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2-Methacryloyloxy ethyl trimethyl ammonium chloride (TMA) is a potent polymeric plasma DNA (pDNA) carrier. The present study shows that TMA/pDNA polyplexes could be internalized into cells efficiently, but could not mediate gene transfection on its own. The transfection process of TMA/pDNA polyplexes is turned on only when ultrasound (US) was applied 4–8 h after incubating TMA/pDNA polyplexes with target cells (with a gene expression 1000 times that of the immediate US group). US is a widely used physical method for gene delivery; its transfection efficiency can be significantly enhanced when combined with cationic polymer vectors. Traditionally, US is given simultaneously with genetic materials, carriers and microbubbles to exert maximal efficacy. The unique on-off phenomenon of TMA/pDNA polyplexes, controlled by US exposure, was found to relate to the endosomal escape effect of US since the polyplexes colocalized well with the lysosome marker if no US was given or was given at inappropriate times. The proposed delivery system using US and TMA carriers has potential in many pharmaceutical applications requiring precise temporal and spatial release control.  相似文献   

3.
We previously developed artificial promoters that were activated in response to X-ray irradiation. Sonication with 1.0 MHz ultrasound that causes intracellular oxidative stress was found to activate some of these promoters though to lesser degrees. The most sensitive one among these promoters showed intensity- and duration-dependent activations by sonication. In addition, its activation by sonication was attenuated when N-acetyl cysteine was present, suggesting the involvement of intracellular oxidative stress in the activation mechanism. Improved promoters for sensitivity to X-ray irradiation were also found more sensitive to sonication. The most improved one showed 6.0 fold enhancement after sonication with 1.0 MHz ultrasound at 1.0 W/cm2 for 60 s. This enhancement was also attenuated with the presence of N-acetyl cysteine. When stably transfected HeLa cells with the most sensitive promoter were transplanted on to mice and sonicated, luciferase activity by the promoter increased to 1.35 fold in average though it was not statistically significant compared to control. Although gene regulation in vivo by sonication was not clear, this is the first report on artificially constructed promoters responsive to ultrasound.  相似文献   

4.
Efficient and safe gene transfection carriers, especially for hard‐to‐transfect cells, are urgently demanded in basic biological research and gene therapy applications. Many insect cell lines widely used in molecular cell biology exhibit relatively low transfection efficiencies when treated by conventional non‐viral agents. Herein, we develop a novel gene delivery vector by coating graphene oxide (GO) with both polyethylene glycol (PEG) and polyethylenimine (PEI), obtaining a dual‐polymer‐functionalized nanoscale GO (nGO‐PEG‐PEI) to transfect insect cells. While exhibiting remarkably reduced cytotoxicity compared with PEI, nGO‐PEG‐PEI, when used as the plasmid DNA transfection agent to treat Drosophila S2 cells, offers ≈7‐fold and ≈2.5‐fold higher efficiency compared with those achieved by using bare PEI and Lipofectamine 2000, a widely used commercial transfection agent, respectively. Interestingly, the advantages of nGO‐PEG‐PEI are even more dramatic when transfecting cells with lower‐quality linearized DNA. It is revealed that nGO‐PEG‐PEI/pDNA complexes enter insect cells via a unique pathway working even at a low temperature, rather different from their entry into mammalian adherent cells. Our results encourage the development of nano‐GO‐based gene carriers to treat special types of hard‐to‐transfect cells (e.g., insect cells), and indicate that nanomaterials would enter cells by cell‐type‐dependent mechanisms, which merit significantly more future attentions.  相似文献   

5.
Since fecal coliforms was introduced as a standard indicator of pollutants in effluents of municipal wastewater treatment plants in China in 2003, chlorine had been widely used in many wastewater treatment plants. However, concerns about the disinfection by-products (DBPs) of chlorine have been increasing. One of the effective way to reduce the production of DBPs is to reduce the effective chlorine dosage by improving the utilization rate of disinfectant. Ultrasound (US) is proved to be effective in wastewater treatment for its multiple chemical and physical effects produced by cavitation, which could favor the disinfection process accordingly. For the purpose of improving disinfection efficiency with the help of US, following points are addressed in the current study: (1) investigate the enhancement effects of US on the disinfection efficiency of sodium hypochlorite (NaClO) for real secondary effluents of municipal wastewater treatment plants; (2) evaluate the possibility of using US specific energy consumption (kJ/L) as an parameter for disinfection efficiency evaluation; and (3) quantify the reduction in chlorine-DBPs through US application. Results demonstrated that sonication could reduce two-thirds (US pretreatment) or one-third (simultaneous US and NaClO disinfection) of the required concentrations of NaClO (available chlorine) for 4 log reduction of fecal coliforms, which could meet the Class 1A (fecal coliforms less than 1000 CFU/L) discharge standard of China. In addition, US pretreatment with NaClO disinfection performed better enhancement in disinfection efficiency compared with simultaneous US and NaClO disinfection. Furthermore, analysis on DBPs showed that US application as pretreatment could obviously reduce the contents of trichloromethane (TCM) and trichloroacetic acid (TCAA) by more than 85% and 50%, respectively, compared with NaClO disinfection alone for the same disinfection efficiency. Meanwhile, the experimental results also showed that the disinfection efficiency and DBPs concentration were only slightly affected under a constant US specific energy consumption, although input power density and irradiation time changed, indicating that specific energy consumption (kJ/L) could be considered as a better control parameter for disinfection efficiency evaluation.  相似文献   

6.
To identify the factors involved in the inhibition of ultrasound (US)-induced free radical production and cell killing by pre-sonication incubation or by high cell density, we used different densities of U937 cells, and with (up to 2 h) or without pre-sonication incubations, the cell suspensions were exposed to 1 MHz US (10% duty factor at 100 Hz pulse rate; intensities 0.1-0.5 W/cm(2) for 1 min). The intensity 0.3 W/cm(2) was used for cell killing experiments and 0.5 W/cm(2) for free radical experiments. Free radical production was determined by electron paramagnetic resonance (EPR)-spin trapping with DMPO while cell killing was determined by assays for lysis, loss of cell viability, apoptosis and necrosis. The results show that at higher cell densities, CO(2) in the medium rapidly increased, with shorter pre-sonication incubation required to attain complete inhibition of both free radical production and cell killing. Cell killing at 0.3 W/cm(2) and free radical production at 0.5 W/cm(2) were both inhibited at 10 million cells/ml without incubation, and at 2 million cells/ml incubated for 2 h before sonication. Level of CO(2) alone could not account for the inhibition; consumption of gases in the medium is also considered in the inhibitory effect of pre-sonication, while suppression of cavitational activities due to the "viscosity effect" is considered a more important factor in the inhibition by high cell density.  相似文献   

7.
Ultrasonic frequency effects on the removal of Microcystis aeruginosa   总被引:4,自引:0,他引:4  
Algae bloom in source water causes high chemical consumption and deteriorates water quality in waterworks. This paper studied the ultrasonic removal of Microcystis aeruginosa. The results showed that algae cells could be effectively removed by sonication and gas vesicle collapse was the main mechanism. The ultrasonic algae removal followed the first order reaction with a rate constant of 0.023 min(-1) (80 W, 80 kHz). Higher ultrasound frequency benefited algae removal; the algae removal rate constant was 0.114 min(-1) at 1320 kHz and 0.0224 min(-1) at 20 kHz (80 W). Higher ultrasound power also accelerated algae removal; the algae removal rate constant was 0.023 min(-1) at 80 W and 0.007 min(-1) at 32 W (80 kHz). However, high ultrasound power and long irradiation caused microcystins to increase. 80 W, 80 kHz sonication for 5 min increased the extracellular microcystins concentration from 0.87 microg/L to 3.11 microg/L. Sound frequency had little impact on the microcystins release. The chlorophyll a concentration initially decreased and then stabilized after 5 min of sonication.  相似文献   

8.
We fabricated three piezoelectric components (PZT) that can produce ultrasonic waves with various generated power in order to improve the delivery of DNA molecule and polymer/DNA complexes into cells. Two cationic polymers (PEI and PDMAEMA) were interacted with DNA to form nano-scaled DNA/polymer complexes with/without the help of PZT devices. The application of PZT devices under optimal conditions helped to avoid cytotoxicity and greatly increased the transfection (DNA delivery) efficiency of these complexes in mammalian cells. The cytotoxicity and transfection efficiency were found to be correlated with the PZT-generated power, waveforms and duration of ultrasonic treatment. There was no observable cytotoxicity in our experimental models and, a maximum transfection efficiency 700% greater than that of polymer/DNA complexes without applying ultrasound was achieved. The transfection efficiency of plain polymer/DNA complexes (without PZT treatment) corresponded to a 630-fold increase in comparison to the naked DNA. The waveforms of generated ultrasound greatly influenced the transfection efficiency, while cytotoxicity was not significantly affected. This means that, for optimal DNA delivery, duration of the peak voltage (Vmax/Div) also plays a role. In addition, the generated waves from PZT do not cause dissociation of polymer/DNA complexes or a change in the particle sizes of these complexes. In conclusion, these results suggest that the operation of PZT devices can be a tunable/safe way to greatly improve DNA delivery for gene therapy.  相似文献   

9.
Sonication improves kasturi lime (Citrus microcarpa) juice quality   总被引:1,自引:0,他引:1  
Freshly squeezed kasturi lime fruit juice was sonicated (for 0, 30 and 60min at 20°C, 25kHz frequency) to evaluate its impact on selected physico-chemical and antioxidant properties, such as pH, °Brix, titratable acidity, Hunter color values (L(?), a(?), b(?)), ascorbic acid, DPPH radical scavenging activity, total phenolics, antioxidant capacity, flavonoids and flavonols. Additionally, the effect of sonication treatments on the microbial load (TPC, yeast and mold) were also evaluated. Sonication of juice samples for 60min showed enhancement in most of the bioactive compounds compared to samples treated for 30min and control samples (untreated). Significant reductions in the microbial load corresponding to sonication time were also recorded. Results of the present study indicate that sonication may be employed as a suitable technique for kasturi lime juice processing, where antioxidant and other bioactive compound retention or enhancement is desired, along with the achievement of safety and quality standards.  相似文献   

10.
Various inorganic nanoparticle designs have been developed and used as non-viral gene carriers. Magnetic gene carriers containing polyethyleneimine (PEI), a well-known transfection agent, have been shown to improve DNA transfection speed and efficiency in the presence of applied magnetic field gradients that promote particle–cell interactions. Here we report a method to prepare iron oxide nanoparticles conjugated with PEI that: preserves the narrow size distribution of the nanoparticles, conserves magnetic properties throughout the process, and results in efficient transfection. We demonstrate the ability of the particles to electrostatically bind with DNA and transfect human cervical cancer (HeLa) cells by the use of an oscillating magnet array. Their transfection efficiency is similar to that of Lipofectamine 2000?, a commercial transfection reagent. PEI-coated particles were subjected to acidification, and acidification in the presence of salts, before DNA binding. Results show that although these pre-treatments did not affect the ability of particles to bind DNA they did significantly enhanced transfection efficiency. Finally, we show that these magnetofectins (PEI-MNP/DNA) complexes have no effect on the viability of cells at the concentrations used in the study. The systematic preparation of magnetic vectors with uniform physical and magnetic properties is critical to progressing this non-viral transfection technology.  相似文献   

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