RESUMO
Long noncoding RNAs (lncRNAs) have been implicated in the regulation of resistance to radiotherapy in cervical cancer, which is a type of gynecological disease with high mortality in women around the world. Hence, our purpose is to delineate the involvement of LINC00958 in regulating cell sensitivity to radiotherapy in cervical cancer. LINC00958 expression in cervical cancer was assayed, followed by verification of the relationship among LINC00958, microRNA-5095 (miR-5095) and ribonucleotide reductase subunit M2 (RRM2). Hela cells were transduced with up-/downregulation of miR-5095 or RRM2, or LINC00958 silencing, respectively, and then treated with or without a 6 Gy dose of X-ray irradiation. Then the cell proliferation, apoptosis, survival fraction rate, as well as sensitivity to radiotherapy, were assessed. Finally, xenograft tumor in nude mice was established by transplanting Hela cells transfected with sh-LINC00958 and irradiated with 6 Gy of X-ray. High expression of LINC00958 was revealed in The Cancer Genome Atlas and Gene Expression Profiling Interactive Analysis, as well as in radiation-resistant patients, which was associated with lower sensitivity to radiotherapy in cervical cancer. Moreover, cervical cancer patients with higher LINC00958 expression exhibited a shorter overall survival according to Kaplan-Meier analysis. In addition, LINC00958 could regulate the expression of RRM2 by competing for miR-5095. A combination of radiotherapy with LINC00958 silencing, RRM2 downregulation or miR-5095 overexpression was found to inhibit cervical cancer cell proliferation and tumor growth, while promoting cell apoptosis both in vitro and in vivo. Collectively, our results suggest that LINC00958 could regulate RRM2 by competing to miR-5095, which regulates cell sensitivity to radiotherapy in cervical cancer.
Assuntos
Regulação Neoplásica da Expressão Gênica/genética , MicroRNAs/genética , RNA Longo não Codificante/genética , Tolerância a Radiação/genética , Ribonucleosídeo Difosfato Redutase/genética , Neoplasias do Colo do Útero/genética , Animais , Apoptose/genética , Proliferação de Células/genética , Feminino , Células HeLa , Xenoenxertos , Humanos , Camundongos , Camundongos NusRESUMO
The discharge of recombinant DNA waste from biological laboratories into the eco-system may be one of the pathways resulting in horizontal gene transfer or "gene pollution". Heating at 100 degrees C for 5-10 min is a common method for treating recombinant DNA waste in biological research laboratories in China. In this study, we evaluated the effectiveness and the safety of the thermo-treatment method in the disposal of recombinant DNA waste. Quantitative PCR, plasmid transformation and electrophoresis technology were used to evaluate the decay/denaturation efficiency during the thermo-treatment process of recombinant plasmid, pET-28b. Results showed that prolonging thermo-treatment time could improve decay efficiency of the plasmid, and its decay half-life was 2.7-4.0 min during the thermo-treatment at 100 degrees C. However, after 30 min of thermo-treatment some transforming activity remained. Higher ionic strength could protect recombinant plasmid from decay during the treatment process. These results indicate that thermo-treatment at 100 degrees C cannot decay and inactivate pET-28b completely. In addition, preliminary results showed that thermo-treated recombinant plasmids were not degraded completely in a short period when they were discharged into an aquatic environment. This implies that when thermo-treated recombinant DNAs are discharged into the eco-system, they may have enough time to re-nature and transform, thus resulting in gene diffusion.
Assuntos
DNA Recombinante/isolamento & purificação , Calefação , Laboratórios , Pesquisa , Gestão da Segurança/métodos , Gerenciamento de Resíduos/métodos , China , DNA Recombinante/metabolismo , DNA Recombinante/toxicidade , Eletroforese , Concentração Osmolar , Plasmídeos , Reação em Cadeia da Polimerase , Fatores de TempoRESUMO
Analysis of Cu2+ adsorption on cell surface of a Gram-negative bacterium P. putida 5-x was carried out. Isolated cell envelopes possess fivefold more Cu2+ adsorption capacity than that of the fresh intact cells. Experimental results also indicate that the content of the peptidoglycan layer, the outer membrane and inner membrane in the cell envelope is in order of inner membrane > outer membrane > PEG layer, while their Cu2+ adsorption capacity is in order of PEG layer > outer membrane > inner membrane. The total contribution of the separated PEG layer material to Cu2+ adsorption is no more than 15% part, and the cell outer membrane and inner membrane contribute about 30%-35% and 25%-30% part, respectively, to Cu2+ adsorption by the cell envelope. The relatively high phospholipid content in the outer wall might result in its higher adsorption capacity to Cu2+, and leads to a higher adsorption capacity of the cell envelope of P. putida 5-x.