Valvular fibroelastomas complicated by Coxiella burnetti endocarditis.

Cardiac papillary fibroelastoma (CPF) is a valvular tumor that may be mistaken for infective endocarditis (IE). We describe a case of CPF complicated by Coxiella burnetti IE. According to Duke's criteria, a diagnosis of IE was repeatedly considered as excluded or established during the clinical course, highlighting the criteria limitations.

MiR-21, EGFR and PTEN in non-small cell lung cancer: an in situ hybridisation and immunohistochemistry study.

AIMS: To analyse microRNA (miR)-21 distribution and expression at the cellular level in non-small cell lung cancer (NSCLC). MiR-21 is an oncogenic microRNA overexpressed in NSCLC. In previous studies, overexpression of miR-21 was evaluated from the tumour bulk by quantitative reverse transcription PCR with results expressed on average across the entire cell population. METHODS: We used in situ hybridisation and immunohistochemistry to assess the correlation between miR-21 levels and the expression of markers that may be possible targets (epidermal growth factor reaction) or may be involved in its upregulation (phosphatase and tensin homolog (PTEN), p53). The Pearson's χ2 tests was used to assess correlation with clinicopathological data and with miR-21 expression both in tumour and tumour stroma. RESULTS: Cytoplasmic staining and expression of Mir-21 were detected in the tumours and in associated stromal cells. Expression was highest in the stroma immediately surrounding the tumour cells and decreased as the distance from the tumour increased. No expression of miR-21 was found in normal lung parenchyma and a significant association was found between tumour localised miR-21 and PTEN. CONCLUSIONS: Presence of miR-21 in both cell tumour and stromal compartments of NSCLC and the relationship with PTEN confirms miR-21 as a microenvironment signalling molecule, possibly inducing epithelial mesenchymal transition and invasion by targeting PTEN in the stromal compartment possibly through exosomal transport. In situ immunohistochemical studies such as ours may help shed light on the complex interactions between miRNAs and its role in NSCLC biology.

Upper and lower genital tract Zika virus screening in a large cohort of reproductive-age women during the Americas epidemic.

RESEARCH QUESTION: To determine whether there is a risk of localized Zika virus (ZIKV) infection in the upper genital tract, specifically the oocytes, follicular fluids and endometrium, in exposed and/or recently infected reproductive-age women. ZIKV is an Aedes mosquito-borne Flavivirus that can lead to birth defects and to developmental anomalies when it infects pregnant women. DESIGN: Controlled observational clinical study following 179 female patients undergoing oocyte vitrification cycles in an academic fertility centre during the ZIKV epidemic in the French territories of the Americas. At the time, the French Ministry of Health issued a ban on medically-induced pregnancies. Oocyte vitrification cycles were the only means of preserving fertility options and ensuring Zika-free oocyte cryopreservation for currently exposed and/or recently infected patients. Samples of serum, urine, lower genital tract, endometrium, follicular fluid and immature oocytes were tested for ZIKV RNA (vRNA) by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Serological analysis for ZIKV antibodies was performed in succession for the duration of the study. The follow-up protocol was set up for more than 6 months post-exposure or post-onset. RESULTS: No vRNA was detected in the various samples from exposed patients. Furthermore, no vRNA was found in the upper genital tracts of women with a recent (3 months) history of acute infection. CONCLUSION: These findings represent evidence of a lack of vRNA persistence in the reproductive tract in ZIKV exposed and/or recently infected reproductive-age women and could help simplify current guidelines.

Kinetics of anti-ZIKV antibodies after Zika infection using two commercial enzyme-linked immunoassays.

High performance assays are essential for the serological diagnosis of recent and past Zika virus (ZIKV) infections but few are presently available. We used two commercially available NS1 antigen-based enzyme-linked immunoassays to study the kinetics of anti-ZIKV IgM and IgG in 15 ZIKV-infected patients for up to 180days after clinical onset. The Diapro assay detected anti-ZIKV IgM reactivity more frequently (100%) and for longer (median 53days) than did the Euroimmun assay (60%; 13days, P<0.005). Both assays detected anti-ZIKV IgG reactivity 11days after clinical onset in all subjects. ZIKV IgG reactivity decreased in 3 subjects, suggesting long-term false-negative results with the Euroimmun assay. Existing anti-Dengue antibodies seem to modify the detection of ZIKV IgG but the specificity of the immunoassays was not assessed. These enzyme-linked immunoassays were user-friendly and provided results rapidly in our hands but they need further assessment before being widely used for diagnosis or public health surveys.

Effect of acute Zika virus infection on sperm and virus clearance in body fluids: a prospective observational study.

BACKGROUND: Evidence of human sexual transmission during Zika virus emergence is a matter of concern, particularly in procreation, but to date, kinetics of seminal shedding and the effects of infection on human reproductive function have not been described. To investigate the effects of Zika virus infection on semen and clearance of Zika virus from semen and body fluids, we aimed to study a cohort of Zika virus-infected men. METHODS: This prospective observational study recruited men presenting with acute Zika virus infection at Pointe-à-Pitre University Hospital in Guadeloupe, French Caribbean, where a Zika virus outbreak occurred between April and November, 2016. Blood, urine, and semen were collected at days 7, 11, 20, 30, 60, 90, and 120 after symptom onset, and semen characteristics, such as total sperm count, sperm motility, vitality, and morphology, and reproductive hormone concentrations, such as testosterone, inhibin, follicle-stimulating hormone, and luteinising hormone, were assessed. At days 7, 11, and 20, semen was processed to isolate motile spermatozoa. Zika virus RNA was detected by RT-PCR using whole blood, serum, urine, seminal plasma, semen cells, and motile spermatozoa fractions. Zika virus was isolated from different sperm fractions on Vero E6 cultures. FINDINGS: 15 male volunteers (mean age 35 years [SD 5; range 25-44) with acute Zika virus infection and positive Zika virus RNA detection in blood or urine were enrolled. Total sperm count was decreased from median 119 × 106 spermatozoa (IQR 22-234) at day 7 to 45·2 × 106 (16·5-89·6) at day 30 and 70 × 106 (28·5-81·4) at day 60, respectively, after Zika virus infection. Inhibin values increased from 93·5 pg/mL (IQR 55-162) at day 7 to 150 pg/mL (78-209) at day 120 when total sperm count recovered. In motile spermatozoa obtained after density gradient separation, Zika virus RNA was found in three of 14 patients at day 7, four of 15 at day 11, and four of 15 at day 20, and replication-competent virus was found in the tested patient. Seminal shedding kinetics seemed heterogeneous among patients. Whole blood was the fluid most frequently positive for Zika virus RNA (62 of 92 samples) and three patients remained positive at day 120. INTERPRETATION: Semen alterations early after acute Zika virus infection might affect fertility and could be explained by virus effects on the testis and epididymis. Frequency of shedding and high viral load in semen, together with the presence of replicative virus in a motile spermatozoa fraction, can lead to Zika virus transmission during sexual contact and assisted reproduction procedures. Whole blood seems to be the best specimen for Zika virus RNA detection, diagnosis, and follow-up. FUNDING: Agence de la Biomédecine/Agence Régionale de Santé de la Guadeloupe/Inserm-REACTing.

Hexavalent chromium at low concentration alters Sertoli cell barrier and connexin 43 gap junction but not claudin-11 and N-cadherin in the rat seminiferous tubule culture model.

Exposure to toxic metals, specifically those belonging to the nonessential group leads to human health defects and among them reprotoxic effects. The mechanisms by which these metals produce their negative effects on spermatogenesis have not been fully elucidated. By using the Durand's validated seminiferous tubule culture model, which mimics the in vivo situation, we recently reported that concentrations of hexavalent chromium, reported in the literature to be closed to that found in the blood circulation of men, increase the number of germ cell cytogenetic abnormalities. Since this metal is also known to affect cellular junctions, we investigated, in the present study, its potential influence on the Sertoli cell barrier and on junctional proteins present at this level such as connexin 43, claudin-11 and N-cadherin. Cultured seminiferous tubules in bicameral chambers expressed the three junctional proteins and ZO-1 for at least 12days. Exposure to low concentrations of chromium (10µg/l) increased the trans-epithelial resistance without major changes of claudin-11 and N-cadherin expressions but strongly delocalized the gap junction protein connexin 43 from the membrane to the cytoplasm of Sertoli cells. The possibility that the hexavalent chromium-induced alteration of connexin 43 indirectly mediates the effect of the toxic metal on the blood-testis barrier dynamic is postulated.

[European regulation REACH and the assessment of testicular toxicity]. / Analyse de la spermatogenèse ex vivo : application à l'analyse de la toxicité testiculaire.

Several studies suggest that exposure to environmental pollutants is partly responsible for testicular pathologies that have considerably increased over the last decades (cryptorchidism, hypospadias, cancer, decrease in the number of ejaculated spermatozoa). However, the cellular and molecular mechanisms involved in this reprotoxicity remain mostly unknown. One of the challenges of the european regulation REACH is to improve the knowledge on the chemical, toxic and ecotoxic properties of substances used in everyday life. As for the testicular toxicity, the few in vivo models used are not always the most appropriate for mechanistic studies. Our laboratory has developed and validated on a physiological point of view, coculture systems of germ cells in bicameral chambers, which reproduce a blood-testis barrier, allowing the determination of the mechanisms responsible for the toxicity of organic or mineral compounds on spermatogenesis, while reducing greatly the number of animals required.

HIV-1 or hepatitis C chronic infection in serodiscordant infertile couples has no impact on infertility treatment outcome.

The objective was to evaluate the viral infection effects on infertility treatment outcome in HIV-1 or hepatitis C (HCV) monoinfected infertile serodiscordant couples, in a retrospective case-controlled, university-based study. Clinical pregnancy rate for HIV-1 or HCV infertile serodiscordant couples was not significantly different from that for seronegative controls.

The Aurora Kinase C c.144delC mutation causes meiosis I arrest in men and is frequent in the North African population.

Infertility concerns a minimum of 70 million couples worldwide. An important proportion of cases is believed to have a genetic component, yet few causal genes have been identified so far. In a previous study, we demonstrated that a homozygous mutation (c.144delC) in the Aurora Kinase C (AURKC) gene led to the production of large-headed polyploid multi-flagellar spermatozoa, a primary infertility phenotype mainly observed in North Africans. We now want to estimate the prevalence of the defect, to improve our understanding of AURKC physiopathology in spermatogenesis and assess its implication in oogenesis. A carrier frequency of 1/50 was established from individuals from the Maghrebian general population, comparable to that of Y-microdeletions, thus far the only known recurrent genetic event altering spermatogenesis. A total of 62 patients were genotyped, all who had a typical phenotype with close to 100% large-headed spermatozoa were homozygously mutated (n = 32), whereas no AURKC mutations were detected in the others. Two homozygous females were identified; both were fertile indicating that AURKC is not indispensible in oogenesis. Previous FISH results had showed a great chromosomal heterogeneity in these patient's spermatozoa. We demonstrate here by flow cytometry that all spermatozoa have in fact a homogeneous 4C DNA content and are thus all blocked before the first meiotic division. Our data thus indicate that a functional AURKC protein is necessary for male meiotic cytokinesis while its absence does not impair oogenesis.

Analysis of the intratesticular control of spermatogenesis by ex-vivo approaching.

Spermatogenesis involves the realization of a particular genetic program which requires a specific environment ("niche"). Multiplication, differentiation and apoptosis of male germ cells are finely regulated by pituitary hormones (mainly LH and FSH), and by a complex network of factors originating from both the somatic cells and the germ cells of the testis. It is becoming clear that hormones and intra-testicular regulatory factors can compensate, at least in part, for the absence of some hormones or factors including FSH and LH or androgen receptors. Since, most of the growth factors, cytokines and neurotrophins produced within the testis are widely expressed in the organism, the attempts to understand their role in spermatogenesis by "classical" knock-out strategies have been often disappointing. Therefore an important aspect of our previous work was to settle and characterize carefully two systems of cocultures of testicular germ cells with somatic cells in bicameral chambers. For instance, we showed for the first time that the whole meiotic step could be performed in vitro in a mammalian species (the rat). Moreover, all our data indicate that our co-culture systems enable to highlight mechanisms pertinent to the physiological processes. Sperm parameters have been deteriorated considerably during the past 4-5 decades. There is now evidence that chemical exposure is at least partly responsible for these testicular diseases. If a large number of environmental pollutants are able to affect male fertility and to exert carcinogenic effects, their cellular and molecular mechanisms are still unidentified. The cultures in bicameral chambers that we settled can be used to study the effects of a toxicant when added in the basal compartment of the culture chamber, which appears relevant to the in vivo situation. Taken together our results indicate that our in vitro culture systems, which allow screening for the effect of biological activity of different physiological factors, can be also helpful to study that of any chemicals on both survival and multiplication/differentiation of somatic and/or spermatogenic cells on a relatively long time period.

Impact of early cleaved zygote morphology on embryo development and in vitro fertilization-embryo transfer outcome: a prospective study.

OBJECTIVE: To evaluate the impact of the first division morphology on embryo development and IVF-embryo transfer outcome. DESIGN: Prospective study. SETTING: Teaching hospital, France. PATIENT(S): All zygotes from 201 couples were checked for early cleavage. We defined as "even," early cleaved (EC) zygotes with 2 cells of even size; as "uneven," EC zygotes with 2 cells of uneven size; and as "fragmented," EC zygotes with more than 20% fragmentation rate. Day 2 embryo quality was assessed as "top" embryo or "non-top," with the evaluation of multinucleated blastomeres. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Day 2 embryo quality, pregnancy and implantation rates. RESULT(S): Among EC zygotes, 59.1% were even, 13.0% were uneven, and 27.9% were fragmented. Even EC yielded more "top" embryos and less multinucleated blastomere embryos than uneven EC (77.0% vs. 46.3%) and fragmented EC (77.0% vs. 13.9%). The 125 double embryo transfers that comprised at least one embryo derived from even EC zygote led to higher pregnancy rate (PR) (64.0% vs. 43.4%) and implantation rate (42.0% vs. 27.6%) compared to the 76 double embryo transfers with embryos derived from breakdown or 2PN zygotes. CONCLUSION(S): The morphology of the early cleaved zygote is involved in embryo development. Evaluation of this morphology is an effective and valuable method of assessing the embryo quality.

Adequate ovarian follicular status does not prevent the decrease in pregnancy rates associated with high sperm DNA fragmentation.

OBJECTIVE: Potential reparation of sperm DNA fragmentation in the oocyte may disturb any relationship between DNA-damaged sperm and the implantation ability of resulting embryos. To rule out this factor, we analyzed the consequences of sperm DNA fragmentation on IVF-ET outcome in women with healthy ovarian function. DESIGN: Prospective study. SETTING: Teaching hospital, France. PATIENT(S): All 117 women were <38 years old, who combined normal serum day-3 FSH and inhibin B levels with an adequate response to controlled ovarian hyperstimulation. INTERVENTION(S): The DNA fragmentation rate was determined in the raw sperm used for conventional IVF by flow cytometric terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay. Cycles were sorted into two groups according to whether DNA fragmentation exceeded (high fragmentation [HF], n = 52) or did not exceed (low fragmentation [LF], n = 65) the 50th percentile of values (35%). MAIN OUTCOME MEASURE(S): D2 embryo quality and implantation and ongoing pregnancy rates. RESULT(S): Patients' characteristics, raw semen parameters, fertilization rates, and embryology data were similar in HF and LF groups. Clinical (37.5% vs. 62.5%) and ongoing (23.5% vs. 57.8%) pregnancy rates per ET and implantation rates (24.5% vs. 42.4%) were lower in the HF group than in the LF group. CONCLUSION(S): High sperm DNA fragmentation spares fertilization and top embryo morphology rates but is associated with decreased IVF-ET outcome.