Deregulation of miR-1245b-5p and miR-92a-3p and their potential target gene, GATA3, in epithelial-mesenchymal transition pathway in breast cancer.

BACKGROUND: MicroRNAs (miRNAs) are small molecules that have prominent roles in tumor development and metastasis and can be used for diagnostic and therapeutic purposes. This study evaluated the expression of miR-92a-3p and miR-1245b-5p and their potential target gene, GATA3 in patients with breast cancer (BC). MATERIALS AND METHODS: In the search for BC-related microRNAs, miR-124b-5p and miR-92a-3p were selected using Medline through PubMed, miR2disease, miRcancer and miRTarBase. Moreover, target gene GATA3 and their possible interaction in the regulating epithelial-mesenchymal transition (EMT) and invasion was evaluated using in silico tools including miRTarBase, TargetScan, STRING-db, and Cytoscape. The expression level of miR-92a-3p, miR1245b-5p, and GATA3 were assessed on extracted RNAs of tumor and nontumor tissues from 36 patients with BC using qPCR. Additionally, clinical-pathologic characteristics, such as tumor grade, tumor stage, lymph node were taken into consideration and the diagnostic power of these miRNAs and GATA3 was evaluated using the ROC curve analysis. RESULTS: In silico evaluation of miR-92a-3p and miR-1245b-5p supports their potential association with EMT and invasion signaling pathways in BC pathogenesis. Comparing tumor tissues to nontumor tissues, we found a significant downregulation of miR-1245b-5p and miR-92a-3p and upregulation of GATA3. Patients with BC who had decreased miR-92a-3p expression also had higher rates of advanced stage/grade and ER expression, whereas decreased miR-1245b-5p expression was only linked to ER expression and was not associated with lymph node metastasis. The AUC of miR-1245b-5p, miR-92a-3p, and GATA3 using ROC curve was determined 0.6449 (p = .0239), 0.5980 (p = .1526), and 0.7415 (p < .0001), respectively, which showed a significant diagnostic accuracy of miR-1245b-5p and GATA3 between the BC patients and healthy individuals. CONCLUSION: MiR-1245b-5p, miR-92a-3p, and GATA3 gene contribute to BC pathogenesis and they may be having potential regulatory roles in signaling pathways involved in invasion and EMT pathways in BC pathogenesis, as a result of these findings. More research is needed to determine the regulatory mechanisms that they control.

DNAH5 gene and its correlation with linc02220 expression and sperm characteristics.

BACKGROUND: Numerous pieces of evidence show that many environmental and genetic factors can cause male infertility. Much research in recent years has investigated the function of long non-coding RNAs (lncRNAs) in fertility. The main objective of the current study was to investigate the expression of Dynein Axonemal Heavy Chain 5 (DNAH5) as a gene that plays an essential role in sperm motility in individuals with asthenozoospermia and terato-asthenozoospermia. Alterations in linc02220 expression (located close to the DNAH5 gene), its action potential in DNAH5 regulating, and the correlation between their expression and normal sperm morphology and motility were also examined. METHOD AND MATERIAL: This study examined the semen of 31 asthenozoospermia individuals (AZ), 33 terato-asthenozoospermia (TAZ) individuals, and 33 normospermia (NZ) individuals with normal sperm as a control group. The expression levels of DNAH5 and linc02220 in the sperm samples were analyzed by real-time PCR. RESULTS: Gene expression analysis revealed a significant association between DNAH5 expression and sperm motility and morphology (p < 0.0001). The DNAH5 expression levels in the TAZ and AZ groups were also significantly reduced; however, linc02220 was significantly upregulated in both TAZ and AZ groups compared to the NZ group (p < 0.0001). DNAH5 expression in the TAZ and AZ groups was negatively correlated with linc02220 expression, thus, DNAH5 downregulation was associated with linc02220 overexpression (p < 0.05). CONCLUSIONS: The gene linc02220 could be a potential regulatory target for DNAH5, and both could affect sperm's normal motility and morphology.

The impact of DAPK1 and mTORC1 signaling association on autophagy in cancer.

BACKGROUND: The autophagy pathway is used by eukaryotic cells to maintain metabolic homeostasis. Autophagy has two functions in cancerous cells which could inhibit tumorigenesis or lead to cancer progression by increasing cell survival and proliferation. METHODS AND RESULTS: In this review article, Web of Science, PubMed, Scopus,  and Google Scholar were searched and summarized published studies to explore the relationship between DAPK1 and mTORC1 signaling association on autophagy in cancer. Autophagy is managed through various proteins including the mTOR, which is two separated structural and functional complexes known as mTORC1 and mTORC2. MTORC1 is an important component of the regulatory pathway affecting numerous cellular functions including proliferation, migration, invasion, and survival. This protein plays a key role in human cancers. The activity level of mTORC1 is regulated by the death-associated protein kinases (DAPks) family, especially DAPK1. In many cancers, DAPK1 acts as a tumor suppressor which can be attributed to its ability to suppress cellular transformation and to inhibit metastasis. CONCLUSIONS: A deep investigation not only will reveal more about the function of DAPK1 but also might provide insights into novel therapies aimed to modulate the autophagy pathway in cancer and to achieve better cancer therapy.

MiR-182-5p, MiR-192-5p, and MiR-493-5p Constitute a Regulatory Network with CRISP3 in Seminal Plasma Fluid of Teratozoospermia Patients.

Numerous evidences suggested that microRNAs (miRs) could play an active and significant role during spermatogenesis. Cysteine-rich secretory protein (CRISP3) has a role in inflammatory response and is extremely over-expressed in adolescents with varicocele seminal plasma and modified semen analysis. Nowadays, the miRs expression's association with their target genes is well recognized. The aim of this study was evaluating the association of CRISP3 and four candidate miRs among teratozoospermia (TZ) infertile men. First, we have selected four miRs, miR-182-5p, miR-192-5p, miR-204-5p, and miR-493-5p bioinformatically. After that, RNA was extracted from semen samples of 21 TZ patients and 20 normozoospermia (Norm). Then, their expression levels were assessed using real-time polymerase chain reaction method. In the next step, we quantified the expression of two CRISP3 protein isoforms, targeted by these miRs, using western blotting. According to our results, up-regulation of miR-182-5p, miR-192-5p, and miR-493-5p was observed. MiR-182-5p, miR-192-5p, and miR-493-5p showed good AUC values which can be introduced as possible biomarkers of TZ. In addition, the expression level of the CRISP3 glycosylated (31 kDa) isoform was significantly lower in TZ patients than Norm ones. Notably, in TZ patients, there was a possibly positive correlation of glycosylated CRISP3 expression with normal sperm morphology. According to our results, CRISP3 protein can play a significant role in male infertility especially in maturation formation of spermatozoa. Also, deregulation of the studied miRs, miR-182-5p, miR-92-5p, and miR-493-5p, can suggest a regulatory network between these miRs and CRISP3 isoforms and suggest their regulatory roles in male infertility.

Apoptotic effects of valproic acid on miR-34a, miR-520h and HDAC1 gene in breast cancer.

Identifying miRNAs involved in cancer and devising strategies to control their expression is a new therapeutic approach. Valproic acid (VPA) has attracted a lot of interest in cancer research. We evaluated the impact of VPA on the expression of miR-34a, miR-520h, and their target gene histone deacetylase 1 (HDAC1), as well as their relationship with apoptosis in breast cancer. First, through bioinformatics analyses, the possible target genes of miR-34a and miR-520h and their roles in apoptosis regulation were investigated. Then, miR-34a, miR-520h, and HDAC1 gene expression in tissues of breast cancer patients were determined using the qRT-PCR method. The anticancer impact of VPA on apoptosis and the expression levels of miR-34a, miR-520h, and HDAC1 gene were measured in MCF-7 and MDA-MB-231 cell lines. The bioinformatics analyses indicated that miR-34a and miR-520h might make a unique contribution in regulating the apoptosis pathway. The relative expression of miR-34a and miR-520h significantly decreased in cancer tissues, while the relative expression of HDAC1 increased. In the in vitro study, VPA led to apoptosis induction and increased lipid peroxidation products in breast cancer cells. Moreover, VPA increased the expression of miR- 34a and miR-520h and decreased HDAC1 expression in MCF-7 cells. In MDA-MB-231 cells, VPA decreased the expression of these miRNAs and increased the expression of HDAC1. It can be concluded that miR-34a and miR-520h are implicated in the apoptosis pathways, and thus, VPA can recruit as a possible option in breast cancer research due to its interference with epigenetic processes.

Study of Linc00574 Regulatory Effect on the TCTE3 Expression in Sperm Motility.

We investigated the relationship of t-complex-associated-testis-expressed 3 (TCTE3) and linc00574 expression levels with sperm motility and morphology in patients with asthenozoospermia (AZ) and terato-asthenozoospermia (TAZ). The study population consisted of 31 AZ patients, 31 TAZ patients, and 32 normozoospermia (NZ) as controls. Quantitative real-time PCR was conducted to evaluate the expression levels of TCTE3 and linc00574. Bioinformatics investigations were performed using databases to find molecular pathway. TCTE3 expression was reduced significantly in AZ and TAZ patients (P < 0.05). Linc00574 expression level increased only in the AZ patients (P < 0.05). The subsequent analyses showed a significantly positive correlation between TCTE3 and linc00574 expression levels (P < 0.05). In addition, a significantly positive relationship was observed between TCTE3 expression level and sperm motility and morphology (P < 0.05). The present study suggests that TCTE3 expression is regulated by linc00574 through a negative self-regulating mechanism and therefore may affect the flagella structure and function.

Investigating the regulatory function of the ANO1-AS2 on the ANO1 gene in infertile men with asthenozoospermia and terato-asthenozoospermia.

Long non-coding RNAs (lncRNAs) have a particular expression in the testicular tissue and exhibit a regulatory function on the reproduction system. ANO1-AS2 (linc02584), as an lncRNA is located near the anoctamin1 (ANO1) gene. ANO1 is an important component of the transmembrane system exhibiting expression modifications in the idiopathic infertile men. Therefore, the present study was conducted to investigate the relationship between ANO1-AS2 and ANO1 gene expression with sperm motility and morphology in the patients with asthenozoospermia (AZ) and terato- asthenozoospermia (TAZ). The study population included 32 patients with AZ, 35 patients with TAZ, and 34 people with normozoospermia (NZ, control). The expression levels of ANO1 gene and ANO1-AS2 in the spermatozoa were measured by the quantitative real-time polymerase chain reaction (PCR). Docking analysis was performed to investigate the interactions of the ANO1 gene promoter and intermediate elements with ANO1-AS2. ANO1 gene expression was significantly (P < 0.05) downregulated in the patients however; ANO1-AS2 expression was significantly upregulated (P < 0.05). The subsequent analysis confirmed the inverse correlation between ANO1 and ANO1-AS2. ANO1 gene expression level was significantly positively correlated with sperm motility and morphology (P < 0.05). Moreover, ANO1-AS2 expression showed an inverse correlation with sperm motility and morphology (P < 0.05). Docking analysis confirmed that ANO1-AS2 could stably interact with ANO1 gene promoter. In conclusion, ANO1-AS2 is likely to downregulate the ANO1 gene by interacting with ANO1 gene promoter, which can influence the sperm motility and morphology.

Hepcidin and HFE Polymorphisms and Ferritin Level in ß-Thalassemia Major.

Background: Thalassemia patients need repeated transfusion that lead to increased blood ferritin level and iron overload in the heart and liver. Because the roles of hepcidin antimicrobial peptide (HAMP) and hemocromatosis protein (HFE) in iron metabolism have been confirmed, this study investigated the effects of these gene's polymorphisms on blood ferritin levels and iron overload in the heart and liver in patients with beta thalassemia major Materials and Methods: This cross-sectional study was conducted on 91 patients referring to the Hajar Hospital in Shahrekord, Iran in 2015. After the blood samples were collected, the ferritin levels were measured, DNA was extracted from the blood cells, and the types of polymorphisms were determined using PCR-RFLP. Data of MRI T2* in the heart and liver were drawn from the patients' medical files. Data analysis was conducted by t-test, chi-square test, Fisher's exact test, and Pearson correlation coefficient. Results: There was no significant correlation between blood ferritin level and c.-582 A>G polymorphisms of hepcidin gene (p=0.58), and H63D of HFE gene (p=0.818). In addition, there was no significant association between the polymorphisms and heart and liver MRI, but there was a significant association between blood ferritin level and qualitative heart and liver MRI (r=-0.34, p=0.035 and r=-0.001, p=0.609, respectively). Conclusion: In patients with ß-thalassemia major, the presence of c.-582A>G HAMP and H63D HFE polymorphisms is not effective on blood ferritin level and iron overload in the heart and liver in the studied region.

Valproic Acid Promotes Apoptosis and Cisplatin Sensitivity Through Downregulation of H19 Noncoding RNA in Ovarian A2780 Cells.

Cisplatin resistance is one of the main limitations in the treatment of ovarian cancer, which is partly mediated by long noncoding RNAs (lncRNAs). H19 is a lncRNA involving in cisplatin resistance in cancers. Valproic acid (VPA) is a commonly used drug for clinical treatment of seizure disorders. In addition, this drug may display its effects through regulation of noncoding RNAs controlling gene expression. The aim of the present study was the investigation of VPA treatment effect on H19 expression in ovarian cancer cells and also the relation of the H19 levels with apoptosis and cisplatin resistance. Briefly, treatment with VPA not only led to significant increase in apoptosis rate, but also increased the cisplatin sensitivity of A2780/CP cells. We found that following VPA treatment, the expression of H19 and EZH2 decreased, but the expression of p21 and PTEN increased significantly. To investigate the involvement of H19 in VPA-induced apoptosis and cisplatin sensitivity, H19 was inhibited by a specific siRNA. Our results demonstrate that H19 knockdown by siRNA induced apoptosis and sensitized the A2780/CP cells to cisplatin-induced cytotoxicity. These data indicated that VPA negatively regulates the expression of H19 in ovarian cancer cells, which subsequently leads to apoptosis induction, cell proliferation inhibition, and overwhelming to cisplatin resistance. The implication of H19→EZH2→p21/PTEN pathway by VPA treatment suggests that we could repurpose an old drug, valproic acid, as an effective drug for treatment of ovarian cancer in the future.

MiR-221/222 promote chemoresistance to cisplatin in ovarian cancer cells by targeting PTEN/PI3K/AKT signaling pathway.

Cisplatin resistance is one of the main limitations in the treatment of ovarian cancer, and its mechanism has not been fully understood. The objectives of this study were to determine the role of miR-221/222 and its underlying mechanism in chemoresistance of ovarian cancer. We demonstrated that miR-221/222 expression levels were higher in A2780/CP cells compared with A2780 S cells. An in vitro cell viability assay showed that downregulation of miR-221/222 sensitized A2780/CP cells to cisplatin-induced cytotoxicity. Moreover, we found that knockdown of miR-221/222 by its specific inhibitors promoted the cisplatin-induced apoptosis in A2780/CP cells. Using bioinformatic analysis and luciferase reporter assay, miR-221/222 were found to directly target PTEN. Moreover, knockdown of miR-221/222 in A2780/CP cells significantly upregulated PTEN and downregulated PI3KCA and p-Akt expression. In conclusion, our results demonstrated that miR-221/222 induced cisplatin resistance by targeting PTEN mediated PI3K/Akt pathway in A2780/CP cells, suggesting that miR-221/222/PTEN/PI3K/Akt may be a promising prognostic and therapeutic target to overcome cisplatin resistance and treat ovarian cancer in the future.

Lower expression of miR-218 in human breast cancer is associated with lymph node metastases, higher grades, and poorer prognosis.

Breast cancer is considered as the most prevalent malignancy in women worldwide. Despite emergence of several prognosticators for better management of patients, there are still limitations for their clinical application due to the complexity of breast tumors, and therefore, new biomarkers for better prognosis of clinical outcomes would be of the great essence. MicroRNAs are highly conserved small non-coding regulatory RNAs involved in post-transcriptional regulating of gene expression during different cellular mechanisms. Accumulating studies suggest that miR-218 plays a multifunctional role in various cancer types and different stages. Here, to address prognostic significance of miR-218 in breast cancer, we investigate the expression profile of miR-218 and B-cell-specific Moloney murine leukemia virus integration site 1 ( BMI1) gene, as one of the putative targets of miR-218, in 33 paired breast tumors and their adjacent normal tissues with respect to the clinicopathological features of patients using quantitative real-time polymerase chain reaction. The correlation of both miR-218 and BMI1 gene expression with overall survival of breast cancer patients was also examined recruiting OncoLNC data portal. Finally, to better understand biological function of miR-218 in breast cancer, we performed in silico Gene Ontology and signaling pathway enrichment analysis on miR-218 targetome. According to our data, significant elevation of the expression of miR-218 and downregulation of BMI1 were observed in clinical breast cancer specimens compared with normal tissues ( p < 0.0001). The lower expression of miR-218 was associated with lymph node metastases, higher grades, and poorer prognosis (logrank p = 0.00988), whereas no significant difference in overall survival was observed between patients with higher and lower expression of BMI1 (logrank p = 0.254). These findings suggest that miR-218 expression profiling might be clinically applicable as a prognostic biomarker in breast cancer. In addition, our in silico enrichment analyses revealed that the association of miR-218 expression with breast cancer prognosis might be through its involvement in endocytosis and gap junction biological pathways.

Association between Interleukin-10-1082 G/A and Tumor Necrosis Factor-α 308 G/A Gene Polymorphisms and Respiratory Distress Syndrome in Iranian Preterm Infants.

Cytokine polymorphisms may contribute to the prevalence of respiratory distress syndrome. The present study was done to investigate the frequency of interleukin- (IL-) 10 and tumor necrosis factor- (TNF-) α gene polymorphisms and their association with the risk of RDS in preterm infants. One-hundred and nineteen patients with RDS and 119 healthy preterm infants were enrolled. PCR restriction fragment length polymorphism was used to determine the frequency of IL-10 and TNF-α genotypes at -1082 A and -308 A, respectively. One-hundred and nineteen out of 238 infants had RDS (50%). The age of the mothers and gestational age ranged 17-45 (mean: 28.6 ± 5.3) years and 24-34 (mean: 34.3 ± 2.38) weeks, respectively. Totally, 23 deaths were recorded in the RDS group. Incidence of TNF-α-308 A/A and TNF-α-308 G/A was 84% and 16%, respectively. TNF-a-308 G/G was not found in both groups. Prevalence of IL-10-1082 G/G and IL-10-1082 G/A variants was 65.5% and 34.5%, respectively. IL-10-1082 A/A was not found in both groups. The incidence of the allele G in the IL-10-1082 polymorphism was lower in RDS group (P < 0.05). We found that the risk of RDS was correlated to sex, gestational age, and IL-10-1082.

Quantitative Gene Expression of ERG9 in Model Saccharomyces cerevisiae: Chamomile Extract For Human Cancer Treatment.

INTRODUCTION: Over expression of squalene synthase gene causes induction of growth tumour and reduction of apoptosis. This gene which is conserved between Saccharomyces cerevisiae yeast and humans, is named (ERG9). AIM: In this work, we studied the effect of Matricaria recutita extract on ERG9 gene (squalene synthase) expression in S.cerevisiae which was used as organism model in cancer therapy. MATERIALS AND METHODS: S. cerevisiae was cultured in YPD medium plus 0,250, 1000 and 3000 µg/ml of Matricaria recutita extract and we evaluated the (ERG9) gene expression by Real-time RT-PCR method after 24 hours. STATISTICAL ANALYSIS USED: At least 3 independent experiments were done. Data were analyzed using One-way ANOVA and Dunnett's test. A p-value of less than 0.01 was considered as significant. RESULTS: We found that 250, 1000 and 3000 µg/ml of Matricaria recutita extract could reduce expression of ERG9 gene significantly (p<0.01). Interestingly, the expression of this gene was completely inhibited in 1000 and 3000 µg/ml concentrations. CONCLUSION: This study predicted that Matricaria recutita extract produced anti-cancer effects in humans, because it could inhibit the expression of an analogue key gene in this malignant disease. Further investigations should be made, to study its molecular mechanism of action at the mammal cell level.

ZAP70 Expression Within del6q21, del11q13 and del17p13 Cytogenetic Subgroups of Iranian Patients with Chronic Lymphocytic Leukemia.

BACKGROUND: B-cell chronic lymphocytic leukemia (B-CLL) is the most common form of leukemia in adults. Some reports showed that expression of ZAP70 gene and chromosomal abnormality are two prognostic factors in management of B-CLL. OBJECTIVES: In this study, we determined ZAP70 mRNA expression level in the del17p13, del6q21 and del11q13 subgroups of Iranian B-CLL patients to investigate prognostic value of ZAP70 expression. PATIENTS AND METHODS: In this cross-sectional study, fluorescence in situ hybridization analysis was carried out on 66 Iranian B-CLL patients. Zap70 mRNA expression was evaluated by using Real Time RT-PCR. RESULTS: Molecular analysis showed that ZAP70 expression increased 2.46 fold in the del11q13 subgroup, 2.87 fold in the del17p13 and 1.87 fold in the del6q21, compared to the 15 patients in the control group. Comparison of standard deviation and mean of the ZAP70 expression profile within the subgroups showed more variability among the cases of the del11q13 and del17p13 versus tight clustering for the del6q21. Therefore, there is a relation between del6q21 aberrations; which has good prognosis with normal levels of ZAP70 expression. CONCLUSIONS: The results of ANOVA test showed that ZAP70 expression gene was significantly increased in del17p13 and del11q13 subgroups compared to control group. Thus, ZAP70 may play an important role in the prognosis of B-CLL patients.

FISH Analysis for del6q21 and del17p13 in B-cell Chronic Lymphocytic Leukemia in Iranians.

BACKGROUND: B-cell chronic lymphocytic leukemia (B-CLL) is the most common leukemia in the Western world. Major progress has been made in assessing typical chromosomal abnormalities and recognition of the correlation of these chromosomal abnormalities with laboratory features and clinical course of the disease. The most frequent genomic changes are deletions at 13q14, 11q22-23 and 17p13 and trisomy of chromosome 12. OBJECTIVES: The aim of this study was to investigate the frequency of chromosomal aberrations in B-CLL patients' peripheral blood and/or bone marrow using a molecular cytogenetic method, interphase fluorescence in situ hybridization (I-FISH) and to evaluate the correlation between these genomic changes and clinical findings. PATIENTS AND METHODS: I-FISH analyses were performed on bone marrow and blood samples of 66 B-CLL patients. RESULTS: Deletion of 17p13 was found in 11 (16.6%) and deletion 6q21 was present in 5 (7.5%). Statistical analyses were performed to investigate the correlation of these molecular-cytogenetic findings with family history, Rai staging and CD38 marker. No clear differences in distribution was noted for del17p13 and del6q21 among patients with and without family history, and no direct correlation was noted between these genomic changes and CD38 marker, but the correlation of del17p13 and Rai stage was significant. There was a high frequency of Rai stage II within del17p13 patients. CONCLUSIONS: It was demonstrated that the presence of del6q21 in B-CLL patients indicates poor prognosis and on the contrary, presence of del17p13 points at the good prognostic value of the disease.

WITHDRAWN: Chromosomal abnormalities and prognostic markers of B-cell chronic lymphocytic leukemia in Iranian patients.

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