Radiosensitization effect of radiofrequency hyperthermia in the presence of PEGylated-gold nanoparticles on the MCF-7 breast cancer cells under 6 MeV electron irradiation.

Purpose: The purpose of the study was to investigate the radiosensitization effect of radiofrequency (RF) hyperthermia in combination with PEGylated gold nanoparticles (PEG-GNPs) on MCF-7 breast cancer cells under electron beam radiotherapy (EBRT) based on the clonogenic assay. Materials and Methods: The cell death of MCF-7 breast cancer cells treated with 13.56 MHz capacitive RF hyperthermia (power: 150W) for 2, 5, 10, and 15 min combined with 6 MeV EBRT, with a dose of 2 Gy, was evaluated in the presence of 20 nm PEG-GNPs with a low nontoxic concentration (20 mg/l). All the treatment groups were incubated for 14 days. Thereafter, survival fractions and viability of the cells were calculated and analyzed against the control group. Results: The presence of PEG-GNPs inside the MCF-7 cancer cells during electron irradiation decreased cell survival significantly (16.7%) compared to irradiated cells without GNPs. Applying hyperthermia before electron irradiation with a capacitive RF system decreased cell survival by about 53.7%, while hyperthermia without irradiation did not show any significant effect on cell survival. Combining the hyperthermia with the presence of PEG-GNPs in the cells decreased the cell survival by about 67% at the electron irradiation, showing their additive radiosensitization effect. Conclusion: Low nontoxic concentration of 20 nm PEG-GNPs increases the radiosensitization effect of combining 6 MeV EBRT and RF hyperthermia on MCF-7 cancer cells. Combining hyperthermia with PEG-GNPs in electron radiotherapy could be an appropriate method for enhancing radiotherapy effectiveness on cancerous cells which can be studied on different cells and electron energies in future research.

Cellular immunity survey against urinary tract infection using pVAX/fimH cassette with mammalian and wild type codon usage as a DNA vaccine

PURPOSE: FimH (the adhesion fragment of type 1 fimbriae) is implicated in uropathogenic Escherichia coli (UPEC) attachment to epithelial cells through interaction with mannose. Recently, some studies have found that UPEC can thrive intracellularly causing recurrent urinary tract infection (UTI). Almost all vaccines have been designed to induce antibodies against UPEC. Yet, the humoral immune response is not potent enough to overcome neither the primary UTI nor recurrent infections. However, DNA vaccines offer the possibility of inducing cell mediated immune responses and may be a promising preventive tool. MATERIALS AND METHODS: In this study, we employed two different open reading frames within mammalian (mam) and wild type (wt) codons of fimH gene. Optimized fragments were cloned in pVAX-1. Expression of the protein in COS-7 was confirmed by western blot analysis after assessing pVAX/fimH(mam) and pVAX/fimH(wt). The constructs were injected to BALB/c mice at plantar surface of feet followed by electroporation. RESULTS: The mice immunized with both constructs following booster injection with recombinant FimH showed increased interferon-gamma and interleukin-12 responses significantly higher than non-immunized ones (p<0.05). The immunized mice were challenged with UPEC and then the number of bacteria recovered from the immunized mice was compared with the non-immunized ones. Decreased colony count in immunized mice along with cytokine responses confirmed the promising immune response by the DNA vaccines developed in this study. CONCLUSION: In conclusion, DNA vaccines of UPEC proteins may confer some levels of protection which can be improved by multiple constructs or boosters.

Cloning of a Recombinant Plasmid Encoding Thiol-Specific Antioxidant Antigen (TSA) Gene of Leishmania majorand Expression in the Chinese Hamster Ovary Cell Line.

BACKGROUND: TSA (thiol-specific antioxidant antigen) is the immune-dominant antigen of Leishmania major and is considered to be the most promising candidate molecule for a recombinant or DNA vaccine against leishmaniasis. The aim of the present work was to express a plasmid containing the TSA gene in eukaryotic cells. METHODS: Genomic DNA was extracted, and the TSA gene was amplified by polymerase chain reaction (PCR). The PCR product was cloned into the pTZ57R/T vector, followed by subcloning into the eukaryotic expression vector pcDNA3 (EcoRI and HindIII sites). The recombinant plasmid was characterised by restriction digest and PCR. Eukaryotic Chinese hamster ovary cells were transfected with the plasmid containing the TSA gene. Expression of the L. major TSA gene was confirmed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and Western blotting. RESULTS: The plasmid containing the TSA gene was successfully expressed, as demonstrated by a band of 22.1 kDa on Western blots. CONCLUSION: The plasmid containing the TSA gene can be expressed in a eukaryotic cell line. Thus, the recombinant plasmid may potentially be used as a DNA vaccine in animal models.

Cloning of a Recombinant Plasmid Encoding Thiol-Specific Antioxidant Antigen (TSA) Gene of Leishmania major and Expression in the Chinese Hamster Ovary Cell Line

Background: TSA (thiol-specific antioxidant antigen) is the immune-dominant antigen of Leishmania major and is considered to be the most promising candidate molecule for a recombinant or DNA vaccine against leishmaniasis. The aim of the present work was to express a plasmid containing the TSA gene in eukaryotic cells. Methods: Genomic DNA was extracted, and the TSA gene was amplified by polymerase chain reaction (PCR). The PCR product was cloned into the pTZ57R/T vector, followed by subcloning into the eukaryotic expression vector pcDNA3 (EcoRI and HindIII sites). The recombinant plasmid was characterised by restriction digest and PCR. Eukaryotic Chinese hamster ovary cells were transfected with the plasmid containing the TSA gene. Expression of the L. major TSA gene was confirmed by sodium dodecyl sulphate–polyacrylamide gel electrophoresis and Western blotting. Results: The plasmid containing the TSA gene was successfully expressed, as demonstrated by a band of 22.1 kDa on Western blots. Conclusion: The plasmid containing the TSA gene can be expressed in a eukaryotic cell line. Thus, the recombinant plasmid may potentially be used as a DNA vaccine in animal models.