Theconstructionofanadvancedinformationsystemsupportingthe workofaclinicisagreatchallenge.Particularly,aftertheinitialdetermination of the functionality of the system, its internal structure must be designed. For the major elements of the system their interaction must be also accurately determined.
Abstract Background: In the present study, we aimed to investigate selected functions of human neutrophils exposed to bisphenol A (BPA) under in vitro conditions. As BPA is classified among xenoestrogens, we compared its action and effects with those of 17β-estradiol (E2). Methods: Chemotaxis of neutrophils was examined using the Boyden chamber. Their phagocytosis and nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) oxidase activity were assessed via Park’s method with latex beads and Park’s test with nitroblue tetrazolium . To assess the total concentration of nitric oxide (NO), the Griess reaction was utilized. Flow cytometry was used to assess the expression of cluster of differentiation (CD) antigens. The formation of neutrophil extracellular traps (NETs) was analyzed using a microscope (IN Cell Analyzer 2200 system). Expression of the investigated proteins was determined using Western blot. Results: The analysis of results obtained for both sexes demonstrated that after exposure to BPA, the chemotactic capacity of neutrophils was reduced. In the presence of BPA, the phagocytic activity was found to be elevated in the cells obtained from women and reduced in the cells from men. Following exposure to BPA, the percentage of neutrophils with CD14 and CD284 (TLR4) expression, as well as the percentage of cells forming NETs, was increased in the cells from both sexes. The stimulatory role of BPA and E2 in the activation of NADPH oxidase was observed only in female cells. On the other hand, no influence of E2 on the expression of CD14 and CD284, chemotaxis, phagocytosis, and the amount of NET-positive neutrophils was found for both sexes. The study further showed that BPA intensified NO production and iNOS expression in the cells of both sexes. In addition, intensified expression of all tested PI3K-Akt pathway proteins was observed in male neutrophils. Conclusions: The study demonstrated the influence of BPA on neutrophil functions associated with locomotion and pathogen elimination, which in turn may disturb the immune response of these cells in both women and men. Analysis of the obtained data showed that the effect of this xenoestrogen on the human neutrophils was more pronounced than E2.
In most mammalian species aromatase is encoded by a single gene ( Cyp19 ), which contains 18 exons, nine of them being translated. In man, the presence of a biologically active aromatase and oestrogen receptors (ERα and ERβ) has been reported in Leydig cells, and also in immature germ cells and ejaculated spermatozoa. Concerning aromatase, the amount of transcript and enzymatic activity are decreased in immotile compared with motile sperm. We have amplified aromatase mRNA by real-time polymerase chain reaction in spermatozoa from asthenospermic, teratospermic and asthenoteratospermic men and recorded, respectively, 44, 52 and 67 per cent decreases of the amount of transcripts compared with fertile donors. A high degree of correlation ( r = −0.64) between the abnormal spermatozoa (especially microcephaly and acrosome malformations) and aromatase/GAPDH transcript ratio has been observed. Idiopathic infertility is a wide health problem and no treatment is currently available. In humans, even if the role of oestrogens in spermatogenesis is still a matter of debate, the observations of decreased sperm number and motility in men genetically deficient in aromatase, together with our data and those reported in the literature, may suggest a role for aromatase/oestrogens not only during the development and maintenance of spermatogenesis but also in the final maturation of spermatozoa.
To study the specific mechanisms through which progesterone and selective progesterone receptor modulators impact the growth, synthesis, and accumulation of the extracellular matrix in uterine leiomyomas.
We compared the effects of different concentrations of raloxifene (1 ,4 and 10 μM) on collagen biosynthesis ,gelatinolytic and prolidase activities and matrix metalloproteinase (MMP) expression (MMP-2 and MMP-9) in estradiol-stimulated (2 nM) breast cancer MCF-7 cells. Raloxifene inhibited in a dose-dependent manner the proliferation of MCF-7 cells ,independently of the presence or absence of estradiol in the growth medium. Raloxifene at concentrations of 1 μM and 4 μM inhibited collagen biosynthesis by about 10-fold and prolidase activity by about 50% ,while at a concentration of 10 μM it inhibited these processes by only about 25%. This phenomenon was accompanied by differences in gelatinolytic activity and MMP (MMP-2 and MMP-9) expression as demonstrated by zymography and Western immunoblot analysis, respectively. In estrogenstimulated MCF-7 cells ,cultured in the presence of 1 μM raloxifene ,a dramatic increase in the activity of both collagenases was found. In contrast ,addition of raloxifene at a concentration of 10 μM to the medium of the cells resulted in restoration of gelatinolytic activity to that found in control cells. Similarly ,but at both doses (1 and 10 μM) ,raloxifene was able to reduce MMP-2 expression in the cells. However ,when used alone (without estradiol) a concentration of 1 μM raloxifene strongly stimulated MMP-2 expression ,while at a concentration of 10 μM the effect was not observed. In the case of MMP-9 ,only trace amounts of this gelatinase were detected ,although in contrast to MMP-2 ,an increase in its expression was noticed at a concentration of 10 μM raloxifene. The data raise the possibility that in estrogen-stimulated MCF-7 cells ,raloxifene at low concentrations (1 and 4 μM) evokes antiestrogenic effect on collagen biosynthesis and prolidase activity on the one hand ,and an estrogenic effect on gelatinolytic activity on the other ,while at higher concentrations (about 10 μM) it evokes an estrogenic effect on collagen biosynthesis and prolidase activity, and an antiestrogenic effect on gelatinolytic activity. Our data suggest that the effects of raloxifene on collagen synthesis ,prolidase and metalloproteinase activities in breast cancer may explain its role in the prevention of breast cancer development.
Background: Primary ovarian insufficiency (POI), a public health problem, affects 1-3.7% of women under 40 yielding infertility and decreased longevity. Most causes are unknown. Recently, genetic causes were identified, mostly in single families. We studied an unprecedented cohort of POI to unravel its molecular pathophysiology.Methods: 318 patients with 66 families were studied using targeted (88 genes) or exome sequencing with pathogenic/likely-pathogenic variant selection. Mitomycin-induced chromosome breakages were studied in patients’ lymphocytes.Findings: A high-yield of 36.5 % supports a clinical genetic diagnosis of POI. We found 77 new variants, involved 13 novel genes: AKT1, LAST2, ELAVL2, NLRP11, CENPE, SYCP3, XPNPEP2, SPATA33, CCDC150, CCDC185, including DNA repair genes MCM81P, HELQ, SWI5 yielding high chromosomal fragility, and confirmed the causal role of BNC1, ERCC6, BMPR1A, BMPR1B, BMPR2, CAV1, SPIDR, RCBTB1, ATG7. In 10.4% of cases, POI is the unique phenotype of a multi-organ genetic disease. New pathways were identified: Hippo-AKT, a novel therapeutic target of POI during in vitro follicular activation (IVA), inflammation, post-transcriptional/translational regulations, mitochondrial autophagy providing future therapeutic targets. Three new genes had been involved in the age of natural menopause supporting a genetic link.Interpretation: We have developed high-performance genetic diagnostic of POI, dissecting the molecular pathogenesis of POI and leading to personalized medicine to i) prevent/cure comorbidities for tumor/cancer susceptibility genes that could affect longevity, or for genetically-revealed syndromic POI, ii) predict residual ovarian reserve. Genetics could select responder patients to IVA, greatly improving its success in treating infertility.Funding Information: This study was supported by Université Paris Sud-Paris Saclay, Hôpitaux Universitaires Paris Saclay (AH, MM), the Institut National de la Santé et de la Recherche Médicale-INSERM (AH, MM), and by the Agence Nationale de Biomédecine (AH, MM) Declaration of Interests: The authors declare no conflict of interest.Ethics Approval Statement: The study was approved by all the institutions involved and by the Agence de Biomedecine (reference number PFS12-002). Written informed consent was received from participants prior to inclusion in the study.
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