Biochemistry (Moscow), Supplement Series B: Biomedical Chemistry (v.4, #2)

Atomic force microscopy detection of serological markers of viral hepatites B and C by Yu. D. Ivanov; P. A. Frantsuzov; T. O. Pleshakova; V. S. Ziborov; S. K. Svetlov; N. V. Krohin; V. A. Konev; O. B. Kovalev; V. F. Uchaikin; O. N. Yastrebova; P. G. Sveshnikov; A. I. Archakov (117-122).
The possibility of detection of serological markers, containing the hepatitis B surface antigen (HBsAg) and hepatitis C virus core-antigen (HCVcoreAg) in human serum, by a new atomic force microscopy (AFM)-based nanotechnological approach has been demonstrated.The antibodies against the hepatitis B virus surface antigen (anti-HBsAg) and the antibodies against the hepatitis C virus core antigen (anti-HCVcoreAg) were immobilized on an AFM-chip. It was shown that such approach enables to detect HBsAg, HCVcoreAg and the viral fragments containing these antigens in the serum. The comparative analysis of detection of HBsAg- and HCVcoreAg-containing particles by the AFM method versus traditional methods (ELISA, PCR) has demonstrated the 75% coincidence of results between the AFM and two other methods.
Keywords: atomic-force microscopy; hepatitis B virus surface antigen (HBsAg); hepatitis C virus core antigen (HCVcoreAg)

Lung cancer is the leading cause of cancer-related mortality in industrialized countries. Unfortunately, most lung cancers are found too late for a cure, therefore early detection and treatment is very important. We have applied proteomic analysis by using two-dimensional gel electrophoresis and peptide mass fingerprinting techniques for examination of cancerous and adjacent non-cancerous lung tissues from the same patient. The aim of the study was to find proteins, which could be used as biomarkers for diagnosis and monitoring of this disease. Indeed, we found differences in expression of several proteins, related to various cellular activities, such as, chaperoning (e,g. GRP96, GRP78, HSP27), metabolism and oxidation stress (e.g. L-fucose, GST), cytoskeleton (e.g., tubulin beta 2/3, beta actin), cell adhesion (e.g. annexin A5/3), binding proteins (e.g. 14-3-3 theta) and signal transduction. These changes may be important for progression of carcinogenesis; they may be used as the molecular-support for future diagnostic markers.
Keywords: proteomics; two-dimensional gel electrophoresis; lung cancer; mass spectrometry

The study of structure-antiarrhythmic activity relationship of N-phenylacetamide derivatives and aromatic carbonic acid amides by V. R. Khairullina; G. P. Tarasov; A. Ya. Gerchikov; F. S. Zarydiy; L. A. Tyurina (130-137).
Using the SARD-21 (Structure Activity Relationship & Design) computer system, structural features of high- and low-effective antiarrhythmic agents have been recognized and the influence of these features on the antiarrhythmic properties has been evaluated. This information has been used for generation of the model to predict antiarrhythmic effectiveness of pharmaceutical preparations at the recognition level of 82% by means of two different approaches. The recognized structural parameters may be successfully used to design new highly effective antiarrhythmic drugs, and also to modify structures of the already-existing anti-arrhythmic drugs in order to increase the effectiveness of their antiarrhythmic action.
Keywords: structure-activity; antiarrhythmic activity; shape recognition theory

The review summarizes data on DNA/RNA aptamers, a novel class of molecular recognition elements. Special attention is paid to the aptamers to proteins involved into pathogenesis of wide spread human diseases. These include aptamers to serine proteases, cytokines, influenza viral proteins, immune deficiency virus protein and nucleic acid binding proteins. High affinity and specific binding of aptamers to particular protein targets make them attractive as direct protein inhibitors. They can inhibit pathogenic proteins and data presented here demonstrate that the idea that nucleic acid aptamers can regulate (inhibit) activity of protein targets has been transformed from the stage of basic developments into the stage of realization of practical tasks.
Keywords: DNA/RNA aptamers; proteins; the SELEX method

The liver transplantation is the most effective method for treating severe acute liver diseases. The hepatocytes transplantation may serve as the perspective means for treating liver failure. This review analyzes the experimental approaches and perspectives on the use of adult hepatocytes for the creation of implanting bioartificial liver module for treatment of hepatic failure.
Keywords: cell therapy; hepatocytes; transplantation; liver failure

Alkyl-type glycerolipids as modulators of tumor cells destruction by S. G. Romanova; V. G. Romanov; G. A. Serebrennikova; A. A. Shtil (161-170).
The review summarizes current information on the biological activity and search of the antineoplastic mechanism of action among alkyl glycerolipids. Special attention is paid to following problems considered in experimental papers published during the last decade: selective ability phosphorus alkyl glycerolipids towards tumor cells using edelfosine and its analogues. The review contains modern information available in the literature, on a possible mechanism cytotoxic effect of such compounds.
Keywords: alkyl glycerolipids; edelfosine; anticancer activity; apoptosis

Phosphorylation of ATM/ATR substrates in eukaryotic cells after infection with Helicobacter pylori by M. O. Anikeenok; Yu. N. Churin; T. F. Meyer; O. N. Ilinskaya (171-176).
Phosphorylation of ATM-kinase substrates in HeLa and AGS cells in response to Helicobacter pylori infection has been characterized. Infection with wild-type (cagPAI-positive) and corresponding isogenic cagPAI negative mutant induced activation of Chk1 and Chk2 kinases. However, only Chk1 was directly activated by ATM-kinase. Using 2D-electrophoresis and mass spectrometry a group of proteins phosphorylated in AGS cells by ATM1/ATR kinases during H. pylori infection has been identified.
Keywords: Helicobacter pylori ; ATM/ATR kinases; phosphorylation; RPA32A; splicing-factor

Biodegradation kinetics of poly(3-hydroxybutyrate)-based biopolymer systems by A. P. Boskhomdzhiev; A. P. Bonartsev; T. K. Makhina; V. L. Myshkina; E. A. Ivanov; D. V. Bagrov; E. V. Filatova; A. L. Iordanskii; G. A. Bonartseva (177-183).
The aim of this study was to evaluate and to compare the long-term kinetics curves of biodegradation of poly(3-hydroxybutyrate) (PHB), its copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate), and a PHB/polylactic acid composite. The total weight loss and the change of average viscosity molecular weight were used as the parameters reflecting the biodegradation degree. The rate of biodegradation was analyzed in vitro in the presence of lipase and in vivo after film implantation in animal tissues. The morphology of the PHB film surface was studied by the atomic force microscopy technique. It was shown that PHB biodegradation involves both polymer hydrolysis and its enzymatic biodegradation. The results obtained in this study can be used for the development of various PHB-based medical devices.
Keywords: polyhydroxyalkanoates (PHA); poly(3-hydroxybutyrate) (PHB); poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV); polylactic acid (PLA); biodegradation; biopolymer systems

A precise and reproducible method of quantitative determination of the photosensitizer (PS) Photolon in liver samples of laboratory animals by means of a spectrophotometric assay with preliminary extraction has been developed. Conditions of the PS extraction have been optimized and validated for the quantitative determination of the PS Photolon by the spectrophotometric assay and the main analytical characteristics have been investigated. Using the method of quantitative determination of the PS Photolon in liver tissue samples a quantitative estimation of in vivo fluorescence of the liver tissue was performed after PS administration to animals. There was a high correlation (R = 0.99) between results obtained by spectrophotometry ex vivo and spectrofluorimetry in vivo. The method of fluorescence detection in vivo is applicable for studies of the pharmacokinetics of different photosensitizers.
Keywords: photodynamic therapy; chlorin e6; Photolon; extraction; fluorescence; spectrophotometry; pharmacokinetics

Development of apoptosis is accompanied by a decrease in transcripts of NADP-isocitrate dehydrogenase (NADP-ICDH, EC 1.1.1.42), and also by alterations of catalytic properties from the rat liver enzyme in comparison with control. Administration of thioctic acid increased the level of expression towards normal values. Molecular weight of homogenous preparations of NADP-ICDH purified from livers of control and experimental rats was the same 112 ± 5.8 kDa, however, K m values and pH-optimum changed during induction of apoptosis. Regulation of NADP-ICDH activity by some intermediates of the tricarboxylic acid cycle also differed in these groups.
Keywords: rat; liver; apoptosis; NADP- isocitrate dehydrogenase; properties; thioctic acid

Earlier it was shown that male mice of the DD/He strain were highly susceptible to ortho-aminoa-zotoluene (OAT) induced hepatocarcinogenesis, and resistant to spontaneous liver tumor development as compared to male mice of the CC57BR/Mv strain. In the present work we have made a comparative investigation of peroxisome proliferator-activated receptor (PPAR), liver X-receptor (LXR) and retinoic X-receptor (RXR) mRNA levels in liver as well as concentrations of corticosterone, glucose, lipids and insulin in blood of male DD/He and CC57BR/Mv mice. Using the multiplex RT-PCR method it was found that PPAR-α, PPAR-γ, RXR-α, and RXR-β mRNA content was significantly lower in the liver of DD mice than in CC57BR mice. No significant interstrain differences were found in the content of LXR-α and LXR-β mRNA. In DD mice there was more than the 3-fold decrease of blood content of corticosterone (involved in PPAR and RXR regulation). DD mice demonstrated a significant decrease in blood serum glucose and insulin concentrations as well as higher reactivity to insulin as compared with ÑÑ57BR mice. Elevated blood total cholesterol and HDL cholesterol levels were found in DD mice whereas triglyceride content was basically the same in both mouse strains. It is known that glucocorticoids, PPAR and RXR play crucial role in transcription regulation of the inflammation response. Therefore our data on decreased corticosterone level in blood, PPAR and RXR mRNA content in the liver of the DD suggest that sensitivity of this strain of mice to the development of OAT-induced hepatocarcinomas may be associated with increased inflammatory reaction to this carcinogen.
Keywords: hepatocarcinogenesis; PPAR; LXR; RXR; corticosterone; interstrain differences

L-Cysteine influx in type 2 diabetic erythrocytes by S. I. Rizvi; N. Srivastava (205-208).
Erythrocyte oxidative stress has been implicated in the pathogenesis of diabetes mellitus, and the deficiency of antioxidant defense by the glutathione (GSH) pathway is thought to be one of the factors responsible for development of complications in diabetes. Erythrocytes require L-cysteine for the synthesis of GSH and the rate of synthesis is determined only by L-cysteine availability. In the present study we have found that the L-cysteine influx in erythrocytes from type 2 diabetic patients was significantly lower compared to age-matched controls. The decreased influx may be one of the factors leading to low GSH concentration observed in type 2 diabetes. Since L-cysteine is the limiting amino acid in GSH synthesis, any strategy aimed to increase L-cysteine influx in erythrocytes may be beneficial for type 2 diabetic patients.
Keywords: Erythrocytes; L-cysteine; diabetes mellitus

The study of RNA markers in blood of patients with malignant tumors of gastrointestinal tract by A. G. Globa; V. S. Demidova; O. N. Dikova; V. A. Vishnevskii; A. I. Schegolev (209-212).
In order to develop a diagnostic panel, mRNA levels of tumor marker genes have been evaluated in capillary blood of patients with various malignant tumors of the gastrointestinal tract (GIT) by means of the method of reverse transcription combined with real-time PCR with detection of reaction products using TaqMan probes. Use of small volumes of capillary blood did not decrease sensitivity of this method. RNA expression of telomerase (mhTERT), alpha-fetoprotein (mAFP), carcinoembryonic antigen (mCEA) and cytokeratin-20 (mCK-20) was higher in most patients with tumors. Blood of donors or non-oncological patients contained much lower (trace) amounts of the RNA markers. The RNA markers are characterized by reasonably high specificity and sensitivity acceptable for diagnostic application. The mhTERT marker was the most universal one and exhibited the highest specificity and sensitivity. Combined determination of several RNA markers increased sensitivity of this method. It is concluded that determination of RNA markers in small volumes of capillary blood may be used for screening, primary diagnostics, and postoperative monitoring.
Keywords: polymerase chain reaction; telomerase; tumor markers; carcinoma