High rate of A2142G point mutation associated with clarithromycin resistance among Iranian Helicobacter pylori clinical isolates

This study aimed to investigate the clarithromycin resistance and its associated molecular mechanisms among Helicobacter pylori isolates from dyspeptic patients in Shiraz, Iran. From January to May 2014, 100 H. pylori strains were isolated from patients with gastroduodenal disorders. The resistance to clarithromycin was quantitatively evaluated, using Epsilometer (E‐test) method. Polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) was performed on all the isolates to detect A2143G and A2142G mutations in 23S rRNA gene. The H. pylori isolation rate was found to be 31.4%. E‐test showed that 20% of isolates were resistant to clarithromycin (MIC ≥ 1 mg/L). MIC of clarithromycin ranged between 0.016 and 24 mg/L. Findings of PCR‐RFLP showed that the A2142G was the most (90%) frequently point mutation, followed by the A2143G (10%). No statistically significant difference was found between H. pylori clarithromycin resistance point mutations and patients’ gender or age. To the best of our knowledge, this is the first report of high frequency of A2142G point mutation in Iran and probably in other regions of the world. Considering the increasing trend of H. pylori resistance to clarithromycin due to these mutations, it is crucial to investigate the new therapeutic approaches against H. pylori infection.     

A Computer-Aided Type-II Fuzzy Image Processing for Diagnosis of Meniscus Tear

Meniscal tear is one of the prevalent knee disorders among young athletes and the aging population, and requires correct diagnosis and surgical intervention, if necessary. Not only the errors followed by human intervention but also the obstacles of manual meniscal tear detection highlight the need for automatic detection techniques. This paper presents a type-2 fuzzy expert system for meniscal tear diagnosis using PD magnetic resonance images (MRI). The scheme of the proposed type-2 fuzzy image processing model is composed of three distinct modules: Pre-processing, Segmentation, and Classification. λ-nhancement algorithm is used to perform the pre-processing step. For the segmentation step, first, Interval Type-2 Fuzzy C-Means (IT2FCM) is applied to the images, outputs of which are then employed by Interval Type-2 Possibilistic C-Means (IT2PCM) to perform post-processes. Second stage concludes with re-estimation of “η” value to enhance IT2PCM. Finally, a Perceptron neural network with two hidden layers is used for Classification stage. The results of the proposed type-2 expert system have been compared with a well-known segmentation algorithm, approving the superiority of the proposed system in meniscal tear recognition.     

Phenotypic and functional maturation of murine dendritic cells induced by 18 alpha- and beta-glycyrrhetinic acid

Various studies have described glycyrrhizin (GL), an active triterpenoic saponin extract of licorice roots, as an anti-inflammatory, antiviral, antimicrobial, anti-tumor and immunomodulating agent. The activity of GL has been mainly attributed to its metabolites, 18-alpha (GA) and 18-beta-glycyrrhetinic acid (GB), which their mechanism of action on the immune system cells is not clearly known. In this study, we have investigated the effects of GA and GB on the immune system by targeting dendritic cells and analyzing phenotypic and functional maturity of murine dendritic cells (DCs) after treatment with these components. Stimulation of DCs with GA and GB resulted in up-regulation of CD40, CD86 and MHC-II molecules indicating their effects on the maturation of DCs. These components induced the allogenic immunostimulatory capacity of DCs by stimulating the proliferation of T cells and production of the T helper (h)1-promoting cytokine, IL-12. They also increased the production of IFN-γ by T cells in mixed-lymphocyte reaction. In conclusion, these results indicate that GA and GB may insert their immunomodulatory effects by enhancing DC maturation and modulating Th1/Th2 response through an increase in Th1 responses, implying their beneficial in host defense against infectious diseases.     

Association between epstein barr virus and tongue squamous cell carcinoma in iranian patients

Detection of Epstein-Barr virus in oral squamous cell carcinoma suggests its involvement in the carcinogenesis of oral cavity. But, there are few studies on the incidence of EBV genome in squamous cell carcinomas at specific locations in the oral cavity like tongue and with different tumor progression. In this study the presence of EBV genome in tongue Squamous Cell Carcinoma (TSCC) in Iranian patients were investigated. Accordingly, a total of 94 cases with TSCC were firstly analyzed for the presence of viral genome through Nested PCR. Patients were divided into different groups based on their gender and the size, nodal involvement, grade and stage of their tumor. Results showed the presence of EBV genome in 72.3% of TSCCs with no significant difference between two genders, although slightly higher in females. Interestingly, PCR products of EBV genome showed a statistically significant higher distribution in TSCCs at IVa stage (p = 0.04), while a considerable low involvement of EBV genome was seen in T1-sized tumors. The result of this study further emphasizes the role of EBV in oral SCCs – mainly at tongue. This is the first investigation to clarify the association between EBV genome and different tumor size and stage in TSCCs; however, more studies in different regions and larger populations should be performed to be able to draw a firmed conclusion.     

Effects of a Healthy Lifestyle Education on the Incidence of Metabolic Syndrome in Children during a 13-Year Follow-up

Purpose

This study aimed to determine effects of community-based healthy lifestyle education on the incidence of metabolic syndrome (MetS) in Tehranian children considering parental factors during a 13-year follow-up.

Methods

This study was conducted within the framework of the Tehran Lipid and Glucose Study (TLGS) on 1603 healthy children, aged 8–18 years, who had complete parental data. Parental factors including MetS, education, age, occupation, and smoking were considered to distinguish parental clusters which could potentially predispose children to MetS. Lifestyle interventions were aimed at achieving healthy dietary patterns and increasing physical activity. Cluster analysis and survival Cox model were used to determine potential low and high risk parental clusters and the effect of intervention on the hazard of MetS in children respectively.

Results

In a model adjusted for children’s age and sex, the hazard of the MetS incidence was 36% higher in children from high-risk parental cluster than those from low-risk parental cluster (HR = 1.36, 95% CI = 1.07–1.73). Moreover, the intervention group showed a 39% lower risk for MetS incidence (HR = 0.61, 95% CI = 0.44–0.85) compared to controls, during the first 6 years after baseline assessment (short term), although the risk lowering effect of the intervention was not maintained long term.

Conclusion

A healthy lifestyle education was successful in reducing the short-term risk of MetS in children. To identify ways of maintaining long-term results, further research is definitely warranted.

Trial Registration

This study was funded by National Research Council and Planning and Management Organization and has been reviewed and approved by the Iranian registry of clinical trials (ISRCTN52588395).

     

Tolerogenic dendritic cells produced by lentiviral‐mediated CD40‐ and interleukin‐23p19‐specific shRNA can ameliorate experimental autoimmune encephalomyelitis by suppressing T helper type 17 cells

Down‐regulation of soluble or membrane‐bound co‐stimulatory molecules by RNAi in dendritic cells can prevent the activation of immune responses. Therefore, this study was designed to evaluate the therapeutic efficacy of bone marrow‐derived DCs (BMDCs) transduced with lentiviral vectors to permanently expressed shRNA specific for CD40 (CD40LV‐DCs) and/or p19 subunit of interleukin (IL)‐23 (p19LV‐DCs) mRNAs in experimental autoimmune encephalomyelitis (EAE). In‐vitro studies showed that double‐transduced BMDCs (CD40⁺p19LV‐DCs) resemble tolerogenic DCs due to profound down‐regulation of CD40, lower expression of proinflammatory cytokines (IL‐6 and IL‐12), increased IL‐10 production and stronger stimulation of myelin oligodendrocyte glycoprotein (MOG)_35–55‐specific T cells for production of IL‐10 compared with CD40LV‐DCs, p19LV‐DCs and BMDCs transduced with control lentiviral vector (CoLV‐DCs). Moreover, injection of transduced CD40⁺p19LV‐ BMDCs in EAE mice resulted in more reduction in clinical score, significant reduction in IL‐17 or increased production of IL‐10 by mononuclear cells derived from the lymph nodes or spinal cord compared with CoLV‐DCs‐treated EAE mice. In conclusion, simultaneous knock‐down of CD40 and IL‐23 production by BMDCs may represent a promising therapeutic tool for the treatment of IL‐17‐dependent autoimmune diseases, including multiple sclerosis.     

HSP65-PRA identification of non-tuberculosis mycobacteria from 4892 samples suspicious for mycobacterial infections

Various molecular methods have been used for the rapid identification of mycobacterial species. In this survey, evaluation of antibiotic resistance and PCR-restriction fragment length polymorphism analysis (PRA) of the hsp65 gene was carried out for identification of non-tuberculosis mycobacteria (NTM) isolates from different clinical specimens. Forty-eight different mycobacterial isolates were selected and followed by the conventional and PRA of hsp65 for species identification. The antibiotic susceptibility test was carried out according to standard methods. A 439 bp PCR product of hsp65 in all selected isolates was amplified and digested with the BstEII and HaeIII restriction enzymes. The restriction fragment length polymorphism (RFLP) patterns were analyzed for species identification. Using PRA for 48 mycobacterial selected isolates, including 15 M. tuberculosis, one M. bovis and all 32 isolates of NTM, revealed 11 different species among the NTM isolates. The most frequent NTM isolates were M. kansasii, M. gordonae III, M. marinum, M. chelonae, M. scrofluaceum and M. gastri. In most cases, the PRA results were perfectly in accordance with the classical biochemical method. Combination of resistance to rifampin and isoniazid was present among M. kansasi, M. gordoniae III, M. scrofluaceum, M. chelonae, M. marinum, M. gastri, M. gordoniae II and M. trivale isolates. A high incidence of co-resistance to six, five, four and three anti-TB drugs was observed in 18.5%, 9.1%, 6.6% and 11.7% of all NTM isolates, respectively. Our results showed that PRA, in comparison with classical methods, is rapid and accurate enough for the identification of mycobacterial species from LJ medium. Additionally, we found that in Iran we have a highly diverse population of NTM isolates among patients suspected of having TB.     

Evaluating the efficacy of extracted squalene from seed oil in the form of microemulsion for the treatment of COVID-19: A clinical study

This study investigates the effect of the nanostructure of squalene in the form of microemulsion on COVID-19 patients. In this blinded clinical trial, a comparison was made between the efficacy of squalene treatment and controls. A total of 30 COVID-19 patients admitted to the emergency department, and the infection ward was equally allocated to case (n = 15) and control (n = 15) groups according to their age and underlying diseases. The baseline characteristics of subjects, including age, gender, time of treatment onset, underlying condition, white blood cells count, and lymphocyte count were similar (p < 0.05). Baseline laboratory tests and computed tomography (CT) scans were performed for the study groups. The treatment group received 5 mg of intravenous squalene twice a day and standard treatment for 6 days, while controls received only standard treatment. After 6 days of treatment, clinical and CT scan changes were evaluated and compared in intervention and control groups. The need for oxygen therapy (p = 0.020), 2 days of no fever (p = 0.025), cough alleviation (p = 0.010), and lung high-resolution computed tomography improvement (p = 0.033) were significantly different between cases and controls within 7 days of admission. No adverse effects were observed in the treatment group. Our data suggest that squalene could be considered as a potential treatment for COVID-19, and further studies are required to confirm the results.     

Paradoxical effects of interleukin-10 on the maturation of murine myeloid dendritic cells

The immunoregulatory cytokine, interleukin-10 (IL-10), has been shown to inhibit the maturation of human myeloid dendritic cells (DC). In the present study, we demonstrate that IL-10 has paradoxical effects on the maturation of murine myeloid bone marrow-derived DC. On the one hand, IL-10 inhibits the maturation of murine myeloid DC. The addition of IL-10 to granulocyte-macrophage colony-stimulating factor (GM-CSF)-supported murine BM-derived DC cultures reduced the frequency of major histocompatibility complex (MHC) class IIbright cells. These IL-10-pretreated DC have a reduced capacity to stimulate T cells in an allogeneic mixed leucocyte reaction. On the other hand, however, and in contrast to the effects of IL-10 on human DC, we found that the addition of IL-10 from the initiation of the culture onwards induced an up-regulation of the expression of the costimulatory molecules CD40, CD80 and CD86 on murine myeloid DC, as compared to DC generated with GM-CSF only. Moreover, a subpopulation of IL-10-pretreated MHC class IIdim DC lacked the capacity to take up dextran-fluorescein isothiocyanate (FITC), a feature of DC maturation. Taken together, our data demonstrate that the generation of murine myeloid DC in the presence of IL-10 results in a population of incompletely matured MHC class IIdim CD80+ CD86+ DC. These DC lack T-cell stimulatory capacity, suggesting a role for IL-10 in conferring tolerogenic properties on murine myeloid DC.     

Metabolism of short-chain ceramide by human cancer cells—Implications for therapeutic approaches

Due to recent use of short-chain ceramides in preclinical studies, we characterized C6-ceramide metabolism in cancer cell lines and assessed metabolic junctures for enhancing efficacy. MDA-MB-231 breast cancer cells decreased the amount of C6-ceramide metabolized to C6-sphingomyelin (C6-SM) and increased the amount metabolized to C6-glucosylceramide (C6-GC) in response to increasing concentrations. A similar trend was seen in DU-145 (prostate cancer), PANC-1 (pancreatic cancer), and LoVo (colorectal cancer) cells. KG-1 leukemia cells favored C6-SM synthesis at low (0.6 μM) and high-dose (12 μM) C6-ceramide. Partnering C6-ceramide with tamoxifen, a P-glycoprotein antagonist that impedes ceramide glycosylation, was an effective regimen for enhancing cytotoxicity in cells. Experiments to assess the mechanism of cell death using KG-1 cells showed that tamoxifen inhibited synthesis of C6-GC and C6-SM from C6-ceramide by 80% and 50%, respectively, which was accompanied by enhanced apoptosis. Radiolabeling of KG-1 cells with [³H]palmitic acid produced a 2-fold increase in ³H-long-chain ceramides when unlabeled C6-ceramide was added and a 9-fold increase when C6-ceramide and tamoxifen were added. The increase in ³H-palmitate radiolabeling of long-chain ceramides was blocked by inclusion of a ceramide synthase inhibitor; however, inhibiting synthesis of long-chain ceramide did not rescue cells. These studies show that tamoxifen enhances the apoptotic effects of C6-ceramide. The proposed mechanism involves blocking short-chain ceramide anabolism to favor hydrolysis and generation of sphingosine. We propose that use of tamoxifen and other P-glycoprotein antagonists can be an effective means for enhancing cytotoxic potential of short-chain ceramides in the treatment of cancer.