2) stable abundance during diapause, followed by a sharp rise at elongation; and (3) gradual increase over development. Proteins in the different clusters were subjected to gene ontology analysis. ‘Cellular detoxification’ in cluster 1 was represented by alcohol dehydrogenase, glutathione S-transferase and peroxiredoxin-2. ATP-citrate synthase, nucleolin, lamin A/C, and purine phosphorylase as cell proliferation regulators were found in cluster 2 and ‘cortical cytoskeleton’, ‘regulation of substrate adhesion-dependent cell spreading’ and ‘melanosome’ were present in cluster 3. Cell cycle promoters were higher abundant at elongation than during diapause, and polyamines presence indicates their role in diapause regulation. This study provides a comprehensive overview of proteins in the roe deer uterine fluid during diapause and forms a basis for studies aiming at understanding the impact of the lack of cell cycle promoters during diapause.