Sevoflurane anesthesia induces apoptosis and cell cycle arrest in NPC-039 nasopharyngeal cells

Purpose: To determine the effects of sevoflurane on NPC-039 nasopharyngeal carcinoma cells.Methods: WST-8 assays and flow cytometry with annexin V/PI staining were used to analyze the effects of sevoflurane on growth and induction of apoptotic changes in NPC-039 cells. Cell viability was performed on a microplate reader.Results: Treatment of NPC-039 cells with 4 – 10 μM sevoflurane significantly inhibited cell growth (p < 0.005) with a median inhibitory concentration of 6 μM. NPC-039 cells incubated with sevoflurane showed prominent nuclear fragmentation and chromatin condensation. NPC-039 cells that exhibited apoptotic symptoms increased from 23.34 to 56.77 % when duration of treatment was increased from 24 to 48 h. Sevoflurane-treated cells also expressed increased levels of Bax and cleaved caspase-3. Reactive oxygen species formation was also higher in sevoflurane-treated NPC-039 cells than in control. Pre-treatment with Z-VAD-FMK followed by incubation with sevoflurane partly reduced the population of apoptotic NPC-039 cells compared with cells treated with sevoflurane alone. The proportion of cells in S- and G0/G1 phases decreased and increased, respectively, when sevoflurane concentration was increased from 2 to 10 μM. Incubating NPC-039 cells with sevoflurane also markedly reduced levels of cyclin-dependent kinases including p15 INK4B, cyclin D1, p16 INK4A, and CDK-6. In contrast, pretreatment with ZVAD-FMK followed by incubation with sevoflurane increased p15 INK4B levels.Conclusion: Sevoflurane inhibits nasopharyngeal carcinoma cell growth, activating apoptosis and inducing cell cycle arrest, thus suggesting its therapeutic potential for the treatment of nasopharyngeal carcinoma.Keywords: Sevoflurane, Nasopharyngeal carcinoma, Chromatin, Condensatio

Stable Agrobacterium-mediated transformation of the halophytic Leymus chinensis (Trin.)

In this study, an efficient procedure for stable Agrobacterium-mediated transformation of Leymus chinensis (Trin.) was established. Agrobacterium tumefaciens strain EHA105, harboring a binary vector pCAMBIA2300, was used for transformation, along with a sweet potato 2-cysteine peroxiredoxin (2-Cys Prx) gene under the control of the stress-inducible sweet potato anionic peroxidase 2 (SWPA2) promoter and the neomycin phosphotransferase (nptII) gene under the control of the cauliflower mosaic virus (CaMV) 35 S promoter. We found that a one-month-old callus derived from mature seeds could be efficiently transformed. Seven-day preculture followed by inoculation with the addition of 100 μmolL-1 acetosyringone (AS) and then a 3 day co-cultivation were performed before selection. Selection of transgenic shoots was done in the presence of 150 mgL-1 kanamycin (KM). An optical density at a wavelength of 600 nm (OD600) of approximately 0.4 for A. tumefaciens infection solution and 20 min of infection time gave the highest transformation efficiency. Polymerase chain reaction (PCR) analysis of KM-resistant plants and newly regenerated rhizomes revealed stable transformation of the 2-Cys Prx gene and the nptII gene, with the highest transformation frequency of 4.93%. RT-RCR analysis was conducted using salt stressed transgenic plants, and the results suggested that 2-Cys Prx had low transcription levels under non-stressed conditions, and increased transcription after 6 h of 200 mM NaCl stress. This gene continued to demonstrate high levels of transcription until 6 h after withdrawal of stress, with a slow recovery. The method reported herein provides a direct opportunity for improvement of the quality traits of L. chinensis via genetic transformation.Keywords: Leymus chinensis, Agrobacterium-mediated transformation, 2-Cys peroxiredoxin, gene transformatio