Reneta Toshkova

Transplacental clastogenic and epigenetic effects of gold nanoparticles in mice.

The broad application of nanotechnology in medicine, biology, and pharmacology is leading to a dramatic increase of the risk of direct contact of nanoproducts, among which gold nanoparticles (AuNP), with the human organism. The present study aimed at evaluating in vivo the genotoxicity of AuNPs with average size of 40 nm and 100 nm. A single intraperitoneal treatment of adult male and female Swiss mice (strain H) with AuNPs, at a dose of 3.3 mg/kg body weight, had no effect on the frequency of micronucleated polychromatic erythrocytes (MN PCEs) in bone marrow. Conversely, the transplacental treatment with AuNP-100 nm, but not with AuNP-40 nm, applied intraperitoneally at a dose of 3.3 mg/kg to pregnant mice on days 10, 12, 14, and 17 of gestation, resulted in a significant increase in the frequency of MN PCEs in both liver and peripheral blood of mouse fetuses. In parallel, the same treatment with AuNP-100 nm, but not with AuNP-40 nm, produced significant changes in microRNA expression. In particular, out of 1281 mouse microRNAs analyzed, 28 were dys-regulated more than two-fold and to a statistically significant extent in fetus lung, and 5 were up-regulated in fetal liver. Let-7a and miR-183 were significantly up-regulated in both organs. The data presented herein demonstrate for the first time the transplacental size-dependent clastogenic and epigenetic effects of AuNPs in mouse fetus, thus highlighting new aspects concerning the putative genotoxicity of AuNPs during a vulnerable period of life.

Electrospun Nanofibrous Mats Containing Quaternized Chitosan and Polylactide with In Vitro Antitumor Activity against HeLa Cells

Nanofibrous materials containing the antitumor drug doxorubicin hydrochloride (DOX) were easily prepared using a one-step method by electrospinning of DOX/poly(L-lactide-co-D,L-lactide) (coPLA) and DOX/quaternized chitosan (QCh)/coPLA solutions. The pristine and DOX-containing mats were characterized by ATR-FTIR and X-ray photoelectron spectroscopy (XPS). The release rate of DOX from the prepared fibers increased with the increase in DOX content. The DOX release process was diffusion-controlled. MTT cell viability studies revealed that incorporation of DOX and QCh in the nanofibrous mats led to a significant reduction in the HeLa cells viability. It was found, that the antitumor efficacy of the DOX-containing mats at 6 h was higher than that of the free DOX. SEM, TEM, and fluorescence microscopic observations confirmed that the antitumor effect of QCh-based and DOX-containing fibrous mats was mainly due to induction of apoptosis in the HeLa cells.

Antitumor activity of quaternized chitosan-based electrospun implants against Graffi myeloid tumor

Nanofibrous implants containing quaternized chitosan (QCh), poly(L-lactide-co-D,L-lactide) (coPLA), and the antitumor drug doxorubicin (DOX) were fabricated by electrospinning. The surface chemical composition and the morphology of the implants were characterized by XPS and SEM. In vitro cell viability studies demonstrated that QCh- and DOX-based implants exhibited high cytotoxicity against Graffi tumor cells. The implants efficiently inhibited the growth of Graffi tumor in hamsters with minimum weight loss. Insertion of QCh/coPLA/DOX implants in the place of removed tumor led to an increase in the animal survival rate and to a decrease in the percentage of recurrences.

A novel glycosylated Cu/Zn-containing superoxide dismutase: production and potential therapeutic effect

The fungal strain Humicola lutea 103 produces a naturally glycosylated Cu/Zn SOD. To improve its yield, the effect of an increased concentration of dissolved oxygen (DO) on growth and enzyme biosynthesis by the producer, cultivated in a 3 l bioreactor, was examined. Exposure to a 20% DO level caused a 1.7-fold increase of SOD activity compared to the DO-uncontrolled culture. Maximum enzyme productivity of SOD was approximately 300 x 10(3) U (kg wet biomass)(-1). The novel enzyme was purified to electrophoretic homogeneity. The presence of Cu and Zn were confirmed by atomic absorption spectrometry. The molecular mass of H. lutea Cu/Zn SOD was calculated to be 31870 Da for the whole molecule and 15936 Da for the structural subunits. The N-terminal sequence revealed a high degree of structural homology with Cu/Zn SOD from other prokaryotic and eukaryotic sources. H. lutea Cu/Zn SOD was used in an in vivo model for the demonstration of its protective effect against myeloid Graffi tumour in hamsters. Comparative studies revealed that the enzyme (i) elongated the latent time for tumour appearance, (ii) inhibited tumour growth in the early stage of tumour progression (73-75% at day 10) and (iii) increased the mean survival time of Graffi-tumour-bearing hamsters. Moreover, the fungal Cu/Zn SOD exhibited a strong protective effect on experimental influenza virus infection in mice. The survival rate increased markedly, the time of survival rose by 5.2 d and the protective index reached 86%. The H. lutea SOD protected mice from mortality more efficiently compared to the selective antiviral drug ribavirin and to commercial bovine SOD. In conclusion, our results suggest that appropriate use of the novel fungal SOD, applied as such or in combination with selective inhibitors, could outline a promising strategy for the treatment of myeloid Graffi tumour and influenza virus infection.

Antitumour activity of novel 1,10-phenanthroline and 5-amino-1,10-phenanthroline derivatives

Synthesis and impact on the tumour growth of palladium(II) complex of 5-amino-1,10-phenanthroline Pd(5-NH(2)-phen)(2)(NO(3))(2) and the protonated dimer (phen)(2)(H(+))(BF(4)(-)) have been described. In the reported experiments a cancerous (100% lethality) myeloid subcutaneous tumour (with Graffi-tumour origin) in hamsters was used. The animals were injected i.p. with different doses of the substances. The longest mean survival time (1.65 times longer than the controls) was achieved when the substance Pd(5-NH(2)-phen)(2)(NO(3))(2) was injected into the animals. One of the animals even survived until the 71st day, which is 2.2 times longer than the controls. The compound (phen)(2)(H(+))(BF(4)(-)) prolonged the mean life-time of the animals 1.4 times in comparison to the controls. On the other hand, the Pd(II) complex of 1,10-phenanthroline, Pd(phen)(2)(H(2)O)(NO(3))(2), did not reveal any antitumour activity. Our experience concerning the effect of other drugs on this tumour has shown a survival time no longer than 4-5d after the death of the controls.

Biological activities of selected polyphenol-rich fruits related to immunity and gastrointestinal health

Small fruits are a rich source of bioactive substances, including polyphenols, and are therefore suitable raw materials for the production of functional foods. In the current work, we studied the antioxidative properties of six fruits: rosehip, chokeberry, hawthorn, blackcurrant, blueberry and rowanberry via different methods (ORAC, TRAP, HORAC and inhibition of lipid peroxidation). Their effect on the production of reactive oxygen species (ROS) by phagocytes, antimicrobial properties against 11 human pathogens, and mitogenic effect on hamster spleen lymphocytes were also tested. Rosehip extract showed the highest antioxidant activity via ORAC, TRAP and HORAC assays, whereas blueberry extract was the most potent inhibitor of lipid peroxidation. All extracts inhibited ROS production of opsonized zymosan-activated phagocytes, indicating that extracts interfere with the signaling cascade of phagocyte activation upstream to the protein kinase C activation. Chokeberry, blackcurrant and rowanberry extracts revealed strong antimicrobial properties against a broad spectrum of microorganisms and also had the highest mitogenic activity.

Antitumor Activity of Glycosylated Molluscan Hemocyanins via Guerin Ascites Tumor

As observed in most molluscan hemocyanins, high-mannose type glycans were identified in hemocyanins from Rapana venosa (RvH), Helix lucorum (HlH) and keyhole limpet (Megatura crenulata). In addition, a glycan with a branching structure containing xylose, fucose and terminal methyl hexose was identified in β-HlH. We have examined the immuno-adjuvant properties of hemocyanins, their derivatives and conjugates associated with the cell mediated immunity in experimental tumor-bearing animals with ascites tumor of Guerin. After immunization of the animals with the experimental vaccine preparations, the highest values of splenic lymphocytes were observed in groups immunized with the conjugates RvH-TAg, β-HlH-TAg and KLH-TAg (42.3%; 40.8% and 40.58%, respectively) than with the native hemocyanins (36.5%; 35.1% and 32.4%, respectively). The immunization of rats with the hemocyanins β-HlH, RvH and KLH and their conjugates, prolonged the median survival time of tumor-bearing animals compared with non-immunized animals (39, 33, 31 and 7 days, respectively). Both hemocyanins β-HlH and RvH activate the immune system of the experimental animals and therefore could be a good alternative for KLH. For this reason they could be included into the composition of non-specific anti-tumor vaccines to enhance their effectiveness.

Cancer Protective Action of Polysaccharide, Derived from Red Microalga Porphyridium Cruentum—A Biological Background

ABSTRACT The cell wall sulfated polysaccharide of the red microalga Porphyridium cruentum (Rhodophyta) (PcrPSH) exhibited strong antitumor activity against Graffi myeloid tumor in hamsters both in vitro and in vivo. When tested in vivo, depending on the concentration and the way of application, this polysaccharide decreased transplantability in all experimental groups till 20 days of observation and mortality rate. The tumor growth was retarded and the mean survival time was prolonged with 10 - 16 days. Applied in in vitro experiments the PcrPSH increased both—spreading and phagocytic ability of peritoneal macrophages from healthy and Graffi tumor bearing hamsters in a dose—dependent manner. Primary Graffi tumor cell culture, cultivated in the presence of PcrPSH showed significant decrease of cell viability, determined by MTT test, while in cells derived from bone marrow it was increased at the same conditions of cultivation and concentration of polysaccharide. Primary Graffi tumor cell culture, treated with PcrPSH showed the appearance of a characteristic DNA ladder on agarose gel electrophoresis, which is a biochemical hallmark of apoptosis. The manifested anticancer activity of PcrPSH could be associated with its immunostimulating action as well as with direct cytotoxic properties. Based on these results, we could suggest that the tested algal PcrPSH is a promising candidate as an antitumor agent. Further studies will be done to clarify the mechanisms of a biological action of PcrPSH.

Poly(3-hydroxybutyrate)/caffeic acid electrospun fibrous materials coated with polyelectrolyte complex and their antibacterial activity and in vitro antitumor effect against HeLa cells.

Abstract The purpose of this work was to investigate the possibility for the preparation of new poly(3-hydroxybutyrate) (PHB)/poly(ethylene glycol) (PEG)-based fibrous materials containing natural phenolic compound caffeic acid (CA) of diverse architectures, as well as to study the impact of the fiber composition on the in vitro CA release profile and on the biological properties of the fibrous materials. The application of the one-pot electrospinning enabled the fabrication of nanofibrous materials from PHB and PEG loaded with the CA. Materials with targeted design were obtained by coating with polyelectrolyte complex of alginate (Alg) and N,N,N-trimethylchitosan (TMCh). Three different processing paths were used to obtain coated mats: (i) with CA incorporated in the PHB/PEG core; (ii) with CA embedded in the Alg layer; and (iii) with CA included in the TMCh layer. The in vitro release of CA was modulated by controlling the composition and the architecture of the nanofibrous mats. The performed microbiological screening and MTT cell viability studies revealed that in contrast to the bare mats, the CA-containing nanofibrous materials were effective in suppressing the growth of the Gram-positive bacteria Staphylococcus aureus and the Gram-negative bacteria Escherichia coli and displayed good cytotoxicity against human cervical HeLa tumor cells. In addition, the proliferation of murine spleen lymphocytes and peritoneal macrophages was increased by the prepared CA-containing nanofibrous materials. The obtained materials are promising for antibacterial wound dressing applications as well as for application in local treatment of cervical tumors.

Quaternized chitosan-coated nanofibrous materials containing gossypol: preparation by electrospinning, characterization and antiproliferative activity towards HeLa cells.

Nanofibrous polylactide-based materials loaded with a natural polyphenolic compound gossypol (GOS) with antitumor properties were prepared by electrospinning. The nanofibrous materials were coated with a thin film of crosslinked quaternized chitosan (QCh). GOS incorporated in the nanofibrous mats was in the amorphous state. GOS release was diffusion-controlled and its in vitro release profiles depended on the mat composition. The nanofibrous materials exhibited high cytotoxicity towards HeLa tumor cells. Interestingly, it was particularly pronounced in the case of fibrous materials, which contain both QCh and GOS. The observed strong antiproliferative effect of the nanofibrous mats was mainly due to induction of cell apoptosis.