MAML1 regulates EMT markers expression through NOTCH-independent pathway in breast cancer cell line MCF7.

Tumor relapse is the main cause of breast cancer related deaths and metastasis due to epithelial-mesenchymal transition (EMT) having a critical role in this process. MAML1 is the main co activator of NOTCH signaling pathway and its role in EMT remains unknown. In this study, this role was evaluated through overexpression and knockdown study of MAML1 in MCF7 and MDA-MB-231 cells. MAML1 overexpression up regulated the epithelial and down regulated the mesenchymal markers. In addition, MAML1 silencing decreased epithelial and increased mesenchymal markers. Notch inhibition using γ-secretase inhibitor resulted in increased E-cadherin expression. MAML1 ectopic expression, further increased E-cadherin expression with inhibition of NOTCH signaling. Wound healing assay showed that MAML1 overexpression decreases the rate of migration, while MAML1 silencing increases this rate significantly. In conclusion, our data indicated that MAML1 negatively regulates EMT markers expression in breast cancer cells.

Comparison of two different media for maturation rate of neural progenitor cells to neuronal and glial cells emphasizing on expression of neurotrophins and their respective receptors.

Neural cells derived from embryonic stem cells (ESCs) have potential usefulness for the treatment of neurodegenerative disorders. Modulation of intrinsic growth factors expression such as neurotrophins and their respective receptors by these cells is necessary to obtain functional neural cells for transplantation. In present study, we compared neural differentiation potential of two different media, NB + 5%ES-FBS + N2B27 and Ko-DMEM + 5%ES-FBS for conversion of mESC derived neural progenitors (NPs) into mature neural cells with emphasis on effect of the these two media on neurotrophins and their respective receptors expression. Immunofluorescence staining, RT-qPCR and western blot analysis showed that the expression of neuronal specific markers, MAP2 and Tuj-1, in NB + 5%ES-FBS + N2B27 medium was significantly higher than the other medium. Western blot assay revealed that the expression of BDNF and NGF increased significantly in mature neural cells obtained from NB + 5%ES-FBS + N2B27 medium but decreased in neural cells from Ko-DMEM + 5%ES-FBS medium compared to mESCs. TrkB protein was not detectable in mESCs but its expression increased in neural cells obtained from both media although its expression in NB + 5%ES-FBS + N2B27 medium was significantly higher than the other medium. In contrast to TrkB, p75NTR protein was detectable in mESCs and is remained constant in neural cells cultured in NB + 5%ES-FBS + N2B27 medium but decreased significantly in the other medium. In conclusion, our results indicated that NB + 5%ES-FBS + N2B27 medium promoted neural differentiation process of mESCs and caused enhancement of neurotrophins protein expression in addition to their cognate receptors.

Assessment of Promoter Hypermethylation and Expression Profile of P14ARF and MDM2 Genes in Patients With Pterygium.

BACKGROUND: Pterygium is a fairly general condition in many regions of the world. The cause of this abnormality is still ambiguous. However, recent findings suggest that pterygium is a benign progressive tissue and not a degenerative disorder. The main goal of our study was to investigate the effects of P14 and MDM2 promoter methylation on the risk of pterygium. MATERIALS AND METHODS: In this study, the DNA of 81 primary pterygium and 75 normal conjunctiva tissues was extracted and modified for the assessment of methylation of P14 and MDM2 promoters by methylation-specific polymerase chain reaction (MSP). We also estimated the mRNA expression levels of these genes in 23 pterygium and 18 normal conjunctiva tissue samples using real-time quantitative reverse transcriptase polymerase chain reaction. RESULTS: The frequency of methylation for P14 was 92.6% for cases and 97.3% for controls. MDM2 gene methylation at the promoter site was 39.5% and 72.0% for pterygium and normal conjunctiva tissues, respectively. So statistically, a significant relationship between MDM2 gene promoter methylation and the risk of disease was found (odds ratio=5.3; 95% confidence limit, 2.6-10.8; P<0.0001). In addition, the expression of MDM2 gene has increased in pterygium (1.371548±0.6727) in comparison with conjunctiva tissues as control (1.20621±1.0) (P<0.05), but it was not significant for P14 gene. CONCLUSION: Our results have indicated that hypomethyaltion and overexpression of MDM2 gene take place in patients with the pterygium. To confirm the presented data, suggesting further studies with a larger sample size in various genetic populations.

Relationship between phosphoinositide-3-kinase genetic polymorphism and schizophrenia.

UNLABELLED: Schizophrenia, with incidence of 1% worldwide, is a common mental disorder. Phosphoinositide-3-kinases (PI3Ks) are a family of enzymes involved in cellular functions such as cell growth, proliferation, differentiation, motility, intracellular trafficking, and survival. These enzymes play an important role in the PI3K/AKT signalling pathway. The PIK3CA gene encodes the alpha catalytic subunit of the PI3K enzyme. The present study analysed the role of three SNPs of the PIK3CA gene (rs6443624 (A/C), rs7640662(C/G) and rs7621329(C/T)) in the development of schizophrenia. METHODS: In this case-controlled study, DNA was extracted from blood samples from 108 patients with schizophrenia and 108 healthy patients as controls. Genotypic analyses of PIK3CA SNPs rs6443624 (A/C), rs7640662(C/G) and rs7621329(C/T) were made using the tetra primer ARMS-PCR technique. RESULTS: The outcome shows significant difference between CT and the combined genotype (CT + TT) of rs7621329 and the risk of schizophrenia (OR = 6.4, 95% CI = 3.023-14.23, p < 0.0001). Outcome showed no significant difference for were for analyses of the rs6443624 and rs7640662 genotypes. CONCLUSIONS: These results indicate an association between PIK3CA gene polymorphism on the rs7621329(C/T) site and the risk of schizophrenia. Further study of the genetic population using a larger sample size is necessary in order to validate these present findings.

Promoter Methylation and BDNF and DAT1 Gene Expression Profiles in Patients with Drug Addiction.

BACKGROUND: Drug addiction is a brain disorder that has negative consequences for individuals and society. Addictions are chronic relapsing diseases of the brain that are caused by direct drug-induced effects and persevering neuroadaptations at the epigenetic, neuropeptide and neurotransmitter levels. Because the dopaminergic system has a significant role in drug abuse, the purpose of this study was to analyze the methylation and expression profile of brain-derived neurotrophic factor (BDNF) and dopamine transporter (DAT1) genes in individuals with drug addiction. MATERIALS AND METHODS: BDNF and DAT1 promoter methylation were investigated with a methylation-specific polymerase chain reaction (PCR) technique in blood samples from 75 individuals with drug addiction and 65 healthy controls. The expression levels of BDNF and DAT1 were assessed in 12 mRNA samples from the blood of patients and compared to the samples of healthy controls (n = 12) with real-time quantitative reverse transcription PCR. RESULTS: No significant differences were found in the methylation of BDNF and DAT1 between patients and controls, but the relative levels of expression of BDNF and DAT1 mRNA differed significantly in the patients compared to controls (p < 0.0001). CONCLUSION: These results showed that the methylation status of the BDNF and DAT1 genes had no significant function in the processes of drug addiction.

Association between promoter methylation and expression of thyroid hormone receptor beta (THRß) gene in patients with gastric cancer in an Iranian population.

BACKGROUND AND AIM: There is evidence that gastric cancer patients suffer from thyroid disorders. However, the relationship between thyroid receptor (TR) expression and gastric cancer remains unknown. The aim of this study was to evaluate the status of promoter methylation and expression of the thyroid hormone receptor beta (THRß) gene in gastric cancer patients in an Iranian population. METHODS: Analysis of THRß promoter methylation was performed on 85 pairs of formalin-fixed, paraffin-embedded (FFPE) tissue samples as cases and controls via methylation-specific polymerase chain reaction (PCR [MSP]). The samples were obtained from tumors and surrounding healthy tissues from resected gastric cancers. The expression assay was also performed with 25 FFPE tissue pairs (tumor and surrounding healthy tissues of the same individual) using real-time PCR. RESULTS: The results of the present study show that there is a statistically significant difference between tumor and adjacent normal tissues regarding promoter methylation status and THRß expression (P = 0.04 and P = 0.036, respectively). CONCLUSION: Therefore, promoter methylation of THRß may be involved in the development of gastric cancer.

Association of glutathione s-transferase gene methylation with risk of schizophrenia in an Iranian population.

BACKGROUND/AIM: It has been believed that epigenetic changes play a critical role in schizophrenia through improper interaction between genome and environmental risk factors. The aim of this case-control study was to investigate the association of the promoter hypermethylation status of glutathione S-transferase T1 (GSTT1) and glutathione S-transferase P1 (GSTP1) genes with the risk of schizophrenia. MATERIALS AND METHODS: Methylation-specific PCR was used to estimate DNA methylation in the blood of 80 patients with schizophrenia and 71 healthy controls. RESULTS: Promoter hypermethylation analysis of GSTT and GSTP indicated a significant difference between individuals with methylated and unmethylated status [odds ratio (OR) = 0.339, 95% confidence interval (95% CI) = 0.14-0.8, p = 0.012 and OR = 0.308, 95% CI = 0.135-0.7, p = 0.005, respectively]. CONCLUSION: The present study supports the hypothesis that impairment in the promoter region of GSTT and GSTP genes by hypermethylation may increase the risk of schizophrenia.

Effect of pro-inflammatory cytokine (IFN-γ +874, IL-18-137 G/C,-607 C/A) genes in relation to risk of vesico-ureteral reflux.

AIM: The aim this work is to estimate whether genetic polymorphisms of +874 of IFN-γ and -137 G/C,-607 C/A of IL-18 genes are implicated in the development of VUR, because a vast literature indicates that genetic variations play a significant role in the pathogenesis of VUR. MATERIALS AND METHODS: The PCR single specific primer (SSP) and amplification refractory mutation system (ARMS) were applied for analyzing the polymorphic sites of -137 G/C,-607 C/A of IL-18 and +874 of IFN-γ genes in 110 healthy controls and 124 VUR children. RESULTS: A significant relationship was found between AT and combined AT + TT genotypes of IFN-γ and highly increased risk of VUR (OR = 4.2, 95% CI, 2.00-9.24; p < 0.0001: OR = 4.00, 95% CI, 1.90-8.70, p < 0.0001, respectively). On the other hand, the genotype frequency of IL18-137 G/C indicated a significant assessment of the decrease risk of VUR for GC and GC + CC genotypes (OR = 0.53, 95% CI, 0.3-0.9; p = 0.02: OR = 0.53, 95% CI, 0.3-0.92 p = 0.01, respectively). No significant association was found between -607 C/A polymorphism of IL-18 and UVR. CONCLUSION: To the author's best knowledge, this is the first data regarding polymorphism of IFN-γ (+874) cytokine genes that highly increased the risk of VUR. To confirm the presented data, further studies should be done in different populations with a larger sample size.

Analysis of methylation and mRNA expression status of FADD and FAS genes in patients with oral squamous cell carcinoma.

BACKGROUND: Apoptosis is an important mechanism that is responsible for the physiological deletion of harmful, damaged, or unwanted cells. Changed expression of apoptosis-related genes may lead to abnormal cell proliferation and finally to tumor genesis. Our aims were to analyze the promoter methylation and gene expression profiles of FADD and FAS genes in risk of OSCC. MATERIAL AND METHODS: we analyze the promoter methylation status of FADD and FAS genes using Methylation - Specific PCR (MSP) in 86 OSCC tissues were kept in paraffin and 68 normal oral tissues applied as control. Also, FADD and FAS genes expression were analyzed in 19 cases and 20 normal specimens by Real-Time Reverse-Transcripts PCR. RESULTS: Aberrant promoter methylation of FADD and FAS genes were detected in 12.79 % (11 of 86) and 60.46 % (52 of 86) of the OSCC cases, respectively, with a significant difference between cases and healthy controls for both FADD and FAS genes (P < 0.001). The gene expression analysis showed statistically significant difference between cases and healthy controls for both FADD (p<0.02) and FAS (p<0.007) genes. CONCLUSIONS: To the best our knowledge, the data of this study are the first report regarding, the effect of promoter hypermethylation of the FADD and FAS genes in development of OSCC. To confirm the data, it is recommended doing further study in large sample sizes in various genetic populations.

Evaluation of polymorphism, hypermethylation and expression pattern of CTLA4 gene in a sample of Iranian patients with schizophrenia.

The cytotoxic T lymphocyte-associated antigen 4 gene (CTLA4) has a crucial role in regulation of T cell proliferation and mediates T cell apoptosis by encoding the T cell receptor. Schizophrenia (SCZ) patients often have abnormalities in terms of the function and development of the immune system. The aim of the present study was to investigate promoter variation and expression profile of the CTLA4 gene in patients with SCZ. We isolated genomic DNA from peripheral blood of 94 individuals with SCZ and 99 healthy control subjects. Genotypic analysis of CTLA4 (-318) was done by Tetra-ARMS-PCR. Methylation-specific polymerase chain reaction (MS-PCR) was used to estimate promoter hypermethylation of the CTLA4 gene. In addition, we investigated CTLA4 mRNA levels in 34 blood samples from cases and healthy controls using real-time reverse transcription PCR. The CT genotype of CTLA4 has a significantly protective effect on the risk to SCZ (OR=0.44; 95% CI 0.18-1.06, P=0.007) in comparison with the wild CC genotype. Promoter methylation of the CTLA4 gene increased the risk of disease statistically (OR=3.82, 95% CI 1.34-10.9, P=0.015) in cases when compared to healthy controls in blood samples. The mRNA expression level results showed statistically significant differences (P<0.0001) between cases (n=17) and healthy controls (n=17). To the best of our knowledge, this is the first evidence showing that promoter methylation of the CTLA4 gene along with transition of C to T was linked to a significantly higher expression of CTLA4 mRNA levels in patients with SCZ.

Different Effects of Low-Level Laser Therapy on the Proliferation of HT29 Cells in Culture and Xenograft Models.

Introduction: Different kinds of treatments have been developed to fight cancers. Low-level laser therapy (LLLT), also known as photobiomodulation therapy (PBMT), is a low-power monochromatic and coherent light that has been used successfully for healing injuries and combating malignancies. However, there are concerns about the application of LLLT to cancers due to the increased proliferation of some cancer cells after LLLT. Methods: This study investigated the effects of 650 nm and 870 nm lasers on the proliferation of HT29 colorectal cancer cell lines in vitro and in vivo. Results: The results showed that the laser with a wavelength of 870 nm did not meaningfully alter the proliferation of cultured cells. However, cell proliferation was promoted when the laser was applied within a wavelength of 650 nm. Treatment of HT29-derived tumors in nude mice with the 650 nm laser resulted in the decline of the tumor progression rate compared to controls. This result was inconsistent with the proliferative effects of the laser on the cultured cells. Conclusion: Cell behavior in response to LLLT might be different between cell culture and xenograft models.

DNA methylation and expression profiles of the brain-derived neurotrophic factor (BDNF) and dopamine transporter (DAT1) genes in patients with schizophrenia.

Methylation and expression profile of CpG islands were examined in the promoters of the brain-derived neurotrophic factor (BDNF) and dopamine transporter (DAT1) genes. These are well known to be involved in the pathophysiology of psychiatric disorders such as schizophrenia. Genomic DNA was extracted from peripheral blood of 80 patients with schizophrenia and 71 healthy controls. Methylation pattern was studied by Methylation-Specific PCR. RNA expression analysis was done on extracted RNA from blood samples from patients suffering from schizophrenia (n = 17) and healthy controls (n = 17). Frequency of the BDNF gene methylation was highlighted as a statistically significant relationship between cases and controls regarding decreased risk of disease in comparison to unmethylated patterns (OR = 0.24; 95 % CI = 1.11-0.50; P = 0.00007). For the DAT1 gene, this relationship was insignificant in 61 cases (76.25 %) and 52 controls (73.23 %) (OR = 1.17; 95 % CI = 0.53-2.61). Estimates of relative gene expression revealed a statistically significant association of the BDNF gene between schizophrenic patients and healthy controls (Mean ± SD: 13.3920 ± 15.19 and 0.437 ± 0.328, P = 0.0001) respectively; however, it was not significant for the DAT1 gene. This first hand evidence, regarding BDNF and DAT1 gene methylation and their expression profile with risk of schizophrenia, indicated a significant function for the BDNF gene in the development of schizophrenia. However, further populations with large sample sizes need to be studied to verify the exact role of BDNF in mental disorders such as schizophrenia.

Association between paraoxonase-1 gene polymorphisms and risk of metabolic syndrome.

Paraoxonase-1 (PON1), a high-density lipoprotein (HDL) associated enzyme, is involved in the metabolism and detoxification of insecticides and pesticides. Three polymorphisms within the PON1 gene affect the enzyme activity. Two of these (L55M and Q192R) are located at the coding region and the third (-107C/T) is in promoter region. We performed a case-control study in order to elucidate the possible contribution of variability within PON1 at three mentioned positions to the risk of MS in a South-East Iranian population. DNA was isolated from peripheral blood of patients (N = 119) with MS and healthy controls (N = 201). Allelic polymorphisms at positions Q192R, L55M and -107C/T in the PON1 gene were studied by Amplification Refractory Mutation System (ARMS)-PCR. It was observed that genotypes RR and QR + RR of Q192R locus significantly increased the risk of MS (OR = 2; 95% CI: 1.17-3.40, P = 0.0001 and OR = 1.62; 95% CI: 1.0-2.63; P = 0.05, respectively). The risk in patients with MM and LM + MM genotypes at the L55M locus was marginal (OR = 1.33; 95% CI: 0.68-1.85; P = 0.34 and OR = 1.12; 95% CI: 0.68-1.85; P = 0.73 respectively). The CC genotype at -107C/T locus also increased the risk of metabolic syndrome, but was not significant. This association was somewhat stronger when combined genotypes at Q192R and L55M loci were analyzed (OR = 3.30; 95% CI: 1.34-8.24; P = 0.007). Our results, in this first study, provide evidence for association of PON1 gene polymorphisms with the risk for metabolic syndrome.

Identification of a TPP1 Q278X Mutation in an Iranian Patient with Neuronal Ceroid Lipofuscinosis 2: Literature Review and Mutations Update.

Neuronal ceroid lipofuscinoses type 2 (CLN2), the most common form of Batten disease, is caused by TPP1 loss of function, resulting in tripeptidyl peptidase-1 enzyme deficiency and cerebral accumulation of lipopigments. Clinical hallmarks include epileptic seizures, vision loss, progressive movement disorder, ataxia, and eventually death. Diagnosis is often delayed due to the rarity of the conditions. Results: Here, we report a case presenting with clinical features of CLN2, carrying a homozygous novel nonsense variant in TPP1 (NM_000391:c.C832T, (p.Q278*), rs1352347549). Moreover, we performed a comprehensive literature review regarding previously identified disease-causing TPP1 mutations and genotype-phenotype correlations. Conclusion: Depending on the type of mutation, different phenotypes are observed in patients with CLN2, suggesting that the severity of phenotypes is related to the genotype of the patients.

Analysis of methylation patterns and expression profiles of p14ARF gene in patients with oral squamous cell carcinoma.

AIMS: To analyze the promoter methylation profile and mRNA expression of the p14ARF gene in oral squamous cell carcinoma (OSCC). METHODS: Promoter methylation of the p14ARF gene was investigated by methylation-specific polymerase chain reaction in paraffin-embedded tissues from 76 patients with OSCC and 57 oral tissues used as healthy controls. Expression of p14ARF mRNA was also determined using real-time quantitative reverse-transcription PCR. The methylation status and mRNA level profile of the gene and their relationship with clinical data were analyzed. RESULTS: Methylation of the p14ARF gene in OSCC was significantly increased compared to normal control tissues (chi(2) = 16.73, p < 0.0001). The relative expression of p14ARF mRNA in OSCC was not significantly different from that in healthy control samples. CONCLUSION: Promoter methylation of p14ARF may be an important mechanism in OSCC, and its determination may be considered an important tool in the early diagnosis and treatment of OSCC.

Association of Genetic Polymorphisms in GSTP1, GSTM1, and GSTT1 Genes with Vesicoureteral Reflux Susceptibility in the Children of Southeast Iran.

BACKGROUND: Vesicoureteral reflux (VUR) disease is the most common type of urinary tract anomalies in children. Genetic risk factors may be associated with the etiology of VUR. The role of the Glutathione S-transferases (GSTs) as multifunctional enzymes is cellular oxidative stress handling. This is the first study aimed at evaluating the relative risk of GSTP1, GSTM1, and GSTT1 polymorphisms in VUR susceptibility in children and provides new important insights into the genetics of affected children. METHODS: The study was done in 2013 in Sistan and Baluchestan University, eastern Iran. Genotyping of three GSTP1, GSTM1, and GSTT1 genes were determined using the multiplex polymerase chain reaction assay in 216 reactions for 72 VUR children and 312 reactions for 104 healthy controls. RESULTS: The presence of GSTT1 deletion was associated with high risk of VUR in children, whereas GSTP1 and GSTM1 genotypes did not show the same effect. Furthermore, the combination of GSTT1/GSTM1 and GSTT1/ GSTP1 genotypes showed a significant influence on lower risk of VUR in children. CONCLUSION: Deletion of GSTT1 functional gene is a genetic risk factor causing VUR in children. Interestingly, the combination of GSTM1 and GSTP1 null genotypes with GSTT1 has shown a protective role against risk of GSTT1 deletion.

No association of TAP1 and TAP2 genes polymorphism with risk of cervical cancer in north Indian population.

BACKGROUND: Transporter associated with antigen processing (TAP), a member of the ATP-binding cassette transporter super family, is composed of two integral membrane proteins, TAP-1 and TAP-2. The TAP gene product is involved in the processing of endogenous peptides that bind to MHC class I molecules. Mutations and/or polymorphism within these genes could alter the efficacy of the immune response which might be relevant for the development of autoimmune diseases and cancer. METHODS: DNA was isolated from peripheral blood sample of 200 patients with cervical cancer and 200 healthy controls. TAP1 and TAP2 allele polymorphism were determined by polymerase chain reaction. RESULT: Significant protective OR (OR = 0.22 95% CI = 0.09-0.51, P < 0.001-OR = 0.47, 95% CI = 0.24-0.92, P = 0.02) was observed for GG and combined AG+GG genotypes of TAP2 in patients with SCC respectively. Similarly, such genotypes (GG, AG+GG) appeared same OR for patient with cervical cancer in study group (OR = 0.12, 95% CI = 0.04-0.39-P < 0.001-OR = 0.5 ,95% CI = 0.25-0.95-P = 0.03). There was decrease risk of cervical cancer in user of oral contraceptive with AG and GG genotypes of TAP2 (OR = 0.55, 95% Cl = 0.41-0.73, P = 0.002, OR = 0.09, 95% CI = 0.02-0.36, P < 0.001) respectively. In case of TAP1 gene all allelic polymorphisms showed a decrease OR in patients with cervical cancer in passive smokers and user of oral contraceptives, though, no significant CONCLUSION: Thus, TAP1 and TAP2 genes polymorphism are not linked to cervical carcinoma, since no association was found between a particular genotype and the disease.

Involvement of the Dysregulation of miR-23b-3p, miR-195-5p, miR-656-5p, and miR-340-5p in Trastuzumab Resistance of HER2-Positive Breast Cancer Cells and System Biology Approach to Predict Their Targets Involved in Resistance.

Resistance to trastuzumab has become a limiting factor for therapeutic efficacy of human epidermal growth factor 2 (HER2)-positive breast cancer. Different expression levels of miRNAs in cancer cells have been associated with poor prognosis and response to chemotherapy. The aim of this study was to evaluate miRNAs that were thought to be associated with HER2-positive breast cancer chemoresistance. In this study, the relative expression of candidate miRNAs to U6 RNA was evaluated in trastuzumab-resistant and trastuzumab-sensitive cells using relative real-time PCR. Our results demonstrated that miR-23b-3p, miR-195-5p, miR-656-5p, and miR-340-5p were significantly dysregulated. For the first time in this study, these miRNAs were identified to be involved in trastuzumab resistance. TargetScan and miRDB were then used for predicting the potential targets of the candidate miRNAs. Our results also revealed that the predicted potential targets of these miRNAs were strongly associated with drug resistance pathways. As a relative expression of candidate miRNAs was statistically different in trastuzumab-resistant and trastuzumab-sensitive cells, their potential targets were involved in drug resistance pathways. We strongly hypothesized the dysregulation of miRNAs as a possible mechanism of trastuzumab resistance. We also assumed that the strategic manipulation of these regulatory networks might be a possible therapeutic strategy to improve the results of chemotherapy for this resistance. However, more research is needed to evaluate the role of these miRNAs in the acquisition of trastuzumab resistance.

Identification of a homozygous GFPT2 variant in a family with asthenozoospermia.

PURPOSE: Asthenozoospermia (ASZ) is a condition characterized by reduced sperm motility in semen affecting approximately 19% of infertile men. Major risk factors, particularly gene mutations, still remain unknown. The main aim of the present study was to identify novel genes and mutations that may influence human sperm motility. METHODS: Whole-exome sequencing (WES) was performed on a large pedigree of infertile men (n = 5) followed by bioinformatics analyses. Candidate pathogenic variants were screened in a control cohort of 400 ancestry-matched Iranian fertile men, 30 unrelated men with idiopathic ASZ, and public databases. RESULTS: A rare mutation in GFPT2 gene (c.1097G > A; p.Arg366Gln) located in the SIS 1 domain was segregated with the phenotype and was consistent with autosomal recessive inheritance. The in silico analyses revealed that the mutation might affect the function of SIS 1 domain and abolish its carbohydrate-binding ability. CONCLUSION: Homozygosity of the GFPT2 p.Arg366Gln mutation was associated with increased levels of reactive oxygen species (ROS) in spermatozoa and decreased sperm motility.

Mimicry and well known genetic friends: molecular diagnosis in an Iranian cohort of suspected Bartter syndrome and proposition of an algorithm for clinical differential diagnosis.

BACKGROUND: Bartter Syndrome is a rare, genetically heterogeneous, mainly autosomal recessively inherited condition characterized by hypochloremic hypokalemic metabolic alkalosis. Mutations in several genes encoding for ion channels localizing to the renal tubules including SLC12A1, KCNJ1, BSND, CLCNKA, CLCNKB, MAGED2 and CASR have been identified as underlying molecular cause. No genetically defined cases have been described in the Iranian population to date. Like for other rare genetic disorders, implementation of Next Generation Sequencing (NGS) technologies has greatly facilitated genetic diagnostics and counseling over the last years. In this study, we describe the clinical, biochemical and genetic characteristics of patients from 15 Iranian families with a clinical diagnosis of Bartter Syndrome. RESULTS: Age range of patients included in this study was 3 months to 6 years and all patients showed hypokalemic metabolic alkalosis. 3 patients additionally displayed hypercalciuria, with evidence of nephrocalcinosis in one case. Screening by Whole Exome Sequencing (WES) and long range PCR revealed that 12/17 patients (70%) had a deletion of the entire CLCNKB gene that was previously identified as the most common cause of Bartter Syndrome in other populations. 4/17 individuals (approximately 25% of cases) were found to suffer in fact from pseudo-Bartter syndrome resulting from congenital chloride diarrhea due to a novel homozygous mutation in the SLC26A3 gene, Pendred syndrome due to a known homozygous mutation in SLC26A4, Cystic Fibrosis (CF) due to a novel mutation in CFTR and apparent mineralocorticoid excess syndrome due to a novel homozygous loss of function mutation in HSD11B2 gene. 1 case (5%) remained unsolved. CONCLUSIONS: Our findings demonstrate deletion of CLCNKB is the most common cause of Bartter syndrome in Iranian patients and we show that age of onset of clinical symptoms as well as clinical features amongst those patients are variable. Further, using WES we were able to prove that nearly 1/4 patients in fact suffered from Pseudo-Bartter Syndrome, reversing the initial clinical diagnosis with important impact on the subsequent treatment and clinical follow up pathway. Finally, we propose an algorithm for clinical differential diagnosis of Bartter Syndrome.