Effect of Silicon Dioxide Nanoparticles on Syrian Hamsters Infected by Opisthorchis felineus: 1H MRS Study of the Brain.

In recent years, silicon dioxide nanoparticles have been widely used in medicine and the pharmaceutical industry, however, their effect on the brain has hardly been studied. We assessed the effects of long-term consumption of 5-nm amorphous silicon dioxide nanoparticles (SiO2-NPs) by Syrian hamsters infected with the trematodes Opisthorchis felineus on the hippocampus and frontal cortex. Spectroscopic determination of brain neurometabolites, performed using a horizontal Magnetic Resonance Imaging system at 11.7 Tesla magnetic field, has shown that the ratio of the excitatory neurotransmitters (glutamate + glutamine + aspartate) to the inhibitory ones (GABA + glycine) was higher in the animals infected with O. felineus. However, pre-consumption of the SiO2-NPs solution prevented this imbalance. In addition, the protective effect of SiO2-NPs on the level of myo-inositol and glycine was found. It is concluded that the use of SiO2-NPs can neutralize the negative effects of infectious factors on the brain.

Genomics and proteomics of the liver fluke Opisthorchis felineus.

The causative agent of opisthorchiasis, the liver fluke Opisthorchis felineus (Rivolta, 1884) is one of the helminths of humans and animals in Russia. Together with closely related species of trematodes O. viverrini (Poirier, 1886) and Clonorchis sinensis (Loos, 1907), O. felineus is a part of a triad of epidemiologically important trematodes in the family Opisthorchiidae. Adult O. felineus worms infest the hepatobiliary system of warm-blooded animals and might provoke the development of severe pathologies, including malignancy of bile duct epithelium. The high medical importance of O. felineus attracts the attention of researchers. This review briefly summarizes the data about O. felineus genomics and proteomics. The review provides a comparative analysis of the number of genes and sizes of nuclear genomes of a number of flatworms, the distribution of intron lengths, as well as results of synteny between the O. felineus, O. viverrini and C. sinensis genomes. Special attention is paid to a particular form of RNA processing known as trans-splicing, widely presented in the opisthorchiid genomes. We also provide the results of a comparative analysis of the xenobiotic metabolizing system between parasitic and free-living flatworms. Moreover, data on parasitic granulins, which are potential promoters of cholangiocyte neoplasia, are also presented. Data on the O. felineus genomics and proteomics provide first insights into the structural and functional organization of the genome of this parasitic flatworm with a complex life cycle as well as provide a significant contribution to our understanding of "host-parasite" interaction and evolution of this group of parasitic flatworms.

Effectiveness of Repeated Administration of Praziquantel with Disodium Glycyrrhizinate and Two Enantiomers of Praziquantel on Opisthorchis felineus (Rivolta, 1884).

BACKGROUND: Nowadays, it is still important to develop effective anti-opisthorchiasis agents. In this work, we tested a complex of praziquantel (PZQ) with a plant origin compound-disodium glycyrrhizinate-in the ratio 1:10 PZQ:Na2GA, containing 11-fold less of the active ingredient. Our aim was to study various ways to treat trematode Opisthorchis felineus with this complex in vitro. Additionally, an in vitro comparison of the anthelmintic action was made among racemic-PZQ, (R)-PZQ, and (S)-PZQ on juvenile and adult maritae of O. felineus. METHODS: Worms extracted from the hamsters were subjected to various regimens of administration of the complex: once a day for 3 days or three times within 1 day. Moreover, mature maritae and juvenile worms of O. felineus were subjected to the comparison the anthelmintic effectiveness of racemic-PZQ, (R)-PZQ, and (S)-PZQ. RESULTS: The O. felineus maritae that received PZQ:Na2GA (1:10) thrice within 1 day were most strongly affected by the drug. Their motility substantially decreased already on the second day after the last dose, and the percentage of live worms by the end of the experimental period was the lowest. These results indicate a cumulative anthelmintic effect of this substance under the regimen "three times within 1 day." For the first time, we report that among the three substances (racemic-PZQ and two enantiomers), (R)-PZQ has the highest anthelmintic activity, toward both juvenile and sexually mature maritae of O. felineus. CONCLUSION: These findings suggest that the development of a supramolecular complex of (R)-PZQ with disodium glycyrrhizinate and administration of this complex three times within 1 day are promising approaches.

Physicochemical Properties and Anti-Opisthorchosis Effect of Mechanochemically Synthesized Solid Compositions of Praziquantel with Glycyrrhizic Acid Disodium Salt.

The mechanochemical preparation of solid compositions of praziquantel with plant saponin (glycyrrhizic acid disodium salt) is described. The study of a number of physicochemical parameters showed that dissolving solid compositions in water is accompanied by the inclusion of praziquantel molecules into micelles, which are formed in the solution of the glycyrrhizic acid disodium salt. Using the opisthorchiasis model caused by Opisthorchis felineus, we found a 4- to 11-fold increase in the anthelmintic activity of praziquantel in the composition as compared to the official praziquantel. According to the pharmacokinetic data, the use of the composition increased the bioavailability of praziquantel 3 times.

Secretion of Thioredoxin Peroxidase Protein of Cat Liver Fluke Opisthorchis felineus during Modeling of Experimental Opisthorchiasis.

Mechanisms of thioredoxin peroxidase secretion by Opisthorchis felineus were studied in vivo and in vitro. Specific antibodies were obtained and used for western blotting and immunohistochemical detection in Syrian hamster model of opisthorchiasis. Secreted thioredoxin peroxidase protein was accumulated in the worm incubation medium under conditions of oxidative stress and in bile duct cells of hamsters with chronic opisthorchiasis.

[Role of opisthorchis felineus on induction of bile duct cancer].

Opisthorchis felineus (Trematoda) is widespread in the Russian Federation, especially in Siberia, and other countries of Europe. Infestation of endemic area population with O. felineus reaches 80%. On animal models of the infection of closely related Opisthorchis viverrini combined with the nitrosamines' intake it has been shown that the parasite induces cholangiocarcinoma. However carcinogenic potential of O. felineus is still poorly studied. The present study is aimed to investigate the role of O. felineus in cholangiocarcinoma carcinogenesis in hamster treated additionally by dimethylnitrosamine (DMN). Golden hamsters were divided into 4 groups (15 specimens in the control group and 20 for other groups): (I) untreated control, (II) 12.5 ppm DMN solution intake, (III) infected with 50 metacercariae of O. felineus and (IV) infected with 50 metacercariae of O. felineus and 12.5 ppm DMN solution intake. According to the histological data, in the. O. felineus-infested group significant hyperplastic and dysplastic biliary changes were found considered as a precancerogenic state. Such pathological changes of bile ducts were more severe in group treated with both factors, with cholangiocarcinoma being found out at 18th week in all the animals of this group. These results demonstrate that O. felineus could play promoting role in two-step model in cholangiocarcinoma carcinogenesis and may be used to define the O.felineus group in the International Agency for Research on Cancer classification of agents, mixtures and exposures (IARC categories).

[Secretome of the adult liver fluke Opisthorchis felineus].

The opisthorchiasis caused by Opisthorchis felineus, the Siberian liver fluke remains a serious public health problem in Russia and Eastern Europe. Proteomic identification of the proteins in the excretory-secretory products (ESPs) released by O. felineus is an important key for the investigation of host-parasite interactions and understanding the mechanisms involved in parasite survival within the host. In the ESP of O. felineus we have identified 37 proteins using high-resolution proteomics approach (LTQ-FT-ICR mass spectrometer). The O. felineus secretes either excretes a complex mixture of proteins including: glycolytic enzymes (enolase, aldolase, fructose-1 ,6-bisphosphatase and other); detoxification proteins (4 isoform of glutathione S-transferases, Cu/Zn superoxide dismutase, thioredoxin peroxidase, thioredoxin); cytoskeletal proteins (beta tubulin and paramyosin); a number of proteases (cathepsin F, B1, leucin aminopeptidase 2); protease inhibitors (putative cys1 protein, leukocyte elastase inhibitor), binding proteins (ferritin, myoglobin, FABP) and other. In the O. felineus ESP we also identified Of-HDM protein belonging to a novel family "helminth defence molecules" (HDMs). The O. felineus proteins identified in this study provide necessary information for the further investigation of molecular mechanisms of opisthorchiasis pathogenesis and some of them would be of interest as potential antigens for vaccine and immunodiagnostics development and as potential new anthelmintic drug targets.

[Oncolytic properties of some orthopoxviruses, adenoviruses and parvoviruses in human glioma cells].

UNLABELLED: Currently one of the most promising approaches in development of cancer virotherapy is based on the ability of oncolytic viruses to selective infection and lysis of tumor cells. AIM: The goal of the study was to identify and evaluate perspective oncolytic viruses capable of selectively destroying human glioma cells. PATIENTS AND METHODS: Original GB2m, GA14m and GB22m glioma cell cultures derived from patients were used for evaluating in vitro oncolytic activity of some typical orthopoxviruses, adenoviruses and parvoviruses. RESULTS: The oncolytic activity in the human glioma cell models was confirmed for LIVP and WR strains of vaccinia virus, Adel2 and Ad2del strains with deletions within E1B/55K gene and derived from human adenoviruses type 2 and 5, respectively. CONCLUSIONS: We consider these oncolytic viruses as promising agents for the treatment of human malignant glioma.

[Oncolytic viruses in the therapy of gliomas].

Despite the advances of modern medicine, malignant glioblastoma cure remains an elusive goal. Both the invasive nature and location in vital areas of the brain make this type of tumors difficult for surgical treatment, while the current adjuvant therapy is not as successful as expected. Frequent recurrence and invasiveness of malignant gliomas is due to resistance of glioma stem cells to conventional radiation and chemotherapy. Technological advances in constructing recombinant viruses have allowed creating strains with high oncolytic activity toward glial tumors. Many of these strains have passed Phase I of clinical trials and demonstrated high safety. Despite the obvious potential of the approach, efficiency of the existing strains is still far from being sufficient for effectively curing the disease and require further improvement. The review summarizes results obtained with the most successful variants of oncolytic viruses that come down to the clinical trials and discusses the prospects for new approaches in virotherapy of malignant gliomas.

[DNA diagnosis of mixed invasions of Opisthorchis felineus and Metorchis bilis by polymerase chain reaction].

Opisthorchiasis is one of the significant naturofocal diseases in Russia. The diagnosis of opisthorchiasis may mask some diseases, such as opisthorchiasis, metorchiasis, and clonorchiasis - biohelminthoses that are induced by various representatives of the family Opisthorchiidae-Opisthorchis felineus/O.viverrini, Metorchis bilis, and Clonorchis sinensis, respectively. Coproovoscopy and serologic methods fail to accurately define the species-specific affiliation; in this connection the identification of opisthorchids, by using DNA diagnostic techniques, becomes urgent. The present paper gives the results of development of DNA diagnosticum, which differentiates parasitic diseases induced by O. felineus and M. bills. The ribosomal RNA gene cluster fragment incorporating internal transcribed spacer 2 (ITS2) was used as a diagnostic marker. A system for diagnosing opisthorchiasis was developed as a multiplex polymerase chain analysis and tested on 37 patients infected with various species of opisthorchids.

Complete Sequencing of the Mitochondrial Genome of Opisthorchis felineus, Causative Agent of Opisthorchiasis.

Opisthorchis felineus, a hepatic trematode, is the causative agent of opisthorchiasis, a dangerous disease in both human beings and animals. Opisthorchiasis is widespread in Russia, especially Western Siberia. The purpose of the present study was to determine the complete mitochondrial DNA sequence of this flatworm. Two parallel methods were employed: (1) capillary electrophoresis to sequence the mitochondrial genome fragments obtained through specific PCR amplification, and (2) high throughput sequencing of the DNA sample. Both methods made possible the determination of the complete nucleotide sequence of the O. felineus mitochondrial genome. The genome consists of a ring molecule 14,277 nt in length that contains 35 genes coding 2 rRNA, 22 tRNA, and 12 proteins: 3 subunits of cytochrome-C-oxidase, 7 subunits of NADH-dehydrogenase, B apocytochrome, and subunit 6 of ATP-synthetase.Like many other flatworms, O. felineus is characterized by the absence of the ATP-synthetase subunit 8 gene. Nineteen out of the 22 tRNAs have a typical "clover leaf" structure. The tRNA(AGC) and tRNA-Cys genes lack DHU-loops, while the tRNA-Ser(UCA) has 2 alternative structures: one with a DHU-loop, and one without it. Analyzing the results obtained from the high throughput sequencing revealed 45 single-nucleotide polymorphisms within the mitochondrial genome. The results obtained in this study may be used in the development of molecular diagnostic methods for opisthorchiasis. This study shows that high throughput sequencing is a fast and effective method for decoding the mitochondrial genome of animals.

[The analysis of structure of insulin-dependent regulatory contours of mature adipocyte].

Insulin is the important regulator of adipose cell metabolism and activates the branched out network of the signaling pathways supervising glucose transport, glycogen synthesis, lipogenesis stimulation, lipolysis inhibition and adipokine secretion. The purpose of our work is the analysis of structure of regulatory contours providing the response of mammalian adipocytes to insulin. With use of computer technology GeneNet adipocyte regulatory-effector network has been reconstructed. The network generalizes experimental data concerning the main insulin-dependent signaling pathways and their targets in a context insulin-sensitive metabolic processes and transcription events. Analysis of the network revealed positive and negative regulatory contours including MAP kinase-, Cbl/TC10- and P13K-dependent signaling pathways. Regulatory contours functioning with participation of transcription factors SREBP-1c, PPARgamma/RXRalpha, C/EBPalpha, FOXO1 are defined also. The major effectors of regulatory contours are glucose carrier GLUT4, and kinase mTOR.

[Real-time polymerase chain reaction and its potential clinical application].

The review introduces real-time polymerase technique, a modern high-tech method, to the medical community. Physico-chemical principles of the method, its main stages, and variants of techniques applying fluorescent reporter platforms are considered. Equations used for quantitative estimation, methods of data processing are given; absolute and relative quantification of specific targets is considered. The special part of the review is dedicated to clinical application of the method by the example of infectious and oncological diseases.

Transcription factor GATA-3 is involved in repression of promoter activity of the human interleukin-4 gene.

GATA-3 was shown to bind to two sites of the IL-4 gene promoter in human T-cell lines PER-117 and Jurkat. A motif located in the region of position -860 and responsible for GATA-3 binding was detected for the first time. Mutation or deletion of this site increased the promoter activity. The findings suggest a direct involvement of GATA-3 in regulation of the human IL-4 gene transcription as a repressor of the promoter activity.

Dexamethasone suppresses human interleukin-5 gene promoter.

Synthetic glucocorticoid dexamethasone suppressed interleukin-5 gene expression in PER-117 human T cells at the level of transcription. The conserved lymphokine element 0 in the interleukin-5 gene promoter context served as a target for dexamethasone.

GATA-3 has dual regulatory functions in human interleukin-5 transcription.

Interleukin-5 (IL-5) is a T-cell cytokine involved in Type 2 diseases and is commonly described as being coordinately regulated with other Type 2 cytokines, such as IL-4 and IL-13. Considering the unique control of eosinophilia by IL-5, such coordinate regulation would be surprising. In fact, the biological specificity of eosinophilia and its control by IL-5 suggests a unique and independent control of IL-5 regulation. In this report we show the binding of GATA-3 to three sites in the human IL-5 promoter in the human T-cell line PER117. The previously identified -70 site and another site at position -152 are shown to positively regulate IL-5 transcription. More importantly, the site located at -400 acts as a powerful repressor of IL-5 transcription with mutagenesis of this site allowing a high level expression of IL-5 without the activation of other factors normally required for IL-5 expression. Whereas GATA-3 has been proposed to be involved in the regulation of the IL-4/IL-5/IL-13 locus, we show here that it has another function in controlling IL-5 transcription that supports the observed unique biological function of this cytokine.

Binding of octamer factors to the murine IL-5 CLE0 in primary T-cells and a T-cell line.

Interleukin-5 (IL-5) is an inducible T-cell derived cytokine with remarkable specificity for the eosinophil lineage. It is controlled at the level of transcription and regulation of the gene is an obvious target for therapy of eosinophil-dependent allergic disorders such as asthma, eczema and rhinitis. Using a T-cell line and primary T-cells we have shown for the first time that the Oct1 and Oct2 transcription factors combine to form a complex with the functionally critical murine IL-5 cis-regulatory element, conserved lymphokine element 0 (CLE0), and contribute to positive regulation of the gene. These results show the increasingly important role of octamer factors in regulation of the IL-5 gene.

Regulation of IL-5 expression.

Interleukin 5 (IL-5) is a cytokine primarily involved in the pathogenesis of atopic diseases. It specifically controls the production, activation and localization of eosinophils, the major cause of tissue damage in atopic diseases. IL-5 belongs to a gene family shared by IL-3, IL-4 and granulocyte-macrophage colony-stimulating factor (GM-CSF) and is predominantly regulated at the transcriptional level. A variety of stimuli and modulators have been identified as regulating production of IL-5 both in vivo and in vitro, indicating a highly complex series of control mechanisms. However, a better understanding of the biology of IL-5 and the regulation of its expression is crucial for the development of new therapeutic agents for allergic disease. This review covers the major molecular aspects of IL-5 research.