A novel variant of C12orf4 linked to autosomal recessive intellectual disability type 66 with phenotype expansion.

BACKGROUND: Intellectual disability (ID) is a hallmark of many rare disorders that are highly heterogeneous and complex. A large number of specific genes are involved in development of this heterogeneity, and each of these genes is only found in a small number of patients. This weakens the definition of the predominant genotype and the phenotypic characteristics associated with that gene. Autosomal recessive ID type 66 (OMIM #618221) is one of these very rare diseases created by defects in the C12orf4 gene. METHODS: The present study included two patients from an Iranian family with initial diagnosis of non-syndromic ID, aiming to identify the possible genetic cause(s), and whole-exome sequencing (WES) was performed for the proband. The obtained variant was confirmed by Sanger sequencing and co-segregated in the family. RESULTS: The patients carried a novel pathogenic splicing variant called c.1441-1G>A in exon 12 of the C12orf4 gene (NM_001304811). They predominantly manifested ID, behavioral problems, speech impairment and dysmorphic facial features, some of which had not been reported in previous studies. CONCLUSIONS: A novel pathogenic splicing variant was identified named c.1441-1G>A in the C12orf4 gene. To date, only seven families have been reported with defects in this gene. Previous studies have not highlighted the exact clinical manifestations of these patients; thus, the present study could contribute to a better delineation of the genotype-phenotype correlation and interpretation of very rare variants of the gene.

Association of -604G/A and -501A/C Ghrelin and Obestatin Prepropeptide Gene Polymorphisms with Polycystic Ovary Syndrome.

Ghrelin hormone has an important role in a wide range of metabolic and non-metabolic processes. Polymorphisms of ghrelin gene could be associated with a large number of diseases. The aim of this study was to evaluate the association of -604G/A and -501A/C polymorphisms in ghrelin and obestatin prepropeptide gene (GHRL) with polycystic ovary syndrome (PCOS) in a sample of Iranian women. One hundred and fifty-two women with PCOS and 162 age-matched apparently healthy women as control group were enrolled in this study. The study subjects were genotyped for polymorphisms in the ghrelin gene using polymerase chain reaction-restriction fragment length polymorphism-based methods. Biochemical parameters, serum prolactin, luteinizing hormone, follicle stimulating hormone, estradiol, and testosterone were estimated by chemiluminescence assay. Serum lipids and lipoproteins were determined by standard enzymatic methods. The association between the risk of PCOS and ghrelin gene polymorphisms was examined using Multivariate analysis. The frequency of the -604G/A and -501A/C polymorphisms was not statistically different between patients and the control group of women (p = 0.12 and p = 0.21, respectively). A significantly higher level of LDL-C was found in the wild-type AA genotype compared with CC genotype of -501A/C polymorphism (p = 0.02). Our findings indicate that neither -604G/A and nor -501A/C polymorphisms of ghrelin gene are associated with PCOS, but suggest a relation between the presence of polymorphic allele of -501A/C polymorphism and LDL-C level in a sample of Iranian women.

Lack of association between LIPC-514 C/T polymorphism of hepatic lipase and endometriosis in Iranian women.

AIM: Patients with endometriosis may suffer from dyslipidemia. Hepatic lipase (HL) is involved in the metabolism of lipoproteins and has an important role in reverse cholesterol transport. The aim of this study was to investigate the association between the LIPC-514 C/T polymorphism in the HL gene and the risk of endometriosis in a group of Iranian women. METHODS: Ninety-seven patients with endometriosis and 107 women who were negative for endometriosis after diagnostic laparoscopy, as control group, were enrolled in this cross-sectional study. Samples were analyzed for polymorphism of the HL gene using polymerase chain reaction restriction fragment length polymorphism. RESULTS: Multivariate analysis was used to examine the association between the risk of endometriosis and LIPC-514 C/T polymorphism. There was no statistically significant difference in the frequency of the LIPC-514 C/T polymorphism between patients and the controls (60.7% CC, 34.6% CT, 4.7% TT versus 68.4%, 27.4%, 4.2%, respectively, P = 0.52). CONCLUSION: The present study suggested that the LIPC-514 C/T polymorphism of the HL gene has no significant association with the risk of endometriosis in the studied Iranian women.

Identification of a Novel Variant in CC2D1A Gene Linked to Autosomal Recessive Intellectual Disability 3 in an Iranian Family and Investigating the Structure and Pleiotropic Effects of this Gene.

Objectives: Intellectual disability (ID) represents a significant health challenge due to its diverse and intricate nature. A multitude of genes play a role in brain development and function, with defects in these genes potentially leading to ID. Considering that many of these genes have yet to be identified, and those identified have only been found in a small number of patients, no complete description of the phenotype created by these genes is available. CC2D1A is one of the genes whose loss-of-function mutation leads to a rare form of non-syndromic ID-3(OMIM*610055), and four pathogenic variants have been reported in this gene so far. Materials & Methods: n the current study, two affected females were included with an initial diagnosis of ID who were from an Iranian family with consanguineous marriage. Whole-exome sequencing was used to identify the probable genetic defects. The Genotypic and phenotypic characteristics of the patients were compared with a mutation in the CC2D1A gene, and then the structure of the gene and its reported variants were investigated. Results: The patients carried a novel homozygous splicing variant (NM_017721, c.1641+1G>A) in intron 14, which is pathogenic according to the ACMG guideline. Loss-of-function mutations in CC2D1A have severe phenotypic consequences such as ID, autism spectrum disorder (ASD), and seizures. However, missense mutations lead to ASD with or without ID, and in some patients, they cause ciliopathy. Conclusion: This study reports the fifth novel, probably pathogenic variant in the CC2D1A gene. Comparing the clinical and molecular genetic features of the patients with loss-of-function mutation helped to describe the phenotype caused by this gene more precisely. Investigating the CC2D1A gene's mutations and structure revealed that it performs multiple functions. The DM14 domain appears more pivotal in triggering severe clinical symptoms, including ID, than the C2 domain.

I405V polymorphism of CETP gene and lipid profile in women with endometriosis.

Genetic factors have an important role in the pathophysiology of endometriosis. In addition, abnormalities in lipid profile and intrinsic inflammatory status are associated with disease progression. The purpose of this study was to evaluate the effect of the I405V polymorphism of cholesteryl ester transfer protein (CETP) gene and lipid profile with the risk of endometriosis in women. Ninety-seven women with laparoscopy-diagnosed endometriosis were recruited for this study, and 107 patients with no evidence of endometriosis confirmed by laparoscopy served as controls. Samples were analyzed for polymorphism of the CETP gene using polymerase chain reaction-restriction fragment length polymorphism-based methods. After adjustment for body mass index, high-density lipoprotein-C and low-density lipoprotein-C, the risk of endometriosis in patients with normal genotype homozygous was more of the rare allele (p < 0.001, odds ratio = 0.21, 95% confidence interval = 0.09-0.45). Our results suggest that I405V polymorphism of CETP gene plays an important role as independent factor in the risk of endometriosis in women.

Clinical and Genetic Characteristics of Splicing Variant in CYP27A1 in an Iranian Family with Cerebrotendinous Xanthomatosis.

Background: Cerebrotendinous xanthomatosis (CTX) is a rare congenital lipid-storage disorder, leading to a progressive multisystem disease. CTX with autosomal recessive inheritance is caused by a defect in the CYP27A1 gene. Chronic diarrhea, tendon xanthomas, neurologic impairment, and bilateral cataracts are common symptoms of the disease. Methods: Three affected siblings with an initial diagnosis of non-syndromic intellectual disability were recruited for further molecular investigations. To identify the possible genetic cause(s), whole exome sequencing was performed on the proband. Sanger sequencing was applied to confirm the final variant. The clinical and molecular genetic features of the three siblings from the new CTX family and other patients with the same mutations, as previously reported, were analyzed. The CYP27A1 gene was also studied for the number of pathogenic variants and their location. Results: We found a homozygous splicing mutation, NM_000784: exon6: c.1184+1G>A, in CYP27A1 gene, which was confirmed by Sanger sequencing. Among the detected pathogenic variants, the splice site mutation had the highest prevalence, and the mutations were mostly found in exon 4. Conclusion: This study is the first to report the c.1184+1G>A mutation in Iran. Our findings highlight the other feature of the disease, which is the lack of relationship between phenotype and genotype. Due to nonspecific symptoms and delay in diagnosis, CYP27A1 genetic analysis should be the definitive method for CTX diagnosis.

The effects of quercetin on the gene expression of the GABAA receptor α5 subunit gene in a mouse model of kainic acid-induced seizure.

The flavonoid quercetin has recently been reported to have neuroprotective effects, and the role of the gamma-aminobutyric acid A alpha 5 subunit (GABAA α5) receptor has been determined in some nervous system disorders. The aim of this study was to identify the molecular mechanism of the effect of quercetin administered at anticonvulsive doses on the expression of the GABAA α5 receptor gene in kainic acid (KA)-induced seizures in mice. The experimental animals were divided into four groups: control, KA, and KA + quercetin at 50 or 100 mg/kg, respectively. The results showed a dose-dependent reduction in the behavioral seizure score with quercetin pre-treatment in the KA mouse model. Two hours after the end of the 7-day treatment regimen, expression of the GABAA α5 receptor gene in the hippocampus was found to be increased in the KA group, but this increase was reduced in the KA + quercetin 50 or 100 mg/kg treatment groups. These results suggest that expression of the GABAA α5 receptor could be a mechanism for reducing seizure severity or may be a marker of seizure severity. Further studies are necessary to clarify quercetin's mechanism of action and the relation of GABAA α5 receptor gene expression to seizure severity.

Modulation of the Expression of the GABAA Receptor ß1 and ß3 Subunits by Pretreatment with Quercetin in the KA Model of Epilepsy in Mice: -The Effect of Quercetin on GABAA Receptor Beta Subunits.

OBJECTIVES: Quercetin is a flavonoid and an important dietary constituent of fruits and vegetables. In recent years, several pharmacological activities of quercetin, such as its neuroprotective activity and, more specifically, its anti-convulsant effects in animal models of epilepsy, have been reported. This study evaluated the role of quercetin pretreatment on gene expression of γ-amino butyric acid type A (GABAA) receptor beta subunits in kainic acid (KA)-induced seizures in mice. METHODS: The animals were divided into four groups: one saline group, one group in which seizures were induced by using KA (10 mg/kg) without quercetin pretreatment and two groups pretreated with quercetin (50 and 100 mg/kg) prior to seizures being induced by using KA. Next, the messenger ribonucleic acid (mRNA) levels of the GABAA receptor ß subunits in the hippocampus of each animal were assessed at 2 hours and 7 days after KA administration. Quantitative real-time polymerase chain reaction (RT-PCR) assay was used to detect mRNA content in hippocampal tissues. RESULTS: Pretreatments with quercetin at doses of 50 and 100 mg/kg prevented significant increases in the mRNA levels of the ß 1 and the ß 3 subunits of the GABAA receptor at 2 hours after KA injection. Pretreatment with quercetin (100 mg/kg) significantly inhibited ß 1 and ß 3 gene expression in the hippocampus at 7 days after KA injection. But, this inhibitory effect of quercetin at 50 mg/kg on the mRNA levels of the ß 3 subunit of the GABAA receptor was not observed at 7 days after KA administration. CONCLUSION: These results suggest that quercetin (100 mg/kg) modulates the expression of the GABAA receptor ß 1 and ß 3 subunits in the KA model of epilepsy, most likely to prevent compensatory responses. This may be related to the narrow therapeutic dose range for the anticonvulsant activities of quercetin.

Evaluation of mRNA Contents of YBX2 and JHDM2A Genes on Testicular Tissues of Azoospermic Men with Different Classes of Spermatogenesis.

OBJECTIVE: Animal model studies have shown that MSY2 and JHDM2A genes have an important role in spermatogenesis process and fertility of male mice. But the potential role of these genes in human spermatogenesis and fertility is not known yet. Therefore, we evaluated expression ratios of these genes in testis tissues of men with normal and impaired spermatogenesis. MATERIALS AND METHODS: In this experimental study, after RNA extraction and cDNA syn- thesis from 50 non-obstructive azoospermic and 12 normal testis tissues, the expression ratios of genes were evaluated by real time polymerase chain reaction (PCR) technique. Hematoxcylin and eosin (H&E) staining was used for histological classification of testis tissues. For statistical analysis, one way analysis of variance (ANOVA) test was carried out. RESULTS: Our results showed a significant reduction in mRNA level of YBX2 in samples with impaired spermatogenesis (p<0.001) compared to samples with qualitatively normal spermatogenesis and normal spermatogenesis; however, in JHDM2A gene, despite sensible reduction in gene expression level in men with impaired spermatogenesis, no significant differences were shown (p>0.05). Furthermore in YBX2, a significant negative correlation was demonstrated between the efficiency score of spermatogenesis and the threshold cycle (CT) (r=-0.7, p<0.0001), whereas in JHDM2A, this negative correlation was not significant (r=-0.4, p=0.06). CONCLUSION: Generally, these data indicated that YBX2 and JHDM2A genes may play an important role in male infertility, and suggested that these molecules can act as useful biomarkers for predicting male infertility.

Mutations of the phenylalanine hydroxylase gene in Iranian patients with phenylketonuria.

Phenylketonuria (PKU) is an autosomal recessive disease which results from mutations in the phenylalanine hydroxylase (PAH) gene. The aim of this study was the identification of sixteen different mutations in Iranian patients with hyperphenylalaninemia. The mutations were detected during the characterization of PAH genotypes of 39 PKU patients from Qazvin and Zanjan provinces of Iran. PAH mutations have been analyzed by PCR and direct sequencing of PCR products of the promoter region and all 13 exons of PAH gene, including the splicing sites. A mutation detection rate of 74.3 % was realized. Two mutations were found at high frequencies: R176X (10.25 %) and p.P281L (10.25 %). The frequencies of the other mutations were: IVS2+5G>A (2.56 %), IVS2+5G>C (2.56 %), p.L48S (2.56 %), p.R243Q (2.56 %), p.R252Q (5.12 %), p.R261Q (7.69 %), p.R261X (5.12 %), p.E280K (2.56 %), p.I283N (2.56 %), IVS9+5G>A (2.56 %), IVS9+1G>A (1.28 %), IVS11+1G>C (1.28 %), p.C357R (1.28 %), c.632delC (2.56 %). The present results confirm the high heterogeneity of the PAH locus and contribute to information about the distribution and frequency of PKU mutations in the Iranian population.

Association between upstream purine complexes of human caveolin-1 gene and schizophrenia in qazvin province of iran.

BACKGROUND: Caveolin is a multifunctional and scaffolding membrane protein, which involves cholesterol trafficking to plasma lipid microdomain. It organizes and targets synaptic parts of the neurotransmitter and neurotrophic receptor signaling pathways. Caveolins are encoded by CAV-1, 2 and 3 genes. Disruption of the CAV1 would likely ruin the neuronal signaling, which leads to symptoms of schizophrenia in predisposed individuals. OBJECTIVES: The upper area of CAV-1 gene is highly conserved and can have a regulatory role in neurodegenerative diseases. This study was designed to find out the possible association of polymorphisms of this area and schizophrenia. PATIENTS AND METHODS: In a case-control study, 254 blood samples were obtained from 127 patients with schizophrenia and 127 well matched controls referred to 22 Bahman Hospital of Qazvin University of Medical Sciences (QUMS) in Qazvin province, Iran, using simple random sampling method. After extracting DNA, the upper region of the human CAV1- gene was amplified by PCR in all collected samples. The products were visualized by silver staining in 10% polyacrylamide gel and then sequenced. RESULTS: We detected nine homozygotes in patients and 15 in control subjects. Homozygosity was 7.08% and 11.8% in cases and control, respectively. Nine types homozygote haplotype were detected in upper region of the CAV1 gene in cases and controls. Three haplotypes were common in cases and controls; four haplotypes were seen in controls only and two in cases. CONCLUSIONS: Our findings implied a significant correlation between some haplotypes of upper region of CAV1 gene and schizophrenia. Existence of some haplotypes and lack of another in CAV1 upstream can suggest a significant correlation between schizophrenia and some haplotypes.

Induction of Apoptosis and Growth Suppression by Homeobox Gene TGIFLX in Prostate Cancer Cell Line Lncap.

BACKGROUND: TGIFLX, a Homoproteins cluster member located on the X chromosome, has a critical role in male reproduction and prostate development. Previous studies have shown the erratic expression of TGIFLX gene in a large proportion of prostate tumors. However TGIFLX function in prostate development remains unknown. The purpose of this study was to evaluate the consequences of TGIFLX expression on prostate cancer cell lines (LNCaP). METHOD: Inducible Tet-On gene expression system was used with a regulatory capability by doxycycline induction. In this system, stable LNCaP cells with TGIFLX tet-on plasmid were able to induce TGIFLX expression by doxycycline treatment. TGIFLX gene expression was confirmed by RT-PCR. RESULTS: Induction of gene expression caused cell proliferation decrement and apoptosis increment in LNCaP TGIFLX cells compared with control cells (P<0.01). Also, by using PEGFPN1 plasmid protein in this study localization was shown in nucleus. The gene was cloned in the plasmid and transfected to LNcap cells with plasmid PEGFPN1 TGIFLX and the plasmid was PEGFPN1. The TGIFLX expression was confirmed by RT-PCR and fluorescent microscopy. CONCLUSION: TGIFLX expression demonstrated a tumor suppressor characterization in a prostatic cancer cell line with low grade of tumorigenicity (LNCaP). More cell lines with different level of tumorogenicity need to be investigated for further clarification of the TGIFLX gene function.