Evaluation of therapeutic potency of human papillomavirus-16 E7 DNA vaccine alone and with interleukin-18 as a genetic adjuvant / Avaliação da potência terapêutica da vacina de DNA do papilomavírus humano-16 E7 isolada e com interleucina-18 como adjuvante genético

AIMS: Despite the existence of effective preventive vaccines for human papillomavirus (HPV), therapeutic vaccines that trigger cell-mediated immune responses are required to treat established infections and malignancies. The aim of this study was to evaluate the therapeutic potency of HPV-16 E7 deoxyribonucleic acid (DNA) vaccine alone and with interleukin (IL)-18. METHODS: In vitro expressions of IL-18 were performed on human embryonic kidney 293 cells and confirmed it by Western blotting methods. DNA vaccine was available from a previous study. A total of 45 female C57BL/6 mice divided into five groups (DNA vaccine, DNA vaccine adjuvanted with IL-18, pcDNA3.1, and phosphate buffer saline) were inoculated with murine tissue culture-1 cell line of HPV related carcinoma, expressing HPV-16 E6/E7 antigens. They were then immunized subcutaneously twice at a seven-day interval. The antitumor and antigen specific-cellular immunity were assessed by lymphocyte proliferation (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide: MTT) assay, lactate dehydrogenase release assay, IL-4 assay and interferon-gamma (IFN-γ) assay. Tumor size was followed for 62 days. RESULTS: MTT assay, which measures the lymphocyte proliferation in response to the specific antigen, increased in the co-administration and the DNA vaccine groups as compared to control and genetic adjuvant groups (p<0.001). The mice immunized with the co-administration generated significantly more IFN-γ and IL-4 than other immunized mice (p<0.001). Reduction of the tumor size in the co-administration and the DNA vaccine groups was significantly more pronounced than in the control and genetic adjuvant groups (p<0.001), but no statistically significant difference between DNA vaccine and co-administration groups (p=0.15) occurred. CONCLUSIONS: IL-18 as a genetic adjuvant and E7 DNA vaccine alone enhanced immune responses in mouse model systems against cervical cancer. However, using of IL-18 as a genetic adjuvant with E7 DNA vaccine had no significant synergistic effect on the immune responses in vivo.

OBJETIVOS: Apesar da existência de vacinas preventivas eficazes contra o papilomavírus humano (HPV), são necessárias vacinas terapêuticas que desencadeiem respostas imunes mediadas por células para tratar infecções e malignidades estabelecidas. O objetivo deste estudo foi avaliar a potência terapêutica da vacina de ácido desoxirribonucleico (DNA) HPV-16 E7 isolada e com interleucina (IL)-18. MÉTODOS: Expressões in vitro de IL-18 foram realizadas em células renais embrionárias humanas 293 e confirmadas por Western blotting. A vacina de DNA foi disponibilizada em um estudo anterior. Um total de 45 camundongos fêmeas C57BL/6 divididos em cinco grupos (vacina de DNA, vacina de DNA adjuvada com IL-18, pcDNA3.1 e solução salina tamponada com fosfato) e foram inoculados com linhagem murina-1 de carcinoma relacionado ao HPV, expressando antígenos E6 / E7 do HPV-16. Os animais foram então imunizados por via subcutânea duas vezes no intervalo de sete dias. A imunidade antitumoral e antígeno-celular específica foi avaliada pela proliferação de linfócitos (ensaio de brometo de 3- [4,5-dimetiltiazol-2-il] -2,5-difeniltetrazólio: MTT), ensaio de liberação de lactato desidrogenase, ensaio de IL-4 e ensaio de interferon-gama [IFN-γ]. O tamanho do tumor foi seguido por 62 dias. RESULTADOS: O ensaio MTT, que mede a proliferação de linfócitos em resposta ao antígeno específico, aumentou nos grupos de coadministração e de vacina de DNA em comparação aos grupos controle e adjuvante genético (p <0,001). Os camundongos imunizados com a coadministração geraram significativamente mais IFN-γ e IL-4 do que os outros camundongos imunizados (p<0,001). A redução do tamanho do tumor nos grupos de coadministração e de vacina de DNA foi significativamente mais acentuada do que nos grupos controle e adjuvante genético (p<0,001), mas não houve nenhuma diferença estatisticamente significativa entre os grupos vacina de DNA e coadministração (p=0,15). CONCLUSÕES: A IL-18 como adjuvante genético e a vacina de DNA E7 aumentaram as respostas imunes em sistemas modelo de camundongos contra o câncer cervical. No entanto, o uso de IL-18 como adjuvante genético com a vacina de DNA E7 não teve efeito sinérgico significativo nas respostas imunes in vivo.

[Comparing Black and Green tea [Camellia sinensis L] extracts effects on the growth inhibition and biofilm formation of Enterobacteriacea]

Extracts of leaves from Camellia sinensis L contains polyphenolic components with antimicrobial activity. In this investigation biofilm inhibitory effects of black and green tea extracts were defined for five members of enterobacteriacea family including: Enteropathogenic Escherichia coli, Shigella flexneri, Proteus mirabilis, Klebsiella pneumoniae and Salmonella enterica serovar Typhimurium. Because tea is the most widely drunk beverage in Iran, therefore investigation of its effects on enterobacterial biofilm formation and colonization is very important. In this experimental study after extraction of samples in water/ methanol solution, further extraction took place in Ethyl acetate phase. The extracts preserved in 4°C refrigerator after sterilization by 0.44 micro filters. Well diffusion [Kirby Bauer] and broth dilution methods were used for evaluation of minimum inhibitory concentration of biofilm formation in black and green tea extracts treated cultures. Evaluation of biofilm formation was assayed by observation of colony forming unit of cultured bacteria per milliliter by sampling from Erlenmeyer flask wall scratching onto Tripticase soy agar medium and comparing the results with controls. Analysis of data was done using analysis of variance. Biofilm inhibitory effects of black tea were greater than green tea. The concentration of 4.5 mg/ml of black tea and 5mg/ml of green tea had bactericidal effects against examined bacteria. On Mueller Hinton agar, Proteus mirabilis was more sensitive to black tea; EPEC was more sensitive to green tea and Klebsiella pneumoniae showed more resistance to both extracts. Due to the fact that gastrointestinal tract is directly affected with consumed beverage, the high concentration of tea entered in lumen can reduce the number of enterobacteriaceae and can reduce their carcinogenic amine products. Thus it plays an important role in inhibition of gastrointestinal lymphoma and colon carcinoma. Also application of tea polyphenols as a food preservative can be useful