[Bioinformatic evaluations for locating the microRNA suppressing PI3K/AKT pathway and analysis in prostate cancer cell lines]

Objective: Prostate cancer is the fifth most common cancer. In 2012, it was the second leading cause of cancer death for men worldwide. The PI3K/AKT pathway plays an essential role in pathogenesis of prostate cancer; the key role of this pathway in cancer progression makes it an attractive target for prostate cancer therapy. MicroRNAs [miRNAs] that regulate gene expression have a special ability to simultaneously control multiple genes and pathways which make them candidates for therapeutics. This study aims to determine miRNAs which target the PI3K/AKT pathway and evaluate them in prostate cancer cell lines

Methods: In order to determine an effective miRNA for the PI3K/AKT pathway, we assessed six genes from this pathway which have been proposed as drug targets in ten different prediction algorithms. Next, the candidate miRNAs were analyzed in expression profile and pathway analysis databases. Expression of candidate miRNAs in control and prostate cancer cell lines were subsequently evaluated

Results: According to bioinformatics, the miR-29 family could target the most genes from this list. Other bioinformatic estimates confirmed these results. The miR-29 family showed significant downregulation in prostate cancer cell lines LNCAP, PC3 and DU-145 compared to control samples

Conclusion: These results propose the possibility of using the miR-29 family to inhibit the PI3K/AKT pathway in prostate cancer

Expression and Purification of Recombinant ROP1 of Toxoplasma gondii in Bacteria

Toxoplasmosis is a worldwide-distributed infection which is mostly asymptomatic but can cause serious health problems in congenitally-infected newborns and immunecompromised individuals. Research is under- going both to improve toxoplasma serological tests, which play the main role in laboratory diagnosis of the infection, and develop an effective vaccine to prevent the infection. Some studies showed usefulness of rhoptry protein 1 [ROP1] antigen of Toxoplasma gondii [T. gondii] in serodiagnosis of the infection and induction of protective immunity. The purpose of this study was to produce recombinant ROP1 and evaluate its antigenicity against human in-fected sera. DNA encoding ROP1, amino acids 171 to 574, was obtained from T. goncff/RH strain by polymerase chain reaction amplification and cloned in probaryotic expression plasmid pET-15b. rROPl was expressed in Escherichia coli [E coli] and purified in a single step by immobilized metal ion affinity chromatography. DNA sequencing showed 99% similarity between the cloned sequence and the corresponding sequence in Gene bank. Results indicated the proper antigenicity of rROPl. Sera from Toxoplasma infected individuals specifically recognized rROPl in Western blotting. rROPl is antigenic toward human infected sera and can be used in studies for development of both a Toxoplasma serological test and a protective vaccine

Development of a high-resolution melting method for screening R188h polymorphism in XRCC2 gene

The High Resolution Melting [HRM] method is a new scanning method for detecting unknown changes in DNA and its advantages have persuaded researchers to recruit it as a screening method. Here, we developed a HRM method to screen R188H SNP [rs3218536] of XRCC2 and compared the results with a well known PCR-RFLP technique. Genomic DNA samples from 350 healthy individuals were obtained. PCR-HRM analysis and PCR-RFLP method were performed simultaneously. Three different melting profiles corresponding to three different genotypes recognized by HRM analysis. The results of PCR-RFLP showed no discrepancy. We concluded that the HRM technique can be used as a screening method for rapid discrimination of R188H genotypes in XRCC2 gene.

Analysis of relation between C677T genotype in MTHFR gene and prostatic cancer in Iranian males

Methylenetetrahydrofolate reductase [MTHFR] enzyme is one of the most important enzymes with a pivotal role in the folate metabolism and DNA synthesis pathways. Single nucleotide polymorphism [SNPs] in the coding gene has been related to many medical diseases as well as diverse malignancies including the prostate cancer which is the leading cause of the cancer deaths in men and one of the major public health problems. The goal of this study is to determine the relationship between the MTHFR C677T SNP and the prostate adenocarcinoma in Iranian males attending to the Labbafi-nezhad hospital in Tehran. In this Case-control unmatched study, 67 and 75 paraffinized tissue samples were taken out of the specimens diagnosed previously as the prostatic adenocarcinoma and nodular prostatic hyperplasia for the case and control groups respectively. MTHFR C677T genotyping was done by the use of multiplex ARMS-PCR and frequencies of the alleles were compared between the case and control groups as well as calculating the deviation from Hardy-Weinberg equilibrium and Odds Ratio for the "T" allele regarding the prostatic carcinoma. The observed rates in the control group were not too different from that of expected from Hardy- Weinberg equilibrium [P=0.407]. Frequencies of the possible genotypes were as follows: CC, 43.28% vs. 42.67%; CT, 49.25% vs. 52% and CT, 7.46% vs. 5.33% in the case and control groups respectively [P=0.85]. 1.37 times increased risk was found for the homozygote carriers of C677T variant [OR: 1.37, 95% CI: 0.33- 5.6; P=0.653] which is however statistically not significant. No association has been evident between the MTHFR 677C>T polymorphism and the risk of prostatic carcinoma in this study confirming the findings of some of the previous attempts; however, [OR: 1.37, 95% CI: 0.33-5.6] implies a slight effect of the homozygote on the carcinogenesis. Thus larger studies especially with a greater number of the smaples are recommended

role of Ile3434Thr XRCC7 gene polymorphism in differentiated thyroid cancer risk in an Iranian population

The aim of this study was to understand any association between differentiated thyroid carcinoma [DTC] and Ile3434Thr XRCC7 gene polymorphism [GenBank accession number: rs7830743]. DTC is the most prevalent thyroid neoplasm, which includes papillary and follicular cell carcinoma. XRCC7 gene encodes a protein that functions in non-homologous end joining DNA repair pathway. Non-synonymous polymorphisms in this gene may alter DNA repair capacity of the cell and change the risk of developing cancers. DTC patients [n = 173] and cancer free individuals [n = 204] were enrolled in a case-control study. The Ile3434Thr polymorphic alleles were discriminated by using amplification refractory mutation system-PCR method. The frequencies of this single nucleotide polymorphism in case and control groups were compared. Also, risk ratio for developing DTC in dichotomized genotypes was estimated by multivariate logistic regression analysis. Dichotomized genotypes into those with and without the 3434Thr allele showed that this allele was associated with DTC [OR [odd ratio]: 1.89, 95% confidence interval [CI] = 1.29-2.79, P<0.001]. Also, TC genotype was significantly associated with increased risk of DTC [OR: 2.42, 95% CI = 1.55-3.81, P = 0.0001] in individuals carrying this genotype. Allele 3434Thr in XRCC7 gene might be associated with differentiated thyroid cancer risk. Further studies with larger samples are needed to verify these initial findings

Association of Arg194Trp, Arg280His and Arg399Gln polymorphisms in x-ray repair cross-complementing group 1 gene and risk of differentiated thyroid carcinoma in Iran

X-ray repair cross-complementing group 1 [XRCC1] gene is a DNA repair gene and its non-synonymous single nucleotide polymorphisms [SNP] may influence DNA repair capacity which has been considered as a modifying risk factor for cancer development. A case-control study was conducted to investigate impact of three frequently studied polymorphisms [Arg194Trp, Arg280His and Arg399Gln] on developing differentiated thyroid carcinoma [DTC]. Increased risks for DTC were shown in homozygous [odds ratio [OR]: 3.66, 95% confidence interval [CI]: 0.38-35.60] and in dominant trait [OR: 1.22, 95% CI: 1.64-2.32] of Arg194Trp genotype. Also, for Arg280His genotype, an increased risk for DTC was shown in dominant trait [OR: 1.42, 95% confidence interval [CI]: 0.76-2.68], while a mildly reduction of risk for DTC [OR: 0.77, 95% [CI]: 0.50-1.17] was estimated in dominant Gln genotype of Arg399Gln. Considering combinatory effects of Arg194Trp and Arg280His genotypes on DTC, the calculated OR and 95% CI for being heterozygous for one of Arg194Trp or Arg280His genotypes were 1.57 and 0.90-2.74, respectively. Genotyping of codons 194, 280 and 399 in XRCC1 gene may use in risk assessment of DTC

Radioiodine therapy for hyperthyroidism

Radioiodine therapy is the safest, simplest, least expensive and most effective method for treatment of hyperthyroidism. The method employed in this research was a systematic bibliographic review, in which only valid studies or the clinically detailed enough open-labeled studies using validated scales were used. Iodine-131 [I-131] acts by the destructive effect of short-range beta radiation on thyroid cells. Indications for radioiodine therapy include toxic nodules [in which I-131 is the first choice of treatment], recurrent hyperthyroidism after antithyroid treatment or surgery, intolerance to antithyroid therapy due to side-effects and patient preference. Due to difficulties in previous methods for dose determination, fixed dose method of I-131 is now considered the best practical method for radioiodine therapy in primary hyperthyroidism. Absolute contraindications for radioiodine treatment are pregnancy and lactation. In pediatric patients, radioiodine therapy can be used, but is mainly considered in recurrent toxic goiter and when antithyroid medication is ineffective. There is no clear evidence indicative of carcinogenic or teratogenic effect of this agent

[Optimization and comparison of the poly fect gene delivery method in three different kinds of mesenchymal stem cell types]

The aim of this study was optimization of the PolyFect gene delivery method of pcDNA3.1 expression vector transfected with the mouse pdx-1 gene in three different kinds of mesenchymal stem cells and Hepa cells as well as comparison of transfection efficiency leading to expression of the mentioned gene in the cell types used. Rat bone marrow-derived mesenchymal stem cells, C57 mouse bone marrow-derived mesenchymal stem cells, human synovium derived mesenchymal stem cells and Hepa cells were used in this study. After culturing of the mentioned cells, mouse pdx-1 gene were transfected into them using the Qiagen PolyFect kit. 72 hours later, the cells were treated with anti-mouse Pdx-1 antibody and immunocytochemically analyzed using a fluorescent inverted microscope. Transfection conditions were optimized in each of these cells by changing different lipofection parameters such as DNA concentration, PolyFect reagent concentration and cell density. The results demonstrated that for transfection of these cells, the best concentrations of DNA and PolyFect reagent are 400 ng/IL and 6000 ng/IL respectively. For maximum transfection efficiency, the best cell density in 12-well plates was 105 cells in Hepa cells, 1.3105 cells in rat bone marrow-derived mesenchymal stem cells, 1.5105 cells in human synovium-derived mesenchymal stem cells and 105 cells in C57 mouse bone marrow-derived mesenchymal stem cells. Under the mentioned optimized conditions, the maximum efficiency of transfection was determined to be 50% for Hepa cells, 40% for rat bone marrow-derived mesenchymal stem cells, 21% for human synovium-derived mesenchymal stem cells and 10% for C57 mouse bone marrow-derived mesenchymal stem cells. These findings implicate that the most important factor extremely influencing transfection efficiency in mesenchymal stem cells is the cell derivation origin. Results of this study can be used in basic and clinical studies dealing with gene therapy in mesenchymal stem cells