Constitutional mismatch repair deficiency syndrome: clinical description in a French cohort.

BACKGROUND: Constitutional mismatch repair deficiency (CMMRD) syndrome is a childhood cancer predisposition syndrome involving biallelic germline mutations of MMR genes, poorly recognised by clinicians so far. METHODS: Retrospective review of all 31 patients with CMMRD diagnosed in French genetics laboratories in order to describe the characteristics, treatment and outcome of the malignancies and biological diagnostic data. RESULTS: 67 tumours were diagnosed in 31 patients, 25 (37%) Lynch syndrome-associated malignancies, 22 (33%) brain tumours, 17 (25%) haematological malignancies and 3 (5%) sarcomas. The median age of onset of the first tumour was 6.9 years (1.2-33.5). Overall, 22 patients died, 9 (41%) due to the primary tumour. Median survival after the diagnosis of the primary tumour was 27 months (0.26-213.2). Failure rate seemed to be higher than expected especially for T-cell non-Hodgkin's lymphoma (progression/relapse in 6/12 patients). A familial history of Lynch syndrome was identified in 6/23 families, and consanguinity in 9/23 families. PMS2 mutations (n=18) were more frequent than other mutations (MSH6 (n=6), MLH1 (n=4) and MSH2 (n=3)). CONCLUSIONS: In conclusion, this unselected series of patients confirms the extreme severity of this syndrome with a high mortality rate mostly related to multiple childhood cancers, and highlights the need for its early detection in order to adapt treatment and surveillance.

Interactions between oocytes and cumulus cells during in vitro maturation of porcine cumulus-oocyte complexes in a chemically defined medium: effect of denuded oocytes on cumulus expansion and oocyte maturation.

The aim of the present study was to clarify interactions between oocytes and cumulus cells (CCs) on the level of cumulus expansion and oocyte maturation during IVM of cumulus-oocyte complexes (COCs) in a chemically defined medium using a system that allows individual tracking of oocytes. Especially, the influence of oocyte-secreted factors was investigated by the aid of addition of denuded oocytes (DOs) as a possible approach to improve the IVM system. The basic maturation medium was porcine oocyte medium with addition of gonadotropins only during the first 20 hours of IVM. During IVM, COCs were kept fixed to the bottom of culture dish by adhesive Cell-Tak coating, which enabled individual tracking of COCs during IVM. Size changes in COCs during IVM were measured by digital image analysis. Cumulus expansion in a porcine oocyte medium of intact COCs increased in a typical manner until 20 hours and decreased in size subsequently until 48 hours of IVM (P < 0.05). Removal of oocytes from COCs by oocytectomy allowed the expansion of CCs to some extent, although their expansion ability was lower than that of COCs (P < 0.05). Addition of DOs (COCs to DOs ratio of 9:16) did not improve cumulus expansion and oocyte maturation rates of intact COCs (P > 0.05) but did enhance cumulus expansion of oocytectomized complexes (P < 0.05). Furthermore, removal of CCs before IVM increased oocyte maturation rates compared with COCs (52.3% and 32.9%, respectively) (P < 0.05) and a similar effect was observed in COCs when the gap junction inhibitor carbenoxolone was added to the IVM medium: carbenoxolone repressed the expansion of COCs at 20 hours of IVM. In conclusion, the porcine oocyte enhances cumulus expansion both by gap junctional communications and presumably by oocyte-secreted factor production. Nevertheless, the presence of oocytes is not a prerequisite for this process. In return, CCs maintain meiotic arrest in cumulus-enclosed oocytes during the initial culture through gap junctions. On the basis of these findings, future research could investigate if coculture with DOs during IVM is beneficial for fertilization and embryo development.

The colposcopic atlas of schistosomiasis in the lower female genital tract based on studies in Malawi, Zimbabwe, Madagascar and South Africa.

BACKGROUND: Schistosoma (S.) haematobium is a neglected tropical disease which may affect any part of the genital tract in women. Female genital schistosomiasis (FGS) may cause abnormal vaginal discharge, contact bleeding, genital tumours, ectopic pregnancies and increased susceptibility to HIV. Symptoms may mimic those typical of sexually transmitted infections (STIs) and women with genital schistosomiasis may be incorrectly diagnosed. An expert consensus meeting suggested that the following findings by visual inspection should serve as proxy indicators for the diagnosis of schistosomiasis of the lower genital tract in women from S. haematobium endemic areas: sandy patches appearing as (1) single or clustered grains or (2) sandy patches appearing as homogenous, yellow areas, or (3) rubbery papules. In this atlas we aim to provide an overview of the genital mucosal manifestations of schistosomiasis in women. METHODOLOGY/PRINCIPAL FINDINGS: Photocolposcopic images were captured from women, between 1994 and 2012 in four different study sites endemic for S. haematobium in Malawi, Zimbabwe, South Africa and Madagascar. Images and specimens were sampled from sexually active women between 15 and 49 years of age. Colposcopic images of other diseases are included for differential diagnostic purposes. SIGNIFICANCE: This is the first atlas to present the clinical manifestations of schistosomiasis in the lower female genital tract. It will be freely available for online use, downloadable as a presentation and for print. It could be used for training purposes, further research, and in clinical practice.

Notifying partners of patients with early syphilis in Madagascar: case-finding effectiveness and public health implications.

OBJECTIVE: To evaluate the case-finding effectiveness of a clinic-based partner notification effort for early syphilis in Madagascar. METHODS: We asked index cases who had proven early syphilis to identify and provide contact information of recent sex partners (in the past 3, 6, and 12 months for primary, secondary, and early latent syphilis, respectively). Named sex partners were contacted by index cases (patient notification) or, if approved by the index case, clinic staff members (provider notification); notified of their potential exposure to syphilis; and asked to come to the clinic for evaluation. We assessed case-finding effectiveness and calculated the 'brought-to-treatment' index (number of newly-diagnosed syphilis cases per number of index cases interviewed). RESULTS: Of 565 index cases, 534 reported recent sex with at least one sex partner. A total of 3167 sex partners were reported, of whom 276 were contactable (9% of 3167). Providers notified 76% and cases notified 24% of these partners. 270 partners were contacted (98% of 276), and of these, 199 presented to the clinic for evaluation (74% of 270). A total of 99 partners tested positive for syphilis and received treatment (50% of 199). The 'brought-to-treatment' index was 0.18 (99 diagnoses per 565 index cases). CONCLUSION: Partner notification was possible in this setting, resulting in treatment of syphilis-infected individuals who otherwise would likely have remained untreated. However, given <10% of the partners reported by index cases were contactable; the results highlight the limitations of partner notification and the need for additional sexually transmitted infection control strategies.

Human papillomavirus and cervical neoplasia among female sex workers in Madagascar.

INTRODUCTION: Human papillomavirus (HPV) prevalence and type distribution were estimated among 90 female sex workers (FSWs) aged 18 to 58 years in Antananarivo, Madagascar. METHODS: A total of 90 FSWs aged 18 to 58 years in Antananarivo, Madagascar, were included in this study. Information on sexual and behavioral characteristics was obtained via a questionnaire. Exfoliated cervical cell specimens were collected for conventional cytologic examination and HPV DNA testing by polymerase chain reaction. The prevalence rates of HPV DNA and their corresponding 95% confidence intervals were stratified into the following 3 age groups: younger than 25, 25 to 34, and 35 years or older. To assess the association between HPV DNA positivity and sociodemographic and sexual behavioral factors, age-adjusted odds ratios and 95% confidence intervals were calculated using unconditional logistic regression. RESULTS: The HPV prevalence in exfoliated cervical cell specimens was 36.7%. The most common HPV types found were HPV-52 (11.1%), HPV-31 and -39 (each at 5.6%), and HPV-16 and -83 (each at 3.3%). The prevalence of low-grade squamous intraepithelial lesions was 3.3%, and that of atypical squamous cells of undetermined significance was 18.9%. No high-grade lesion was found. Although associations were imprecise, the HPV prevalence was higher among women who reported younger age at the first intercourse, contraceptive use, a history of cervical lesions, and no history of condom use. DISCUSSION: The prevalence rates of HPV and cervical lesions among FSWs in Madagascar appear higher than among FSW populations from other African countries with a relatively higher population-based prevalence of human immunodeficiency virus infection.

Nucleolar re-activation is delayed in mouse embryos cloned from two different cell lines.

Aim of this study was to evaluate and compare embryonic genome activation (EGA) in mouse embryos of different origin using nucleolus as a marker. Early and late 2-cell and late 4-cell stage embryos, prepared by in vitro fertilization (IVF), parthenogenetic activation (PG), and nuclear transfer of mouse embryonic fibroblast (MEF) and mouse HM1 embryonic stem cells (HM1), were processed for autoradiography following (3)H-uridine incubation (transcriptional activity), transmission electron microscopy (ultrastructure) and immunofluorescence (nucleolar proteins; upstream binding factor, UBF and nucleophosmin, B23). All early 2-cell embryos showed transcriptional activity only in nucleoplasm, not over nucleolar precursor bodies (NPBs). UBF was diffusely localized to cytoplasm and B23 to cytoplasm and nucleoplasm. Late 2-cell IVF and PG embryos displayed transcription over nucleoplasm and NPBs. Ultrastructurally, the latter were developing into functional nucleoli. NT-MEF and NT-HM1 embryos displayed transcription over nucleoplasm, but not over NPBs. Development of NPBs into nucleoli was lacking. UBF was in both groups localized to nucleoplasm or distinctly to presumptive NPBs. B23 was distinctly localized to NPBs. All 4-cell embryos presented nucleoplasmic transcription and developing fibrillo-granular nucleoli. UBF and B23 were distinctly localized to nucleoli. However, whereas fully transformed reticulated fibrillo-granular nucleoli were found in IVF and PG embryos, NT-MEF and -HM1 embryos displayed early NPBs transformation. In conclusion, despite normal onset of EGA in cloned embryos, activation of functional nucleoli was one cell cycle delayed in NT embryos. NT-MEF embryos displayed normal targeting but delayed activation of nucleolar proteins. Contrary, in NT-HM1 embryos, both of these processes were delayed.

A simplified biopsy method for precompacted mouse embryos: a technical report.

This article presents a new, simple and rapid embryo biopsy method. The blastomere for genetic analysis can be separated from a precompacted mouse embryo after a partial zona digestion with the use of a holding pipette. For the micromanipulation only two microcapillaries and micromanipulators are needed. The development of the biopsied embryos was studied during in vitro culture and in utero following embryo transfer. There was no significant difference between the treated and the control groups in the ratio of embryos that developed to the blastocyst stage, although the biopsied embryos were delayed in their development because they contained significantly fewer cells compared to the control ones at the same stage. Although there was no difference in the ratio of implantation, the development of the biopsied embryos in utero was also delayed 12-24 hours on the 9th day of pregnancy. No difference in development was visible from the 13th day of pregnancy. Statistically, no differences were found in the developmental ratio (number of developed fetuses/transferred embryos) of the control and treated embryos during gastrulation (9th day of pregnancy), at the beginning of organogenesis (13th day of pregnancy) and before birth (19th day of pregnancy). The embryo biopsy method presented here can be a new and useful tool for preimplantation genetic diagnosis.

Effect of swim up and Percoll treatment on viability and acrosome integrity of frozen-thawed bull spermatozoa.

The aim of this study was to investigate the effect of the swim up and Percoll methods to select frozen-thawed bull spermatozoa with high quality membrane and acrosomal integrity and final concentration. Semen samples from six Holstein-Friesian bulls were examined. The whole experiment was repeated three times. Before and after both treatments, spermatozoa were subjected to a double-staining method and evaluated by brightfield light microscope using 40x dry, or 100x oil immersion objectives. The concentration of spermatozoa evaluated by haemocytometer was 8.8 x 10(7)/ml after thawing, and the percentage of live cells with intact acrosome was 45.8%. Both treatments significantly increased the proportion of live spermatozoa compared with no treatment, and the use of Percoll gradient resulted in a significantly higher percentage of living cells with an intact acrosome (88.2%) than the swim up method (69.4%). The concentration of spermatozoa after Percoll separation (9.3 x 10(6)/ml) was higher than that after the swim up method (5.8 x 10(6)/ml). These results indicate that spermatozoa with a higher viability and acrosome integrity can be obtained by Percoll separation than by the swim up method. Therefore the use of Percoll-treated spermatozoa in IVF systems can be more expedient.

Preimplantation genetic diagnosis in cattle: a review.

Preimplantation Genetic Diagnosis (PGD) is reviewed and novel fields where it may be applied are investigated. Technical advances of PGD in cattle embryos have already enabled its integration as a part of the MOET (Multiple Ovulation Embryo Transfer) breeding system. PGD for well-defined selection targets can enhance cattle breeding and embryo trade. It allows embryo selection according to their sex, and it may be used to breed special cow lines, or top bulls, by selecting embryos for valuable production traits using Marker Assisted Selection (MAS). A good allelic profile and/or the insertion of a transgene can be detected by PGD. This review article presents the technical requirements for PGD, and shows that this biotechnological method has great economic potential.

Accumulation of the proteolytic marker peptide ubiquitin in the trophoblast of mammalian blastocysts.

Ubiquitination is a universal protein degradation pathway in which the molecules of 8.5-kDa proteolytic peptide ubiquitin are covalently attached to the epsilon-amino group of the substrate's lysine residues. Little is known about the importance of this highly conserved mechanism for protein recycling in mammalian gametogenesis and fertilization. The data obtained by the students and faculty of the international training course Window to the Zygote 2000 demonstrate the accumulation of ubiquitin-cross-reactive structures in the trophoblast, but not in the inner cell mass of the expanding bovine and mouse blastocysts. This observation suggests that a major burst of ubiquitin-dependent proteolysis occurs in the trophoblast of mammalian peri-implantation embryos. This event may be important for the success of blastocyst hatching, differentiation of embryonic stem cells into soma and germ line, and/or implantation in both naturally conceived and reconstructed mammalian embryos.

Comparison of different treatments for parthenogenetic activation of bovine oocytes matured in vitro.

In vitro matured bovine oocytes were treated by three parthenogenetic activation methods. Basic treatment with Ca-ionophore and cytochalasin D was combined with cycloheximide (Group 2), electric impulse (Group 3), and cycloheximide and electric pulse (Group 1) treatments, respectively. Survival and the in vitro development of parthenogenetic embryos to hatched blastocyst stage were compared. Rates of the first cleavage and morula development significantly differed among the treatment groups. Further development rates of the treated embryos up to blastocyst stage did not differ. The best results were obtained by the combination of cycloheximide and electric treatments (Group 1; 14% blastocyst, 7% hatched blastocyst). The results suggest that the combined treatment for oocyte activation is the most efficient and should be applied for cloning procedures.