Association of combined GIF290T>C heterozygous mutation/FUT2 secretor variant with neural tube defects.

Folate and vitamin B12 are needed for the proper embryo-fetal development possibly through their interacting role in the 1-carbon metabolism. Folate fortification reduces the prevalence of complex birth defects, and more specifically neural tube defects (NTDs). GIF and FUT2 are 2 genes associated with the uptake and blood level of vitamin B12. We evaluated GIF and FUT2 as predictors of severe birth defects, in 183 aborted fetuses compared with 375 healthy newborns. The GIF290C allele frequency was estimated to 0.4% in healthy newborns and to 8.1% in NTD fetuses (odds ratio 17.8 [95% confidence interval CI: 4.0-77.6]). The frequency of FUT2 rs601338 secretor variant was not different among groups. The GIF 290C heterozygous/FUT2 rs601338 secretor variant combined genotype was reported in 6 of the 37 NTD fetuses, but not in other fetuses and healthy newborns (P < .0001). This GIF/FUT2 combined genotype has been previously reported in children with congenital gastric intrinsic factor (GIF) deficiency, with respective consequences on B12 binding activity and GIF secretion. In conclusion, a genotype reported in congenital GIF deficiency produces also severe forms of NTD. This suggests that vitamin B12 delivery to neural tissue by the CUBN/GIF pathway could play a role in the neural tube closure mechanisms.

Elaboration of Sterically Stabilized Liposomes for S-Nitrosoglutathione Targeting to Macrophages.

S-nitrosoglutathione (GSNO) is a potential therapeutic for infectious disease treatment because of its pivotal role in macrophage-mediated inflammatory responses and host defense in addition to direct antibacterial activities. In this study, sterically stabilized cationic liposomes (SSCL) and sterically stabilized anionic liposomes (SSAL) were developed as nanocarriers for macrophage targeting. Elaborated liposomes were characterized in terms of size, zeta potential, morphology, encapsulation efficiency, in vitro drug release behavior and cytotoxicity. Their versatility in targeting monocytes/macrophages was determined by confocal laser scanning microscopy and transmission electron microscopy. Flow cytometry revealed that cellular uptake of both SSCL and SSAL was governed by several endocytic clathrin- and caveolae-dependent mechanisms. Quantitative assessments of intracellular nitric oxide demonstrated highly efficient uptake of GSNO-loaded SSCL that was twenty-fold higher than that of GSNO-free molecules. GSNO-loaded SSCL displayed strong bacteriostatic effects on Staphylococcus aureus and Pseudomonas aeruginosa, which can be involved in pulmonary infectious diseases. These results reveal the potential of liposomal GSNO as an anti-infective therapeutic due to its macrophage targeting capacity and direct antibacterial effects.

[Azoospermia and 45,X/46,XY chromosomal mosaicism: a case report]. / Azoospermie et mosaïque 45,X/46,XY : à propos d'un cas.

Chromosomal abnormalities are common in patients with oligozoospermia or azoospermia. We report the case of a 32-year patient, with male phenotype, and without hormonal or morphological abnormalities, with a severely reduced spermatogenesis. It was revealed a 45,X/46,XY gonadal dysgenesis. We have reviewed the various problems inherent in the discovery of this rare gonadal dysgenesis, including genetic, cancer and fertility risks.

[In search of the surgical sperm cell: do we have to perform a testicular biopsy systematically?]. / À la recherche du spermatozoïde chirurgical : doit-on effectuer une biopsie testiculaire systématique ?

INTRODUCTION: Testicular biopsies are diagnostic and therapeutic tools involved in male infertility care. However, this surgery is invasive and not systematically successful. We studied the preoperative clinical and hormonal factors allowing to predict the obtaining of sperm cells. PATIENTS AND METHODS: A retrospective study was conducted on 209 patients who all had a testicular biopsy for procreation medically assisted (PMA). The studied criteria were: the age at the time of the surgery, the male cause of the infertility, the testicular volume, the tobacco smoking exposure, the concentrations of estradiol, FSH, LH, prolactin, and testosterone. The comparison of both groups (successful biopsy versus failed biopsy) was made in bivariate analysis then in multivariate analysis. RESULTS: The testicular volume average and the cause were the two only factors which had a real influence on the negativity of the biopsy. In it was added in bivariate analysis a statistically significant correlation of the smoking exposure and the FSH with the failed biopsy. DISCUSSION: The existence of these factors, and their accumulation, was strongly predictive of a failure of the biopsy. However, we found germ cells in patients exposed to the studied factors, letting think that it is systematically necessary to propose the surgery at the risk of a limited profit.

Sperm cephalic vacuoles: new arguments for their non acrosomal origin in two cases of total globozoospermia.

To add new arguments concerning the origin of the sperm-head vacuoles observed under high magnification with interference contrast microscopy, we carried out in two patients with total globozoospermia confirmed using transmission electron microscopy (TEM), a detailed sperm morphometric analysis with high magnification (×6000) under Nomarski contrast, an acrosomal status analysis (using fluorescent labelling with peanut agglutinin (PNA) lectins and anti-CD46 antibodies) and a nuclear status analysis (using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling assay TUNEL, sperm chromatin structure assay SCSA and aniline blue staining). Our two patients with globozoospermia had relative sperm vacuole areas of 6.3% and 5%, similar to those observed in a reference population of 12 fertile men (5.9%). TUNEL and SCSA assays gave normal results in both patients, although the percentage of immature nuclei using aniline blue staining was increased (27 and 46% for patient 1 and 2 respectively). Cytofluorescence and TEM analysis evidence differences between the two patients: although no acrosomal neither Golgi residue could be detected in patient 1, patient 2 had positive PNA lectin labelling for 9% of spermatozoa and Golgi residues were seen using electron microscopy. Unlike patient 1, a live birth could be obtained after intracytoplasmic sperm injection (ICSI) for patient 2. This descriptive study of two patients with total globozoospermia confirmed using TEM argue in favour of a deep analysis of total globozoospermia before assisted reproductive technology and provides further information on the non-acrosomal origin of the sperm-head vacuoles observed under high magnification.

Pre-pregnancy body mass index and weight gain during pregnancy: relations with gestational diabetes and hypertension, and birth outcomes.

To study the relationship between pre-pregnancy body mass index (BMI) and weight gain during pregnancy with pregnancy and birth outcomes, with a focus on gestational diabetes and hypertension and their role in the association with fetal growth. We studied 1,884 mothers and offspring from the Eden mother-child cohort. Weight before pregnancy (W1) and weight after delivery (W2) were collected and we calculated BMI and net gestational weight gain (netGWG = (W2 - W1)/(weeks of gestation)). Gestational diabetes, hypertension gestational age and birth weight were collected. We used multivariate linear or logistic models to study the association between BMI, netGWG and pregnancy and birth outcomes, adjusting for center, maternal age and height, parity and average number of cigarettes smoked per day during pregnancy. High BMI was more strongly related to the risk of giving birth to a large-for-gestational-age (LGA) baby than high netGWG (odds ratio OR [95% CI] of 3.23 [1.86-5.60] and 1.61 [0.91-2.85], respectively). However, after excluding mothers with gestational diabetes or hypertension the ORs for LGA, respectively weakened (OR 2.57 [1.29-5.13]) for obese women and strengthened for high netGWG (OR 2.08 [1.14-3.80]). Low in comparison to normal netGWG had an OR of 2.18 [1.20-3.99] for pre-term birth, which became stronger after accounting for blood pressure and glucose disorders (OR 2.70 [1.37-5.34]). Higher net gestational weight gain was significantly associated with an increased risk of LGA only after accounting for blood pressure and glucose disorders. High gestational weight gain should not be neglected in regard to risk of LGA in women without apparent risk factors.

Drug delivery by polymeric nanoparticles induces autophagy in macrophages.

Drug delivery nanosystems are currently used in human therapy. In preliminary studies we have observed that Eudragit RS nanoparticles, prepared by nanoprecipitation or double emulsion techniques, are cytotoxic for NR8383 rat macrophages. In this study, we expand our previous analysis and suggest that unloaded Eudragit RS nanoparticles prepared by nanoprecipitation (NP/ERS) may induce important morphological and biochemical cellular modifications leading to cellular death. In NR8383 rat macrophages cell line exposed to doses varying from 15 to 100 µg/mL, NP/ERS nanoparticles are internalized inside the cells, reach the mitochondria and alter the structure of these organelles. In addition, the exposure to nanoparticles induces cellular autophagy as demonstrated by electron microscopy analysis, microchip array, qRT-PCR and Western blot assays. Although toxicity of nanoparticles has already been evidenced, it is the first time that results show clearly that the toxicity of polymeric nanovectors may be related to an activation of autophagy.

A retrospective study by oligonucleotide array-CGH analysis in 50 fetuses with multiple malformations.

OBJECTIVE: To retrospectively define the frequency and the nature of submicroscopic chromosomal imbalances among fetuses with multiple congenital anomalies (MCA). METHODS: We used oligonucleotide arrays to perform comparative genomic hybridization after termination of pregnancy in 50 polymalformated fetuses with a normal karyotype. These fetuses presented with at least three significant malformations (42 cases) or a severe brain anomaly (eight cases). RESULTS: We identified a deleterious copy number variation (CNV) in five fetuses (10%). De novo genomic imbalances identified in this study consisted of a 6qter deletion in a fetus with brain and renal malformations, a mosaicism for a 8p tetrasomy in a fetus with agenesis of corpus callosum, growth retardation, mild facial dysmorphic features, and vertebral anomalies, a 17p13.3 deletion in a fetus with a complex brain malformation, and a partial 11p trisomy in a fetus with severe growth retardation and oligoamnios. In one case, we identified a partial 17q trisomy resulting from malsegregation of a cryptic-balanced translocation. CONCLUSIONS: This study shows that array comparative genomic hybridization (aCGH) is particularly effective for identifying the molecular basis of the disease phenotype in fetuses with multiple anomalies. Our study should help to define clinical relevant regions that would need to be included in targeted arrays designed for prenatal testing.

Refining the clinicopathological pattern of cerebral proliferative glomeruloid vasculopathy (Fowler syndrome): report of 16 fetal cases.

Cerebral proliferative glomeruloid vasculopathy (PGV) is a severe disorder of brain angiogenesis, resulting in abnormally thickened and aberrant perforating vessels, forming glomeruloids with inclusion-bearing endothelial cells. This peculiar vascular malformation was delineated by Fowler in 1972 as a stereotyped lethal fetal phenotype associating hydranencephaly-hydrocephaly with limb deformities, called Fowler syndrome (FS) or "proliferative vasculopathy and hydranencephaly-hydrocephaly" or "encephaloclastic proliferative vasculopathy" (OMIM#225790). In PGV, the disruptive impact of vascular malformation on the developing central nervous system (CNS) is now well admitted. However, molecular mechanisms of abnormal angiogenesis involving the CNS vasculature exclusively remain unknown, as no genes have been localized nor identified to date. We observed the pathognomonic FS vascular malformation in 16 fetuses, born to eight families, four consanguineous and four non-consanguineous. A diffuse form of PGV affecting the entire CNS and resulting in classical FS in 14 cases, can be contrasted to two cases with focal forms, confined to restricted territories of the CNS. Interestingly in PGV, immunohistological response to a marker of pericytes (SMA, Smooth in PGV Muscle Actin), was drastically reduced as compared to a match control. Our studies has expanded the description of FS to additional phenotypes, that could be called Fowler-like syndromes and suggest that the pathogenesis of PGV may be related to abnormal pericyte-dependent remodelling of the CNS vasculature, during CNS angiogenesis. Gene identification will determine the molecular basis of PGV and will help to know whether the Fowler-like phenotypes are due to the same underlying molecular mechanisms.

Congenital dyserythropoietic anaemia, type I, in a Caucasian patient with retinal angioid streaks (homozygous Arg1042Trp mutation in codanin-1).

A congenital dyserythropoietic anaemia (CDA) was recognised in a French Caucasian male patient. Blood smears showed a pronounced aniso-poikilocytosis. Bone marrow light microscopy showed signs of dyserythropoesis, but no internuclear chromatin bridges. Electron microscopy disclosed erythroblast nuclei with the Swiss cheese aspect and the presence of cytoplasmic organelles, assessing the diagnosis of CDA I. The presence of internuclear chromatin bridges may thus be missing in CDA I. The patient proved to be homozygous for the Arg1042Trp mutation in codanin-1 (the 'Bedouin mutation'). By the age of 25, the patient's vision started to deteriorate as a result of retinal angioid streaks and macular abnormalities. Evolution was controlled and the patient, being nearly 50 yr old now, still has a partial use of his eyes. This second case of retinal angioid streaks reported in CDA I adds to the non-haematological features likely to be associated with this condition.

Prenatal diagnosis of mosaicism for 11q terminal deletion.

The phenotype of 11q terminal deletion also known as Jacobsen syndrome is a clinically well known entity whose diagnosis in infancy and childhood is based on clinical examination, hematological and cytogenetic findings. Hematological features in Jacobsen syndrome are very similar to those reported in Paris-Trousseau syndrome (PTS) which is also associated with11q terminal deletion. Karyotype analysis shows a variable terminal deletion from 11q23 sub-band extending to the telomere. Most often in patients with Jacobsen syndrome, this chromosomal deletion is present in all metaphases. We report on the identification of a distal 11q deletion in mosaic (20% of deleted cells) in a fetus ascertained after amniocentesis for maternal serum screening test indicative for Down syndrome. The present case is the third prenatal diagnosis of a mosaic for a distal 11q deletion with the lowest mosaicism rate. The 2D-ultrasound examination and cord blood hematological studies were useful to estimate the prognosis at term, considering the contribution of the mosaicism rate to the phenotypic variability in Jacobsen syndrome. The identification of mosaicism for distal 11q deletion is a very rare event in prenatal diagnosis. This case illustrates the complexity in genetic counselling for prenatally ascertained partial monosomy 11qter in mosaic.

Quantitative analysis of the elastic fibres in the human temporomandibular articular disc and its attachments.

A quantitative study of the elastic fibres found in the human temporomandibular disc and its attachments was performed. Seven left discs from 57- to 82-year-old subjects, without macroscopic evidence of a TMJ disorder, were analysed and prepared in parasagittal sections. The surface amount was measured, thresholded and expressed from 0 to 1, using microscopic digitized views after Weigert's resorcin-fuchsin staining of elastic fibres. Fibre density rates ranged from 0 to 0.687. The mean density was 0.1532 (sigma=0.1150) in the upper bilaminary zone, 0.1097 (sigma=0.1159) in the lower bilaminary zone, 0.0474 (sigma=0.0782) in the anterior band, 0.0180 (sigma=0.0603) in the posterior band and null in the intermediate zone. The difference in density rate between the structures was significant, except for the posterior band and the intermediate zone. The elastic fibre density rates in central and medial locations of the upper and lower bilaminary zones were twice as big as in the lateral locations. In the anterior band, the elastic fibre density was less abundant medially than in its lateral part. These quantitative results support the current elastic fibre distribution scheme, and confirm the necessity of studying their orientation, taking into account age and temporomandibular joint health parameters.

Ultrasound-guided radiofrequency-driven umbilical cord occlusion: an in-vitro study.

OBJECTIVES: To assess the ability and safety of radiofrequency (RF) to induce cord thermal lesions using in-vitro perfused umbilical cords. METHODS: Nineteen human term umbilical cords were cannulated at both ends and perfused continuously with saline serum in a saline serum bath (37 degrees C). The RF electrode was then inserted into the cord close to the umbilical vein. Different RF power and temperature controls were applied to determine the optimal RF procedure in terms of cord tissue injury and safety in nine experiments. The safety of RF procedures was investigated in ten cords by measuring temperature changes at different sites close to the RF electrode insertion and the impact of RF on cord narrowing was evaluated by continuous monitoring of intraluminal pressure. Subsequent histopathological analysis was carried out in all cases. RESULTS: The optimal RF procedure reached a temperature of 100 degrees C in 10 min. RF produced a significant increase in intraluminal pressure (from 54.2 +/- 16.4 mmHg at baseline to 118.3 +/- 42.7 mmHg after 10 min, P < 0.05). There was no significant increase in temperature next to the site of insertion during the RF procedure. Histopathological analysis confirmed a > 30% decrease in cord and vein diameter. Cord tissue lesions were characterized by damage in the vessel walls and in the surrounding Wharton's jelly. CONCLUSION: Our results suggest that RF might be a feasible and safe technique to induce occlusion of umbilical vessels. Further in-vivo experiments are needed to assess its ability to induce a complete occlusion of the umbilical cord.

[Negative impact of cigarette smoking on male fertility: from spermatozoa to the offspring]. / Impact négatif du tabac sur la fertilité masculine: des spermatozoïdes à la descendance.

Cigarette smoking has negative effects on male fertility. Recent studies showed an active transfer of several components of cigarettes through the blood-testis barrier. The presence of these components in the seminal plasma may induce a degradation of sperm parameters and nuclear quality of spermatozoa, and compromise the chances of pregnancy. Moreover, smoking may have a negative impact on the smokers'offspring: poor quality embryos, development of childhood cancers. Oxidative stress-induced DNA damage seems to be one of the major causes of sperm quality alteration. Several methods are now available to analyze the degree of DNA fragmentation. In order to optimize the success rate of assisted reproduction technologies, the deleterious effects of smoking on male fertility and the necessity of cessation have to be explained in detail to these patients.

Expression of IL-2alpha and IL-2beta receptors on the membrane surface of human sperm.

Cytokines are secreted proteins that act as local immunological mediators. Increased seminal cytokine concentrations are associated with fertility problems. The purpose of the present study was to investigate the presence of IL-2alpha, and IL-2beta receptors on fresh and isolated sperm by flow cytometry and transmission electron microscopy. Twenty sperm samples from oligospermic men were incubated with CD25, a mouse monoclonal antibody specific for IL-2alpha-chain receptor, and CD122, a mouse monoclonal antibody specific for IL-2beta-chain receptor. The strong initial fluorescence intensity and, subsequently, a labeling index yielded by CD25 and CD122 decreased in sperm centrifuged on a Percoll gradient (p < .05). The expression of CD25 and CD122 correlated negatively with fresh sperm concentration, but in sperm centrifuged on a Percoll gradient there was no correlation. Labeling with CD25 and CD122 antibody was evident on the head and the middle piece in fresh sperm, while in sperm centrifuged on a Percoll gradient a weak labeling was observed only on the principal piece. The authors have identified and localized cytokine receptors on human sperm for the first time. Cytokine receptors may be involved in the regulation of pathophysiological events in sperm cell functions and male infertility. The exact pathway involved in modulation of these receptors requires further investigation. These results contribute to the understanding of cytokine-sperm relationships.

Choroid plexus and ageing in rats: a morphometric and ultrastructural study.

Choroid plexuses (CP) are intraventricular structures involved in the production of cerebrospinal fluid (CSF) and in the synthesis and transport of numerous CSF components. Age-related modifications of the CP structure are still ill defined. We performed an ultrastructural and morphometric study of ageing CP in nine Sprague-Dawley rats 6, 18 and 30 months of age. Epithelial cells of CP villi were cubic in shape at 6 months, more dome-like at 18 months, and significantly flattened at 30 months of age. Epithelial basement membranes were thin and regular at 6 months, significantly thicker at 18 months and thicker and irregular at 30 months. Intravillous stroma increased nonhomogeneously with age. The ageing of CP in rats is characterized morphologically by epithelial atrophy, irregular fibrosis of the stroma and a thickening of epithelial basement membranes. These modifications suggest an alteration of secretory and filtrating functions in ageing CP.

Protected renal stenting with the PercuSurge GuardWire device: a pilot study.

PURPOSE: To evaluate the feasibility and safety of renal artery angioplasty and stenting utilizing a distal protection device to reduce the risk of intraprocedural atheroembolism. METHODS: Twenty-eight hypertensive patients (18 men; mean age 71.3 +/- 8.6 years, range 49-87) with atherosclerotic renal artery stenosis (4 bilateral) underwent angioplasty and stenting with distal protection in 32 renal arteries (29 ostial lesions). The lesion was crossed with a GuardWire temporary occlusion balloon, which was inflated to provide parenchymal protection. Generated debris was aspirated and analyzed. Blood pressure and serum creatinine levels were followed. RESULTS: Immediate technical success was 100%. All lesions were stented, either directly (14 ostial lesions), after predilation (15 ostial lesions), or owing to suboptimal angioplasty (3 nonostial lesions). Visible debris was aspirated from all patients. Mean particle number and diameter were 98.1 +/- 60.0 per procedure (range 13-208) and 201.2 +/- 76.0 microm (range 38-6206), respectively. Mean renal artery occlusion time was 6.55 +/- 2.46 min (range 2.29-13.21). Mean follow-up was 6.7 +/- 2.9 months (range 2-17). Systolic and diastolic blood pressure declined from 167.0 +/- 15.2 and 103.0 +/- 12.0 mm Hg, respectively, to 154.7 +/- 12.3 and 93.2 +/- 6.8 mm Hg after the procedure. The mean creatinine level dropped from 1.34 +/- 0.35 mg/dL preprocedurally to 1.22 +/- 0.36 mg/dL at 24 hours and remained constant. At 6-month follow-up, renal function did not deteriorate in any patient, whereas 5 patients with baseline renal insufficiency improved after the procedure. CONCLUSIONS: These preliminary results suggest the feasibility and safety of distal balloon occlusion during renal interventions to protect against atheroembolism. This technique's beneficial effects should be evaluated by randomized studies.

Eosinophil count in nasal secretions of subjects with and without nasal symptoms.

The aim of this paper, based on a cross-sectional study of 129 patients with nonallergic chronic nasal symptoms and 40 healthy controls, was to examine the leucocyte differential count in nasal secretions as a diagnostic test. Nasal secretions were collected using preweighed suction glass canulas under controlled conditions (-100Pa, 30 sec). Leucocyte and differential counts were performed using a Thoma hemocytometer and on cytospin slides after May-Grünwald-Giemsa staining. The percentage of eosinophils (Eo) was significantly higher in patients (mean +/- SEM: 15.1 +/- 2.3%) than in controls (5 +/- 2.6%) (p < 0.04). Comparison of the frequency distribution of the percentage of Eo in patients and controls clearly showed a subgroup of patients presenting with nasal secretion hypereosinophilia, and allowed us to set the positivity criterion at Eo = 20%. Diurnal variations in Eo count in 11 controls and 8 patients confirmed the value of the cutoff point. In 28 patients with nasal polyposis who underwent surgery, a correlation was found between secretion and tissue eosinophelia (r = 0.58, p = 0.001). Patients with nasal secretion hypereosinophilia had no more leucocytes in their secretions than healthy controls, the increase in eosinophils being balanced by a decrease in neutrophils. In patients without hypereosinophilia, the number of leucocytes per milligram of secretion was four times higher (8672 +/- 2521) than in the controls (2020 +/- 823) (p = 0.06) (cut-off point = 2500 leu/mg). These data show that the nasal cytogram can be modified either in qualitative or quantitative way, probably depending on the underlying inflammatory process.

Monocyte-derived IL-10-secreting dendritic cells in choroid plexus epithelium.

Choroid plexuses form an interface between peripheral blood and cerebrospinal fluid. Dendritic-like cells have been reported in a few studies of choroid plexuses in man. Here we used electron microscopy and immunophenotyping to precise the morphologic features and phenotype of these cells. Examination of 10 human choroid plexuses evidenced intra-epithelial dendritic cells with a clear cytoplasm, reniform nucleus and long expansions. These cells express MHC Class II, CD11b, CD14, CD32, CD68 and IL-10, but not CD40, CD80 or CD86, suggesting an immunosuppressive role for these dendritic cells. Their sentinel position could make them participate to the immunological silence of the brain.