Evaluation of expression ratio of HOXB13:IL17BR in patients with breast cancer by qRT-PCR method using SYBR Green dye

Background and aims: Studies have shown that a HOXB13:IL17BR expression ratio index predicts clinical outcome in ER-positive, lymph node-negative breast cancer patients that treated with adjuvant tamoxifen. All of these experiments were conducted with qReal Time RT-PCR using TaqMan probes. The aim of this study was to determine the ratio using SYBR Green I qReal Time RT-PCR. Methods: In this case- control study, expression levels of HOXB13:IL17BR was measured in 40 paired formalin-fixed, paraffin-embedded primary breast tumor specimens. After extracting RNA from the tissues, cDNA synthesis and amplification with the polymerase chain reaction to obtain the optimum annealing temperature, the expression levels was measured by SYBR Green I qReal Time RT-PCR. To determine and normalize the expression levels, BestKeeper software was used to obtain the BestKeeper Index using the geometric mean of expression levels of housekeeping genes. Comparison of mean expression of genes between tumoral and non-tumoral tissues was performed by t-test and association between patient grouping (high/low risk) and time for disease free survival was assessed by Fisher`s Exact test. Results: HOXB13:IL17BR expression value did not show significant difference between tumoral and non-tumoral tissues. The results showed that there was a direct and significant association between patient grouping (based on HOXB13:IL17BR ratio) and disease free survival status. Conclusion: Results in the current study showed that in spite of using SYBR Green dye (instead of TaqMan probes), there is still a significant correlation between HOXB13:IL17BR ratio and disease free survival status

Hypothesis: A challenge of overexpression Zfp521 in neural tendency of derived dental pulp stem cells

Neurodegenerative diseases have now become a major challenge, especially in aged societies. Most of the traditional strategies used for treatment of these diseases are untargeted and have little efficiency. Developments in stem cell investigations have given much attention to cell therapy as an alternative concept in the regeneration of neural tissues. Dental pulp stem cells (DPSCs) can be readily obtained by noninvasive procedures and have been shown to possess properties similar to well-known mesenchymal stem cells. Furthermore, based on their neural crest origin, DPSCs are considered to have a good potential to differentiate into neural cells. Zfp521 is a transcription factor that regulates expression of many genes, including ones involved in the neural differentiation process. There for based on neural crest origin of the cell and high expression of neural progenitor markers, we speculate that sole overexpression of Zfp521 protein can facilitate differentiation of dental stem cells to neural cells and researchers may find these cells suitable for therapeutic treatment of neurodegenerative disease

SLUG transcription factor : a pro-survival and prognostic factor in gastrointestinal stromal tumour

Background: The SLUG transcription factor has been linked with the KIT signalling pathway that is important for gastrointestinal stromal tumour (GIST) tumourigenesis. Its clinical significance in GIST is unknown. Methods: Influence of SLUG expression on cell proliferation and viability were investigated in GIST48 and GIST882 cell lines. The association between tumour SLUG expression in immunohistochemistry and recurrence-free survival (RFS) was studied in two clinical GIST series, one with 187 patients treated with surgery alone, and another one with 313 patients treated with surgery and adjuvant imatinib. Results: SLUG downregulation inhibited cell proliferation, induced cell death in both cell lines, and sensitised GIST882 cells to lower imatinib concentrations. SLUG was expressed in 125 (25.0%) of the 500 clinical GISTs evaluated, and expression was associated with several factors linked with unfavourable prognosis. SLUG expression was associated with unfavourable RFS both when patients were treated with surgery alone (HR = 3.40, 95% CI = 1.67-6.89, P = 0.001) and when treated with surgery plus adjuvant imatinib (HR = 1.83, 95% CI = 1.29-2.60, P = 0.001). Conclusions: GIST patients with high tumour SLUG expression have unfavourable RFS. SLUG may mediate pro-survival signalling in GISTs.Peer reviewe

Novel roles for class II Phosphoinositide 3-Kinase C2 beta in signalling pathways involved in prostate cancer cell invasion

Phosphoinositide 3-kinases (PI3Ks) regulate several cellular functions such as proliferation, growth, survival and migration. The eight PI3K isoforms are grouped into three classes and the three enzymes belonging to the class II subfamily (PI3K-C2a, ß and ?) are the least investigated amongst all PI3Ks. Interest on these isoforms has been recently fuelled by the identification of specific physiological roles for class II PI3Ks and by accumulating evidence indicating their involvement in human diseases. While it is now established that these isoforms can regulate distinct cellular functions compared to other PI3Ks, there is still a limited understanding of the signalling pathways that can be specifically regulated by class II PI3Ks. Here we show that PI3K-C2ß regulates mitogen-activated protein kinase kinase (MEK1/2) and extracellular signal-regulated kinase (ERK1/2) activation in prostate cancer (PCa) cells. We further demonstrate that MEK/ERK and PI3K-C2ß are required for PCa cell invasion but not proliferation. In addition we show that PI3K-C2ß but not MEK/ERK regulates PCa cell migration as well as expression of the transcription factor Slug. These data identify novel signalling pathways specifically regulated by PI3K-C2ß and they further identify this enzyme as a key regulator of PCa cell migration and invasion

Comparison of TBX3 Gene Expression among Patients with Squamous Esophageal Cancer and Normal Individuals

Background & aim: Esophageal cancer is the eighth most prevalent cancer worldwide and the second in Iran. Annually, esophageal cancer makes up 3.2% of all new cancer cases. Altered expression of stem cells self-renewal genes is of great importance in the tumorigenesis. The aim of this study was to compare the TBX3 gene expression in tumoral samples of patients with squamous esophageal cancer and normal tissue samples of healthy individuals.   Methods: The present cross-sectional study was performed on 40 samples, including 20 samples of tumor esophageal cancer tissue and 20 samples of normal tissue samples from Imam Khomeini Hospital in Ahwaz, Iran. After extracting RNA from FFPE tissues and synthesizing cDNA, expression levels of TBX3 and GUSB (as internal control) genes were relatively quantified using qRT-PCR. Data were analyzed using t-test.   Results: The results of the qRT-PCR analyses showed that the average relative gene expression in tumor samples was higher than normal samples. Furthermore, TBX3 gene expression exhibited greater expression in the tumor samples of the patients in the 26-50 years age range in compared with the 51-81 years age range   Conclusion: According to the cancer stem cell theory, many tumors originated from stem cells leading to the inhibition of apoptosis and resistance to chemotherapy. Considering the role of TBX3 gene in self-renewal of stem cells and its increased expression in patients with squamous esophageal cancer, it seems that TBX3 gene might be used as a biomarker in the diagnosis and prognosis of esophageal cancer in future.   &nbsp

Molecular assessment and bioinformatic analysis of two common mutations of phenylalanine hydroxylase (PAH) gene by HRM

BACKGROUND AND OBJECTIVE: Phenylketonuria (PKU) is the most prevalent inborn error of amino acid metabolism in the world and its incidence rate is increasing in Iran. The aim of the present study was to assess the efficiency of HRM technique as a fast and suitable method in identifying common mutations of phenylalanine hydroxylase gene including IVS10-11G>A and P281L in order to improve the early detection of the disease to prevent the occurrence of it. METHODS: In this case-control study, 20 DNA samples including one sample with IVS10-11G>A mutation , one sample with P281L mutation and 18 control samples were extracted from peripheral blood collected in Medical Genetic Center of Isfahan and were genotyped with HRM technique. To validate the mutations, the mutant samples were genotyped using sequencing. Bioinformatic analyses were used for determining structural and functional effects of P281L mutation on the of PAH protein. FINDINGS: HRM analysis identified IVS10-11G>A and P281L mutations with a sensitivity and specificity of 100% and the mutant and normal samples differentiated well in normalized and difference plots. Bioinformatic analyses demonstrated instability and pathological effects of PAH protein containing P281L mutation. CONCLUSION: HRM is a simple and fast technique detecting the two IVS10-11G>A and P281L mutations with 100% sensitivity and specificity

Synthetic biology based on genetic logic circuit, using the expression of drug resistance, bcrp pump in mcf-7 cancer cell line

Biological circuits are developed as biological parts within a cell to carry out logical functions resembling those studied in electronics circuits. These circuits can be performed as a method to vary cellular functions, to develop cellular responses to environmental conditions, or to regulate cellular developments. This research explored the possibility of synthetic biology based on the genetic logic circuit A and (not B) using the inducible expression of the both BCRP drug resistance pump and its specific shRNA in MCF-7 cancer cell line utilizing the third generation of lentiviral vectors. The accuracy of the output of the proposed circuit for living cells, was confirmed by the results of the Real-Time PCR and flow cytometry at the RNA and protein levels. At the RNA level, the effect of the inducers on the BCRP gene expression and silencing were investigated by real-time PCR. Furthermore, at the protein level, induction of the expression of the BCRP pump resulted in driving out of the substrate from inside the cells leading to the decrease of the fluorescent emission from the transfected cells. We successfully designed and implemented the genetic logic circuit A and (not B) using the inducible expression of the both BCRP drug resistance pump and its specific shRNA in MCF-7 cancer cells. © 2020, Iranian Journal of Pharmaceutical Research. All rights reserved

Design and evaluation of biological gate circuits and their therapeutic applications in a model of multidrug resistant cancers

Objectives: Synthetic biology is primarily an emerging research field that consists of designing new synthetic gene circuits dedicated to targeted functions and therapies such as cancer therapy. In this study, a genetic logic NOT-IF gate is used to reduce the multidrug resistance and facilitate the malignant cancer therapy. MCF7 cancer cells were cultured in RPMI-1640 medium and transfected with lentiviral vectors including MDR1 gene and the corresponding shRNA against MDR1 with controllable promoters. Transcript levels and protein levels of MDR1 gene were quantified. Results: Our results showed that when doxycycline (DOX) and sodium butyrate were present and IPTG was absent, these led to a 74,354-fold increase in MDR1 gene expression. Upon IPTG treatment, the MDR1 gene expression was not detected due to the lack of the inducer. In addition, following IPTG induction in the presence of DOX and sodium butyrate and expressing shRNA, there was a 75 reduction in MDR1 gene expression compared to those cells treated only with sodium butyrate and DOX. Conclusions: We successfully designed and implemented the genetic logic NOT-IF gate at the transcriptional level using the inducible expression of both MDR1 drug resistance pump and its specific shRNA in MCF7 cancer cells, using the third generation lentiviral vectors. © 2020, Springer Nature B.V

Clinical and metabolic response to probiotic supplementation in patients with rheumatoid arthritis: a randomized, double-blind, placebo-controlled trial

Objective: This study was performed to determine the effects of probiotic supplementation on clinical and metabolic status of patients with rheumatoid arthritis (RA). Methods: Sixty patients with RA aged 25�70 years were assigned into two groups to receive either probiotic capsules (n = 30) or placebo (n = 30) in this randomized, double-blind, placebo-controlled trial. The patients in the probiotic group received a daily capsule that contained three viable and freeze-dried strains: Lactobacillus acidophilus (2 � 109 colony-forming units CFU/g), Lactobacillus casei (2 � 109 CFU/g) and Bifidobacterium bifidum (2 � 109 CFU/g) for 8 weeks. The placebo group took capsules filled with cellulose for the same time period. Fasting blood samples were taken at the beginning and the end of the study to quantify related markers. Results: After 8 weeks of intervention, compared with the placebo, probiotic supplementation resulted in improved Disease Activity Score of 28 joints (DAS-28) (�0.3 ± 0.4 vs. �0.1 ± 0.4, P = 0.01). In addition, a significant decrease in serum insulin levels (�2.0 ± 4.3 vs. +0.5 ± 4.9 μIU/mL, P = 0.03), homeostatic model assessment-B cell function (HOMA-B) (�7.5 ± 18.0 vs. +4.3 ± 25.0, P = 0.03) and serum high-sensitivity C-reactive protein (hs-CRP) concentrations (�6.66 ± 2.56 vs. +3.07 ± 5.53 mg/L, P < 0.001) following the supplementation of probiotics compared with the placebo. Subjects who received probiotic capsules experienced borderline statistically significant improvement in total- (P = 0.09) and low-density lipoprotein-cholesterol levels (P = 0.07) compared with the placebo. Conclusion: Overall, the results of this study indicated that taking probiotic supplements for 8 weeks among patients with RA had beneficial effects on DAS-28, insulin levels, HOMA-B and hs-CRP levels. © 2016 Asia Pacific League of Associations for Rheumatology and John Wiley & Sons Australia, Lt