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Aim: Cell-free DNA in the plasma is known to be a potential biomarker for noninvasive diagnosis of oncogenic mutations. The authors aimed to design an optimized padlock probe-based hyperbranched rolling circle amplification biosensor to detect the KRAS G12D mutation using fluorescence and colorimetric methods. Methods: Single-factor experiments, Plackett–Burman design and response surface methodology were applied to optimize the padlock probe-based hyperbranched rolling circle amplification reaction. Results: The maximum fluorescence intensity was achieved at a padlock probe concentration of 1.5 pM and target concentration of 9 pM at 38°C ligation temperature. The proposed biosensor has a low detection limit of 60 fM of target DNA and a linear response in the concentration range of 60 fM to 0.2 pM. Conclusion: The results indicated the power of these assays to detect KRAS point mutations in liquid state reactions.

Radiation can lead to various damages in the process of spermatogenesis that lead to a decrease in the number of sperm, an increase in spermatogenesis disorders, and defective sperm function. Radioprotectors are considered a good approach to reducing the damage caused by radiation. The goal of this work was to study how X-ray radiation affects testicular tissue and the process of spermatogenesis, as well as the radioprotective effects of selenium nanoparticles (SeNPs) and Lactobacillus casei (L. casei) as probiotic compounds, given alone or together. This study included 64 adult Syrian male mice weighing approximately 20 ± 5 g and aged 10 ± 1 weeks. Animals were randomly divided into eight groups: control group, SeNPs, probiotic, SeNPs and probiotic, X-ray radiation, SeNPs (X-ray), probiotic (X-ray), and SeNPs and probiotic (X-ray). Histology parameters and levels of oxidative stress biomarkers such as catalase, malondialdehyde, superoxide dismutase, and glutathione peroxidase were examined. In addition, the level of apoptosis was measured in testicular cells that had been treated with SeNPs and L. casei as a probiotic. The results showed that the administration of SeNPs or probiotic diminished the effects of X-ray radiation. These compounds induced a significant decreased in malondialdehyde, caspase 3, and caspase 9 gene levels and a remarkable increased in catalase, superoxide dismutase, and Catsper gene expression. SeNPs and probiotic exhibited a potent antioxidant effect and elevated the mean number of spermatogonia cells, sperm cell count, spermatogenesis percentage, and sperm motility percentage. The prescribed compound exhibited an ideal radioprotective effect with the ability to reduce the side effects of ionizing radiation and to protect normal tissues. SeNPs and probiotic inhibit testicular injury and improve the antioxidant state in male mice.

International audience; Aim: The effects of nutrition and exercise on autophagy are not well studied. This study aimed to investigate the combined effects of high-fat diets (HFD) and exercise training (ET) on autophagy in white adipose tissue of mice.Materials and methods: Male C57BL/6 mice were assigned into four groups of 7 mice per group: (1) Control, (2) high-fat diet-induced obesity (HFD-Ob), (3) exercise training (ET), and (4) high-fat diet with exercise training (HFD-ET). The HFD-Ob group was fed a high-fat diet for 14 weeks, while the ET group continuously ran on a treadmill for five sessions per week for seven weeks, and the HFD-ET group had both HFD and exercise training. qReal-time-PCR and western blot were used to measure the mRNA and protein levels of autophagy markers in white adipose tissue.Results: Mice from the HFD group showed higher levels in autophagy-related gene5 (ATG5, p = 0.04), ATG7 (p = 0.002), cathepsin B (CTSB, p = 0.0004), LC3-II (p = 0.03) than control. Mice in the ET group displayed higher levels of genes for ATG7 (p = 0.0003), microtubule-associated protein1-light chain 3 (LC3, p = 0.05), lysosome-associated membrane protein 2 (LAMP2, p = 0.04) and cathepsin L (CTSL, p = 0.03) than control. Mice from the HFD-ET group had higher levels of genes for ATG7 (p = 0.05) and CTSL (p = 0.043) and lower levels of genes for CTSB (p = 0.045) compared to the HFD group and lower levels of LAMP2 (p = 0.02) compared to the ET group.Conclusion: There were increases in autophagosome formation in the white adipose tissue from mice in the HFD and ET groups. A combination of HFD and ET enhances autophagosome formation and modulates lysosomal degradation in white adipose tissue.

Background. Some studies have established a relationship between obesity and the autophagic process in adipose tissue. This study aimed to investigate the effect of exercise training on the autophagic process in white adipose tissue (WAT) of high fat diet-induced obese mice.Methods and Results. C57BL/6 mice were assigned into three groups included: 1) Control 2), High-Fat Diet-induced Obesity (HFD-Ob), and 3) High-Fat Diet with Exercise Training (HFD-Ex). The subjects of HFD-Ob were fed a high-fat diet for 14 weeks. The mice of HFD-Ex had eight weeks of endurance training on a treadmill in addition to having the HFD. The Real-Time–PCR and western blot methods were used to measure the mRNA and protein levels of markers of the autophagic process. HFD caused an upregulation in the factors of the autophagosome formation, including ATG5 and ATG7, LC3, and the exercise training could augment the upregulation. Further, the training program prevented the change in LAMP2 expression (a marker of autophagolysosome), which being reduced by HFD. The lysosomal clearance factors (CTSB and CTSL) were raised in HFD-Ob and differently changed in HFD-Ex.Conclusion. HFD-induced obesity promoted the early and last steps of autophagy whereas defected the intermediate-step of it. Interestingly, the exercise training enhanced the early phase of autophagy, which being increased by HFD. Further, the training program could modify the rising effect of HFD on the last step of autophagy. It seems that a part of the protective effect of exercise training on obesity-related complications may be mediated by modulating the autophagic process in white adipose tissue.

Despite advances in the treatment of acute myocardial infarction, due to the non-proliferative nature of adult cardiomyocytes, the injured myocardium is mainly replaced by fibrotic tissue, which ultimately causes heart failure. To prevent heart failure, particularly after myocardial infarction, exosome-based therapy has emerged as one of the most promising strategies to regenerate cardiac function. Exosomes can carry microRNAs in support of neovascularization, anti-inflammatory, and endogenous cardiac regeneration. This study demonstrated that animal rat models' combination treatment with microRNA-126 and microRNA-146a mimics in exosomes is desirable for cardiac regeneration after myocardial infarction. The exosomes isolated from stem cells and loaded with microRNAs were characterized their impacts in cell migration, tube formation, and vascular endothelial growth factor degree. In the following, the usefulness of loaded microRNAs in exosomes and their encapsulation within alginate derivative hydrogel was analyzed in myocardial infarction for an animal model. Exosomes isolated and loaded with microRNAs showed the synergetic impact on cell migration, tube formation, and promoted vascular endothelial growth factor folding. Moreover, microRNAs loaded exosomes and encapsulated them in alginate hydrogel could help in reducing infarct size and improving angiogenesis in myocardial infarction. The angiogenesis markers including CD31 and connexion 43 upregulated for myocardial infarction models treated with alginate-based hydrogels loaded with exosomes and microRNAs-exosomes. Histological analysis indicated that myocardial infarction model rats treated with alginate hydrogel loaded with microRNAs-exosomes possessed lower and higher degrees of fibrosis and collagen fiber, respectively. These findings have important therapeutic implications for a myocardial infarction model through angiogenesis and vascular integrity regulation.

Early detection is one of the ways to deal with DNA virus widespread prevalence, and it is necessary to know new diagnostic methods and techniques. Colorimetric assays are one of the most advantageous methods in detecting viruses. These methods are based on color change, which can be seen either with the naked eye or with special devices. The aim of this study is to introduce and evaluate effective colorimetric methods based on amplification, nanoparticle, CRISPR/Cas, and Lateral flow in the diagnosis of DNA viruses and to discuss the effectiveness of each of the updated methods. Compared to the other methods, colorimetric assays are preferred for faster detection, high efficiency, cheaper cost, and high sensitivity and specificity. It is expected that the spread of these viruses can be prevented by identifying and developing new methods.

Background: DNA probes have been widely used as diagnostic tools for chromosomal translocations in malignancies. PCR-based methods often fail to detect translocations such as MYC/TRD in chronic lymphocytic leukemia. In addition, microscopic techniques cannot be helpful due to size detection limitations. This study sought to design a screening tool using immobilized ssDNA probes on a nitrocellulose membrane followed by 3C library fragments hybridization. Results: Hence, we focused on developing a suitable 27 bp specific probe for the juxtaposed region of MYC and TRD. Colloidal gold nanoparticles (AuNP) functionalized translocation fragments of the MYC gene with a thiol group (MYC-AuNP-probe). Then TRD-probes were immobilized on nitrocellulose surface to detect TRD/MYC translocation in the SKW3 cells. Hybridization between DNA probes and 3C-library fragments of SKW3 cells was determined by color intensity. Optimal hybridization of the 3C library sample of the cell line to TRD-probe and MYC-AuNP-probe showed higher color intensity due to their convenient proximity to the juxtaposed region compared with normal cells. Conclusions: Our results demonstrated that DNA hybridization colorimetric assay could be a helpful technique in chromosomal rearrangements screening. Accordingly, the combination of 3C based techniques and DNA-DNA hybridization can identify cancer cells with high specificity and sensitivity.

A motile, Gram-stain-positive, rod-shaped, non-sporing, tolerate up to 5% NaCl, grew at 0–25 °C, designated Exiguobacterium sp. HA2 was isolated from the soil of the Ilam Mountains of Iran during October 2016. The major isoprenoid quinone is MK-7 and in the smaller amount are MK-6 and MK-8. Polar lipids included diphosphatidylglycerol, phosphatidylglycerol, phosphatidylserine, phosphatidylinositol, phosphatidylethanolamine. Major fatty acids (>10 %) are isoC13:0, isoC15:0 and C16:0. The bacterial cell wall peptidoglycan layer was lysine-glycine. The 16S rRNA sequence was analyzed at the phylogenetic levels. Also, A supplemental comparison was made between five other genes including csp, gyrB, hsp70, rpoB, and citC. According to the results of genotypic and phenotypic characteristics, the strain was categorized in the genus Exiguobacterium. This bacterium had the closest relation with Exiguobacterium undae, and thus was dubbed Exiguobacterium sp. HA2. The different in the Phenotypic, functional characteristics and phylogenetic indicated Exiguobacterium sp. HA2 can be regarded as representing considered a novel species within the genus Exiguobacterium.

Background Metastasis accounts for ninety percent of breast cancer (BrCa) mortality. Cortactin, Ras homologous gene family member A (RhoA), and Rho-associated kinase (ROCK) raise cellular motility in favor of metastasis. Claudins (CLDN) belong to tight junction integrity and are dysregulated in BrCa. Thus far, epidemiologic evidence regarding the association of different pro-metastatic genes with pathological phenotypes of BrCa is largely inconsistent. This study aimed to determine the possible transcriptional models of pro-metastatic genes incorporate in holding the integrity of epithelial cell–cell junctions (CTTN, RhoA, ROCK, CLDN-1, CLDN-2, and CLDN-4), for the first time, in association with clinicopathological features of primary BrCa. Methods In a consecutive case-series design, 206 newly diagnosed non-metastatic eligible BrCa patients with histopathological confirmation (30–65 years) were recruited in Tabriz, Iran (2015–2017). Real-time RT-PCR was used. Then fold changes in the expression of target genes were measured. Results ROCK amplification was associated with the involvement of axillary lymph node metastasis (ALNM; ORadj. = 3.05, 95%CI 1.01–9.18). Consistently, inter-correlations of CTTN-ROCK (β = 0.226, P RhoA-ROCK (β = 0.311, P + BrCa. In addition, the overexpression of CLDN-4 was frequently observed in tumors identified by ALNM+ or grade III (P CTTN, CLDN-1, and CLDN-4 genes was correlated positively with the extent of tumor size. CTTN overexpression was associated with the increased chance of luminal-A positivity vs. non-luminal-A (ORadj. = 1.96, 95%CI 1.02–3.77). ROCK was also expressed in luminal-B BrCa tumors (P RhoA-ROCK (β = 0.280, P ROCK-CLDN-2 (β = 0.267, P CLDN-1-CLDN-4 (β = 0.451, P Conclusions For the first time, our findings suggested that the inter-correlations of CTTN-ROCK and RhoA-ROCK were significant transcriptional profiles determined in association with ALNM involvement; therefore the overexpression of ROCK may serve as a potential molecular marker for lymphatic metastasis. The provided binary transcriptional profiles need more approvals in different clinical features of BrCa metastasis.

Additional file 3: Table S2. General characteristics of patients with invasive breast cancer (N = 206).

Additional file 4: Table S3. Associations between lymphatic invasion and the involvement of axillary lymph node metastasis (ALNM).

Additional file 1: Fig S1. The overexpression of pro-metastatic genes were associated with clinicopathologic features of breast cancer. CLDN claudin, CTTN cortactin, HGPRT hypoxanthine–guanine phosphoribosyltransferase, NTC non-template control, RhoA ras homolog gene family member A, ROCK rho-associated kinase, N normal, T tumor.

Additional file 2: Table S1. Primer sequences used for qRT-PCR amplification.

Type 1 diabetes is a metabolic disorder caused by the loss or dysfunction of β-cells in the pancreas. Organ shortage is a critical concern of diabetic patients in need of beta islet transplantation. Tissue engineered islets are promising alternatives to traditional organ transplantation. Recent progress in stem cell biology and gene cloning techniques has raised hopes for the generation of insulin producing cells (IPCs) without the need of immunosuppression. The purpose of this study was to produce IPCs using human adipose-derived stem cells (hADSCs) and human endometrial-derived stem cells (hEnSCs) and also to compare the level of insulin secretion by these cells in 2D and 3D culture systems on fibrin scaffolding. Stem cells differentiation was carried out through transduction with an insulin over expression lentiviral vector. Real-time PCR and immunocytochemistry confirmed the successful transduction of both cell types. Both cell types showed comparable insulin secretion by ELISA.3D culture resulted in higher amounts of insulin secretion of the two cell types versus 2D as control. This study showed that insulin gene delivery to the stem cells could be an efficient method for producing IPCs and fibrin encapsulation enhances the functionality of these cells.

Background: Chromosomal breakpoints are the most common cause of hereditary diseases and cancers. Today, many standard clinical methods such as cytogenetic and PCR based techniques are used which have limitation regarding detection resolution. Chromosome conformation capture is a method for detecting gene proximity and chromosomal rearrangements. Materials and Methods: In this study, SKW3 cell line was used for detecting t(8;14)(q24;q11) using a 3Cbased technique. SKW3 cell line was used for 3C library preparation. For Inverse PCR, two regions were selected in upstream and downstream of the viewpoint locus on chromosome 8-MYC gene based on EcoRI restriction sites. The captured sequence with intra-chromosomal interaction between chr8-c-MYC and chr14-TRD was selected for the translocation PCR primer design. Results: The DNA fragment captured in 3C PCR showed a specific TRD sequence translocated downstream of the MYC gene. Translocation PCR demonstrated the existence of (8; 14) (q24; q11) MYC /TRD in both library and genomic DNA. Conclusion: This result demonstrated 3C- based method could be used as a useful low-cost easy operating technique in chromosomal rearrangements detection. In this study, the integration of whole genome library monitoring and PCR method was used as a high- through put method in chromosomal breakpoints detection.

Background: There exists a dramatic rise in liver failure and numerous patients undergo liver transplant for life-saving reasons annually. Introducing alternatives to allo-graft transplantation is necessary due to present limitations. Recently, a noninvasive stem cell population from Menstrual blood-derived Stem Cells (MenSCs) has been identified. There is an increasing interest in the application of MenSCs in tissue engineering; however, the fact that these gender-specific stem cells are safe for use in male sex is still not well defined. Methods: In this research, a model of acute liver failure was created in male and female immunocompetent Balb-C mice through intraperitoneal injection of Carbon tetrachlo-ride (CCl4 ) and MenSCs were transplanted intravenously 48 hrs after induction of liver injury to evaluate their therapeutic potential. All mice were sacrificed on days 1, 7, and 30 post-transplantation to examine biochemical and molecular markers and pathological appearances. Results: Results showed the liver engraftment of MenSCs by immunofluorescence staining using anti-human mitochondrial antibody in both male and female treated groups. The restoration of serum markers of liver injury, aspartate aminotransferase and ala-nine aminotransferase, as well as expression levels of liver-specific genes, tyrosine aminotransferase and cholesterol 7 alpha-hydroxylase, were more significant in the female treated group compared with the male treated group on day 7 (p

B-cell lymphoma 2 (BCL2) family proteins play a critical role in tuning cell death processes. Almost in half of all human cancers, a dysregulation in BCL2 family gene expression has been shown which made it an impressive target for human gene therapy as a novel approach in cancers. In this study we will optimize lentiviral-mediated RNA interference (RNAi), recombinant lentiviruses accommodating anti-BCL2 micro adaptive short hairpin RNAs (shRNAs), to downregulate BCL2 in human embryonic kidney 293T (HEK293T) cells to produce stable cell lines. We tested 4 different Dharmaconâ„¢ GIPZâ„¢ shRNAmir lentiviral vectors targeting BCL2 in different positions and a pGIPZ non-silencing shRNAmir lentiviral vector (as a negative control). Lentivirus packaging was performed by the calcium phosphate precipitation method. HEK293T cells were transduced by each type of recombinant lentiviruses individually and selected by puromycin within 10 days. The relative mRNA level and protein expression were assayed by using real-time polymerase chain reaction (PCR) technic and western blotting, respectively. Lentivirus (LV) packaging was performed in high efficiency (transfection rate was > 90%). Recombinant viruses of 4 expression vector addition to a control vector were produced then transduced to HEK293T cells successfully. All the 4 cell groups showed a significant down regulation of BCL2 gene (~90-95%) at mRNA level compared to the control group (p 0.05). We showed that the lentivirus-mediated RNAi technique is an efficient method to establish HEK293 cell lines with stable down-regulation of BCL2 gene.

Background and Objectives: Inhibin is a glycoprotein hormone commonly found in the circulation, but its level increases in some diseases, and its measurement by serological method using anti-inhibin monoclonal antibodies, can help in diagnosis of some genetic diseases. This study was performed with the objective of cloning, expression, and purification of α, βa, and βb subunits of human inhibin protein in Escherichia coli. Methods: For each Inhibin subunit gene. a primer pair was designed. Then, the gene sequence of each of the subunits, was obtained from human genomic DNA using polymerase chain reaction (PCR) method, and then was cloned into pET22b vector after enzymatic digestion. Recombinant vector associated with each subunit, was transferred to the host cell E. coli (strain BL21). The transformed cells were cultured in LB culture medium containing ampicillin, and positive colonies were isolated for mass production of recombinant protein. After mass culture and induction of transformed strains by isopropyl β-D-thiogalactopyranoside (IPTG), the produced recombinant protein, was isolated using nickel column chromatography. Results: The inhibin hormone subunit genes, was correctly cloned in pET22b vector and its expression in E. coli, was considerably high. The results of the expression of inhibin hormone also showed that in the absence of codon optimization, the inhibin expression in the prokaryote host increases significantly. The protein subunits α, βa, and βb of inhibin hormone, were purified, respectively, with molecular weights of 13.7, 12, and 12 kDa. Conclusion: protein subunits of inhibin hormone, can be expressed in the prokaryotic host due to their small size, short gene sequence, and minor post-transcriptional modifications. &nbsp

Background aims The application of menstrual blood stem cells (MenSCs) in regenerative medicine is gaining increasing attention. The aim of this study was to investigate the therapeutic potential of MenSCs compared with bone marrow–derived stem cells (BMSCs) in an animal model of CCl4-induced acute hepatic failure. Methods Injured Balb/C mice were divided into multiple groups and received MenSCs, BMSCs or hepatocyte progenitor-like (HPL) cells derived from these cells. Results Tracking of green fluorescent protein–labeled cells showed homing of cells in injured areas of the liver. In addition, the liver engraftment of MenSCs was shown by immunofluorescence staining using anti-human mitochondrial antibody. Microscopically examination, periodic acid-Schiff and Masson's trichrome staining of liver sections demonstrated the considerable liver regeneration post–cell therapy in all groups. Assessment of serum parameters including aspartate aminotransferase, alanine aminotransferase, total bilirubin, urea and cholesterol at day 7 exhibited significant reduction, such that this downward trend continued significantly until day 30. The restoration of liver biochemical markers, changes in mRNA levels of hepatic markers and the suppression of inflammatory markers were more significant in the MenSC-treated group compared with the BMSC-treated group. On the other hand, HPL cells in reference to undifferentiated cells had better effectiveness in the treatment of the acute liver injury. Conclusions Our data show that MenSCs may be considered an appropriate alternative stem cell population to BMSCs for treatment of acute liver failure.

Oligonucleotide aptamers are short, synthetic and single-stranded DNA or RNA molecules capable of binding to a wide range of molecules, from small molecules to large cells. Nowadays, aptamers are valuable tools in research, clinical diagnosis and treatment. Their small size and high specificity in addition to their lack of immunogenicity make them great alternatives to other diagnosing candidates such as antibodies. In this study, we have introduced a new method based on competitive Enzyme-Linked Aptamer Sorbent Assay (ELASA) using single-stranded DNA (ssDNA) aptamers to measure cystatin-c levels in serum samples. To this aim, through a Systematic Evolution of Ligands by Exponential Enrichment (SELEX) process a number of aptamers were selected from which an aptamer with a Kd (dissociation constant) value of 65.5 ± 0.007 nM was chosen for further analyses. The limit of detection (LoD) was found to be 216.077 pg/ml. The results of the analytical application of this method in serum samples were comparable to those of commonly used commercial kits.

PURPOSE: Methylated cell-free fetal DNA (cffDNA) in maternal plasma can potentially be used as a biomarker for accurate noninvasive prenatal testing (NIPT) of fetal disorders. Recovery and purification of cffDNA are key steps for downstream applications. In this study, we aimed to developed and evaluated different aspects of an optimized method and compared its efficiency with common methods used for extraction of methylated cffDNA. METHODS: Single factor experiments, Plackett-Burman (PB) design, and response surface methodology (RSM) were conducted for conventional Triton/Heat/Phenol (cTHP) method optimization. The total cell-free DNA (cfDNA) was extracted from pooled maternal plasma using the optimized method called the Triton/Heat/Phenol/Glycogen (THPG), cTHP method, a column-based kit, and a magnetic bead-based kit. In the next step, methylated cfDNA from the extracted total cfDNA was enriched using a methylated DNA immunoprecipitation (MeDIP) kit. Real-time quantitative polymerase chain reaction was performed on the RASSF1 gene and hyper region to determine the genomic equivalents per milliliter (GEq/ml) values of the methylated cfDNA and cffDNA, respectively. RESULTS: The optimum values of the significant factors affecting cfDNA extraction from 200 μl of plasma were 3% SDS, 1% Triton X-100, 0.9 μg/μl glycogen, and 0.3 M sodium acetate. The GEq/ml values of methylated cffDNA extracted using the THPG method were significantly higher than for the tested extraction methods (p

Table S2. Dietary characteristics across tertile (T) of three identified protein patterns (N = 172). (DOCX 20 kb)

Table S1. Factor loadings for identified protein patterns (N = 177). (DOCX 13 kb)

Objectives: In this study, a simple electrochemical nano-genosensor has been developed for the rapid and sensitive detection of methylated SEPT9 DNA as a useful biomarker for early colorectal cancer detection or screening. Methods: The process consists of three main steps: (i) the surface modification of screen-printed carbon electrode (SPCEs) with a poly(l-Arg)/RGO composite film followed by immobilizing anti-5-methylcytosine antibody (ii) preparation of probe-modified Fe3O4/TMC/Au nanocomposites for the hybridization with complementary DNA sequences, (iii) capturing methylated DNA target by antibody-modified SPCEs and subsequent electrochemical detection through redox peak currents of gold nanoparticles which generated a concentration-dependent response. Results: The surface modification of the electrode and hybridization with the methylated target were confirmed by cyclic voltammetry (CV) method and differential pulse voltammetry (DPV) was employ for quantitative evaluation of methylated target DNA. Conclusion: The assay showed a wide linear range from 0.01 pM to 1000 pM with a low detection limit of 0.01pM.

BACKGROUND DCLK1, as the most potential colorectal cancer stem cell (CSC) marker has been the core of many recent investigations. However, no study has been performed to evaluate the circulating cellular DCLK1 protein (CCDP) that might reflect the presences of colorectal CSC in circulation. OBJECTIVES We aimed to evaluate CCDP in the peripheral blood mononuclear cells (PBMCs) of colorectal cancer (CRC) patients applying immunoassay based methods including PLA, IPCR and ELISA in order to introduce the method of choice for clinical detection of CCDP. METHODS PBMCs were extracted from blood samples of 58 CRC patients along with 58 blood samples of tumor free controls. Total protein of PBMC was extracted and the CCDP level was evaluated. The results of three applied immunoassay tests were compared and the best approach for clinical application was introduced, accordingly. In addition, the correlation of CCD Plevel with clincopathologic findings of CRC patients was assessed. RESULTS The results of three immuneassay methods confirmed each other. Based on our finding, ELISA could be the most judicious method for clinical evaluation of CCDP considering its simplicity for clinical implications. Our results also showed a significant higher amount of CCDP in peripheral blood of CRC patients compared to control group which was also correlated with patients' clinicopathologic finding such as stage, grade and neoadjuvant history. CONCLUSION CCDP could be applied for monitoring purposes in CRC patients. However, its application needs to be more elucidated in future investigations implementing larger samples.

In this study, the enzyme streptokinase (thrombolysis agent) and chitosan (Cs) nanoparticles were prepared by self-assembly. Using experimental design, chitosan concentration, solution pH and stirring time were studied as independent variables to identify their effects on size, polydispersity index (PDI) and loading efficiency of nanoparticles. Results showed that pH and concentration have a direct effect on size. Additionally, minimum PDI was observed at lowest values of concentration and highest values of stirring time. pH-5.6 was also necessary to obtain the smallest PDI and highest loading efficiency values. The model predicted that to obtain maximum loading efficiency and minimum size along with low PDI, optimum values are 0.5 mg/mL, 5.18 and 30 min for the Cs concentration, solution pH and stirring time, respectively. The corresponding mean ± SD values for experimentally prepared nanoparticles were 43 ± 10%, 526 ± 121 nm, 0.3 ± 0.2, respectively. MTT and euglobulin clot lysis assays on the optimized nanoparticles showed that chitosan/streptokinase nanoparticles have slightly toxic effect on human fetal lung fibroblast cells (Mrc-5), compared with chitosan and streptokinase alone as a control. Also, thrombolytic activity of encapsulated streptokinase in nanoparticles is decreased slightly in comparison with free streptokinase. However, the preparation keeps a good potency for use as a thrombolytic agent in vivo.

This study investigated the correlation between the hepatic level of miR-122 and the extent of liver tissue regeneration in CCl4 induced liver injury mice model following transplantation of menstrual blood-(MenSCs) and bone marrow-derived stem cells (BMSCs). Hepatic miR-122 levels were significantly up-regulated following administration of CCl4 (P

International audience

Cancer stem cell (CSC) markers have attracted considerable attention in tumor diagnostic, prognostic, and therapeutic implications. Detection of cancer stem cells in circulating blood using cancer stem cell markers has received remarkable attention recently. In this study, we aimed to investigate the messenger RNA (mRNA) expression level of Lgr5 and DCLK1 as most proposed colorectal CSC markers in blood circulation also determine the subsequent association to patients' clinical and pathological findings. Peripheral blood mononuclear cells (PBMCs) of 58 patients with colorectal cancer at stage I-IV with 33 out of 58 patients undergoing preoperative chemoradiotherapy (CRT), as well as 58 healthy controls have been isolated and the extracted RNAs were analyzed using real-time PCR. The mRNA expression pattern of CSC markers of patients and controls was compared using ΔΔCt method. The expression level of Lgr5 was significantly higher in colorectal cancer (CRC) patients comparing to healthy group (4.8-fold change, p

Psoriasis is a T-cell-mediated skin disease with autoimmune nature that is generally not observed in animals, this lack of a relevant experimental animal model of psoriasis has hindered the investigation of pathogenesis of disease. Application and systemic delivery of small interfering RNAs offer many effective therapeutic advantages for gene regulation in the skin. In this study, we present an IMQ animal model of psoriasis and designed a safe fusion peptide carrier, spherical nucleic acid gold nanoparticles conjugate, to improve penetration of the siRNA into the cells and skin and their targeting ability to gene regulation. We evaluated the model of psoriasis and EGFR siRNA treatment (as spherical nucleic acid nanoparticles), phenotypically (signs of erythema, scaling, inflammation and thickening), microscopic evaluation of cell proliferation and immunohistochemically evaluation of CD3, CD4, and CD8 markers. Also, we monitored suppression of EGF&EGFR genes after treatment of A431 cells by SNA-NCs. The expression of genes was validated by qRT-PCR in human skin cells. The results showed that the SNA-NCs were stable and non-toxic. In vitro experiments indicated that EGF&EGFR siRNAs conjugated with spherical nucleic acid gold nanoparticles can significantly reduce gene expression in cells. In vivo experiments showed that the topical application of siRNAs delivered by SNA-NCs through the skin can significantly inhibit the proliferation of cells. Microscopic evaluation of mice back skin and immunohistochemistry process approved Inhibitory effect of SNA-NCs siRNA in the mouse model of psoriasis. Since the proliferation of T cells was crucial for the development of a psoriatic phenotype. These results demonstrate that topical application of SNA-NCs siRNA may improve psoriatic-like skin lesions by suppressing gene expression and functional activity of T cell production.

Context: To date, CSCs have been identified in a variety of hematopoietic and solid tumors. Applying CSC in clinical implication still depends on future studies to remove complexities including CSC heterogeneity and CSC similarity to normal stem cells. However, several potential clinical applications including therapeutic, diagnostic and prognostic implications have been introduced for cancer stem cells (CSC). In this review, we discuss previously considered and unconsidered potential clinical application of CSCs including how CSCs could be applied for pan-specific cancer screening and therapy. Evidence Acquisition: We will first discuss the previously proposed CSC clinical implications using a brief review of the literature. Subsequently, we will discuss some theoretical potential CSC implications which have not been discussed before including pan-specific cancer screening and therapy, and present confirmatory references for each part of our hypothesis. Results: We hypothetically demonstrated the presence of similar markers in the CSC subset of different tumors and introduced it as a way to simultaneously screen several cancers using one CSC marker. Conclusions: Simultaneous screening of several cancers applying one CSC marker could be regarded as a novel high-value cost-conscious cancer screening approach which might evolve cancer screening concept. However, this application remains to be explored in the future instigations.

Nanosuspensions, as a promising strategy to improve the solubility and bioavailability of poorly water soluble drugs, have been widely investigated in recent years. However, no comprehensive work so far has detailed the effect of independent processing/formulation parameters on the quality of the prepared nanosuspension. In the present study, the relations between solvent flow rate, stirring rate of antisolvent and surfactant concentration (i.e., inputs) on size, and polydispersity index (PDI) (i.e., outputs) of an N-acetylcysteine nanosuspension were investigated using artificial neural networks (ANNs). The response surfaces, generated as 3D graphs after ANNs modeling, demonstrated that all the three factors have a reverse effect on size and PDI. The dominant factor appeared to be the concentration of surfactant. Overall, it was found that the optimum formulation (i.e., minimum size and PDI value) is obtained at high values of surfactant concentration, solvent flow rate, and stirring rate (i.e., >0.9 mg/ml and 120 ml/h and 500 rpm, respectively).

The aim of this study was to determine the seroprevalence of Toxoplasma gondii infection in dogs, horses, and donkeys from west of Iran. Blood samples were collected randomly from 270 dogs, 120 horses, and 100 donkeys from Hamedan province. Using Modified Agglutination Test (MAT),10.7 %, 13.3 %, and 47 % of samples were seropositive in dogs, horses, and donkeys, respectively. There were statistical differences among dogs infection and age groups, and type of dogs, unlike to gender. In donkeys, there were statistical differences in age groups and gender, contrast to horses. This study is the first report of T. gondii infection in donkeys from Iran. Therefore, it is necessary to take integrated strategies for prevention and control of infection in animals, which could help to reduce human infections in this region.

Nanoparticles of ultra low molecular weight chitosan (ULMWCS)/hyaluronic acid (HA) were prepared by ion gelation. Three independent variables, namely, ratio of concentration of ULMWCS to HA (CS/HA), pH of solution and stirring time were studied to identify their effects on size, polydispersity and zeta potential of prepared nanoparticles using a Box-Behnken design. Results showed that pH and CS/HA have a direct effect on size, while increase of stirring time decreases the size of nanoparticles. Additionally, it was shown that all the independent parameters have direct effects on zeta potential. Also, the minimum polydispersity index was observed at lowest values of CS/HA. The model also predicted that the optimum values are 4.15, 4.14 and 180 (min) for the CS/HA, solution pH and stirring time, respectively. The obtained preparation had a size of 200 nm, polydispersity index of 0.37, and zeta potential of 13.0 mV.

The aim of this study was to determine the seroprevalence of Neospora caninum infection in dogs and cattle from Hamedan province (West of Iran). Blood samples were collected from 1,046 cattle and 270 dogs in this area. Cattle and dog samples were tested and analyzed using ELISA and IFAT, respectively. IgG-antibodies to N. caninum were found in 27 and 17.4 % of dogs and cattle samples, respectively. In cattle study, The association between infection and type of cattle was statistically significant (P = 0.004). Also, significant statistical differences were observed regarding to stray canids presence in farm (P

Problems commonly associated with using nanoemulsions are their cytotoxic effects and low stability profiles. Here, for the first time, concentrations of ingredients of a nanoemulsion system were investigated to obtain the most stable nanoemulsion system with the least cytotoxic effect on MCF7 cell line. Artificial neural networks (ANNs) were used to model the experimentally obtained data. Surfactant concentration was found to be the dominant factor in determining the stability - surfactant concentration above a critical point made the preparation unstable, while it appeared not to be influencing the cytotoxicity. Concentration of oil showed a direct relationship to the cytotoxicity with a minimum value required to provide an acceptable safety profile for the preparation. Co-surfactant appeared not to be considerably effective on neither stability nor cytotoxicity. To obtain the optimum preparation with maximum stability and minimum cytotoxicity, surfactant and oil values need to be kept at their maximum and minimum possible, respectively.

In vivo and in vitro tests are the two major ways of identifying the triggering allergens in sensitized individuals with allergic symptoms. Both methods are equally significant in terms of sensitivity and specificity. However, in certain circumstances, in vitro methods are highly preferred because they circumvent the use of sensitizing drugs in patients. In current study, we described a highly sensitive immuno-PCR (iPCR) assay for serum IgE specific to Bermuda allergens. Using oligonucleotide-labelled antibody, we used iPCR for the sensitive detection of serum IgE. The nucleotide sequence was amplified using conventional PCR and the bands were visualized on 2.5% agarose gel. Results demonstrated a 100-fold enhancement in sensitivity of iPCR over commercially available enzyme-linked immunosorbent assay (ELISA) kit. Our iPCR method was highly sensitive for Bermuda-specific serum IgE and could be beneficial in allergy clinics.

Brain oxidative status is a crucial factor in the development of sporadic Alzheimer's disease (AD). Klotho, an anti-aging protein, diminishes oxidative stress by the induction of manganese superoxide dismutase (MnSOD). Thus, the substances that increase klotho expression could be considered as a potential treatment for Alzheimer's disease when the oxidative imbalance is present. Statins are suggested to up-regulate klotho expression. We examined the effect of simvastatin (5mg/kg, daily for 3weeks) on hippocampal klotho and MnSOD expression in the cognitive declined animal model induced by intracerebroventricular (ICV)-streptozotocin (STZ) administration. Cognitive assessment was performed by the Morris Water Maze (MWM) test. The results indicated that mean escape latency and distance were prolonged in the ICV-STZ group compared with the control group. The expression of klotho and MnSOD were also down regulated in the hippocampus. Furthermore, improved spatial performance was observed in simvastatin-treated animals. This effect could be related to increase in oxidative stress tolerance as evidenced by klotho and MnSOD up-regulation. Our current study indicates that klotho upregulation may be a neuroprotective mechanism of simvastatin against cognitive decline in AD.