Carbosilane dendrimers are a non-viral delivery system for antisense oligonucleotides: characterization of dendriplexes.

The success of gene therapy depends on the development of suitable carriers, and because of their architecture dendrimers are promising tools for gene delivery. This research concerns the use of second generation carbosilane dendrimers as carriers for anti-HIV oligodeoxynucleotides (ODNs). The aim was to characterize complexes formed by positively charged dendrimers and negatively charged oligonucleotides using a fluorescence method, laser Doppler electrophoresis, dynamic light scattering (DLS), atomic force microscopy (AFM), transmission electron microscopy (TEM) and molecular modeling. The zeta-potential of ODNs increased from -25 mV to positive values after the addition of dendrimers. DLS and TEM revealed that the diameters of dendriplexes ranged from 75 to 240 nm and from 50 to 260 nm, respectively, and this was dependent on the type of dendrimer and the molar ratios of the complexes formed; complexes were stable for between 100 and 300 minutes. AFM measurements and molecular modeling studies were carried out to determine the structure and size of dendriplexes. The physicochemical properties of the dendriplexes studied and data from previous research suggest that carbosilane dendrimers are good candidates for nucleic acid delivery.

Characterization of complexes formed by polypropylene imine dendrimers and anti-HIV oligonucleotides.

Current anti-HIV therapies are capable of controlling viral infection but do not represent a definitive cure. They rely on the administration of antiretroviral nucleoside analogues, either alone or in combination with vectors. Dendrimers are branched, synthetic polymers with layered architectures, promising non-viral vectors in gene therapy. The aim of the paper was to study the interactions between three anti-HIV antisense oligonucleotides (ODNs): SREV, ANTI TAR, GEM91 and different generation polypropylene imine dendrimers (PPI) by monitoring changes in the fluorescence polarization of fluorescein attached to the ends of the ODNs when increasing concentrations of dendrimers were added. Laser Doppler electrophoresis, dynamic light scattering (DLS) and transmission electron microscopy (TEM) were used to characterize, respectively, zeta potential, particle size and morphology of dendriplexes formed in different molar ratios. Antisense oligonucleotides interacted with polypropylene imine dendrimers in different molar ratios depending on generation. Zeta potential of dendriplexes varied from (-25 to -21) mV to -5 mV (for PPIG3 and PPIG4 complexes) and to zero (for PPIG2 complexes). The structures presented a polydisperse size from about 50 nm to even 700-800 nm by TEM and about 250 nm by DLS. It means that besides single dendriplexes, aggregates were also present.

The effects of the anthocyanin-rich extract from red cabbage leaves on Allium cepa L. root tip cell ultrastructure.

The effect of exogenously applied 250 µM anthocyanin-rich (ATH-rich) extract from red cabbage leaves on the ultrastructure of Allium cepa root meristematic cells was investigated. The tested extract slightly affected mitochondria, endoplasmic reticulum (ER), Golgi apparatus and vacuoles. In the presence of ATH, 62% of mitochondria converted to condensed type. In addition swollen, circular ER cisternae were sporadically observed. In the ATH-treated roots, one third of Golgi structures was characterized by the reduced number of vesicles. Moreover in 54% of vacuoles, the electron-dense granular and circular material appeared. Additionally, in the cytoplasm, the presence of numerous multivesicular bodies (MVB) was noticed. The observed ultrastructural modifications of mitochondria, and presumably also ER, probably resulted from the ability of an ATH to affect mitochondrial respiratory activity. The other changes in A. cepa root meristematic cell ultrastructure were connected with the transport of exogenously applied ATH into vacuoles. It seems that they are transported from the plasmolemma to the vacuole by multivesicular bodies (MVB), and there trapped by anthocyanic vacuolar inclusions (AVIs). However, none of the observed ultrastructural changes seemed to disturb cell functions, therefore the ATH-rich extract from red cabbage leaves may be regarded as cell-friendly and can be safely used as a detoxifying agent against heavy metal poisoning, as it is more and more often postulated.

Use of polyamidoamine dendrimers to engineer BDNF-producing human mesenchymal stem cells.

We report the use of polyamidoamine (PAMAM-NH(2)) dendrimers along with other non-viral vehicles for the in vitro transfection of human bone marrow mesenchymal stem cells (hMSCs) and for engineering MSCs to secrete brain-derived neurotrophic factor (BDNF). Different generations of cationic polyamidoamine dendrimers (generations 3-6) were tested on HEK 293T cells. hMSCs were then transfected with PAMAM-NH(2) G4 dendrimers and Lipofectamine 2000, which elicited the expression of GFP reporter in around 6 and 20% of the cells, respectively. Both vehicles were then shown to elicit the expression of BDNF in MSCs from a bicistronic cassette. Non-virally induced neurotrophin expression may be a safe and easy method for adapting autologous stem cells for therapeutic treatment of diseases and neural system injuries.

Effects of anthocyanin-rich extract from red cabbage leaves on meristematic cells of Allium cepa L. roots treated with heavy metals.

The incubation of Allium cepa L. roots in Pb(NO3)2, Cd(NO3)(2)x4H2O or Cr(NO3)(3)x9H2O solution at the concentration of 100 microM lowered the mitotic index (MI) value in meristem by 58%, 39%, 48%, respectively. The proportion of mitotic phases (mainly prophases and telophases) in MI value was also changed. Moreover, mitotic disturbances such as: c-metaphases, sticky and lagging chromosomes, chromosome bridges, binucleate cells, micronuclei, "budding" nuclei and nucleoli partly outside nuclei were induced in the presence of the tested heavy metals, most frequently after Pb treatment. Pre-incubation in the ATH-rich extract from red cabbage leaves caused 2.5%, 1.8% or 1.6% increase in MI value as compared to the meristematic cells of A. cepa L. roots treated only with Cd, Pb, or Cr, respectively. Additionally, the ATH-rich extract was responsible for changing phase index values towards the control level in the material incubated in Pb or Cd. Moreover, the total number of mitotic abnormalities induced by the tested metals was reduced due to the preincubation in the ATH-rich extract, most effectively in the roots treated with Cd (by 2/3) while by half in Cr presence. These data suggest a protective action of the ATH-rich extract from red cabbage leaves against heavy metal toxicity.