NOS3 gene variants and male infertility: Association of 4a/4b with oligoasthenozoospermia.

Results of recent studies confirmed that oxidative stress negatively affects sperm motility and causes sperm DNA damage. Produced by nitric oxide synthase 3 (NOS3), nitric oxide is considered to be one of the important mediators of oxidative stress in testis tissue. The aim of this study was to assess the possible association of three genetic variants (rs2070744, rs1799983 and intron variant 4a/4b) in NOS3 gene and infertility occurrence in two groups of infertile men (idiopathic azoospermia and oligoasthenozoospermia) and fertile controls. Genotypes for the single-nucleotide genetic variants rs1799983 and rs2070744 were determined by PCR-RFLP, while genotyping of intron 4 variant 4a/4b was performed by gel electrophoresis of PCR products. Statistical analysis was performed by SNPStats software. No significant association between the three genetic variants of the NOS3 gene and infertility risk was determined comparing allele and genotype frequencies among group of patients diagnosed with azoospermia and the control group. Nevertheless, there was a significant positive association between 4a/4b and infertility in the group of males diagnosed with oligoasthenozoospermia, under overdominant genetic model. Our findings suggest that tandem repeat variant within intron 4 of the NOS3 gene is associated with an increased risk of infertility in men diagnosed with idiopathic oligoasthenozoospermia.

Genetic variants in RNA-induced silencing complex genes and prostate cancer.

PURPOSE: The purpose of this study is to evaluate the potential association between genetic variants in genes encoding the components of RNA-induced silencing complex and prostate cancer (PCa) risk. Genetic variants chosen for this study are rs3742330 in DICER1, rs4961280 in AGO2, rs784567 in TARBP2, rs7813 in GEMIN4 and rs197414 in GEMIN3. METHODS: The study involved 355 PCa patients, 360 patients with benign prostatic hyperplasia and 318 healthy controls. For individuals diagnosed with PCa, clinicopathological characteristics including serum prostate-specific antigen level at diagnosis, Gleason score (GS) and clinical stage were determined. Genotyping was performed using high-resolution melting analysis, PCR-RFLP, TaqMan SNP Genotyping Assay and real-time PCR-based genotyping assay using specific probes. Allelic and genotypic associations were evaluated by unconditional linear and logistic regression methods. RESULTS: The study provided no evidence of association between the analyzed genetic variants and PCa risk. Nevertheless, allele A of rs784567 was found to confer the reduced risk of higher serum PSA level at diagnosis (P = 0.046; Difference = -66.64, 95 % CI -131.93 to 1.35, for log-additive model). Furthermore, rs4961280, as well as rs3742330, were shown to be associated with GS. These variants, together with rs7813, were found to be associated with the lower clinical stage of PCa. Also, rs3742330 minor allele G was found to be associated with lower PCa aggressiveness (P = 0.036; OR 0.14, 95 % CI 0.023-1.22, for recessive model). CONCLUSIONS: According to our data, rs3742330, rs4961280 and rs7813 qualify for potentially protective genetic variants against PCa progression. These variants were not shown to be associated with PCa risk.

Pregnancy associated with melanoma and fetal anomalies: a case report and review of literature.

The estimated incidence of melanoma complicating pregnancy has ranged from 0.1 to 2.8 per 1,000 pregnancies. Here the Authors present a case of a 40-year-old pregnant woman, who was admitted to the Clinic of Obstetrics and Gynaecology in 26 weeks of gestation, with diagnosis of melanoma and suspected with fetal anomaly, as possible bowel obstruction, and polyhydraminos. The melanoma was asported with a wide local excision under local anesthesia. Histological evaluation revealed melanoma Stage Ib (Clark IV, Breslow thickness 1.2 mm, pT2a). Lymph node sonography of neck, axilla, inguinum, abdomen, and pelvis as well as chest radiography did not demonstrate any evidence of metastatic disease. After vaginal delivery at 37 weeks of gestation, the female preterm hypotrophic newborn was transferred to the Institute for Neonatology and underwent resection of duodenojejunal atresia with tapering duodenoplasty and duodenojejunal termino-terminal anastomoses due to intestinal obstruction. No evidence of the melanoma was found in the placenta. Two years later the child was healthy and the mother was disease-free.

Discrimination and Romani health: a validation study of discrimination scales among Romani women in Macedonia and Serbia.

OBJECTIVES: Scales used to assess discrimination in public health research have rarely been validated outside of high income countries. Our objective was to validate the Experiences of Discrimination (EOD) scale and the Everyday Discrimination Scale (EDS) among 410 Romani women in Macedonia and Serbia. METHODS: Romani female interviewers conducted interviews in 2012-2013. We used a multiple indicator multiple cause approach to test a one-factor model for each scale and to assess differential item functioning (DIF) by age, wealth, country, and education. We also measured associations between the EOD and EDS with smoking in the past year and psychological distress. RESULTS: Three items of the EOD were conceptually irrelevant. Two items of the EDS were not conditionally independent. DIF was found by country for one item in each scale. After excluding these items, all scales exhibited good model fit and were associated with smoking (EOD beta = 0.40, 95% CI = 0.18, 0.63; EDS beta = 0.33, 95% CI = 0.12, 0.54) and psychological distress (EOD beta = 0.26, 95% CI = 0.15, 0.37; EDS beta = 0.26, 95% CI = 0.04, 0.47). CONCLUSIONS: Discrimination scales can be adapted for use among Romani women and are associated with both smoking and psychological distress.

Assessment of association between genetic variants in microRNA genes hsa-miR-499, hsa-miR-196a2 and hsa-miR-27a and prostate cancer risk in Serbian population.

Due to their potentially functional significance, genetic variants within microRNA genes have been recognized as candidates for cancer-related genetic biomarkers. Among the most extensively studied so far are rs3746444, rs11614913 and rs895819. Nevertheless, only few previous studies in Asian population analyzed the association of rs3746444 and rs11614913 with prostate cancer (PCa) risk, while rs895819 was not evaluated in relation to this issue. The aim of this study was to assess the possible association between these genetic variants and PCa risk and progression in Serbian population. 355 samples of peripheral blood were obtained from the patients with PCa and 353 samples from patients with benign prostatic hyperplasia (BPH). 312 volunteers derived from general population who gave samples of buccal swabs were included in the control group. Genotyping of rs3746444, rs11614913 and rs895819 was performed by using PCR-RFLP method, HRM analysis and allele-specific PCR, respectively. Allelic and genotypic associations were evaluated by unconditional linear (for serum PSA level in PCa patients) and logistic regression method with adjustment for age. Minor allele C of rs895819 was found to be associated with the increased risk of developing PCa under dominant (P=0.035; OR=1.38, 95%CI 1.02-1.86) and overdominant (P=0.04; OR=1.37, 95%CI 1.01-1.85) genetic model. Same genetic variant was found to be associated with the clinical stage of localized PCa, as well as with the presence of distant metastases. Allele G of rs3746444 was also shown to be associated with the decreased risk of PCa progression. According to our data, rs3746444 qualifies for a genetic variant potentially associated with PCa aggressiveness in Serbian population. Furthermore, our study provided the first evidence of association between rs895819 and PCa risk, as well as for its genetic association with the presence of distant metastases among PCa patients.

Vulvar neoplasm and reconstruction of a lesion with a vertical rectus abdominis myocutaneous flap: case report.

A 41-year-old nulliparous woman was admitted to the Institute of Gynecology and Obstetrics in 2007 with a painless tumor mass invading the whole vulvoperineal and gluteal region. Pathohistological biopsy revealed FIGO Stage II squamocellular invasive carcinoma, but clinically it was Stage IVa verucal vulvar carcinoma because of malignant infiltration to the distal part of the vagina and bilateral gluteal region. The patient underwent radical vulvectomy with bilateral inguinofemoral lymphadenectomy and partial vaginectomy. Reconstruction was performed after three weeks using a distally based vertical rectus abdominis myocutaneous flap (VRAM). A new entrance of the vagina was created. The patient was further treated with radiation therapy.

Detection of genetically modified organisms in processed meat products on the Serbian food market.

The addition of soybean proteins to processed meat products has significantly increased in recent years due to the interesting functional and nutritional properties of these vegetable proteins. Since the Roundup Ready (RR) soybean is the only transgenic soybean line approved for market in EU this work was aimed at monitoring its presence in meat products on the Serbian food market. The extracted DNA was analyzed using duplex polymerase chain reaction (PCR) with primer pairs aimed at the lectin gene and 35S promoter. Samples positive for the presence of GM soybean were subjected to a real-time quantification of the percentage of RR soya. The results indicated that out of fifty processed meat products examined, twelve gave positive results with 35S promoter and all contained RR soya below 0.1%.

Upward penetration of grains through a granular medium.

We study experimentally the creeping penetration of guest (percolating) grains through densely packed granular media in two dimensions. The evolution of the system of the guest grains during the penetration is studied by image analysis. To quantify the changes in the internal structure of the packing, we use Voronoï tessellation and a certain shape factor which is a clear indicator of the presence of different underlying substructures (domains). We first consider the impact of the effective gravitational acceleration on upward penetration of grains. It is found that the higher effective gravity increases the resistance to upward penetration and enhances structural organization in the system of the percolating grains. We also focus our attention on the dependence of the structural rearrangements of percolating grains on some parameters like polydispersity and the initial packing fraction of the host granular system. It is found that the anisotropy of penetration is larger in the monodisperse case than in the bidisperse one, for the same value of the packing fraction of the host medium. Compaction of initial host granular packing also increases anisotropy of penetration of guest grains. When a binary mixture of large and small guest grains is penetrated into the host granular medium, we observe size segregation patterns.

[The tunneled supraclavicular island flap--a fasciocutaneous flap for head and neck reconstructions].

Reconstructive procedures in the head and neck region use a wide range of flaps for defect closure. The methods range from local, mostly myocutaneous flaps and skin grafts to free microsurgical flaps. To ensure a satisfactory functional and aesthetic result, good texture and color of the flap are always essential. Moreover, the donor-site defect needs to be reduced, with no resulting functional or aesthetic impairment. We have found that the shoulder is a region providing an optimum skin texture match to the neck and face. Fasciocutaneous island flap, nourished by the supraclavicular artery is used purely as a subcutaneously tunneled island flap. The tunneling maneuver significantly improves the donor site by reducing scarring. The flap is characterized by a long subcutaneous pedicle of up to 20 cm. The pivot point is in the supraclavicular region and allows the flap to be used in the upper chest, neck, chin, and cheek. In this article we present clinical cases in reconstructive procedures with supraclavicular flap and its expanded indications.

Taxol activates inducible nitric oxide synthase in rat astrocytes: the role of MAP kinases and NF-kappaB.

Taxol is a microtubule-stabilizing agent that has recently been shown effective in the treatment of experimental autoimmune encephalomyelitis, an animal model of multiple sclerosis. As astrocytes could modulate central nervous system (CNS) autoimmunity through inducible nitric oxide synthase (iNOS)-mediated production of immunoregulatory free radical nitric oxide (NO), we investigated the effect of taxol on NO synthesis in rat astrocytes. Taxol, either alone or in combination with interferon-gamma, induced NO generation in primary astrocytes and astrocytoma C6 cells in a dose- and time-dependent manner. Accordingly, the drug markedly up-regulated the expression of both iNOS mRNA and protein in astrocytes. The observed effect of taxol was mediated through induction of iNOS transcription factors NF-kappaB and IRF-1, and required the activation of p38 MAP kinase and JNK. Finally, NO release by taxol-stimulated astrocytes was blocked with the microtubule-depolymerizing agent colchicine, suggesting the involvement of a microtubule-stabilizing activity of taxol in the observed effect.

Regulation of cyclin D1 and p16(INK4A) is critical for growth arrest during mammary involution.

A coordinated growth arrest during mammary involution completes the dramatic changes in mammary cell proliferation seen during pregnancy and lactation. Signals regulating this arrest are poorly understood, despite their potential relevance to oncogenesis. Here we report that the arrest involves a unique pulse of p16(INK4A) expression in vivo, which accompanies decreased cyclin D1 expression and a shift to an active repressor E2F4 complex. We used INK4A/ARF-/- mice as well as cyclin D1 and p16(INK4A) transgenic strains to examine the physiological significance of these patterns. p16(INK4A) directly regulated the in vivo transition from E2F3 to E2F4 as the major E2F DNA binding activity, and its contribution to growth arrest was independent of cyclin D1. Transgenic cyclin D1 expression prevented normal terminal differentiation by ablating the p16(INK4A) pulse, abolishing the shift from E2F3 to E2F4, derepressing E2F target genes, and expanding a stem cell population. The effects of cyclin D1 were reversed by restoring p16(INK4A) but were not seen in INK4A/ARF-/- mice. Our results indicate that cyclin D1 may contribute to tumorigenesis by altering cell differentiation and demonstrate a significant function for p16(INK4A) in development in vivo. These regulatory mechanisms used during mammary involution offer a potential explanation for the protective effect of pregnancy against breast cancer.

The murine GABA(B) receptor 1: cDNA cloning, tissue distribution, structure of the Gabbr1 gene, and mapping to chromosome 17.

GABA (gamma-aminobutyric acid) is a major inhibitory neurotransmitter in the central nervous system (CNS) which activates both ionotropic (GABA(A)/GABA(C)) and metabotropic (GABA(B)) receptor systems. We identified two alternatively spliced cDNA variants of the murine GABA(B) receptor 1 that are predominantly expressed in the CNS. Deduced protein structures are highly homologous to the previously characterized rat and human receptors. Comparison of the genomic structures of mouse and human revealed that alternative splicing occurred at the same position, whereas the mouse gene has an additional 5' exon. Radiation hybrid mapping, combined with database searches, indicated that the GABA(B) receptor gene (Gabbr1) is located on mouse chromosome 17, adjacent to the marker D17Mit24 in a region homologous to human chromosome 6p21.3.

Pregnancy is not associated with altered morphology of the femoral artery.

While pregnancy is associated with adjustments in cardiovascular function, the morphology of the vascular system during pregnancy has been generally viewed as being very stable. However, recently we have demonstrated that pregnancy remodels the aorta and the carotid artery. In the present study, we assessed the morphological characteristics of the guinea-pig femoral artery during different stages of pregnancy using light and electron microscopy. There were no significant differences between external and internal diameters, wall thickness, total cross-sectional area and cross-sectional areas of lumen, intima, media, and adventitia of femoral arteries from non-pregnant and early-, mid- and late-pregnant guinea-pigs (n = 8-10). In previous studies, we have shown that the morphology of vascular smooth muscle and endothelial cells in the aorta and the carotid artery may be altered by pregnancy. Therefore, to test this possibility we measured diameters as well as cross-sectional areas of femoral arterial muscle and endothelial cells using electron microscopy. These parameters, at the electron microscopy level, were also not significantly changed by pregnancy (n = 8-10). It is concluded that the morphology of the guinea-pig femoral artery is not altered during pregnancy. In this regard, this study demonstrated that pregnancy-induced vascular remodelling varies between blood vessels that undergo the same functional alterations. Therefore, this may suggest that pregnancy-induced changes in blood flow through different vascular beds are not the most important factor involved in vascular remodelling observed during pregnancy. Rather, it is possible that haemodynamic-independent factors regulate pregnancy-mediated structural changes of the vascular wall.

Effect on neonatal oestrogen and hCG treatment on the genital organs of adult female rats.

The ovaries and the uterine as well as vaginal mucous membranes of 80-, 180- and 365-day-old intact female rats and females neonatally treated with a single dose of oestrogen and repeated doses of human chorionic gonadotropin (hCG) were studied. Numerous follicles, interstitial cells and corpora lutea (CL) were present on the ovary of intact females from 80 up to 365 days of age. The number of primary and secondary follicles decreased in the intact female rats between Day 180 and Day 365 of life. On the ovaries of 180- and 365-day-old female rats neonatally treated with oestrogen, interstitial glandular cells and cystic follicles predominated. No CL were present on these ovaries. The height of epithelial cells of the uterine and vaginal mucous membranes increased in intact female rats from 80 to 365 days of age, whereas in oestrogen-treated females the height of epithelium decreased. From 80 up to 365 days of age, the height of epithelial cells of uterine and vaginal mucous membranes of rats neonatally treated with repeated doses of hCG was similar to that in the corresponding control animals.

Predicting quadriceps muscle activity during gait with an automatic rule determination method.

It has been suggested that control using a skill-based expert system can be applicable to gait restoration. Rule-based systems have several advantages for this application: they generate a fast response (they are not computationally intensive) and they are easy to comprehend and implement. A major problem with using such systems is the inability of users to determine its rules. In this study, an automatic method for obtaining the production rules from a set of examples is described. The rule base was automatically induced from a model which used external sensor signals as inputs and electromyogram (EMG) patterns as outputs. The method is based on the minimization of entropy. A production rule estimated the muscle activity pattern using the sensor information. The algorithm was tested using data recorded from six able-bodied individuals during ground level walking, with and without ankle-foot orthoses. The data showed that gait variability will increase in able-bodied subjects when the motion of ankle joints is restricted, thus, providing a good test for generalization. The experimental results illustrate performance of the production rule that estimates quadriceps muscle group activity pattern for ground level walking in able-bodied subjects.

The human glycine receptor subunit alpha3. Glra3 gene structure, chromosomal localization, and functional characterization of alternative transcripts.

The neuronal glycine receptor is a ligand-gated chloride channel composed of ligand binding alpha and structural beta polypeptides. Homology screening of a human fetal brain cDNA library resulted in the identification of two alternative splice variants of the glycine receptor alpha3 subunit. The amino acid sequence predicted for the alpha3L variant was largely identical to the corresponding rat subunit. In contrast, the novel splice variant alpha3K lacked the coding sequence for 15 amino acids located within the cytoplasmic loop connecting transmembrane spanning region 3 (TM3) and TM4. Using P1 artificial chromosome (PAC) clones, the structure of the GLRA3 gene was elucidated and its locus assigned to human chromosomal bands 4q33-q34 by fluorescence in situ hybridization. Two transcripts of 2.4 and 9 kilobases, corresponding to alpha3L and alpha3K, respectively, were identified and found to be widely distributed throughout the human central nervous system. Structural analysis of the GLRA3 gene revealed that the alpha3K transcript resulted from a complex splice event where excision of the novel exon 8A comprising the alternative sequence of 45 base pairs coincides with the persistence of a large intronic sequence in the 3'-untranslated region. Functional expression in HEK 293 cells of alpha3L and alpha3K subunits resulted in the formation of glycine-gated chloride channels that differed significantly in desensitization behavior, thus defining the cytoplasmic loop as an important determinant of channel inactivation kinetics.

Quality of life as defined by orthopedic oncology patients.

BACKGROUND: Quality of life (QOL), not just survival, is central to outcomes analysis in musculoskeletal oncology. However, little information exists about the patients' definition of what constitutes QOL. METHODS: Self-administered outcomes questionnaires were given to 201 surgically treated patients with lower extremity tumors. Of these patients, 192 (137 with malignant tumors, 55 with benign tumors) provided a written definition of QOL. Their responses were independently collated and matched with clinical information. RESULTS: For most patients (153, or 80%) the definition of QOL encompassed several attributes. A consistent combination of four major attributes was used in the QOL definition by 44 (32%) of the malignant cases and 19 (35%) of the benign cases. Differences in responses between men and women were idiosyncratic and more common in the benign group. Good family relations and good health were equally important to men and women. Responses varied by patient age. Older patients valued self-sufficiency and freedom from pain, whereas younger patients emphasized happiness, trust in God or church, achieving goals and being successful, and love. Those whose surgery was less extensive cited good family relations, the ability to function physically and emotionally, and having a good job or work. CONCLUSION: The variation in patients' perspectives and definitions of quality of life must be taken into account when assessing QOL in musculoskeletal oncology patients. Patients often emphasize concerns that are not adequately addressed by current outcomes-measures in orthopedics and general oncology.

Kinematic analysis of limb position during quadrupedal locomotion in rats.

A test of locomotor behavior using the coordinates of ipsilateral limb positions of rats walking on a moving treadmill is described. Specific points on the forelimb and hindlimb were digitized from video records for 20-sec continuous sequences of locomotion, and step periods and step distances were calculated. The extent to which a given limb position would predict its own position--or the position of another limb--at different points in time was mathematically determined by autocorrelation and cross-correlation, respectively. Autocorrelation of position data was performed using a three-step window and the standard formula for correlating phasic data. A novel method of data preparation, which included normalization of the step data to eliminate variability introduced by differences in step period length, was used prior to cross-correlations of forelimb to hindlimb positions. Rats walking at 0.10, 0.15, and 0.25 m/sec had high limb autocorrelations, comparable forelimb/hindlimb phase relationships, and consistently high average cross-correlation coefficients. This analysis has resulted in the quantification of rat locomotor behavior in terms of the degree of limb movement rhythmicity and the strength of the forelimb/hindlimb coordination, and has provided baseline data for comparisons with spinal cord-injured rats that have retained or recovered alternating hindpaw movements.

Hanhart's syndrome with facial asymmetry: case report.

An unusual variant of Hanhart's syndrome associated with facial asymmetry in a female patient is presented together with surgical treatment of her facial anomalies. Following reconstruction which included bimaxillary osteotomies and osteopericranial and pericranial flaps, a good aesthetic result was achieved which enabled the psychological and social rehabilitation of the patient.

Longitudinal growth of leg and wing bones of chickens reared in cages and on the floor.

The histomorphometry of proximal growth plate and rate of longitudinal growth was estimated in tibia and humerus of chickens reared in the cages and on the floor, during 0, 2, 4, 6 and 8 weeks of age. The floor vs. the cage area was 3:1. All chicks were fed ad libitum standard broilers mixture: starter (0-4 weeks) with 0.9% Ca; 0.7% P and grower (4-8 weeks) with 0.8% Ca; 0.5% P. The rearing system showed no significant effect on macroscopy estimated tibia and humerus growth in length, just as in case of bone's proximal growth plate microscopy of floor reared chickens. However, our results suggested that wing bones are more affected by the housing system than that of leg bones. The proliferative zone, hypertrophic zone and total growth plate of the humerus in cage reared birds were significantly higher than in those reared on the floor.