Gene network reveals LASP1, TUBA1C, and S100A6 are likely playing regulatory roles in multiple sclerosis.

Introduction: Multiple sclerosis (MS), a non-contagious and chronic disease of the central nervous system, is an unpredictable and indirectly inherited disease affecting different people in different ways. Using Omics platforms genomics, transcriptomics, proteomics, epigenomics, interactomics, and metabolomics database, it is now possible to construct sound systems biology models to extract full knowledge of the MS and recognize the pathway to uncover the personalized therapeutic tools. Methods: In this study, we used several Bayesian Networks in order to find the transcriptional gene regulation networks that drive MS disease. We used a set of BN algorithms using the R add-on package bnlearn. The BN results underwent further downstream analysis and were validated using a wide range of Cytoscape algorithms, web based computational tools and qPCR amplification of blood samples from 56 MS patients and 44 healthy controls. The results were semantically integrated to improve understanding of the complex molecular architecture underlying MS, distinguishing distinct metabolic pathways and providing a valuable foundation for the discovery of involved genes and possibly new treatments. Results: Results show that the LASP1, TUBA1C, and S100A6 genes were most likely playing a biological role in MS development. Results from qPCR showed a significant increase (P < 0.05) in LASP1 and S100A6 gene expression levels in MS patients compared to that in controls. However, a significant down regulation of TUBA1C gene was observed in the same comparison. Conclusion: This study provides potential diagnostic and therapeutic biomarkers for enhanced understanding of gene regulation underlying MS.

LINC00467: A key oncogenic long non-coding RNA.

The significance of long non-coding RNAs (lncRNAs) in the development and progression of human cancers has attracted increasing attention in recent years of investigations. Having versatile interactions and diverse functions, lncRNAs can act as oncogenes or tumor-suppressors to actively regulate cell proliferation, survival, stemness, drug resistance, invasion and metastasis. LINC00467, an oncogenic member of long intergenic non-coding RNAs, is upregulated in numerous malignancies and its high expression is often related to poor clinicopathological features. LINC00467 facilitates the progression of cancer via sponging tumor-suppressive microRNAs, inhibiting cell death cascade, modulating cell cycle controllers, and regulating signalling pathways including AKT, STAT3, NF-κB and Wnt/ß-catenin. A growing number of studies have revealed that LINC00467 may serve as a novel prognostic biomarker and its inhibitory targeting has a valuable therapeutic potential to suppress the malignant phenotypes of cancer cells. In the present review, we discuss the importance of LINC00467 and provide a comprehensive collection of its functions and molecular mechanisms in a variety of cancer types.

Overexpression of hsa-miR-30a-5p and non-obstructive azoospermia: A case-control study.

Background: Some previous human and animal studies have supported the idea that KDM3A down-regulation might be the main cause of male infertility, especially in non-obstructive azoospermia (NOA). The regulatory role of micro-RNAs (miRNA) has been investigated in the development of male infertility. Objective: The expression level of hsa-miR-30a-5p in azoospermia was evaluated to reveal its possible association with the etiology of male infertility. Materials and Methods: In this case-control study, 30 men with azoospermia (19 of whom had NOA) were selected as the case individuals, and 11 men with obstructive azoospermia (OA) were selected as control individuals. The best miRNA with the strongest ability to target the KDM3A gene was detected via comprehensive bioinformatics analysis. Reverse transcriptase quantitative polymerase chain reaction was used to assess the expression level of hsa-miR-30a-5p. After analyzing the data, the expression level of hsa-miR-30a-5p wascompared between men with NOA and men with OA. Results: The findings supported the idea that hsa-miR-30a-5p is the miRNA with the best ability to target the KDM3A transcript. The expression analysis of hsa-miR-30a-5p indicated a significant overexpression (p = 0.04) in men with NOA compared to in men with OA. Conclusion: Hsa-miR-30a-5p was overexpressed in men with NOA compared to in control individuals. Hsa-miR-30a-5p could target the KDM3A transcript and may suppress its expression.

Rosmarinic acid inhibits DNA glycation and modulates the expression of Akt1 and Akt3 partially in the hippocampus of diabetic rats.

Non-enzymatic glycation of DNA and the associated effects are among pathogenic factors in diabetes mellitus. Natural polyphenols have anti-diabetic activity. Herein, the protective role of one of the phytochemicals, rosmarinic acid (RA), was evaluated in glycation (with fructose) of human DNA and expression of Akt genes in the hippocampus of diabetic rats. In-vitro studies using fluorescence, agarose gel electrophoresis, fluorescence microscopy, and thermal denaturation analyses revealed that glycation causes DNA damage and that RA inhibits it. In-vivo studies were performed by induction of diabetes in rats using streptozotocin. The diabetic rats were given RA daily through gavage feeding. The expression of Akt genes (inhibitors of apoptosis) in the hippocampus was evaluated using RT-qPCR. In diabetic rats, Akt1 and Akt3 were significantly down-regulated compared to the control group. Treating the diabetic rats with RA returned the expression of Akt1 and Akt3 relatively to the normal condition. Past studies have shown that diabetes induces apoptosis in the hippocampal neurons. Given that glycation changes the genes expression and causes cell death, apoptosis of the hippocampal neurons can be due to the glycation of DNA. The results also suggest that RA has reliable potency against the gross modification of DNA under hyperglycemic conditions.

Identification of Novel miRNAs in the F8 Gene Via Bioinformatics Tools.

BACKGROUND: Hemophilia A is an X-linked bleeding disorder resulting in a deficiency of plasma clotting factor VIII and caused by mutations in the FVIII gene (F8 gene). MicroRNAs (miRNAs) in body fluids are promising biomarker candidates for Hemophilia A, due to their stability in body fluids and accessibility by non- or minimally-invasive procedures. Therefore; Advances in miRNA analysis methods resulted in a wide range of publications on miRNAs as putative biomarkers. OBJECTIVE: Here we tried to scan the F8 gene region to predict a novel miRNA and identify it as a regulator of the F8 gene. MATERIALS AND METHODS: To this aim, the ability to express novel miRNAs in F8 locus was assessed via reliable bioinformatics databases such as SSCprofiler, RNAfold, miREval, FOMmiR, MaturBayes, miRFIND, UCSC genome browser, Deep Sequencing, and miRBase. RESULTS: Data analysis from the relevant databases offers one stem-loop structure that is predicted to express a novel miRNA. CONCLUSIONS: The diagnosis of Hemophilia A with the help of these types of biomarkers is a non-invasive procedure that has been demonstrated to have a significant role in the early diagnosis of the disease. Hopefully, the proposed candidate sequence will be confirmed in vitro and become a non-invasive biomarker in the near future.

Experimental validation of a predicted microRNA within human FVIII gene.

Hemophilia A is an X-linked bleeding disorder that occurs due to the deficiency of Factor VIII (FVIII) protein clotting activity. The mutations in the F8 gene, which encodes FVIII coagulating protein have been widely reviewed. However, there is a wide range of criteria that in addition to F8 gene mutations, different molecular mechanisms may be associated with hemophilia A. Various functions of FVIII could be related to the hypothetical small non-coding RNAs, located within the F8 gene sequence. Therefore, miRNAs that can post-transcriptionally regulate gene expression might confer susceptibility to developing hemophilia A. Here, we have selected a bioinformatically predicted hairpin structure sequence in the first intron of the F8 gene that has the potential to produce a real miRNA (named put-miR1). We tried to experimentally detect the predicted miRNA via RT-PCR following its precursor overexpression in HEK 293 cell lines. Despite the accuracy of miRNA prediction, according to the reliable bioinformatics studies, we couldn't confirm the existence of considered mature miRNA in transfected cells. We hope that through changing experimental conditions, designing new primers, or altering cell lines and expression vectors, the exogenous and endogenous expression of the predicted miRNA will be confirmed.

MicroRNA and Hemophilia-A Disease: Bioinformatics Prediction and Experimental Analysis.

OBJECTIVE: Hemophilia-A is a common genetic abnormality resulted from decreased or lack of factor VIII (FVIII) pro-coagulant protein function caused by mutations in the F8 gene. Majority of molecular studies consider screening of mutations and their relevant impacts on the quality and expression levels of FVIII. Interestingly, some of the functions in FVIII suggest a probable involvement of small non-coding RNAs embedded within the sequence of F8 gene. Therefore, microRNAs which are encoded within the F8 gene might have a role in hemophilia development. In this study, miRNAs production in the F8 gene was investigated by bioinformatics prediction and experimental validation. MATERIALS AND METHODS: In this experimental study, bioinformatics tools have been utilized to seek the novel microRNAs inserted within human F8 gene. The ability to express new microRNAs in F8 locus was studied through reliable bioinformatics databases such as SSCProfiler, RNA fold, miREval, miR-FIND, UCSC genome browser and miRBase. Then, expression and processing of the predicted microRNAs were examined based on bioinformatics methods, in the HEK293 cell lines. RESULTS: We are unable to confirm existence of the considered mature microRNAs in the transfected cells. CONCLUSION: We hope that through changing experimental conditions, designing new primers or altering cell lines as well as the expression of vectors, exogenous and endogenous expressions of the predicted miRNA will be confirmed.

A Novel CAR Expressing NK Cell Targeting CD25 With the Prospect of Overcoming Immune Escape Mechanism in Cancers.

For many years, high-affinity subunit of IL-2 receptor (CD25) has been considered as a promising therapeutic target for different pathologic conditions like allograft rejection, autoimmunity, and cancers. Although CD25 is transiently expressed by newly-activated T cells, it is the hallmark of regulatory T (Treg) cells which are the most important immunosuppressive elements in tumor microenvironment. Thus, Tregs can be considered as a potential target for chimeric antigen receptor (CAR)-based therapeutic approaches. On the other hand, due to some profound adverse effects pertaining to the use of CAR T cells, CAR NK cells have caught researchers' attention as a safer choice. Based on these, the aim of this study was to design and develop a CAR NK cell against CD25 as the most prominent biomarker of Tregs with the prospect of overcoming immune escape mechanism in solid and liquid cancers. In the current study, an anti-CD25 CAR was designed and evaluated by comprehensive in silico analyses. Then, using lentiviral transduction system, NK-92 cell line was engineered to express this anti-CD25 CAR construct. In vitro functional analyses of anti-CD25 CAR for its reactivity against CD25 antigen as well as for cytotoxicity and cytokine production assays against CD25 bearing Jurkat cell line were done. In silico analyses demonstrated that the anti-CD25 CAR transcript and scFv protein structures were stable and had proper interaction with the target. Also, in vitro analyses showed that the anti-CD25 CAR-engineered NK-92 cells were able to specifically detect and lyse target cells with an appropriate cytokine production and cytotoxic activity. To conclude, the results showed that this novel CAR NK cell is functional and warrant further investigations.

Computational study for suppression of CD25/IL-2 interaction.

Cancer recurrence presents a huge challenge in cancer patient management. Immune escape is a key mechanism of cancer progression and metastatic dissemination. CD25 is expressed in regulatory T (Treg) cells including tumor-infiltrating Treg cells (TI-Tregs). These cells specially activate and reinforce immune escape mechanism of cancers. The suppression of CD25/IL-2 interaction would be useful against Treg cells activation and ultimately immune escape of cancer. Here, software, web servers and databases were used, at which in silico designed small interfering RNAs (siRNAs), de novo designed peptides and virtual screened small molecules against CD25 were introduced for the prospect of eliminating cancer immune escape and obtaining successful treatment. We obtained siRNAs with low off-target effects. Further, small molecules based on the binding homology search in ligand and receptor similarity were introduced. Finally, the critical amino acids on CD25 were targeted by a de novo designed peptide with disulfide bond. Hence we introduced computational-based antagonists to lay a foundation for further in vitro and in vivo studies.

Hsa-miR-27a-3p overexpression in men with nonobstructive azoospermia: A case-control study.

BACKGROUND: The role of KDM3A and its downstream genes in male fertility has been approved in animal models. Additionally, the expression shrinkage of KDM3A is significantly correlated with human azoospermia phenotype. Aberrant expression of micro-RNAs could mislead spermatogenesis and mostly lead to diverse phenotypes of male infertility. OBJECTIVE: The aim of this study was to evaluate the expression level of hsa-miR-27a-3p in azoospermic men to reveal its possible association with infertility. MATERIALS AND METHODS: This case-control study was conducted on 30 azoospermic men, of whom, 19 had non obstructive azoospermia (NOA) and 11 obstructive azoospermia (OA) according to the pathological examinations. Comprehensive bioinformatics investigations were performed securely and hsa-miR-27a-3p was selected afterward. Reverse Transcriptase-quantitative polymerase chain reaction (RT-qPCR) method was used and statistical analysis was performed to compare the expression level of hsa-miR-27a-3p in both OA and NOA individuals. RESULTS: In silico analysis suggested hsa-miR-27a-3p, with its potential binding ability to target KDM3A transcripts. The expression analysis of candidate hsa-miR-27a-3p indicated its significant overexpression in NOA men. CONCLUSION: The hsa-miR-27a-3p was overexpressed in NOA men compared to OA-control individuals. As a consequence, the overexpressed micro-RNA could downregulate directly KDM3A and indirectly TNP1 and PRM1. Therefore, spermatogenesis could be misled and male infertility could be developed.

Correction: A compound downregulation of SRRM2 and miR-27a-3p with upregulation of miR-27b-3p in PBMCs of Parkinson's patients is associated with the early stage onset of disease.

[This corrects the article DOI: 10.1371/journal.pone.0240855.].

A compound downregulation of SRRM2 and miR-27a-3p with upregulation of miR-27b-3p in PBMCs of Parkinson's patients is associated with the early stage onset of disease.

Parkinson's disease (PD) is diagnosed when motor symptoms emerges, which almost 70% of dopamine neurons are lost. Therefore, early diagnosis of PD is crucial to prevent the progress of disease. Blood-based biomarkers, which are minimally invasive, potentially used for diagnosis of PD, including miRNAs. The aim of this study was to assess whether SRRM2 and miR-27a/b-3p could act as early diagnostic biomarkers for PD. Total RNAs from PBMCs of 30 PD's patients and 14 healthy age and gender matched subjects was extracted. The expression levels of respective genes were assessed. Data were presented applying a two-tailed unpaired t-test and one-way ANOVA. We observed significant down-regulation of SRRM2 (p = 0.0002) and miR-27a-3p (p = 0.0001), and up-regulation of miR-27b-3p (p = 0.02) in PBMCs of Parkinson's patients. Down-regulation of miR-27a-3p is associated with increasing disease severity, whereas the up-regulation of miR-27b-3p was observed mostly at HY-1 and disease duration between 3-5 years. There was a negative correlation between SRRM2 and miR-27b-3p expressions, and miR-27a-3p positively was correlated with miR-27b-3p. Based on functional enrichment analysis, SRRM2 and miR-27a/b-3p acted on common functional pathways. miR-27a/b-3p could potentially predict the progression and severity of PD. Although both miRs had no similarity on expression, a positive correlation between both miRs was identified, supporting their potential role as biomarkers in clinical PD stages. Of note that SRRM2 and miR-27a-3p were able to distinguish PD patients from healthy individuals. Functional analysis of the similarity between genes associated with SRRM2 and miR-27a/b-3p indicates common functional pathways and their dysfunction correlates with molecular etiopathology mechanisms of PD onset.

Factor VIII: Perspectives on Immunogenicity and Tolerogenic Strategies for Hemophilia A Patients.

A major complication in treating hemophilia A is the development of neutralizing antibodies (inhibitors) against therapeutic administered factor VIII (FVIII), which occurs in approximately 20-30% of patients with severe disease. These inhibitors render FVIII replacement therapy ineffective and increase the morbidity and mortality risk. The currently accepted method to eradicate inhibitors is immune tolerance induction (ITI), and frequent intensive administration of FVIII until inhibitor titers drop. Current ITI protocols are extremely costly and not effective in all patients. During the last decade, many types of research have been accomplished to clarify the mechanisms that mediate immune tolerance induction. Novel experimental therapies including monoclonal antibodies, viral vector-mediated gene therapy, regulatory T cell induction using immunosuppressive drugs, and nanoparticle-based immune modulation show promising results in hemophilia A clinical trials. This review focuses on treatment options towards the anti-FVIII immune responses and current novel therapies in clinical trials.

Association between Different Polymorphic Markers and ß-Thalassemia Intermedia in Central Iran.

ß-Thalassemia intermedia (ß-TI) is a clinical condition characterized by moderate, non transfusional anemia and hepatosplenomegaly. The main objective of this study was to determine the molecular basis of the clinical phenotype of ß-TI in Iran. To elucidate the mild phenotype of many patients with ß-TI, we screened for three prevalent ß-globin gene mutations [IVS-II-1 (G>A) HBB: c.315+1G>A, IVS-I-110 (G>A) HBB: c.93-21G>A and IVS-I-5 (G>C) [HBB: c.92+5G>C], deletions on the α-globin genes, XmnI polymorphisms and restriction fragment length polymorphism (RFLP) haplotypes on the ß-globin gene cluster in 50 ß-TI patients. Fifty-eight percent of the patients (29 cases) were associated with the mentioned mutations. We showed that the HBB: c.315+1G>A mutation is linked to haplotype [+ - + +] (57.69%). This haplotype is in linkage disequilibrium with the XmnI polymorphism (NG_000007.3: g.42677C>T) and has been associated with increased expression of Hb F in ß-TI patients. The XmnI polymorphism is defined in association with this prevalent mutation. Two patients had a single α-globin gene deletion [-α3.7 (rightward) deletion]. The main genetic factor in mild phenotype ß-TI patients is the linkage of an XmnI polymorphism (NG_000007.3: g.42677C>T) with the HBB: c.315+1G>A (80.76%), which is associated with increased production of Hb F and coinheritance of haplotype [+ - + +] with ß-TI, especially with the homozygous HBB: c.315+1G>A mutation. Molecular basis of ß-TI could be explained by the involvement of different factors that tend to develop the disease phenotype.

Integral membrane protein expression of human CD25 on the cell surface of HEK293 cell line: the available cellular model of CD25 positive to facilitate in vitro developing assays.

Typically, CD25 is expressed on the cellular surface of regulatory T (Treg) cells. These cells are significant in regulating the self-tolerance and also preventing the immune system from attacking a person's own tissues and cells. They promote the cancer progression by playing an important role in evading the immune system. Thus, the experimental procedures was aimed to clone and express human CD25 in HEK293 cell line, as the available cellular model, for the purpose of developing assays to facilitate and enhance the studies on an available CD25 positive cell. The secondary RNA structure of CD25 was evaluated by in silico analysis. Then, cDNA of human CD25 were synthesized from isolated total mRNA of cultured and stimulated PBMCs from blood donors. After cloning the cDNA of CD25 into a pcDNA3.1(+) plasmid, using the effective transfection of the recombinant pcDNA3.1(+) in HEK293, qRT-PCR and flow cytometry methods were used to quantitatively evaluate CD25 transcripts and protein level. There was a 4.8 fold increase in transcripts and a 76.2% increase in protein levels of CD25 when comparing the transfected and control cell lines. The genetically engineered HEK293 cell line expressing Treg cell surface marker of CD25 was introduced in this study for the first time. This cell line can be used to overcome the problematic issues for studying Treg cells including low population of Tregs in peripheral blood, low recovery methods for Treg isolation, time-consuming and non-cost benefit methods in the conditions of in vitro cell culture experiments for the studies focused on the binding of IL-2 to CD25.

MiR-26a and miR-26b downregulate the expression of sucrase-isomaltase enzyme: A new chapter in diabetes treatment.

Type II diabetes is a metabolic disease that has affected 460 million people around the globe and become a heavy burden on health care system. Diabetic patients suffer from hyperglycemia and hyperinsulinemia which can damage vital organs in body like heart, kidneys, eyes and nervous system. Different strategies have been introduced to control or lessen these diabetic complications in which one of the most promising approaches is the inhibition of intestinal sucrase-isomaltase (SI). Inhibition of this enzyme will block the release of glucose into bloodstream and lead to reduced postprandial hyperglycemia. MicroRNAs are small regulatory molecules that play critical roles in different cellular pathways and molecular mechanisms. It is proved that microRNAs have significant effects on cellular mechanisms involved in diabetes and can be used as biomarkers for diagnosis of this metabolic disease. Based on bioinformatics analysis miR-26a and miR-26b can interact with a conserved 3'-UTR region of SI mRNA which lead to a hypothesis that these miRs may have negative regulatory effect on this enzyme. In this study, we investigated the impact of high glucose conditions on expression of sucrase-isomaltase, miR-26a and miR-26b in caco-2 cell line. It is proved that in a simulated diabetic condition there is a reverse correlation between the expression pattern of these miRs and SI. QRT-PCR method was used to evaluate the expression of our target molecules. Interestingly, transfection of miR-26a and miR-26b in caco-2 cell line reduced the transcription of SI mRNA and decreased the sucrase and maltase activity of its active sites. To sum up, our results demonstrate the first evidence of the significant effect of miR-26a and miR-26b on SI expression and activity. We proved that these microRNAs may directly inhibit this enzyme and can be used as a new scaffold in search of finding novel treatments for type II diabetes.

Investigation of RFLP Haplotypes ß-Globin Gene Cluster in Beta-Thalassemia Patients in Central Iran.

Introduction: Beta-thalassemia is one of the most prevalent inherited blood diseases among Iranians. The aim of this study was to elucidate the chromosomal background of beta-thalassemia mutations in Esfahan province, Iran. Materials and Methods: In this study, we investigated three frequent mutations (c.315+1G>A, c.93-21G>A and c.92+5G>C in ß-globin gene, the frequency of RFLP haplotypes, and LD between markers at ß-globin gene cluster) in 150 beta-thalassemia patients and 50 healthy individuals. The molecular and population genetic investigations were performed on RFLP markers HindIII in the c.315+1G>A of Gγ (HindIIIG) and Aγ (HindIIIA) genes, AvaII in the c.315+1G>A of ß-globin gene and BamHI 3' to the ß-globin gene. All statistical analyses were performed using Power Marker software and SISA server. Results: Fifty percent of beta-thalasemia patients were associated with these mutations. Haplotype I was the most prevalent haplotype among beta-thalassemia patients (39.33%) and normal individuals (46%). The commonest c.315+1G>A mutation in our population was tightly linked with haplotype III (43.75%) and haplotype I (31.25%). The second prevalent mutation, c.92+5G>C, was 90%, 6.66%, and 3.33% in linkage disequilibrium with haplotypes I, VII, and III, respectively. The c.93-21G>A mutation indicated a strong association with haplotype I (80%). Conclusion: Our study participants like beta-thalassemia patients from Kermanshah province was found to possess a similar haplotype background for common mutations. The emergence of most prevalent mutations on chromosomes with different haplotypes can be explained by gene conversion and recombination. High linkage of a mutation with specific haplotype is consistent with the hypothesis that chromosomes carrying beta-thalassemia mutations experienced positive selection pressure, probably because of the protection against malaria experienced by beta-thalassemia carriers.

Maltase-glucoamylase inhibition potency and cytotoxicity of pyrimidine-fused compounds: An in silico and in vitro approach.

The prevalence of diabetes mellitus has been incremented in the current century and the need for novel therapeutic compounds to treat this disease has been significantly increased. One of the most promising approaches is to inhibit intestinal alpha glucosidases. Based on our previous studies, four pyrimidine-fused heterocycles (PFH) were selected as they revealed satisfactory inhibitory action against mammalian α-glucosidase. The interaction of these compounds with both active domains of human maltase-glucoamylase (MGAM) and their effect on human Caco-2 cell line were investigated. The docking assessments suggested that binding properties of these ligands were almost similar to that of acarbose by establishing hydrogen bonds especially with Tyr1251 and Arg526 in both C-terminal and N-terminal MGAM, respectively. Also, these compounds indicated a stronger affinity for C-terminal of MGAM. L2 and L4 made tightly complexes with both terminals of MGAM which in turn revealed the importance of introducing pyrimidine scaffold and its hinge compartment. The results of molecular dynamics simulation analyses confirmed the docking data and showed deep penetration of L2 and L4 into the active site of MGAM. Based on cell cytotoxicity assessments, no significant cell death induction was observed. Hence, these functional MGAM inhibitors might be considered as new potential therapeutic compounds in treatment of diabetes and its complications.

Isothermal Amplification Methods for the SNP Genotyping.

The demands for genotyping techniques with acceptable precision, accuracy, cost-effectiveness in high throughput formats made driving forces for continuous development of novel technologies. A wide range of mutation detection techniques based on polymerase chain reaction (PCR) have been introduced. The best alternatives were the isothermal amplification technologies that those did not require a thermal cycler. In this review, we aimed to describe the most known isothermal amplification techniques for SNP genotyping.

Evaluation of miRNA-9 and miRNA-34a as potential biomarkers for diagnosis of breast cancer in Iranian women.

PURPOSE: MicroRNAs are involved in diverse biological processes and their dysregulation is a common event in various diseases including breast cancer. Breast cancer is a major threat to women's health. This study was designed to examine the expression levels of miR-9 and miR-34a in breast tumor tissue samples and plasma of breast cancer patients, compare their expression pattern between tissue samples and plasma samples of patients and analyze their relationship with tumor clinical features. Also, the potential of these miRNAs as diagnostic biomarkers for breast cancer was investigated. MATERIALS AND METHODS: The expression levels of miR-9, miR-34a and CDH1 were measured by real-time reverse transcription polymerase chain reaction and ΔΔct method. Data were analyzed using t-test and one-way ANOVA. The sensitivity and specificity of miRNAs were determined by receiver operating characteristic (ROC) curve. RESULTS AND DISCUSSION: The expression levels of miR-9 and miR-34a were significantly down-regulated in tumor tissues compared to healthy tissues (fold change = 0.26, p = 0.0051 for miR-9 and fold change = 0.55, p = 0.021 for miR-34a). While no significant difference was observed in the expression levels of miR-9 (p = 0.205) and miR-34a (p = 0.132) in plasma samples of patients compared to normal plasma. CDH1 expression in tumor tissue was not significantly different from normal tissue (p = 0.33). We found that expression level of miR-9 in patients with tumor size larger than 5 cm (p = 0.026) and expression level of miR-34a in patients with higher stage (lll & lV, p = 0.03) were significantly down-regulated. Also miR-34a expression level was positively correlated with patient's age (p = 0.03). CONCLUSION: According to the ROC curves, the area under the curve (AUC) of miR-9 in tissue was 0.71 (p = 0.009) with sensitivity 83.33% and specificity 70.37%. The AUC for miR-34a in tissue was 0.72 (p = 0.007) with sensitivity 72% and specificity 76%. Thus miR-9 and miR-34a have the capability for distinguishing tumor tissues from healthy tissues and the study of their expression levels in tissue may be used as a biomarker for the diagnosis of breast cancer patients from healthy women.