RESUMO
The occurrence of high-level tigecycline resistance tet(X) variant genes represents a new transferable resistance crisis to food safety and human health. Here, we investigated the abundance of tet(X)-variant genes [tet(X), tet(X1) to tet(X6)] in 33 samples collected from layer manures, manured/un-manured soils, and corresponding lettuce from three provinces in China. The results showed the occurrence of tet(X)/(X2), tet(X3), and tet(X4) in 24 samples. The detection rate of tet(X)/(X2) (23/24) is higher than that of tet(X3) (7/24) and tet(X4) (2/24), and tet(X)/tet(X2) and tet(X3) were found to be enriched and more abundant in most manured soil and several lettuce samples from manured soils than that from manure samples. Twenty six tigecycline-resistant bacteria were isolated, and tet(X)-variant genes were found to be disseminated not only by bacterial clone spreading but also via multidrug resistance plasmids. The total concentrations of tet(X)-variant genes showed significantly positive correlations (R = 0.683, p < 0.001) with ISCR2. Two veterinary tetracyclines (tetracycline and oxytetracycline) and other classes of antimicrobials (enrofloxacin, azithromycin, thiamphenicol, and florfenicol) showed significant correlations with the total concentrations of tet(X)-variant genes (R = 0.35-0.516, p < 0.05). The findings indicate the transmission of tet(X)-variant genes from layer manures to their receiving environmental soils and lettuce and highlight the contribution of veterinary antimicrobials to the spread of tet(X)-variant genes.
Assuntos
Esterco , Solo , Antibacterianos/farmacologia , China , Fazendas , Genes Bacterianos , Lactuca/genética , Esterco/análise , Microbiologia do Solo , Resistência a TetraciclinaRESUMO
Cancer Targeting Gene-Viro-Therapy (CTGVT) is a promising cancer therapeutical strategy that strengthens the anti-tumour effect of oncolytic virus by expressing inserted foreign anti-tumour genes. In this work, we constructed a novel adenoviral vector controlled by the tumour-specific survivin promoter on the basis of the ZD55 vector, which is an E1B55KD gene deleted vector we previously constructed. Compared with the original ZD55 vector, this new adenoviral vector (ZD55SP/E1A) showed much better ability of replication and reporter gene expression. We then combined anti-tumour gene interleukine-24 (IL-24) with an RNA polymerase III-dependent U6 promoter driving short hairpin RNA (shRNA) that targets M-phase phosphoprotein 1 (MPHOSPH1, a newly identified oncogene) by inserting the IL-24 and the shRNA of MPHOSPH1 (shMPP1) expression cassettes into the new ZD55SP/E1A vector. Our results demonstrated excellent anti-tumour effect of ZD55SP/E1A-IL-24-shMPP1 in vitro on multiple cancer cell lines such as lung cancer, liver cancer and ovarian caner. At high multiplicity-of-infection (MOI), ZD55SP/E1A-IL-24-shMPP1 triggered post-mitotic apoptosis in cancer cells by inducing prolonged mitotic arrest; while at low MOI, senescence was induced. More importantly, ZD55SP/E1A-IL-24-shMPP1 also showed excellent anti-tumour effects in vivo on SW620 xenograft nude mice. In conclusion, our strategy of constructing an IL-24 and shMPP1 dual gene expressing oncolytic adenoviral vector, which is regulated by the survivin promoter and E1B55KD deletion, could be a promising method of cancer gene therapy.
Assuntos
Adenoviridae/genética , Genes Supressores de Tumor , Terapia Genética/métodos , Vetores Genéticos , Vírus Oncolíticos/genética , Animais , Apoptose , Linhagem Celular Tumoral , Feminino , Deleção de Genes , Regulação da Expressão Gênica , Humanos , Proteínas Inibidoras de Apoptose/genética , Proteínas Inibidoras de Apoptose/metabolismo , Interleucinas/genética , Interleucinas/metabolismo , Neoplasias Hepáticas/patologia , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Ovarianas/patologia , RNA Interferente Pequeno/genética , Proteínas Repressoras , Survivina , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Aberrant JAK/STAT3 pathway has been reported to be related to hepatocellular carcinoma (HCC) in many cell lines. In this study, a double-regulated oncolytic adenovirus vector that can replicate and induce a cytopathic effect in alpha-fetoprotein (AFP)-positive HCC cell lines with p53 dysfunction was successfully constructed. Two therapeutic genes, suppressor of cytokine signaling 3 (SOCS3) and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), were chosen and incorporated into this vector system, respectively. The combined treatment of AFP-D55-SOCS3 and AFP-D55-TRAIL (2:3 ratio) exhibited potent antitumor activity in AFP-positive HCC cell lines compared with any other treatment both in vitro and in vivo. Specific replication and low progeny yield in AFP-positive HCC cell lines rendered these double-regulated oncolytic adenoviruses remarkably safe. Our data demonstrated that restoration of SOCS3, which inhibits the JAK/STAT3 pathway, by AFP-D55-SOCS3 not only could antagonize HCC therapeutic resistance to TRAIL and adenoviruses, but could also induce cell cycle arrest in HCC cell lines. SOCS3 could down-regulate Cyclin D1 and anti-apoptotic proteins such as XIAP, Survivin, Bcl-xL, and Mcl-1, which are responsible for the synergistic inhibitory effects of AFP-D55-SOCS3 and AFP-D55-TRAIL. Dual gene and double-regulated oncolytic adenoviruses may provide safety and excellent antitumor effects for liver cancer, which is the advantage of a cancer-targeting gene virotherapy strategy.
Assuntos
Adenoviridae/genética , Carcinoma Hepatocelular/terapia , Neoplasias Hepáticas/terapia , Vírus Oncolíticos/genética , Proteínas Supressoras da Sinalização de Citocina/genética , Ligante Indutor de Apoptose Relacionado a TNF/genética , Animais , Apoptose/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Efeito Citopatogênico Viral , Feminino , Expressão Gênica , Terapia Genética , Vetores Genéticos/administração & dosagem , Vetores Genéticos/toxicidade , Células HEK293 , Células HT29 , Células HeLa , Células Hep G2 , Humanos , Estimativa de Kaplan-Meier , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Terapia Viral Oncolítica , Regiões Promotoras Genéticas , Proteínas Supressoras da Sinalização de Citocina/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Carga Tumoral/genética , Replicação Viral , Ensaios Antitumorais Modelo de Xenoenxerto , alfa-Fetoproteínas/genéticaRESUMO
Interleukin (IL)-24 is an excellent therapeutic gene for cancer therapy. In this work, IL-24 was inserted into Ad.sp-E1A(Delta24), an oncolytic adenovirus with a 24-bp deletion in the E1A gene, which was driven by the survivin promoter to form Ad.sp-E1A(Delta24)-IL-24. Ad.sp-E1A(Delta24)-IL-24 has an excellent antitumor effect in vitro for human nasopharyngeal, liver, lung, and cervical carcinoma cell lines but does no or little damage to normal cell lines L-02 and WI38. Furthermore, it achieved nearly complete inhibition (although not elimination) of NCI-H460 lung carcinoma growth in nude mice. The antitumor efficacy of Ad.sp-E1A(Delta24)-IL-24 on NCI-H460 cells was clearly mediated by apoptosis, because it induced caspase-3 and poly(ADP-ribose) polymerase cleavage. This is the first report of Ad.sp-E1A(Delta24)-IL-24 with such an excellent, broad, and specific antitumor effect in vitro and nearly complete inhibition of lung tumor growth in vivo.
Assuntos
Adenoviridae/genética , Interleucinas/genética , Proteínas Associadas aos Microtúbulos/genética , Vírus Oncolíticos/genética , Regiões Promotoras Genéticas/genética , Proteína do Retinoblastoma/genética , Ensaios Antitumorais Modelo de Xenoenxerto , Adenoviridae/fisiologia , Animais , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Efeito Citopatogênico Viral/imunologia , Terapia Genética , Humanos , Proteínas Inibidoras de Apoptose , Interleucinas/uso terapêutico , Camundongos , Terapia Viral Oncolítica , Vírus Oncolíticos/fisiologia , Survivina , Replicação ViralRESUMO
The sorption characteristics of high-use tetracycline veterinary antibiotics-chlortetracycline (CTC) on pig manure and chicken manure were investigated in laboratory according to the OECD guideline 106. The results showed that CTC was strongly and rapidly adsorbed to the two manures and Freundlich model could describe preferably the CTC nonlinear sorption isotherms. The adsorbed CTC was difficult to be desorbed by rain with desorption H of 1.522 for pig manure and 1.329 for chicken manure, respectively. Only 18.3%-20.4%, 18.7%-19.4% and 55.7%-57.6% adsorbed CTC were desorbed by methanol, 3 mol/L NaCl and 3 mol/L MgCl2. The sorption of CTC on manure were affected by solution pH and ionic strength, the amount of adsorbed CTC on manure decreased with an increase of pH value and ionic strength, and greater sorption effect in the Ca systems than the Na systems was observed. These results suggest that organic matter and cation exchange mainly contribute to the sorption of CTC on manure.