Growth hormone reduces aneuploidy and improves oocytes quality by JAK2-MAPK3/1 pathway in aged mice.

BACKGROUND: The global delay in women's reproductive age has raised concerns about age-related infertility. The decline in oocyte quality is a limiting factor of female fertility, yet there are currently no strategies to preserve oocyte quality in aged women. Here, we investigated the effects of growth hormone (GH) supplementation on aneuploidy of aged oocytes. METHODS: For the in vivo experiments, the aged mice (8-month-old) were intraperitoneally injected with GH daily for 8 weeks. For the in vitro experiments, germinal vesicle oocytes from aged mice were treated with GH during oocyte maturation. The impacts of GH on ovarian reserve before superovulation was evaluated. Oocytes were retrieved to assess oocyte quality, aneuploidy and developmental potential characteristics. Quantitative proteomics analysis was applied to investigate the potential targets of GH in aged oocytes. RESULTS: In this study, we demonstrated that GH supplementation in vivo not only alleviated the decline in oocyte number caused by aging, but also improved the quality and developmental potential of aged oocytes. Strikingly, we discovered that GH supplementation reduced aneuploidy in aged oocytes. Mechanically, in addition to improving mitochondrial function, our proteomic analysis indicated that the MAPK3/1 pathway may be involved in the reduction in aneuploidy of aged oocytes, as confirmed both in vivo and in vitro. In addition, JAK2 may also act as a mediator in how GH regulates MAPK3/1. CONCLUSIONS: In conclusion, our research reveals that GH supplementation protects oocytes against aging-related aneuploidy and enhances the quality of aged oocytes, which has clinical significance for aged women undergoing assisted reproduction technology.

Near-infrared fluorophore IR-61 delays postovulatory aging of mouse oocytes through suppressing oxidative stress mediated by mitochondrial protection.

Postovulatory aging can trigger deterioration of oocyte quality and subsequent embryonic development, and thus reduce the success rates of assisted reproductive technology (ART). The molecular mechanisms underlying postovulatory aging, and preventative strategies, remain to be explored. The near-infrared fluorophore IR-61, a novel heptamethine cyanine dye, has the potential for mitochondrial targeting and cell protection. In this study, we found that IR-61 accumulated in oocyte mitochondria and reduced the postovulatory aging-induced decline in mitochondrial function, including mitochondrial distribution, membrane potential, mtDNA number, ATP levels, and mitochondrial ultrastructure. In addition, IR-61 rescued postovulatory aging-caused oocyte fragmentation, defects in spindle structure, and embryonic developmental potential. RNA sequencing analysis indicated that the postovulatory aging-induced oxidative stress pathway might be inhibited by IR-61. We then confirmed that IR-61 decreased the levels of reactive oxygen species and MitoSOX, and increased GSH content in aged oocytes. Collectively, the results indicate that IR-61 may prevent postovulatory aging by rescuing oocyte quality, promoting successful rate in ART procedure.

Near-infrared fluorophore IR-61 improves the quality of oocytes in aged mice via mitochondrial protection.

Maternal aging is associated with a decline in oocyte quality, which leads to the decreased fertility. Therefore, developing approaches to reduce aging-induced deterioration of oocyte quality in older women is important. Near-infrared cell protector-61 (IR-61), a novel heptamethine cyanine dye, has the potential for antioxidant effects. In this study, we found that IR-61 can accumulate in the ovaries and improved ovarian function of naturally aged mice; it also increased the oocyte maturation rate and quality by maintaining the integrity of the spindle/chromosomal structure and reducing the aneuploidy rate. In addition, the embryonic developmental competence of aged oocytes was improved. Finally, RNA-sequencing analysis indicated that IR-61 might perform the beneficial effects on aged oocytes by regulating mitochondrial function, this was confirmed by immunofluorescence analysis of mitochondrial distribution and reactive oxygen species. Taken together, our findings demonstrate that IR-61 supplementation in vivo can increase oocyte quality and protect oocytes from aging-induced mitochondrial dysfunction, and thus could improve the fertility of older women and efficiency of assisted reproductive technology.

Irisin: circulating levels in serum and its relation to gonadal axis.

Irisin is an exercise-induced myokine/adipokine in mice and humans that plays an important role in 'browning' of white adipose tissue and has shown great potential as a treatment for some metabolic diseases, such as obesity, insulin resistance, and inflammation. The circulating irisin level is reported to be associated with exercise, obesity, diet, diseases, and exposure to different pharmacological agents. Several studies have attempted to characterize the role of irisin in PCOS and other reproductive diseases, but contradictory results have been reported. Our previous study showed that irisin may serve further functions in folliculogenesis and fertility. In this review, we present the current knowledge on the physiology of irisin and its role in gonadal axis. Firstly, we describe irisin circulating levels and speculate on the potential mechanisms involved in irisin secretion and regulation. Then, we focus on the irisin levels in PCOS, and explore the relationships between, BMI, insulin resistance, and hyperandrogenism. Finally, we present the results from animal interventional studies and in vitro experiments to investigate the relationship between irisin and gonadal axis, indicating its novel effects on reproduction and fertility.

Irisin deletion induces a decrease in growth and fertility in mice.

BACKGROUND: Irisin, which is cleaved from fibronectin type III domain-containing protein 5 (Fndc5), plays an important role in energy homeostasis. The link between energy metabolism and reproduction is well known. However, the biological actions of irisin in reproduction remain largely unexplored. METHODS: In this study, we generated Fndc5 gene mutation to create irisin deficient mice. Female wild-type (WT) and Fndc5 mutant mice were fed with standard chow for 48 weeks. Firstly, the survival rate, body weight and fertility were described in mice. Secondly, the levels of steroid hormones in serum were measured by ELISA, and the estrus cycle and the appearance of follicles were determined by vaginal smears and ovarian continuous sections. Thirdly, mRNA-sequencing analysis was used to compare gene expression between the ovaries of Fndc5 mutant mice and those of WT mice. Finally, the effects of exogenous irisin on steroid hormone production was investigated in KGN cells. RESULTS: The mice lacking irisin presented increased mortality, reduced body weight and poor fertility. Analysis of sex hormones showed decreased levels of estradiol, follicle-stimulating hormone and luteinizing hormone, and elevated progesterone levels in Fndc5 mutant mice. Irisin deficiency in mice was associated with irregular estrus, reduced ratio of antral follicles. The expressions of Akr1c18, Mamld1, and Cyp19a1, which are involved in the synthesis of steroid hormones, were reduced in the ovaries of mutant mice. Exogenous irisin could promote the expression of Akr1c18, Mamld1, and Cyp19a1 in KGN cells, stimulating estradiol production and inhibiting progesterone secretion. CONCLUSIONS: Irisin deficiency was related to disordered endocrinology metabolism in mice. The irisin deficient mice showed poor growth and development, and decreased fertility. Irisin likely have effects on the expressions of Akr1c18, Mamld1 and Cyp19a1 in ovary, regulating the steroid hormone production. This study provides novel insights into the potential role of irisin in mammalian growth and reproduction.

Natural attenuation of lead by microbial manganese oxides in a karst aquifer.

Lead is a toxic environmental contaminant associated with current and historic mine sites. Here we studied the natural attenuation of Pb in a limestone cave system that receives drainage from the ancient Priddy Mineries, UK. Extensive deposits of manganese oxides were observed to be forming on the cave walls and as coatings in the stream beds. Analysis of these deposits identified them as birnessite (δ-MnO2), with some extremely high concentrations of sorbed Pb (up to 56 wt%) also present. We hypothesised that these cave crusts were actively being formed by microbial Mn(II)-oxidation, and to investigate this the microbial communities were characterised by DNA sequencing, enrichment and isolation experiments. The birnessite deposits contained abundant and diverse prokaryotes and fungi, with ~5% of prokaryotes and ~ 10% of fungi closely related to known heterotrophic Mn(II)-oxidisers. A substantial proportion (up to 17%) of prokaryote sequences were assigned to groups known as autotrophic ammonia and nitrite oxidisers, suggesting that nitrogen cycling may play an important role in contributing energy and carbon to the cave crust microbial communities and consequently the formation of Mn(IV) oxides and Pb attenuation. Enrichment and isolation experiments showed that the birnessite deposits contained Mn(II)-oxidising microorganisms, and two isolates (Streptomyces sp. and Phyllobacterium sp.) could oxidise Mn(II) in the presence of 0.1 mM Pb. Supplying the enrichment cultures with acetate as a source of energy and carbon stimulated Mn(II)-oxidation, but excess organics in the form of glucose generated aqueous Mn(II), likely via microbial Mn(IV)-reduction. In this karst cave, microbial Mn(II)-oxidation contributes to the active sequestration and natural attenuation of Pb from contaminated waters, and therefore may be considered a natural analogue for the design of wastewater remediation systems and for understanding the geochemical controls on karst groundwater quality, a resource relied upon by billions of people across the globe.

Disordered metabolism in mice lacking irisin.

Irisin is a product of fibronectin type III domain-containing protein (Fndc5) and is involved in the regulation of adipokine secretion and the differentiation of osteoblasts and osteoclasts. In this study, we aimed to determine whether irisin lacking affects glucose/lipid and bone metabolism. We knocked out the Fndc5 gene to generate irisin-lacking mice. Remarkable, irisin lacking was related to poor 'browning response', with a bigger size of the intraperitoneal white adipose cell and decreased a number of brown adipose cells in brown adipose of interscapular tissue. The irisin lacking mice had hyperlipidemia and insulin resistance, reduced HDL-cholesterol level, increased LDL-cholesterol level, and decreased insulin sensitivity. The lacking of irisin was associated with reduced bone strength and bone mass in mice. The increased number of osteoclasts and higher expression of RANKL indicated increased bone resorption in irisin lacking mice. The level of IL-6 and TNF-α also increased in irisin lacking mice. The results showed that irisin lacking was related to decreased 'browning response', glucose/lipid metabolic derangement, and reduced bone mass with increased bone resorption. Further studies are needed to confirm these initial observations and explore the mechanisms underlying the effects of irisin on glucose/lipid and bone metabolism.

The correlation between UDP-glucuronosyltransferase polymorphisms and environmental endocrine disruptors levels in polycystic ovary syndrome patients.

BACKGROUND: In recent years, there has been an interest in whether environmental endocrine disruptors (EEDs) may contribute to the endocrine disorders in patients with polycystic ovary syndrome (PCOS). The clearance of EEDs from the human body is regulated by the glucuronidation of UDP-glucuronosyltransferases (UGT). This study aimed to analyze the relationship of UGT1A1, UGT2B7, and UGT2B15 polymorphisms with the metabolism of EEDs in patients with PCOS. METHODS: A total of 357 Chinese women (119 PCOS cases and 238 controls) were genotyped for polymorphisms of UGT1A1, UGT2B7, and UGT2B15. The plasma concentrations of EEDs were measured by the gas chromatography-mass spectrometry method. The association between UGT polymorphisms and the serum level of EEDs in patients with PCOS was analyzed. RESULTS: The UGT2B7 single nucleotide polymorphism was associated with an increased risk of PCOS. The homozygous polymorphism (TT) of UGT2B7 showed higher bisphenol A and PAEs concentrations in serum. However, a single nucleotide polymorphism on UGT2B15 expression was associated with a decreased risk of PCOS. Subjects homozygous for the T allele of UGT2B15 had a significant effect on phthalates in the blood. In addition, our results also showed that the homozygous polymorphism (TT) of UGT2B7 and UGT2B15 was associated with the capacity of the excretion of androgen in patients with PCOS. CONCLUSIONS: Our study reported the novel associations between the UGT polymorphisms and EEDs concentrations in patients with PCOS, supporting the relevance of genetic differences in EEDs metabolism, which might be considered as an etiology of PCOS.

Effects of the Dibutyl Phthalate (DBP) on the Expression and Activity of Aromatase in Human Granulosa Cell Line KGN.

In recent years, environmental endocrine disruptors (EEDs) have received extensive attention because of their hormone-like or anti-hormone effects. Dibutyl phthalate (DBP) is not only one of the most widely-used phthalates but also a member of EEDs with the estrogenic property. Although some studies have revealed the negative effect of DBP on the reproductive system, the underlying mechanisms are still elusive. Here the effect of DBP on P450 aromatase, a rate-limiting enzyme stimulated by FSH in the estradiol synthesis, was investigated in human granulosa cell line KGN. Cultured cells were treated with FSH and various doses of DBP (0.1µM, 1µM, 10µM, 50µM, or 100µM) for 24hr. Then the expression of aromatase was assessed, and the synthesis of estradiol was detected to reflect aromatase activity. As shown by the results, all concentrations of DBP could up-regulate the mRNA as well as protein levels of aromatase, and 0.1µM DBP increased the production of estradiol significantly. Furthermore, the ovary-specific promoter of aromatase, promoter II, was activated by 0.1µM DBP, and the expression of FSH receptor (FSHR) was increased by DBP from 0.1µM to 100µM. The study results show that DBP can affect aromatase from both quantitative and functional aspects, and this process may involve the activation of aromatase promoter II and upregulation of FSHR in KGN. Additionally, low-concentration DBP, near human serum concentration, has a more robust effect. This study suggests that DBP may affect the steroidogenic capacity in human ovaries and contributes to our understanding of the effects of DBP on the female reproductive system.

Evaluation of body composition in POF and its association with bone mineral density and sex steroid levels.

The study aims to investigate the body composition and bone mineral density (BMD) characteristics and discuss the relationships among body composition, BMD and sex steroid level in POF. A total of 240 POF patients, 240 normal women, and peri/postmenopausal women (Peri-M/Post-M) (260 patients in each group) were included. Compared to the control group, POF patients? strength of left/right lower limb (SLL/SRL), muscle distributing coefficient of lower limbs (MD) decreased however, waist circumference (WC) and hip circumference (HC) increased. The weight, WC, HC, whole body fat percentage (BF%), average fat distribution (FD), MD of POF patients were lower than those among Peri-M and Post-M and BMD were lower than the Peri-M, yet still higher than Post-M. Moreover, BMD were significantly positively correlated with BF%, FD, SLL, MD and estradiol (E2). The factors associated with L2-L4 BMD were E2, SRL, FD and age. For the FN BMD, the factors were FD, E2 and SLL. Therefore, we conclude that maintenance of appropriate weight, physical exercise and hormone replacement treatment (HRT) may have positive effects on increasing BMD, improving muscle mass and muscle strength, preventing osteoporosis.

Irisin promotes proliferation but inhibits differentiation in osteoclast precursor cells.

The receptor activator of NF-κB ligand-induced osteoclast differentiation has a critical role in the process of bone metabolism. Overactivation of osteoclastogenesis may result in a series of diseases. Irisin, a novel myokine, which was first reported in 2012, has been proposed to mediate the beneficial metabolic effects of exercise. Studies have demonstrated that irisin targets osteoblasts by promoting osteoblast proliferation and differentiation; however, the underlying mechanism regarding the effect of irisin on osteoclasts remains elusive. Using 2 types of osteoclast precursor cells, RAW264.7 cells and mouse bone marrow monocytes, we showed that irisin promoted osteoclast precursor cell proliferation but inhibited osteoclast differentiation. Irisin down-regulated the expression of osteoclast differentiation marker genes, including receptor activators of NF-κB, nuclear factor of activated T cells, cytoplasmic 1, cathepsin K, and tartrate-resistant acid phosphatase (TRAP), as well as decreasing the number of TRAP-positive multinucleated cells and hydroxyapatite resorption pits. Furthermore, we showed that irisin suppressed the NF-κB signaling pathway, but activated the p38 and JNK signaling pathways. In the presence of an inhibitor of p38 and JNK, irisin-induced promotion of RAW264.7 cell proliferation was attenuated. However, irisin-induced inhibition of osteoclast differentiation was not affected by either the p38 or JNK signaling pathway. Our study suggested the direct effect of irisin on osteoclastogenesis and revealed the mechanism responsible for the therapeutic potential of irisin in bone metabolism disease.-Ma, Y., Qiao, X., Zeng, R., Cheng, R., Zhang, J., Luo, Y., Nie, Y., Hu, Y., Yang, Z., Zhang, J., Liu, L., Xu, W., Xu, C. C., Xu, L. Irisin promotes proliferation but inhibits differentiation in osteoclast precursor cells.

The effect of His-tag and point mutation on the activity of irisin on MC3T3-E1 cells.

Irisin is a myokine secreted from the cleavage of fibronectin type III domain-containing protein 5 (FNDC5) and has an effect on bone formation. There are limited studies about the structure of irisin and its functional unit. In order to clarify the candidate domain responsible for irisin action, we constructed several irisin variants and tested their influence on the proliferation and osteogenesis of MC3T3-E1 cells. On the one hand, His-tag was added to the N terminal or C terminal of irisin. On the other hand, the flexible region or salt bridge site were chosen as the candidate for point mutation. Alkaline phosphatase (ALP), Runt related transcription factor 2 (Runx2) and collagen type I alpha 1 (COL1α1) were chosen to test the differentiation efficiency. We found point mutation on flexible regions, Glu-57 and Ile-107, and adding His-tag on the C-terminal of irisin did affect its action. The osteogenic potential of irisin E57K, irisin I107F and irisinC-His decreased about 90.1%, 88.8% and 96.6% activity of recombinant-irisin (r-irisin) (P < 0.05), respectively. Point mutation on the salt bridge, Arg-75, partly decreased the effect of irisin (45 ± 11.3% of r-irisin) (P < 0.05). N-terminal His-tag showed almost no effect (93.5 ± 25.7% of r-irisin) (P = 0.658). This study suggested that the flexible region of residues 55-58 and 106-108, and C-terminal of irisin are vital for its activity. Disrupting the dimerization of irisin might result in a partly reduced effect on cell differentiation.

Association between maternal, fetal and paternal MTHFR gene C677T and A1298C polymorphisms and risk of recurrent pregnancy loss: a comprehensive evaluation.

PURPOSE: Numerous studies have investigated the associations between methylenetetrahydrofolate reductase (MTHFR) gene C677T and A1298C polymorphisms and risk of recurrent pregnancy loss (RPL); however, the results remain controversial. The aim of this study is to drive a more precise estimation of association between MTHFR gene polymorphisms and risk of RPL. METHODS: We searched PubMed, EMBASE, Cochrane library, Web of Science and China Knowledge Resource Integrated Database for papers on MTHFR gene C677T and A1298C polymorphisms and RPL risk. The pooled odds ratios (ORs) with 95 % confidence intervals (CIs) were used to assess the strength of association in the homozygous model, heterozygous model, dominant model, recessive model and an additive model. The software STATA (Version 13.0) was used for statistical analysis. RESULTS: Overall, 57 articles were included in the final meta-analysis. In maternal group the MTHFR C677T polymorphism showed pooled odds ratios for the homozygous comparison [OR = 2.285, 95 % CI (1.702, 3.067)] and the MTHFR A1298C polymorphism showed pooled odds ratios for recessive model [OR = 1.594, 95 % CI (1.136, 2.238)]. In fetal group the MTHFR C677T polymorphism showed pooled odds ratios for dominant model [OR = 1.037, 95 % CI (0.567, 1.894)] and the MTHFR A1298C polymorphism showed pooled odds ratios for dominant model [OR = 1.495, 95 % CI (1.102, 2.026)]. CONCLUSIONS: In summary, the results of our meta-analysis indicate that maternal and paternal MTHFR gene C677T and A1298C polymorphisms are associated with RPL. We also observed a significant association between fetal MTHFR A1298C polymorphism and RPL but not C677T.