Evaluation and comparison of cytotoxicity, genotoxicity, and apoptotic effects of poly-l-lysine/plasmid DNA micro- and nanoparticles.

The principal impediment to gene therapy is the development of efficient, nontoxic gene carriers that can handle and deliver foreign genetic materials into various cell types, including healthy and cancerous cells. Poly-l-lysine (PLL) polymers are one of the most favorable gene carriers among nonviral vectors, and PLL had low transfection and safety issues. The purpose of this study was to measure cellular toxicity, DNA damage, and apoptotic effects of PLL nanoparticles. Neuro2A mammalian cells were cultured and exposed to PLL/DNA complexes at different polymer/DNA ratios (C/P ratio 2 and 6) for 24 h. To evaluate metabolic activity, genotoxicity, and apoptotic influences of PLL nanoparticle, the following experimental methods were employed, in order: 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), DNA damage (COMET analysis) assay, and sub-G1 peak apoptosis assay. Our data indicate that toxicity is concentration dependent and a high concentration of polymer declined the metabolic activity. In addition, largest complexes (C/P 6 in HEPES buffered saline buffer) have slighter negative impact on metabolic activity. In agreement with our cytotoxicity data, apoptotic assay result represented that increase in size of PLL/DNA complexes decrease the number of apoptotic cells. Also, there was a remarkable increase in percent tail DNA of Neuro2A cells treated with higher concentration of PLL and its polyplexes. The present study demonstrated that PLL/DNA complexes caused cytotoxic, apoptotic, and genotoxic effects in a dose-dependent and weight ratio-dependent manner, which also affected the size of polyplexes.

Sequential or multiplex electrochemical detection of miRs based on the p19 function relative to three sandwiches of different structural hybrids on the liposomal sensor.

In this work, we designed a liposomal electrochemical sensor with DOTAP-DOPE liposome, chimeric probes, p19 as a caliper molecule, and the competitor structural hybrid (just RNA) for detection of three micro-RNAs in one SPCE/GNP electrode. The sensor is stabled when the cationic spherical DOTAP-DOPE liposomes sandwich with hybrids of the different sandwiched of probes (T-M-linear, Stem) and 21-124a-221miRs. With the addition of P19, in the presence of a sandwiched competitor (T-linear/21miR), the system is stable (ON) and is shut off in the presence of structural sandwiched hydrides of M-linear+124a/Stem+221 miR due to the lack of adequate access to segments of RNA-miRs of chimeric probes. For the first time in this study, three probes were sandwiched on the separate liposome for sequential identification of 21-124a-221 or multiplex detection of miRs (221 or 124a with 21) with high specificity and sensitivity (as low as 0.1 fM). Electrochemical impedance (EIS) were performed for sensing three miRs in PBS containing 1 mM [Fe(CN)6]-3/-4 which DOTAP-DOPE liposome acted as an enhancing intermediate layer in the electrochemical reactions. Transmission Electron Microscopy (TEM)), Atomic Force Microscopy (AFM), Dynamic Light Scattering (DLS) and Ultraviolet-Visible (UV) spectroscopic techniques are used to understand the interactions between the DOPE-DOTAP, AuNP, different probes, miRs and p19.

Author Correction: Electrochemical Sensor for Detection of miRs Based on the Differential Effect of Competitive Structures in The p19 Function.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Electrochemical Sensor for Detection of miRs Based on the Differential Effect of Competitive Structures in The p19 Function.

The present study aim to design a liposomal electrochemical sensor using 1, 2-dioleoyl-3-trimethylammoniumpropane (DOTAP) and dioleoylphosphatidylethanolamine(DOPE), chimeric probes and p19, it has been considered as a caliper molecule as well. Also the competitor structural hybrid (RNA) was used to detect three types of miRs in one screen printed electrode modified by gold nanoparticle (SCPE/GNP). In this purpose, the sensor signal stabilized when the cationic DOTAP-DOPE with hybrids of the chimeric probes (Stem, M-linear) sandwiched in order to detect 221-124a miRs. Given the lack of accessibility to RNA-miRs segments of chimeric probes, p19 inhibited the electrochemical reaction and shifted signal to off. After that p19 connected with the free hybrid of T-linear/21miR (just RNA) as competing for structure and the signal was shifted to ON, again. In this study, the electrochemical measurements were performed between the potentials at -0.4 V and +0.4 V with 1 mM [Fe(CN)6]-3-/4 which DOTAP-DOPE acted as an enhancer layer in the electrostatically reaction. This sensor determines as low as 0.4 fM of miRNA with high selectivity and specificity for sequential analysis of 124a-221-21 miRs in just 2 h.