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Designing and evaluation of MERS-CoV siRNAs in HEK-293 cell line.
MERS-CoV sirna在HEK-293细胞系中的设计和评价。
- 影响因子:2.09
- DOI:10.1016/j.jiph.2020.12.018
- 作者列表:"Sohrab SS","El-Kafrawy SA","Mirza Z","Hassan AM","Alsaqaf F","Azhar EI
- 发表时间:2021-02-01
Abstract
BACKGROUND:The MERS-CoV was identified for the first time from Jeddah, Saudi Arabia in 2012 from a hospitalized patient. This virus has now been spread to 27 countries with a total of 858 deaths and 2494 confirmed cases and has become a serious concern for the human population. Camels are well known for the transmission of the virus to the human population. METHODS:In this report, we have discussed the designing, prediction, and evaluation of potential siRNAs against the orf1ab gene of MERS-CoV. The online software was used to predict and design the siRNAs and finally, total twenty-one siRNA were filtered out from four hundred and sixty-two sIRNAs as per their scoring and specificity criteria. We have used only ten siRNAs to evaluate their cytotoxicity and efficacy by reverse transfection approach in HEK-293-T cell lines. RESULTS:Based on the results and data generated; no cytotoxicity was observed for any siRNAs at various concentrations in HEK-293-T cells. The ct value of real-time PCR showed the inhibition of viral replication in siRNA-1, 2, 4, 6, and 9. The data generated provided the preliminary information and encouraged us to evaluate the remaining siRNAs separately as well as in combination to analyses the replication of MERS-CoV inhibition in other cell lines. CONCLUSION:Based on the results obtained; it is concluded that the prediction of siRNAs using online software resulted in the filtration of potential siRNAs with high accuracy and strength. This technology can be used to design and develop antiviral therapy not only for MERS-CoV but also against other viruses.
摘要
背景: MERS-CoV是2012年首次从一名住院患者中发现的,来自沙特阿拉伯吉达。这种病毒现已传播到27个国家,确诊病例共有858人死亡,2494人死亡,已成为人类严重关切的问题。众所周知,骆驼将病毒传播给人类。 方法: 在本报告中,我们讨论了针对MERS-CoV orf1ab基因的潜在sirna的设计、预测和评估。使用在线软件预测和设计siRNA,最后,根据它们的评分和特异性标准,从462个siRNA中过滤出总共21个siRNA。我们仅使用10个sirna通过反向转染方法在HEK-293-T细胞系中评估其细胞毒性和功效。 结果: 基于产生的结果和数据; 在HEK-293-T个细胞中没有观察到任何不同浓度的sirna的细胞毒性。实时PCR的ct值显示siRNA-1、2、4、6和9对病毒复制的抑制。产生的数据提供了初步信息,并鼓励我们单独评估剩余的sirna以及组合分析MERS-CoV抑制在其他细胞系中的复制。 结论: 基于所获得的结果; 结论是使用在线软件预测sirna导致了具有高准确性和强度的潜在sirna的过滤。该技术可用于设计和开发不仅针对MERS-CoV而且针对其他病毒的抗病毒疗法。
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METHODS::The ATP binding-cassette superfamily corresponds the mostly transmembrane transporters family found in humans. These proteins actively transport endogenous and exogenous substrates through biological membranes in body tissues, so they have an important role in the regulation of many physiological functions necessary for human homeostasis, as well as in response regulation to several pharmacological substrates. The development of multidrug resistance has become one of the main troubles in conventional chemotherapy in different illnesses including cancer, being the increased efflux of antineoplastic drugs the main reason for this multidrug resistance, with a key role of the ABC superfamily. Likely, the interindividual variability in the pharmacological response among patients is well known, and may be due to intrinsically factors of the disease, genetic and environmental ones. Thus, the understanding of this variability, especially the genetic variability associated with the efficacy and toxicity of drugs, can provide a safer and more effective pharmacological treatment, so ABC genes are considered as important regulators due to their relationship with the reduction in pharmacological response. In this review, updated information about transporters belonging to this superfamily was collected, the possible role of these transporters in cancer, the role of genetic variability in their genes, as well as some therapeutic tools that have been tried to raise against main transporters associated with chemoresistance in cancer.
METHODS:BACKGROUND:Cholinergic neurotransmission regulates neuroinflammation in Parkinson disease (PD). RESEARCH DESIGN AND METHODS:The authors conducted a delayed-start study of donepezil for cognitive decline in non-demented PD patients. The study consisted of a 96-week randomized placebo-controlled double-blind phase 1, followed by a 24-week donepezil extension phase 2. The primary outcome measure was a change in the Mini-Mental State Examination (MMSE) at week 120. RESULTS:A total of 98 patients were randomly allocated to the early-start (donepezil-to-donepezil) and delayed-start (placebo-to-donepezil) groups. Mean (SD) of the baseline MMSE was 27.6 (2.0) and 28.0 (2.1), respectively. MMSE change at week 120 was better in the early-start group than in the delayed-start group, but the difference was not significant. The MMSE declined in apolipoprotein ε4 carriers, but not in non-carriers, and the factor interaction (intervention × ε4 genotype) was highly significant (P < 0.001). Analyzed with the interaction, the difference was significant (group difference 1.95 [0.33 to 3.57], P = 0.018). The MMSE decline slope in phase 1 was significantly better in the early-start group than in the delayed-start group (P = 0.048). CONCLUSIONS:Cognitive function deteriorated in ε4 carriers, but not in non-carriers, and early-start donepezil may postpone cognitive decline in the former.
METHODS::Since the discovery of dental pulp stem cells, a lot of teams have expressed an interest in dental pulp regeneration. Many approaches, experimental models and biological explorations have been developed, each including the use of stem cells and scaffolds with the final goal being clinical application in humans. In this review, the authors' objective was to compare the experimental models and strategies used for the development of biomaterials for tissue engineering of dental pulp with stem cells. Electronic queries were conducted on PubMed using the following terms: pulp regeneration, scaffold, stem cells, tissue engineering and biomaterial. The extracted data included the following information: the strategy envisaged, the type of stem cells, the experimental models, the exploration or analysis methods, the cytotoxicity or viability or proliferation cellular tests, the tests of scaffold antibacterial properties and take into account the vascularization of the regenerated dental pulp. From the 71 selected articles, 59% focused on the "cell-transplantation" strategy, 82% used in vitro experimentation, 58% in vivo animal models and only one described an in vivo in situ human clinical study. 87% used dental pulp stem cells. A majority of the studies reported histology (75%) and immunohistochemistry explorations (66%). 73% mentioned the use of cytotoxicity, proliferation or viability tests. 48% took vascularization into account but only 6% studied the antibacterial properties of the scaffolds. This article gives an overview of the methods used to regenerate dental pulp from stem cells and should help researchers create the best development strategies for research in this field.