Tumor necrosis factor Alpha -308 G/A single nucleotide polymorphism and risk of sperm abnormalities in Iranian males

Background: Signaling molecules such as cytokines regulate spermatogenesis during the maturation of germ cells and sperm apoptosis. Tumor necrosis factor alpha [TNF alpha] is one of the most-documented cytokines that is involved in spermatogenesis. We investigated the association of the TNF alpha -308 G/A single nucleotide polymorphism with sperm abnormalities in Iranian males

Materials and Methods: This case-control study included 180 infertile men who referred to Yazd Research and Clinical Center for Infertility and 100 healthy normospermic controls. Infertile men were classified into four groups of azoospermia [n=91], oligospermia [n=26], teratospermia [n=30] and asthenoteratospermia [n=33]. After sperm analysis, DNA was extracted from blood and polymerase chain reaction-restriction fragment length polymorphism [PCR-RFLP] was carried out for the genotyping of TNF alpha-308 G/A

Results: The A allele was significantly associated with sperm abnormality in our population [[P<0.001, odds ratios [OR] 95% confidence interval [CI]=2.31]. In addition, the A allele was also associated with azoospermia [P<0.001, OR [95% CI]=2.484], oligospermia [P=0.005, OR [95% CI]=2.51] and teratospemia [P<0.001, OR [95% CI]=3.385] but not with asthenoteratospermia [P=0.623]

Conclusion: Our data suggest that this single nucleotide polymorphism [SNP] maybe associated with the risk of sperm abnormality in infertile men of Iranian origin

novel approach on drug delivery: investigation of a new nano-formulation of liposomal doxorubicin and biological evaluation of entrapped doxorubicin on various osteosarcoma cell lines

Objective: In this study we prepared a novel formulation of liposomal doxorubicin [L-DOX]. The drug dose was optimized by analyses of cellular uptake and cell viability of osteosarcoma [OS] cell lines upon exposure to nanoliposomes that contained varying DOX concentrations. We intended to reduce the cytotoxicity of DOX and improve characteristics of the nanosystems

Materials and Methods: In this experimental study, we prepared liposomes by the pH gradient hydration method. Various characterization tests that included dynamic light scattering [DLS], cryogenic transmission electron microscopy [Cryo-TEM] imaging, and UV-Vis spectrophotometry were employed to evaluate the quality of the nanocarriers. In addition, the CyQUANT[registered] assay and fluorescence microscope imaging were used on various OS cell lines [MG-63, U2-OS, SaOS-2, SaOS-LM7] and Human primary osteoblasts cells, as novel methods to determine cell viability and in vitro transfection efficacy

Results: We observed an entrapment efficiency of 84% for DOX within the optimized liposomal formulation [L-DOX] that had a liposomal diameter of 96 nm. Less than 37% of DOX released after 48 hours and L-DOX could be stored stably for 14 days. L-DOX increased DOX toxicity by 1.8-4.6 times for the OS cell lines and only 1.3 times for Human primary osteoblasts cells compared to free DOX, which confirmed a higher sensitivity of the OS cell lines versus Human primary osteoblasts cells for L-DOX. We deduced that L-DOX passed more freely through the cell membrane compared to free DOX

Conclusion: We successfully synthesized a stealth L-DOX that contained natural phospholipid by the pH gradient method, which could encapsulate DOX with 84% efficiency. The resulting nanoparticles were round, with a suitable particle size, and stable for 14 days. These nanoparticles allowed for adequately controlled DOX release, increased cell permeability compared to free DOX, and increased tumor cell death. L-DOX provided a novel, more effective therapy for OS treatment

Mutation analysis of exon1 of bone morphogenetic protein-15 gene in Iranian patients with polycystic ovarian syndrome

Background: with the prevalence of 6-10%, polycystic ovarian syndrome [PCOS] is considered the most common endocrinological disorder affecting women in their reproductive age. It has been suggested that genetic factors participate in the development of PCOS. Follicular development has been considered as one of the impaired processes in PCOS. Bone morphogenetic protein-15 [BMP-15] gene is a candidate gene in follicular development and its variants may play role in pathogenesis of PCOS

Objective: to investigate whether BMP-15 gene mutations are present in Iranian women with PCOS

Materials and Methods: in this cross-sectional study 5 ml venous blood samples was taken from 70 PCOS women referring to Afzalipour Hospital, Kerman University of Medical Sciences, Kerman, Iran, between January to December 2014. Genomic DNA was extracted from the blood sample by salting out method. Then a set of PCR reactions for exon1 of BMP-15 gene was performed using specific primers followed by genotyping with direct sequencing

Results: two different polymorphisms were found in the gene under study. In total 20 patients [28.6%] were heterozygote [C/G], and 2 patients [2.86%] were homozygous [G/G] for c.-9C>G in 5´UTR promoter region of BMP-15 gene [rs3810682]. In addition, in the coding region of exon1, three patients [4.3%] were heterozygote [G/A] for c.A308G [rs41308602]. Two PCOS patients [2.86%] appeared to have both c.-9C>G [C/G] and c.A308G [G/A] variants simultaneously

Conclusion: our research detected two polymorphisms of BMP-15 gene among PCOS patients, indicating that even though it cannot be concluded that variants of BMP-15 gene are the principal cause of polycystic ovarian syndrome; they could be involved in pathogenic process in development of PCOS

SULF 1 gene polymorphism, rs6990375 is in significant association with fetus failure in IVF technique

Sulfatase 1 [SULF1] function is to remove the 6-O-sulphate group from heparan sulfate. This action changes the binding sites of extracellular growth factors. SULF1 expression has been reported to be changed in angiogenesis. We hypothesized that single nucleotide polymorphisms [SNPs] of SULF1 would impact clinicopathologic characteristics. Study of SULF1 gene polymorphism with fetus failure in in vitro fertilization [IVF] technique. We studied one common [minor allele frequency >0.05] regulatory SNP, rs6990375, with polymerase chain reaction and restriction fragment length polymorphism method, in 53 infertile women with fetus failure in IVF technique and 53 women with at least one healthy child as controls. We found that rs6990375 is significantly associated with an early failure in IVF and frequency of G allele is high in women with fetus failure in IVF technique [p<0.001]. These findings suggest that SULF1genetic variations may play a role in IVF technique fetus failure. Further studies with large sample sizes on SULF1 SNPs may be useful in support of this claim

Mycobacterium tuberculosis infection in women with unexplained infertility

Background: Genital tuberculosis [GTB] is an important cause of female infertility, especially in developing countries. The positive results of polymerase chain reaction [PCR] in endometrial GTB in the absence of tubal damage raise the possibility of the detection of sub-clinical or latent disease, with doubtful benefits of treatment

Objective: To evaluate the mycobacterium tuberculosis infection in endometrial biopsy samples collected from unexplained infertile women attending Yazd Research and Clinical Center for Infertility by using PCR techniques

Materials and Methods: In this cross sectional study, 144 infertile women with unexplained infertility aged 20-35 years old and normal 1-listro-saplango graphy findinus were enrolled. Endometrial biopsy samples from each participant were tested fbr mycobacterium tuberculosis detecting by PCR. In 93 patients, peritoneal fluid was also taken for culture and PCR

Results: The PCR results of endometrial specimens were negative in all cases, demonstrating that there was no GTB infection among our patients

Conclusion: Our results showed that GTB could not he considered as a major problem in women with unexplained infertility. Although, studies have indicated that PCR is a useful method in diagnosing early GTB disease in infertile women with no demonstrable evidence of tubal or endometrial involvement

Detection of Human papillomavirus 16 and 18 types in oral squamous cell carcinoma patients in Yazd, Iran :a case-control study

The aim of this study was to ascertain the prevalence of human papillomavirus [HPV] 16/18 in oral squamous cell carcinoma [OSCC] and normal oral mucosa in Iranian individuals with OSCC and their association with various risk factors. The investigation followed a hospital-based case-control design. Cases were individuals with diagnosed OSCC, age- and sex-matched with controls. Sixty-six paraffin embedded tissue samples [39 OSCC and 27normal oral mucosa] were studied. DNA was purified to amplify HPV-DNA using HPV type specific primers-mediated polymerase chain reaction [PCR] method. Mean age was 64.2 +/-14 years in cases [range 25-81 years], and 63.5 +/-13.5 years in controls [range 22-78 years]. Forty-three percent [17/39] of OSCC but none of normal oral mucosa were positive for HPV 16/18 DNA [p = 0.001]. The most frequent type in patient group was HPV-16 type. The prevalence rate was high older in age [>50 years] and gender [male] and tongue tissue but no significant association was found [p=0.7, p=0.5 and p=0.42 respectively]. We found a significant association of HPV16/18 with oral squamous cell carcinoma. Also, HPV16/18 is a co-factor in oral carcinogenesis. In addition, also it is suggested that HPV is as a risk factor independent of alcohol and tobacco can be effective in creating OSCC

investigation on mitochondrial DNA deletions and telomere shortening during multiple passages of adult stem cells

Limited resources for adult stem cells necessitate their in vitro culture prior to Clinical use. Investigating mitochondrial DNA [mtDNA] and telomere shortening has proved to be important indications of stem cell validity. This study was designed to investigate these indicators in multiple passages of three adult stem cell lines which were produced in our stem cell laboratory. In this study, Dental Pulp Stem Cells [DPSCs], Periapical Follicle Stem Cells [PAFSCs] and Human Foreskin Fibroblast [HFF] cell lines were expanded for 20 passages. After 1, 5, 10, 15 and 20 passages, expanded cells were harvested and DNA was extracted for further studies. Common mtDNA mutation was detected by multiplex PCR and telomere shortening was tested by Southern blot analysis. The common deletion was not detected in any of the stem cells or cell lines after several passages. In addition, Southern blot analysis indicated that the mean difference of telomere length between first and last passage was 0.25 kb in DPSC, 0.1 kb in PAFSC and 0.32 kb in HFF which indicates that the mean telomere length in various passages of the samples showed insignificant changes. Absence of mtDNA mutations in adult stem cell lines indicates good mitochondrial function even after 20 passages. In addition, absence of telomere shortening indicates stem cells validity after multiple passages. It is hoped this information could pave the way for using in vitro expansion of adult stem cells for future Clinical applications

luteinizing hormone beta-subunit exon 3 [Gly102Ser] gene mutation and ovarian responses to controlled ovarian hyperstimulation

Despite extensive progress in IVF techniques, one of the most difficult problems is the variability in the response to controlled ovarian hyperstimulation [COH]. Recent studies show the effects of individual genetic variability on COH outcome. To evaluate the correlation between LHbeta G1502A polymorphisms in exon 3 of the LH gene and ovarian response to COH. A total of 220 women treated with a long protocol for ovarian stimulation were studied. Three genotypes of GG, GA and AA were detected by polymerase chain reaction-restriction fragment length polymorphism [PCR-RFLP] analysis. In total, 34 [17%] patients were poor responders, 154 [77%] were normal responders and 12 [6%] were hyper responders. The most frequent genotype was GA [55.5%] whereas 44.5% of patients showed GG genotype and there was no patient with AA genotype. In total 54.5% of normal responders, 61.8% of poor responders and 50% of hyper responders showed GA genotype. Our results did not establish a significant relationship between this polymorphism and the ovarian response. Therefore it is still very difficult to use the genotype of patients for prediction of the ovarian response to stimulation

frequency of Yq microdeletion in azoospermic and oligospermic Iranian infertile men

About 15% of couples have infertility problems which 40% of them are related to the male factors. Genetic factors are candidate for about 10% of male infertility conditions. Among these, AZFa, AZFb, AZFc and AZFd regions on the Yq are considered most important for spermatogenesis. Microdeletions of these regions are thought to be involved in some cases of azoospermic or oligospermic infertile men. We studied the prevalence of AZF microdeletions among Iranian infertile men with non-obstructive azoospermia and oligospermia. A total of 50 Iranian azoospermic and oligospermic infertile men were selected for case group and 50 men with normal spermogram as control group. The molecular study of Y chromosome microdeletions was done by multiplex polymerase chain reaction [M-PCR] method by using of 13 sequence tagged site [STS] markers from AZF region. Four [8%] patients showed Y chromosome microdeletions among case group, deletion in AZFc region was the most frequent [80%] followed by AZFb [20%], in AZFa and AZFd region we did not detect any deletions. No deletion was detected in control group; the ratio of Y chromosome microdeletion in azoospermic men was higher than this ratio in oligospermic men [19% [3/16] among azoospermic men and 3% [1/34] among oligospermics]. Serum FSH level in men with microdeletions was higher than this level in men with no deletions [p=0.034]. Because of relatively high prevalence of microdeletions on the long arm of Y chromosome among Iranian azoospermic and oligospermic patients, screening of this microdeletion may be advised to infertile men particularly azoospermic and oligospermic men before using assisted reproductive treatments

Characterization of mesenchymal stem cells from human dental pulp, preapical follicle and periodontal ligament

Human dental stem cells have high proliferative potential for self-renewal that is important to the regenerative capacity of the tissue. The aim was to isolate human dental pulp stem cells [DPSC], periodontal ligament stem cells [PDLSC] and periapical follicle stem cells [PAFSC] for their potential role in tissue regeneration. In this experimental study, the postnatal stem cells were isolated from dental pulp, preapical follicle and periodontal ligament .The cells were stained for different stem cell markers by immunocytochemistry. To investigate the mesenchymal nature of cells, differentiation potential along osteoblastic and adipogenic lineages and gene expression profile were performed. For proliferation potential assay, Brdu staining and growth curve tests were performed. Finally, all three cell types were compared together regarding their proliferation, differentiation and displaying phenotype. The isolated cell populations have similar fibroblastic like morphology and expressed all examined cell surface molecule markers. These cells were capable of differentiating into osteocyte with different capability and adipocyte with the same rate. PAFSCs showed more significant proliferation rate than others. Reverse transcriptase PCR [RT-PCR] for nanog, oct4, Alkaline phosphatase [ALP] and glyceraldehydes-3-phosphate dehydrogenease [GADPH] as control gene showed strong positive expression of these genes in all three isolated cell types. PDLSCs, DPSCs and PAFSCs exist in various tissues of the teeth and can use as a source of mesenchymal stem cells for developing bioengineered organs and also in craniomaxillofacial reconstruction with varying efficiency in differentiation and proliferation

Multiplex PCR screening of Y-chromosome microdeletions in azoospermic ICSI candidate men

It has been hypothesized that Y-q microdeletion can account for significant proportion of infertility in men. There are three nonoverlapping regions referred to as the "azoozpermia factors" AZFa, AZFb, and AZFc from proximal to distal part of Y-q. These have been defined as spermatogenesis loci, this region deletions have been shown to be involved in male azoospermic or severe oligoozospermic infertility. Evaluation the rate of Y-chromosome microdeletions in infertile men. In this case-control study, 25 azoospermic infertile men candidate for intracytoplasmic sperm injection [ICSI] were selected as case group. For control group, 25 normoozoospemric men were selected. All cases and controls had normal 46XY karyotype. DNA extraction and molecular analysis were done on blood samples. Multiplex-PCR method was done to identify the presence of microdeletion in AZFa, AZFb or AZFc loci. Eight STS primers that include two controls were selected to determine Y-chromosome microdeletions. 20% [5/25] of all patients have at least one microdeletion in more than one region of AZF loci. Totally 17 microdeletions was observed, one case had deletions in three AZF regions, and 4 cases had deletions in two AZF regions. The rate of deletions was 42% [7/17] for AZFc, 35% [6/17] for AZFa and 23% [4/17] for AZFb. The molecular DNA analysis could help us to know the real cause of infertility and can give good information for good decision for example in men whit microdeletions who want to undertake ICSI procedure the deletions will be passed to their son

No association of GSTM1 null polymorphism with endometriosis in women from central and southern Iran

Endometriosis is one of the most common gynecologic disorders. It is a complex trait and both genetic and environmental factors have been implicated in its pathogenesis. There is growing evidence indicating that exposure to environmental contaminants is a risk factor for endometriosis. Glutathione-S-Transferase M1 [GSTM1] is one of the genes involved in detoxification of endogenous and exogenous compounds. Several studies have indicated an association between GSTM1 null mutation and endometriosis. In this study, the possible association between the GSTM1 gene null genotype and susceptibility to endometriosis in woman from central and southern Iran was investigated. One hundred and one unrelated premenopausal women with endometriosis and 142 unrelated healthy premenopausal women without endometriosis were enrolled in the study. Genomic DNA was extracted from Peripheral blood in all subjects. GSTM1 null genotyping was performed by polymerase chain reaction [PCR]. There was no significant difference between frequencies of GSTM1 null genotype in case and control groups [50.5% Vs. 52.1%, p=0.804]. Furthermore, this genotype was not associated with severity of endometriosis in our sample [p=0.77]. Further studies involving gene-environment and gene-gene interactions, particularly combination of GSTM1 and other GST gene family polymorphisms are needed

Public knowledge and perceptions of biotechnology and genetically modified organisms in Iran

Biotechnology offers a variety of potential environmental, social and economic benefits but, it is the center of extreme public and political debate at present. A major field of discussion in biotechnology are genetically modified [GM] organisms. New medical, political, ethical and religious discussions arise over the production and consumption of these organisms. Many surveys have been conducted in industrialized countries to investigate the public perceptions regarding the risks and benefits of biotechnology, while in developing countries hardly any studies have been done so far. The present descriptive study intends to contribute to a better understanding of public attitudes toward biotechnology and GM foods in Iran. To assess the public's knowledge and opinions on the dangers or opportunities of genetic modification, 300 university students as a sample of educated community and 300 individuals with no university's education as a sample of ordinary people were asked to complete a specially designed questionnaire on the risks and benefits of biotechnology and GM foods. This article summarizes the current situation with regard to biotechnology, with a particular focus on GM foods and discusses the results of the survey. Our results indicated that public's knowledge about biotechnology is low in Iran and more efforts are needed to improve their understanding of different aspects of biotechnology

Cytogenetic analysis in couples with recurrent spontaneous abortion

Recurrent abortion is a difficult medical problem happening in about 1-2% of fertile women. Most spontaneous miscarriages which happen in the first and second trimesters are caused by chromosomal abnormalities. The present study tries to find the rate of chromosomal abnormalities in couples with recurrent pregnancy loss. In total 165 couples were referred to genetic counselling clinic with a history of at least three previous abortions. In all women antibodies against toxsoplasmose, rubella and cytomegalovirus [CMV] were analysed by ELIZA. In 88 couples karyotyping was conducted by analysis of G and/or C banding. Metaphase spreads were made from phytohaemaglutinin-stimulated peripheral lymphocytes using standard cytogenetic techniques. The chromosomal status was analyzed using CytoVision Ultra ver.4.0 from Applied Imaging. The CHI[2]-test and ANOVA were used for statistical evaluation. The level of p<0.05 was considered as significance. Most of the patients had 3 repeated abortions [61.2%]. Cytogenetic analysis performed for 88 couples and karyotypes of 12.5% of them were abnormal. The majority of them had monosomy X [6.82%], followed by balanced translocation [2.27%]. The number of female carries chromosomal abnormality exceeded significantly than of male. Coefficient of inbreeding in more than 50% of couples had fifth degree of relationship [89 out of 165]. Our results showed that 12.5% of the couples with missed abortion had an abnormal karyotype, with no other abnormality. Cytogenetic findings in spontaneous aborted specimens could provide valuable information for genetic counseling and prenatal care in future pregnancies in couples with a history of repeated pregnancy loss

Knowledge and attitude about HIV/AIDS among pregnant women in Yazd, Iran

Since AIDS is not only a vital medical problem, but also a socioeconomic complication, therefore increasing people's knowledge and replacing their unhealthy behavior by a healthy one is of important consideration. Women, specially in reproductive ages of their life, have a special situation regarding the probability of pregnancy and infection of their fetus. The aim of this study was to explore pregnant women's knowledge about HIV/AIDS, their perception of risk, risk behavior and management, and their attitudes towards AIDS. In a cross sectional study, a total of 120 pregnant women, who referred to family health clinics in Yazd, were selected by simple random sampling. Information was collected via a special designed questionnaire containing 22 questions for evaluating knowledge and 6 questions for evaluating attitude. Statistical analysis was performed using Chi-square test with SPSS software. The knowledge of pregnant women about AIDS was not significantly different in different age groups [p=0.151], while it had a significant relationship with their education [p=0.000]. There was a correlation between general knowledge and attitude in pregnant women [p=0.033] [r=0.126]. The attitude about AIDS in pregnant women was fairly good and there was a significant difference in this regard related to their education [p=0.000], while there was not significant difference regarding their age [p=0.410] [Mean =19.8]. There is an urgent need for HIV prevention efforts, such as health education and focusing on the pregnant women in developing world. Health educators should tailor education programs for women at risk, particularly those with lower education, to enhance their knowledge about HIV and to improve their attitude about AIDS

Glutathione S-transferases null genotype in acute myeloid leukemia

The glutathione S-transferase [GST] family of metabolising enzymes plays an important role in the detoxification of mutagens and carcinogens. The expression of many of these cancer susceptibility enzymes is genetically polymorphic. An increased frequency of GST-null genotypes has been associated with several malignancies. To investigate the rate of GSTT1 and GSTM1 null genotypes in AML patients and to determine its importance in prognosis of the disease. DNA was extracted by phenol/chloroform method from peripheral blood or bone marrow of 180 white Caucasian patients. A multiplex PCR method was used simultaneously to amplify regions of GSTM1, GSTT1, and beta-globin genes in genomic DNA. The survival curves were analyzed by the Kaplan-Meier method and compared by the log-rank test [Mantel-Cox] using the SPSS software program. Of the total of 180 patients, 23 cases [12.8%] showed null genotypes in both genes, while in 52 patients [28.9%] both genes were wild-types. GSTM1 null-GSTT1 wild-type was detected in 91 patients [50.6%] and GSTM1 wild-type-GSTT1 null genotype was detected in 14 patients [7.8%]. These rates are within the upper limit of the rates detected in the normal European population. There was no significant difference in the overall survival and in disease free survival between different groups. These observations suggest that the inherited absence of the GSTT1 and GSTM1 carcinogen detoxification pathway may be related to carcinogenesis but it is not an important determinant of prognosis in AML