Vascular endothelial growth factor gene 1154 G/A, 2578 C/A, 460 C/T, 936 C/T polymorphisms and association with recurrent pregnancy losses.

Vascular endothelial growth factor (VEGF) regulates endothelial cell proliferation, migration and differentiation. VEGF plays a critical role in angiogenesis during placenta formation. We investigated whether VEGF gene polymorphisms are associated with recurrent pregnancy loss. Thirty-eight women with recurrent pregnancy loss and 30 control women with live-born children were recruited from 2010 to 2011 in the region of Bursa, Turkey. VEGF gene polymorphisms were assessed with PCR-RFLP analysis of DNA samples obtained from leukocytes. DNA fragments were investigated by using appropriate primers. SNP scanning was performed using MnII, BgIII, BshI2361, Hsp92II restriction enzymes for 1154 G/A, 2578 C/A, 460 C/T, and 936 C/T polymorphisms, respectively. The frequencies of 2578 C/A, 460 C/T, 936 C/T polymorphisms were not significantly different between the controls and women with recurrent pregnancy loss. However, the prevalence of the 1154 G/A polymorphism A/A genotype was significantly higher in the recurrent pregnancy loss group (23.7 vs 3.4%). One of the four common polymorphisms of the VEGF gene was found to be more frequent in women with recurrent pregnancy loss. It is possible that disruption of VEGF function and placental angiogenesis can contribute to pregnancy loss in women with recurrent pregnancy loss.

Effects of Enterococcus faecium supplementation and floor type on performance, morphology of erythrocytes and intestinal microbiota in broiler chickens.

1. The experiment was to study the effects of floor type and probiotic supplementation (Enterococcus faecium) on performance, morphology of erythrocytes and intestinal microbiota of male Ross 308 broiler chickens. 2. The experimental design was a factorial 2 × 2 with 6 replicates. The factors were floor type (wire floor versus wood shaving litter) and the presence or absence of probiotic. 3. Birds housed on wood shavings exhibited significantly improved weight gain and food intake. 4. Addition of E. faecium led to significantly decreased food intake and gizzard weight. Supplementation with E. faecium positively influenced the ileal and caecal microbiota, with a significant decrease in the population of Escherichia coli. 5. Erythrocyte length decreased and erythrocyte width increased in the birds housed on wood shavings.

Influence of a combination of formic and propionic acids added to wheat- and barley-based diets on the performance and gut histomorphology of broiler chickens.

Day-old male chicks were used to test the effects of a combination of formic and propionic acids (CFP) added to wheat- and barley-based diets on the performance and gut histomorphology of broilers. The CFP, containing formic and propionic acids based on an inorganic phyllo-silicate carrier, was added to starter and grower broiler diets of standard (NRC, 1994) or lower nutrient density (LND) using different levels of wheat and barley. Body weight gain (BWG) and feed conversion ratio (FCR) of broiler chicks were significantly (P < 0.001) improved by CFP supplementation at 21 days of age regardless of nutrient density (ND). However, at 35 days of age CFP addition significantly (P < 0.001) increased BWG and FCR. Proventriculus weight was also significantly affected by the dietary treatments. Ileal viscosity was significantly (P < 0.05) affected by ND which was associated with wheat and barley contents, whereas feed viscosity was significantly (P < 0.05) affected by CFP supplementation. Villus height was significantly (P < 0.05) increased by the CFP supplementation. In the case of villus width, an interaction effect was observed: there was a significant (P < 0.05) decrease in the group fed the standard ND diet, but only a numerical decrease in the group fed the LND diet compared to control group. No significant effect of dietary treatment on the thickness of the lamina muscularis mucosae could be detected.

Y chromosome microdeletion in a case with Klinefelter's Syndrome.

In male infertility, the frequency of genetic factors is high. Klinefelter's Syndrome is the most frequent sex chromosomal abnormality detected in male infertility. In this study we report a patient diagnosed with Klinefelter's Syndrome with a deletion of the Yq interval. The patient was 24-years old with primary infertility. Semen analyses carried out in triplicate indicated azoospermia. The plasma leutenizing hormone (LH) and follicle stimulating hormone (FSH) levels were abnormally high and the testosterone level was lower than the usual range. Each of his testes had a volume of 3 cc. Peripheral blood karyotype analysis showed Klinefelter's Syndrome (47, XXY) pattern. Polymerase chain reaction amplification of DNA was performed using the following primers; AZFa (sY81, sY82, sY84), AZFb (sY127, sY142, sY164, RBM1), AZFc (CDY, BPY, sY254, sY255, sY277), AZFd (sY152, sY145, sY153). Analysis revealed a single deletion of AZFa region (sY84). Deletion of the AZFa region may be an additional factor for absolute azoospermia in men with Klinefelter's Syndrome. For individuals with Klinefelter's Syndrome who plan to undergo assisted reproduction techniques, Y chromosome microdeletion screening can diagnostically be convenient.

Natural transmission of AZFb Y-chromosomal microdeletion from father to his three sons.

Microdeletions of the so-called azoospermia factor (AZF) locus of the Y chromosome long arm (Yq) are an etiological factor of severe oligozoospermia or azoospermia. Patients affected are infertile unless assisted reproductive techniques are used. We report the case of an azoospermic patient (proband) and three brothers who inherited a Yq microdeletion from their father through a spontaneous pregnancy. Leukocyte DNA was extracted using a commercially available kit. A total of 15 pairs of sequence-tagged site (STSs) based primers, spanning the AZFa, b and c regions, were used for screening. All brothers and their father carried a Yq microdeletion of the AZFb subregion where the RNA-binding motif (RBM) gene is located. The proband carried additional deletions of the AZFa and AZFb subregions. RBM deletion can be associated with oligozoospermia allowing natural conception and therefore natural transmission of this genetic anomaly.

Clinical, andrological and genetic characteristics of patients with congenital bilateral absence of vas deferens (CBAVD).

Congenital bilateral absence of the vas deferens (CBAVD) is a form of infertility with an autosomal recessive genetic background in otherwise healthy males. In this study, we examined the clinical and cystic fibrosis transmembrane-conductance regulator (CFTR) gene mutations in sixty patients with bilateral absence of vas deferens that applied to andrology clinic due to male factor infertility. Urogenital anomalies of vas deferens, seminal vesicle and epididymis were detected in our patient group. CFTR gene mutations, which are known to be frequent among cystic fibrosis patients, could not be detected in our patient group with that high frequency. Delta F508 mutations were detected in only 6% of patients. IVS8 polyT alleles were positive in 68% of patients. No 1677delTA mutations and M470V variants were detected in our patient group. However, sperm retrieval is almost always possible from CBAVD patients; secondary pathologies may also result defective spermatogenesis.

Genetic anomalies detected in patients with non-obstructive azoospermia and oligozoospermia.

Genetic factors have a major importance in male infertility etiology. Numerical and structural chromosomal abnormalities seem to be frequent inoligospermia and azoospermia cases with unknown etiology. In this study, 819 patients with azoospermia (383) and oligospermia (436) who attended the infertility department between 1995-2005 were evaluated. Spermogram and basic hormone proties (FSH-testosterone) were studied two times in a one month interval from each patient, and all the cases were evaluated cytogenetically. The 47 (12%) of 383 azoospermia patients and the 20 (4%) of 436 oligospermia patients were found to have chromosomal abnormalities. The 9 (19%) of the chromosomal abnormalities found in azoospermia patients were autosomal and the 38 (80%) were gonosomal. In oligospermia cases, the 8 (40%) of the chromosomal abnormalities were autosomal and 12 (60%) were gonosomal. Cytogenetic analysis and genetic counseling would be helpful in infertile males with azoospermia and oligospermia by determining the genetic factors causing infertility and by assessing the genetic risks of the offsprigs provided by assisted reproductive techniques.

Relationship between granular cytoplasm of oocytes and pregnancy outcome following intracytoplasmic sperm injection.

Couples undergoing intracytoplasmic sperm injection (ICSI) for male infertility using oocytes with centrally located granular cytoplasm (CLCG) were evaluated for fertilization, embryo development, implantation and pregnancy rate. CLCG is a rare morphological feature of the oocyte, that is diagnosed as a larger, dark, spongy granular area in the cytoplasm. Severity is based on both the diameter of granular area and the depth of the lesion. Twenty-seven couples with 39 cycles presenting CLCG in >50% of retrieved oocytes were evaluated. A total of 489 oocytes was retrieved, out of which 392 were at MII. CLCG was observed in 258 of the MII oocytes (65. 8%); 66.7% of these oocytes had slight and 33.3% had severe CLCG. The overall fertilization rate was 72.2% and no statistical significant difference was found between normal and CLCG oocytes and between the oocytes representing slight and severe CLCG. The development and quality of embryos was the same in normal and CLCG oocytes. In nine cycles, preimplantation genetic diagnosis was executed to evaluate a possible accompanying chromosomal abnormality. Out of 44 blastomeres biopsied, 23 had chromosomal abnormality (52. 3%). Eleven pregnancies were achieved in 39 cycles (28.2%), six pregnancies resulted in abortion (54.5%). The implantation rate was found to be 4.2%. Only five ongoing pregnancies were achieved in 39 cycles (12.8%). Couples with CLCG oocytes should be informed about poor on-going pregnancy rates even if fertilization, embryo quality and total pregnancy rates are normal. Furthermore, a high aneuploidy rate may be linked to a high abortion rate.

Healthy births and ongoing pregnancies obtained by preimplantation genetic diagnosis in patients with advanced maternal age and recurrent implantation failure.

Preimplantation genetic diagnosis (PGD) and subsequent embryo development was evaluated in 72 couples presenting at our centre for intracytoplasmic sperm injection (ICSI) due to severe male factor. The embryo biopsies were performed in Ca(2+)/Mg(2+)-free medium. These patients were further divided into those with advanced maternal age (AMA, n = 49) and those with recurrent implantation failure (RIF, n = 23). Fluorescence in-situ hybridization (FISH) was carried out on 329 blastomeres (91.3%) with probes for the X, Y, 13, 18 and 21 chromosomes. The chromosomal abnormality rate was 41.3% with no significant difference between the AMA and RIF groups. Aneuploidy accounted for the majority (72.8%) of chromosomal abnormalities. Out of 329 embryos, 84.2% had cleaved after 24 h and 15.1% had arrested. Embryos were transferred in 70 patients and 22 pregnancies were achieved (31.4% with an ongoing pregnancy rate of 28.5%). There were no significant differences between the pregnancy rates of the AMA and RIF groups (32.5 and 30% respectively). Therefore PGD should be offered to patients with AMA and RIF. Furthermore, the use of Ca(2+)/Mg(2+)-free medium during the blastomere biopsy facilitates the procedure, while further embryo cleavage, ongoing pregnancies and healthy births are possible.