Cold atmospheric plasma is a potent tool to improve chemotherapy in melanoma in vitro and in vivo

Malignant melanoma is a devastating disease. Because of its aggressiveness, it also serves as a model tumor for investigating novel therapeutic avenues. In recent years, scientific evidence has shown that cold atmospheric plasma (CAP) might be a promising modality in cancer therapy. In this study, we aimed to evaluate the effect of CAP generated by an argon plasma jet alone or in combination with dacarbazine (DAC) on melanoma cells in vitro and in vivo. The effects of the CAP on inducing lipid peroxidation and nitric oxide production were higher in B16 melanoma cells in comparison to non-malignant L929 cells. Assays on cell growth, apoptosis, and expression of genes related to, e.g., autophagic processes, showed CAP to have a substantial impact in melanoma cells while there were only minoreffects in L929 cells. In vivo, both CAP monotherapy and combination with DAC significantly decreased tumor growth. These results suggest that CAP not only selectively induces cell death in melanoma but also holds promises in combination with chemotherapy that might lead to improved tumor control. © 2020 by the authors

Multiple Sclerosis Gene Therapy Using Recombinant Viral Vectors: Overexpression of IL-4, IL-10 and Leukemia Inhibitory Factor in Wharton's Jelly Stem Cells in The EAE Mice Model

Objective: Immunotherapy and gene therapy play important roles in modern medicine. The aim of this study is to evaluate the overexpression of interleukin-4 (IL-4), IL-10 and leukemia inhibitory factor (LIF) in Wharton's jelly stem cells (WJSCs) in the experimental autoimmune encephalomyelitis (EAE) mice model. Materials and Methods: In this experimental study, a DNA construction containing IL-4, IL-10 and LIF was assembled to make a polycistronic vector (as the transfer vector). Transfer and control vectors were co-transfected into Human Embryonic Kidney 293 (HEK-293T) cells with helper plasmids which produced recombinant lentiviral viruses (rLV). WJSCs were transduced with rLV to make recombinant WJSC (rWJSC). In vitro protein and mRNA overexpression of IL-4, LIF, and IL-10 were evaluated using quantitative polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA) and western blot (WB) analysis. EAE was induced in mice by MOG-CFA and pertussis toxin. EAE mice were injected twice with 2x10(5) rWJSCs. The in vivo level of IL-4, LIF, IL-10 cytokines and IL-17 were measured by ELISA. Brain tissues were analyzed histologically for evaluation of EAE lesions. Results: Isolated WJSCs were performed to characterize by in vitro differentiation and surface markers were analyzed by flow cytometry method. Cloning of a single lentiviral vector with five genes was done successfully. Transfection of transfer and control vectors were processed based on CaPO4 method with > 90% efficiency. Recombinant viruses were produced and results of titration showed 2-3x10(7) infection-unit/ml. WJSCs were transduced using recombinant viruses. IL-4, IL-10 and LIF overexpression were confirmed by ELISA, WB and qPCR. The EAE mice treated with rWJSC showed reduction of Il-17, and brain lesions as well as brain cellular infiltration, in vivo. Weights and physical activity were improved in gene-treated group. Conclusion: These results showed that gene therapy using anti-inflammatory cytokines can be a promising approach against multiple sclerosis (MS). In addition, considering the immunomodulatory potential of WJSCs, an approach using a combination of WJSCs and gene therapy will enhance the treatment efficacy

Multiple Sclerosis Gene Therapy with Recombinant Viral Vectors: Overexpression of IL-4, Leukemia Inhibitory Factor, and IL-10 in Wharton's Jelly Stem Cells Used in EAE Mice Model.

OBJECTIVES: Immunotherapy and gene therapy play important roles in modern medicine. The aim of this study is to evaluate the overexpression of interleukin-4 (IL-4), IL-10 and leukemia inhibitory factor (LIF) in Wharton's jelly stem cells (WJSCs) in the experimental autoimmune encephalomyelitis (EAE) mice model. MATERIALS AND METHODS: In this experimental study, a DNA construction containing IL- 4, IL-10 and LIF was assembled to make a polycistronic vector (as the transfer vector). Transfer and control vectors were co-transfected into Human Embryonic Kidney 293 (HEK-293T) cells with helper plasmids which produced recombinant lentiviral viruses (rLV). WJSCs were transduced with rLV to make recombinant WJSC (rWJSC). In vitro protein and mRNA overexpression of IL-4, LIF, and IL-10 were evaluated using quantitative polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA) and western blot (WB) analysis. EAE was induced in mice by MOG-CFA and pertussis toxin. EAE mice were injected twice with 2×105 rWJSCs. The in vivo level of IL-4, LIF, IL-10 cytokines and IL-17 were measured by ELISA. Brain tissues were analyzed histologically for evaluation of EAE lesions. RESULTS: Isolated WJSCs were performed to characterize by in vitro differentiation and surface markers were analyzed by flow cytometry method. Cloning of a single lentiviral vector with five genes was done successfully. Transfection of transfer and control vectors were processed based on CaPO4 method with >90% efficiency. Recombinant viruses were produced and results of titration showed 2-3×107 infection-unit/ml. WJSCs were transduced using recombinant viruses. IL-4, IL-10 and LIF overexpression were confirmed by ELISA, WB and qPCR. The EAE mice treated with rWJSC showed reduction of Il-17, and brain lesions as well as brain cellular infiltration, in vivo. Weights and physical activity were improved in gene-treated group. CONCLUSIONS: These results showed that gene therapy using anti-inflammatory cytokines can be a promising approach against multiple sclerosis (MS). In addition, considering the immunomodulatory potential of WJSCs, an approach using a combination of WJSCs and gene therapy will enhance the treatment efficacy

Ninety-six–hour starved peripheral blood mononuclear cell supernatant inhibited LA7 breast cancer stem cells induced tumor via reduction in angiogenesis and alternations in Gch1 and Spr expressions

Introduction The microenvironment of solid tumors such as breast cancer is heterogeneous and complex, containing different types of cell, namely, cancer stem cells and immune cells. We previously reported the immunoregulatory behavior of the human immune cell in a solid tumor microenvironment-like culture under serum starvation stress for 96 h. Here, we examined the effect of this culture-derived solution on breast cancer development in rats. Method Ninety-six–hour starved PBMCs supernatant (96 h-SPS) was collected after culturing human PBMCs for 96 h under serum starvation condition. Breast cancer stem cells, LA7 cell line, was used for in vitro study by analyzing gene expression status and performing cytotoxicity, proliferation, scratch wound healing assays, followed by in vivo tumor induction in three groups of mature female Sprague Dawley rats. Animals were treated with 96 h-SPS or RPMI and normal saline as control, n = 6 for each group. After biochemical analysis of iron, lactate, and pH levels in the dissected tumors, Ki67 antigen expression, angiogenesis, and necrosis evaluation were carried out. Metabolic-related gene expression was assessed using RT-qPCR. Moreover, 96 h-SPS composition was discovered by Nano-LC-ESI-MS/MS. Results 96 h-SPS solution reduced the LA7 cell viability, proliferation, and migration and Gch1 and Spr genes expression in vitro ( p < 0.05), whereas stemness gene Oct4 was upregulated ( p < 0.01). The intracellular lactate was significantly decreased in the 96 h-SPS treated group ( p = 0.007). In this group, Gch1 and Spr were significantly downregulated ( p < 0.05), whereas the Sox2 and Oct4 expression was not changed significantly. The number of vessels and mitosis (Ki67 + cells) in the 96 h-SPS–treated group was significantly reduced ( p = 0.024). The increased rate of necrosis in this group was statistically significant ( p = 0.04). Last, proteomics analysis revealed candidate effectors’ components of 96 h-SPS solution. Conclusion 96 h-SPS solution may help to prevent cancer stem cell mediated tumor development. This phenomenon could be mediated through direct cytotoxic effects, inhibition of cell proliferation and migration in association with reduction in Gch1 and Spr genes expression, angiogenesis and mitosis rate, and necrosis augmentation. The preliminary data obtained from the present study need to be investigated on a larger scale and can be used as a pilot for further studies on the biology of cancer development

Mise en evidence d'une relation fonctionnelle entre les deux types de centrosomes du Myxomycete physarum polycephalum

SIGLECNRS T Bordereau / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Beta Globin Frameworks in Thalassemia Major Patients from North Iran

Objective: Four combinations of five neutral sequence changes at rs713040, rs10768683, rs7480526, rs7946748, and rs1609812 occurring in the human beta globin gene defined as frameworks have been reported in beta globin gene. Here we report for the frequency of these frameworks in thalassemia major patients of North Iran. Methods: Beta globin gene frameworks of 46 thalassemia major patients of North Iran were determined using Denaturing Gradient Gel Electrophoresis. Findings: All these frameworks called framework 1, 2, 3, 3a were present at the frequency of 23.9%, 45.7%, 6.5% and 23.9% respectively. Conclusion: These frameworks may be used for tracking mutant alleles in prenatal diagnosis programs

Effect of Cold Atmospheric Plasma on Gene Expression and Methylation of Genes Involved in Colorectal Cancer

Background and purpose: Colorectal cancer (CRC) is ranked as the second most common cancer in men and the third most common cancer in women worldwide. Routine treatments have many side effects and little efficiency. The aim of this study was to investigate the effects of Cold Atmospheric Plasma (CAP) on epigenetic changes of some genes involved in CRC progression. Materials and methods: Proliferation rate of cells after 40 s treatment was analyzed using MTT assay. Gene expressions of Beclin-1, Runx3, and AIM2 were measured using Real time PCR and methylation alternations of mentioned genes were evaluated with quantitative Methylation-Specific PCR (Q-MSP). Results: The viability of CAP-treated C26 cells significantly decreased compared to that of the L929 cells. The viability of C26 cell line decreased to75%, 52.2%, and 34% following 12, 24, and 48 hours of CAP therapy for 40 seconds. CAP therapy decreased the AIM2 expression (fold change=0.12) and increased the Runx3 gene expression (fold change=2.66) in C26 cell line. Also, treatment with CAP decreased the methylation level of Runx3 gene in CAP-treated cells compared with untreated cells (33.45% vs 63.24%). Conclusion: This study confirmed the selective effect of CAP on cancer cells which is an ultimate goal of cancer therapy approach. Also, CAP treatment was found to affect gene expression and methylation level of critical players of CRC