Glial cell-derived neurotrophic factor gene polymorpisms affect severity and functionality of bipolar disorder.

Glial cell-derived neurotrophic factor and other neurotrophins have important role in the development of mental disorders. Here, we aimed to assess the effects of Single nucleotide polymorphisms at potentially regulated regions of GDNF on severity and functionality of bipolar disorder and GDNF serum levels in bipolar disorder patients and healthy volunteers. Severity and functionality of bipolar disorder were evaluated using the Clinical Global Impression and Global Assessment of Functioning scales in sixty-six bipolar disorder patients. The GDNF serum levels obtained from bipolar disorder patients and healthy volunteers who had been already reported SNPs information by our group. GAF scales were lower and GDNF serum levels were higher in Bipolar disorder patients with T/A genotype at 5:37812784 and 5:37812782 compared to patients with T/T genotype. There were significant difference in severity and functionality scores, but not in GDNF serum levels, between patients with G/G and G/A genotype of rs62360370 G > A SNP.rs2075680 C > A and rs79669773 T > C SNPs had no effect on bipolar disorder severity and functionality scores and GDNF serum levels. The results suggest that some SNPs of GDNF have potential association with severity and functionality of bipolar disorder. In addition, except two SNPs, none of GDNF SNPs had association with GDNF serum levels.

Mutation/SNP analysis in EF-hand calcium binding domain of mitochondrial Ca[Formula: see text] uptake 1 gene in bipolar disorder patients.

Calcium signaling is important for synaptic plasticity, generation of brain rhythms, regulating neuronal excitability, data processing and cognition. Impairment in calcium homeostasis contributed to the development of psychiatric disorders such as bipolar disorder (BP). MCU is the most important calcium transporter in mitochondria inner membrane responsible for influx of Ca[Formula: see text]. MICU1 is linked with MCU and has two canonical EF hands that are vital for its activity and regulates MCU-mediated Ca[Formula: see text] influx. In the current study, we aimed to investigate the role of genetic alteration of EF hand calcium binding motifs of MICU1 on the development of BP. We examined patients with BP, first degree relatives of these patients and healthy volunteers for mutations and polymorphisms in EF hand calcium binding motifs of MICU1. The result showed no SNP/mutation in BP patients, in healthy subjects and in first degree relatives. Additionally, alignment of the EF hand calcium binding regions among species (Gallus-gallus, Canis-lupus-familiaris, Bos-taurus, Mus-musculus, Rattus-norvegicus, Pan-troglodytes, Homosapiens and Danio-rerio) showed exactly the same amino acids (DLNGDGEVDMEE and DCDGNGELSNKE) except in one of the calcium binding domain of Danio-rerio that there was only one difference; leucine instead of Methionine. Our results showed that the SNP on EF-hand Ca[Formula: see text] binding domains of MICU1 gene had no effect in phenotypic characters of BP patients.

TLR-2 Arg753Gln, TLR-4 Asp299Gly, and TLR-4 Thr399Ile polymorphisms in Henoch Schonlein purpura with and without renal involvement.

Infections may trigger or aggravate glomerulonephritidis and renal vasculitis like Henoch Schonlein purpura (HSP). HSP is seen more frequently in patients with familial Mediterranean fever in which TLR-2 Arg753Gln polymorphism frequency is increased. Although renal involvement is the most important factor affecting the prognosis in HSP, it is not known which patients will have renal disease or why some patients have severe renal involvement while some others have mild renal disease. We investigated the role of TLR-2 and TLR-4 polymorphisms on the incidence and severity of renal involvement in HSP patients. We studied HSP patients with and without nephritis (n = 15 for each group) and healthy controls (n = 100). TLR-2 Arg753Gln and TLR-4 Asp299Gly/Thr399Ile polymorphisms were analyzed with polymerase chain reaction-restriction fragment length polymorphism method. The frequency of TLR-2 Arg753Gln, TLR-4 Asp299Gly, and Thr399Ile polymorphisms in healthy controls were 1, 3, and 2%, respectively. The frequencies of these polymorphisms were not different in HSP patients with or without nephritis compared to healthy controls. TLR-2 Arg753Gln, TLR-4 Asp299Gly, and Thr399Ile polymorphisms are not increased in HSP or HSP nephritis patients.

Predicting invasive phenotype with CDH1, CDH13, CD44, and TIMP3 gene expression in primary breast cancer.

We aimed to determine changes in the expression of the genes CDH1, CDH13, CD44, and TIMP3 to look for any relationship between them, HER2 and ESR1 expression at the RNA level, and the histopathological properties of tumors. We also analyzed the expression properties of double-negative (estrogen receptor [ER] and human epidermal growth factor receptor [HER2] both negative) breast tumors. Expression status was studied in fresh tissue at the mRNA level with quantitative PCR using hydrolysis probes. Sixty-two cancer patients and four normal controls were included in the study. When the tumor group was analyzed as a whole, the correlations of ESR1 with CDH1, CDH13, and TIMP3 were P < 0.05, P < 0.005, and P < 0.005, respectively. In ER-positive tumors, CDH1 and CDH13 were correlated directly (P < 0.005) when HER2 was correlated with CDH1, CDH13, and TIMP3 indirectly (P < 0.005, P < 0.005, and P < 0.05, respectively). CDH1 and CD44 had a strong indirect correlation (P < 0.005) in ER-negative tumors. There were significant differences in the expression levels of the CDH13, TIMP3, and CD44 genes (P < 0.005, P < 0.005, and P < 0.05, respectively) between the ER-positive and -negative groups. All four genes were found to be correlated with invasive properties in both ER-positive and -negative tumors. In double-negative tumor samples, only CD44 had a significant and strong correlation with stage, lymph node involvement, and metastasis (P < 0.05, P < 0.005, and P < 0.05, respectively). As a conclusion, a decrease in CDH1, CDH13, and TIMP3 expression levels with an increase in CD44 can be used as an indicator for invasion in both ER-positive and -negative breast tumors. In double-negative tumor tissues, CD44 can be considered a marker for aggressive properties.

Correlation of modified Shimada classification with MYCN and 1p36 status detected by fluorescence in situ hybridization in neuroblastoma.

Neuroblastoma (NB) is a childhood cancer derived from neural crest cells, with a highly variable clinical course and biologic behavior. NB cells harbor complex genetic changes. Also, MYCN amplification is a well-known molecular marker for aggressive progression, and deletion of the short arm of chromosome 1 is frequently observed in NB. The aim of this study was to investigate the correlation between genetic markers and prognostic morphological parameters to address the biology and underlying the clinical complexity of NB. Therefore, we performed fluorescence in situ hybridization analyses of chromosome band 1p36 and MYCN in a series of tumors from 43 cases classified according to the recommendation of International Neuroblastoma Pathology Committee (modification of Shimada classification). The correlations of MYCN amplification status and two distinct types of 1p36 alterations (deletion and imbalance) with Shimada classification and histologic prognostic factors were statistically analyzed. Amplification of MYCN and 1p36 deletion was present in 14 (32.6%) and 18 (41.9%) cases, respectively. Sixteen cases (37.2%) displayed a favorable histology, while 27 (62.8%) had an unfavorable histology. The 1p36 deletion was found to be an independent predictor of unfavorable histology by multivariate analysis (logistic regression test, P = 0.03), but the 1p36 imbalance did not show any significance. Both 1p36 deletion and MYCN amplification showed significant correlation with undifferentiated tumors (chi-square test, P = 0.002 and 0.03, respectively). Highly significant correlation was found between the higher mitotic karyorrhectic index (MKI) and MYCN amplification (chi-square test, P < 0.001), whereas neither 1p36 deletion nor 1p36 imbalance significantly correlated with a higher MKI (chi-square test, P > 0.05). We conclude that 1p36 deletion may be a reliable parameter in determining unfavorable histology and predicting prognosis in NB. Further studies with prognostic data are needed to highlight its clinical significance.

Downregulation of VANGL1 inhibits cellular invasion rather than cell motility in hepatocellular carcinoma cells without stimulation.

AIMS: The Wnt planar cell polarity (PCP) pathway is one of the Wnt pathways which plays a critical role in cell proliferation and fate. The VANGL1 protein is one of Wnt-PCP pathway components. It is known that Wnt-PCP pathway has major roles in cell motility but its role in hepatocellular carcinoma (HCC) progression through invasion and metastasis needs to be clarified. METHODS: We silenced VANGL1 gene expression in the HepG2 HCC cell line by stable transfection with a vector containing siRNA template for VANGL1 and investigated the change in cell invasion and motility. RESULTS: Transfected cells with the siRNA template showed significantly suppressed invasive capacity when compared to controls although cellular motility was only slightly affected. CONCLUSION: Our study showed a basal role for VANGL1 with respect to the invasive capacity of HCC cells. This suggests that the Wnt-PCP pathway may play a role in progression of HCC through cellular invasion but further studies are needed to clarify its role in cell motility.

DNA copy number changes detected by comparative genomic hybridization and their association with clinicopathologic parameters in breast tumors.

The purpose of this study was to use comparative genomic hybridization (CGH) to screen breast tumors for copy number changes: 22 ductal, 9 lobular, 7 mixed, 2 micropapillary carcinomas, and 2 ductal carcinoma in situ were studied and various regional genomic imbalances were detected. The majority of the aberrations identified in this study were in line with previous CGH findings. The most frequent DNA sequence copy number changes were 1q, 8q, and 20q gains. The frequency of 16q losses was significantly higher in lobular carcinomas. The nodal involvement was 10 times higher in cases showing losses of 13q than in cases having normal peak profile at this region. Estrogen receptor positivity was significantly higher in cases displaying 20q gains and 16q losses. Unambiguous high-level DNA amplifications have also been detected. These mapped to 4q31, 6q21 approximately q22, 8q21 approximately q24, 8p11.2 approximately p12, 11q13, 15q24 approximately qter, 20q13.1 approximately qter, and 20q12 approximately qter chromosomal locations. Our results highlight several chromosomal regions that may be important in the molecular genetics of distinct clinicopathologic breast cancer subgroups.

Microsatellite instability is not a common feature in medullary breast cancer.

Microsatellite instability (MSI) is a form of genomic instability associated with defective DNA mismatch repair in tumors. MSI is found in 85-90% of hereditary nonpolyposis colorectal cancer cases; however, its occurrence in breast carcinogenesis still remains to be clarified. In addition, data are limited on the incidence of MSI in the medullary subtype. The purpose of this study was to investigate the occurrence of MSI in medullary breast cancer (MBC). The study included a total of 16 patients with MBC, nine with typical and seven with atypical histology. The incidence of MSI in five microsatellite loci (D2S123, D3S1611, D17S807, D17S796 and Xq11-12) was determined by comparing paired normal and tumor tissue DNA after PCR amplification from paraffin-embedded tissues. All 16 tumors showed stability at five loci. Although the number of microsatellite markers and DNA samples may limit the value of our results, we conclude that the MSI phenotype is uncommon in human MBC.

Microsatellite instability in early-onset breast cancer.

Breast cancer in a young person is considered a rare and very aggressive disease. The theories addressing the underlying genetic mechanisms of this disease are controversial. Therefore, additional genetic concepts playing a possible role in its pathogenesis and prognosis must be investigated. Microsatellite instability (MSI) characterized by a mutational process of insertions or deletions in microsatellite repeats might constitute a sensitive indicator for genomic instability in cancer. MSI has been described in a wide variety of tumors, particularly in hereditary non-polyposis colorectal cancer. The reports regarding its occurrence and prognostic significance in breast cancer are in conflict with each other. The purpose of this study was to investigate MSI in early-onset breast cancer and to correlate its occurrence with clinicopathological prognisticators. In this study, 16 female patients with primary breast cancer under 35 years of age (range 29-34) were investigated for the incidence of MSI in five microsatellite loci (D2S123, D3S1611, D17S807, D17S796 and Xq11-12) by comparing paired normal and tumor tissue DNA after PCR amplification from paraffin-embedded tissues. No instability was found in any of these five microsatellite loci. Although care must be taken not to overstate the importance of this result due to the inadequate number of microsatellite markers and DNA samples studied, this preliminary report indicates that MSI phenotype is uncommon in human early-onset breast cancer. Therefore, it does not appear to be related to the prognosis of disease.

Analysis of cytotoxic T lymphocyte antigen-4 (CTLA-4) exon 1 polymorphism in patients with type 1 diabetes mellitus in a Turkish population.

Recent studies have described linkage and association between cytotoxic T-lymphocyte antigen-4 (CTLA-4) gene polymorphism and type 1 diabetes mellitus (DM1) in some ethnic populations, but not others. This finding suggests that CTLA-4 gene association with DM1 may be influenced by the racial composition of the population. Thus, it is important to study the polymorphism of the CTLA-4 gene in different ethnic groups. In this case-control association study, the CTLA-4 gene exon 1 A/G polymorphism was analyzed in 48 children with DM1 and 80 healthy controls using polymerase chain reaction-restriction fragment length polymorphism analysis. The possible interaction of the CTLA-4 gene polymorphism with the presence of established genetic markers (HLA-DR genotyping) was also evaluated in 29 patients. The results of the present study do not suggest an association of the known polymorphism in exon 1 of the CTLA-4 gene with DM1 in this Turkish population, and G-allele containing CTLA-4 genotypes were not preferentially associated with age at clinical presentation or with presence of other genetic (HLA-DR3 or -DR4) markers of DM1.

Mosaic form (45X/46XX) of Turner's syndrome. A case report.

BACKGROUND: Turner's syndrome (TS) is a sex chromosome disorder occurring in 1 in 2,500 female births and in approximately 50 in 100,000 adult females and is characterized by retarded growth, gonadal dysgenesis and infertility. The 45,X/46,XX chromosomal pattern is the most frequent mosaic type of this disease (36%). CASE: The mosaic form of TS was diagnosed in a 25-year-old, nulliparous woman whose major symptoms were menstrual irregularity and menorrhagia. She had normal development of secondary sexual characteristics and spontaneous menarche despite a very low amount (7%) of normal cells. CONCLUSION: Dysfunctional uterine bleeding is very uncommon in TS. Mosaic forms of TS may have very few features of TS despite a very low range of normal cells.

New alterations at potentially regulated regions of the Glial Derived Neurotrophic Factor gene in bipolar disorder.

INTRODUCTION: Glial Derived Neurotrophic Factor (GDNF) plays an important role in the survival and differentiation of neurons. We examined 5'upstream and 3' untranslated region of the GDNF gene by PCR amplification and direct sequencing to explore the effect of alteration in the potentially regulated part of GDNF in bipolar disorder. MATERIALS AND METHODS: Sixty-six patients with bipolar disorder, 27 first degree relatives of these patients and 56 healthy volunteers were screened for mutations and polymorphisms in GDNF gene. RESULTS: Seven previously reported polymorphisms and additional three novel allele variants of GDNF were detected. Association test of rs2075680 C>A SNP showed significant difference between patients and healthy subjects with higher allele frequency in healthy subjects performing Chi-square test. However, there was no significant difference after multiple test corrections between groups. There were no significant differences in association test of rs2075680 C>A SNP between first degree relatives and healthy volunteers/patients. rs142426358 T>C SNP was seen only in one patient with an early age of illness onset. New T>A alterations were found in chromosome locations 5:37812784 and 5:37812782 in two male bipolar disorder patients with age of illness onset 12 and 24 years. LIMITATIONS: The sample size was relatively small. DISCUSSION: Our study proposes the suggestive association between polymorphisms in the potential regulatory sites of GDNF and bipolar disorder.

Downregulation of E-cadherin expression in breast cancer by promoter hypermethylation and its relation with progression and prognosis of tumor.

Breast cancer is the most common cancer in women around the world, and novel prognosis strategies is needed to control more accurate and effective of this malignant disease. Among the latest prognostic markers is E-cadherin, which mediates cell-cell adhesion by associating with catenins. Loss of E-cadherin gene (CDH1) function by genetic or epigenetic alteration leads to tumorigenesis. The aim of our study was to investigate E-cadherin gene promoter methylation in breast cancer, and its correlation with E-cadherin protein expression. Fifty primary breast cancers tissue with ductal type and 50 normal breast sample from the same patients that was located adjacent to tumor region as controls were provided by Imam Reza-based referral and teaching hospital affiliated to Tabriz University of Medical Sciences, Tabriz, Iran. CDH1 promoter region CpG sites methylation and E-cadherin protein expression were determined by bisulfite-specific polymerase chain reaction and Western blot analysis, and the resulting products were sequenced on an ABI automated sequencer for firm conclusion. CDH1 hypermethylation in breast tumor specimen (ductal type) was observed in 94 % (47 of 50) comparing with normal samples methylation, and the significant difference was (p = 0.000). Protein expression in tumor samples tends to diminish with the CDH1 promoter region methylation. In the group of 50 ductal carcinomas cases, most of the cases showing CDH1 hypermethylation correlated inversely with the reduced levels of expression of E-cadherin proteins (95 % of full-methylated tumor samples had no protein expression, and 4.5 % of them had weak expression levels). Possible association was observed between CDH1 methylation and its protein expression (p = 0.000). The results of methylation analysis in promoter region in ten CpG sites (863, 865, 873, 879, 887, 892, 901, 918, 920, and 940) suggested that abnormal CDH1 methylation occurs in high frequencies in ductal breast tumors probably sounds the process of carcinogenesis progression.

TLR polymorphisms in FMF: association of TLR-2 (Arg753Gln) and TLR-4 (Asp299Gly, Thre399Ile) polymorphisms and myeloid cell TLR-2 and TLR-4 expression with the development of secondary amyloidosis in FMF.

Amyloidosis is the major complication of familial Mediterranean fever (FMF). Toll-like receptors (TLR) are involved in the activation of an innate immune system TLR-2 and TLR-4 recognize lipoteichoic acid and lipopolysaccharides (LPS), respectively. While TLR-2 Arg753Gln polymorphism upregulates, TLR-4 Asp299Gly and Thre399Ile polymorphisms downregulate inflammation. We investigated the effect of these polymorphisms on the development of amyloidosis in FMF patients. We also investigated myeloid cell TLR-2 and TLR-4 expressions in these patients. We studied 26 FMF patients and 13 FMF patients with amyloidosis. TLR-2 Arg753Gln and TLR-4 Asp299Gly and Thr399Ile polymorphisms were analyzed with the polymerase chain reaction-restriction fragment length polymorphism method. Myeloid cell baseline TLR-2 and TLR-4 and LPS-induced TLR-4 expressions were evaluated. The TLR-2 and TLR-4 polymorphism rate was compared with the results of 100 healthy subjects in our previous study. In addition, 13 healthy controls were enrolled for leukocyte TLR-2 and TLR-4 expressions. Serum amyloid A (SAA) levels were measured in these 13 control cases and in FMF patients during attack-free periods. The frequency of TLR-2 Arg753Gln, TLR-4 Asp299Gly, and Thr399Ile polymorphisms in healthy controls in our previous study were 1%, 3%, and 2%, respectively. The frequency of these polymorphisms were not different in FMF patients (with or without amyloidosis) compared to the control group. Likewise, myeloid cell TLR-2 and TLR-4 expressions were not different among the controls and FMF patients. However, LPS-induced TLR-4 expression in granulocytes was more prominent in FMF patients. There was no correlation between TLR-2 and TLR-4 expressions and SAA levels. Neither myeloid cell TLR-2 and TLR-4 expressions nor TLR-2 Arg753Gln, TLR-4 Asp299Gly, and Thr399Ile polymorphisms seem to affect the development of secondary amyloidosis in FMF patients in our study population.

Hyperbilirubinemic serum is cytotoxic and induces apoptosis in murine astrocytes.

BACKGROUND: High levels of unconjugated bilirubin can be neurotoxic and gliotoxic. However, the effect of serum from patients with neonatal indirect hyperbilirubinemia on astrocyte viability has never been investigated. OBJECTIVES: In the present study, we searched for the possible toxic effect of hyperbilirubinemic serum on murine astrocytes. METHODS: Heat-inactivated patient serum was added to astrocyte cultures at different concentrations varying from 1 to 20%, and cultures were incubated for 24, 48, and 72 h. Sera from healthy infants without hyperbilirubinemia were used as controls. Cytotoxicity was evaluated according to the release of lactate dehydrogenase in the culture medium. Apoptotic cell death was determined by anti-single-strand DNA immunostaining. RESULTS: The results of the present study show that hyperbilirubinemic serum induces cytotoxicity and apoptotic astrocyte death in a concentration- and time-dependent manner. CONCLUSIONS: We conclude that serum from patients with neonatal indirect hyperbilirubinemia is cytotoxic to murine astrocytes.