Role of Endocytosis Pathways in Electropermeablization of MCF7 Cells Using Low Voltage and High Frequency Electrochemotherapy.

OBJECTIVE: The cell membrane is a major barrier for delivery of hydrophilic drugs and molecules into the cells. Although low voltage and high frequency electric fields (LVHF) are proposed to overcome the cell membrane barrier, the mechanism of membrane permeabilization is unclear. The aim of study is to investigate endocytosis pathways as a possible mechanism for enhancing uptake of bleomycin by LVHF. MATERIALS AND METHODS: In this experimental study, MCF-7 cells were exposed to bleomycin or to electric fields with various strengths (10-80 V/cm), frequency of 5 kHz, 4000 electric pulse and 100 µs duration in the presence and absence of three endocytosis inhibitors-chlorpromazine (Cpz), amiloride (Amilo) and genistein (Geni). We determined the efficiency of these chemotherapeutic agents in each group. RESULTS: LVHF, depending on the intensity, induced different endocytosis pathways. Electric field strengths of 10 and 20 V/cm stimulated the macropinocytosis route. Clathrin-mediated endocytosis was observed at electric field intensities of 10, 30, 60 and 70 V/cm, whereas induction of caveolae-mediated endocytosis was observed only at the lowest electric field intensity (10 V/cm). CONCLUSION: The results of this study imply that LVHF can induce different endocytosis pathways in MCF-7 cells, which leads to an increase in bleomycin uptake.

Endocytosis induction by high-pulsed magnetic fields to overcome cell membrane barrier and improve chemotherapy efficiency.

Cell membrane acts as a barrier to the entry of impermeable drugs into cells. Recent studies have suggested that using magnetic fields can enable molecules to overcome the cell membrane barrier. However, the mechanism of membrane permeabilization remains unclear. Therefore, we evaluated the increases in bleomycin (CT) uptake, a non-permanent chemotherapy agent, using high-pulsed magnetic fields and investigated whether endocytosis was involved in the process. This study exposed MCF-7 cells to magnetic fields (2.2 T strength, different number of 28 and 56 pulses, and frequency of 1 and 10 Hz) in order to investigate whether this approach could promote the cell-killing efficiency of bleomycin. The involvement of endocytosis as a possible mechanism was tested by exposing cells to three endocytosis inhibitors, namely chlorpromazine, genistein, and amiloride. Our results illustrated that magnetic fields, depending on their conditions, could induce different endocytosis pathways. In such conditions as 10 Hz-28 pulses, 10 Hz-56 pulses, and 1 Hz-56 pulse, clathrin-mediated endocytosis was observed. Moreover, macropinocytosis was induced by the 10 Hz magnetic field and caveolae-mediated endocytosis occurred in all the magnetic field conditions. The findings imply that high-pulsed magnetic fields generate different endocytosis pathways in the MCF-7 cells, thus increasing the efficiency of chemotherapy agents.

Silver nanoparticles and electroporation: Their combinational effect on Leishmania major.

Leishmaniasis is an emerging and uncontrolled disease. The use of routine drugs has been limited due to proven side effects and drug resistance. Interestingly, novel approaches such as nanotechnology have been applied as a therapeutic modality. Silver nanoparticles have shown antileishmanial effects but because of their nonspecific and toxic effects on normal cells, their use has been limited. On the other hand, it has been demonstrated that electric pulses induce electropores on cell membranes resulting in higher entrance of certain molecules into cells. There is a hypothesis proposing that use of electroporation and silver nanoparticles simultaneously can induce greater accumulation of particles in infected cells, besides higher toxicity. In this study, after applying electric pulses with different concentrations of silver nanoparticles (SNPs), cell survival rate was determined by standard viability assays. On the basis of these data, 2 µg/ml of SNPs and 700 V/cm with 100 µs duration of electroporation were selected as the non-lethal condition. Promastigotes and infected macrophage cells received both treatments and the survival percentage and Infection Index were calculated. In parasites and cells receiving both treatments, higher toxicity was observed in comparison to each treatment given individually, showing a synergic effect on promastigotes. Therefore, application of electric pulses could overcome limitations in using the antileishmanial properties of silver nanoparticles.

Silver nanoparticles increase cytotoxicity induced by intermediate frequency low voltages.

Electrical properties of the cells play a key role in biological processes. Intermediate frequencies of electrical fields influence the cells proliferation without heat generation and electrical stimulation. Silver nanoparticle (SNP) as a metallic agent can change the electrical characteristics of the cells. We study the effect of low voltages at an intermediate frequency (300 kHz) on a human breast adenocarcinoma cell line (MCF7) in the presence of SNPs. At first, cell toxicity of SNPs was determined at different concentrations. Then three different voltages were applied to the cells for 15 min, both in the presence and absence of SNPs. The treatments efficiency was evaluated by MTT assay. The results showed that the intermediate frequency-low voltages with SNPs not only provide an additive efficacy on cytotoxicity, but also a synergism was observed between these factors. By increasing the voltage from 3 to 9 V, a rising synergistic rate was observed. It seems that the synergistic effect between SNPs and the 300 kHz low voltages can inhibit cell proliferation and/or increases cell death of MCF-7, and hence increases treatment efficiency of SNPs, effectively.