A Model of Acute Peritonitis.

The biological models used in the study of generalized peritonitis can be subdivided into 5 groups (introduction of foreign bodies, cultures of microorganisms, suspensions of feces, chemicals, and mechanical damage to the gastrointestinal tract) or into 4 groups (introduction of foreign bodies, chemicals, bacterial contamination of the abdominal cavity, and combined methods). After analysis of published reports, the most justified classification of methods of peritonitis modelling is based on the type of peritonitis-inducing agent and the administration route and on the nature of peritonitis developing in the abdominal cavity. The choice of the model maximally close reproducing clinical conditions of peritonitis should be based on the specific objectives of the study, focusing on the etiology, pathogenesis, and severity of the disease course, planned measures aimed at eliminating the process, and other factors.

Some useful ideas for multistate protein design: Effect of amino acid substitutions on the multistate proteins stability and the rate of protein structure formation.

The design of new protein variants is usually confined to slightly "fixing" an already existing protein, adapting it to certain conditions or to a new substrate. This is relatively easy to do if the fragment of the protein to be affected, such as the active site of the protein, is known. But what if you need to "fix" the stability of a protein or the rate of its native or intermediate state formation? Having studied a large number of protein mutant forms, we have established the effect of various amino acid substitutions on the energy landscape of the protein. As a result, we have revealed a number of patterns to help researchers identify amino acid residues that determine the folding rate and the stability of globular proteins states and design a mutant form of a protein with desired properties.

Simulation of Mechanical and Thermal Wounds of Soft Tissues.

Millions of accidental and surgical injuries of soft tissues are registered annually around the world [5]. Untimely and insufficiently effective treatment of wounds in 50-70% leads to the development of purulent-septic infection and the development of septic conditions and fatal outcomes [1-4], which necessitates thorough study of inflammatory and regenerative processes occurring in the injured soft tissues. Various models of mechanical and thermal damage to soft tissues are proposed for studying the inflammatory and reparative processes, for assessing the therapeutic effects and developing new approaches to wound treatment. However, the developed models do not fully meet the requirements of researchers and are not always simple and uniformly reproducible, close to the course of the pathology in humans, and highly reliable. When choosing the model of mechanical and thermal wounds, the experience of other researchers should be taken into consideration due to the need of actualization and improvement of existing models.

Micropattern platform promotes extracellular matrix remodeling by human PSC-derived cardiac fibroblasts and enhances contractility of co-cultured cardiomyocytes.

In native heart tissue, cardiac fibroblasts provide the structural framework of extracellular matrix (ECM) while also influencing the electrical and mechanical properties of cardiomyocytes. Recent advances in the field of stem cell differentiation have led to the availability of human pluripotent stem cell-derived cardiac fibroblasts (iPSC-CFs) in addition to cardiomyocytes (iPSC-CMs). Here we use a novel 2D in vitro micropatterned platform that provides control over ECM geometry and substrate stiffness. When cultured alone on soft micropatterned substrates, iPSC-CFs are confined to the micropatterned features and remodel the ECM into anisotropic fibers. Similar remodeling and ECM production occurs when cultured with iPSC-CMs in a co-culture model. In addition to modifications in the ECM, our results show that iPSC-CFs influence iPSC-CM function with accelerated Ca2+ transient rise-up time and greater contractile strains in the co-culture conditions compared to when iPSC-CMs are cultured alone. These combined observations highlight the important role cardiac fibroblasts play in vivo and the need for co-culture models like the one presented here to provide more representative in vitro cardiac constructs.

Aligned human cardiac syncytium for in vitro analysis of electrical, structural, and mechanical readouts.

Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) have emerged as an exciting new tool for cardiac research and can serve as a preclinical platform for drug development and disease modeling studies. However, these aspirations are limited by current culture methods in which hPSC-CMs resemble fetal human cardiomyocytes in terms of structure and function. Herein we provide a novel in vitro platform that includes patterned extracellular matrix with physiological substrate stiffness and is amenable to both mechanical and electrical analysis. Micropatterned lanes promote the cellular and myofibril alignment of hPSC-CMs while the addition of micropatterned bridges enable formation of a functional cardiac syncytium that beats synchronously over a large two-dimensional area. We investigated the electrophysiological properties of the patterned cardiac constructs and showed they have anisotropic electrical impulse propagation, as occurs in the native myocardium, with speeds 2x faster in the primary direction of the pattern as compared to the transverse direction. Lastly, we interrogated the mechanical function of the pattern constructs and demonstrated the utility of this platform in recording the strength of cardiomyocyte contractions. This biomimetic platform with electrical and mechanical readout capabilities will enable the study of cardiac disease and the influence of pharmaceuticals and toxins on cardiomyocyte function. The platform also holds potential for high throughput evaluation of drug safety and efficacy, thus furthering our understanding of cardiovascular disease and increasing the translational use of hPSC-CMs.

[To the 100th anniversary of the founding of the Pyatigorsk research institute of balneology]. / K 100-letiyu so dnya osnovaniya Pyatigorskogo nauchno-issledovatel'skogo instituta kurortologii.

The article is dedicated to the 100th anniversary of the Pyatigorsk state research Institute of balneology. It presents the main stages of formation and development, scientific achievements of the Institute and highlights the main activities at the present stage.

[Telomerase activity, mTert gene expression and the telomere length in mouse mesenchymal stem cells in the late period after γ- and γ,n-irradiation and in the tumors developed from these cells]. / Aktivnost' telomerazy, ékspressiia gena mTert i dlina telomer v otdalennyi period posle γ- i γ,n-oblucheniia v mezenkhimal'nykh stvolovykh kletkakh myshi i v opukholiakh, obrazovavshikhsia iz étikh kletok.

In proliferating normal and tumor cells, the telomere length (TL) is maintained by high telomerase activity (TA). In the absence of TA the TL maintenance involves a mechanism of alternative lengthening of telomeres (ALT). The aim of this study was to investigate the level of TA, the mTert expression and TL in cultured normal and transformed by γ- and γ,n-irradiation mesenchymal stem cells (MSCs) from mouse bone marrow, in sarcomas that developed after the transplantation of these cells into syngeneic mice, and in fibrosarcoma cell lines obtained from these tumors to find out the role of AT or ALT in maintaining TL in these cells. During prolonged cultivation of normal and transformed under the influence of γ- (1 Gy and 6 Gy) and γ,n-irradiation (0.05 Gy, 0.5 Gy, and 2 Gy) MSCs from mouse bone marrow, a decrease in TA was detected in irradiated cells. Even deeper decrease in TA was found in sarcomas developed after administration of transformed MSCs to syngeneic mice and in fibrosarcoma cell lines isolated from these tumors in which TA was either absent or was found to be at a very low level. TL in three of the four lines obtained was halved compared to the initial MSCs. With absent or low TA and reduced TL, the cells of all the obtained fibrosarcoma lines successfully proliferated without signs of a change in survival. The mechanism of telomere maintainance in fibrosarcoma cell lines in the absence of TA needs further investigation and it can be assumed that it is associated with the use of the ALT. The detected decrease or absence of TA in transformed under the action of irradiation MSCs with the preservation or even an increase in the telomerase gene expression may be associated with the formation of inactive splicing variants, and requires further study. The obtained lines of transformed MSCs and fibrosarcomas with TA and without the activity of this enzyme can be a useful model for studying the efficacy of TA and ALT inhibitors in vitro and in vivo.

[Current trends in bladder cancer diagnosis].

In the Russian Federation, a proportion of bladder cancer in the overall structure of malignant tumors is about 2.8% and among oncological diseases of genitourinary system its incidence is second only to prostate cancer. Bladder cancer ranks as ninth most prevalent in males and as eighteens in females. The most important issue is to determine a recurrence rate of non-muscle invasive bladder cancer, which can reach 80%. In this regard, currently, all over the world much more attention is paid to studying and creation of early detection, including non-invasive, which will be reliable in the early stages. It can possibly lead to a reduction the number of cystoscopy and become as "golden" standard of non-invasive diagnosis bladder cancer.

[Bacteriophage T5 Mutants Carrying Deletions in tRNA Gene Region].

A new series of heat-stable (st) mutants of bacteriophage T5, which contains deletions in the tRNA gene region, has been isolated. An accurate mapping of the deletion boundaries for more than 30 mutants of phage T5 has been carried out. As a result of the analysis of nucleotide sequences flanking the deleted regions in wild-type phage DNA, it has been shown that they all contain short, direct repeats of different lengths (2-35 nucleotide residues), and that only one repetition is retained in the mutant phage DNA. On the basis of the obtained results, it was suggested that deletion mutants of the phage T5 are formed as a result of illegal recombination occurring with the participation of short repeats in DNA (SHDIR). Based on the example of two mutants, it has been shown that the resistance to thermal inactivation depends on the size of the deleted region.

CHARACTERISTICS OF TUMORS THAT DEVELOPED IN MICE AFTER TREATMENT WITH IRRADIATED SYNGENEIC MESENCHYMAL STEM CELLS OF BONE MARROW.

Mesenchymal stem cells (MSCs) are present in almost all organs and tissues of the organism. It is believed, that MSCs could be transformed into cancer stem cells spontaneously or under influence of genotoxic factors and trigger the growth of tumors. The aim of this work was to study the possibility of malignant transformation of cultured MSCs from murine bone marrow (MSCs-BM) after g-irradiation in vitro and characterize of biochemical and histological features of the tumors that developed after transplantation of MSCs-BM into syngeneic mice. Tumors were observed in 3­4 months after MSCs-BM transplantation. After administration of MSCs-BM irradiated at a dose of 1 Gy, tumors were seen in 2 of 5 mice. After transplantation of MSCs-BM irradiated at a dose of 6 Gy, tumors were found in all 5 of 5 mice. In the case of control MSCs-BM, only one tumor appeared in 6 months after transplantation. The telomerase activity was two times higher in the tumor developed from 6 Gy irradiated MSCs-BM than from 1 Gy irradiated MSCs-BM. The tumors developed from control and irradiated MSCs-BM were classified as multicomponent mesenchymomas («mixture of sarcomas¼). Histological examination showed that tumors contained tissue areas of different histogenesis. Thus, MSCs-BM g-irradiated at doses of 1 and 6 Gy and, much less frequently, control MSCs-BM can transform into tumor cells and induce development of multicomponent mesenchymomas.

ASSESSMENT OF EFFICACY OF THE METHOD OF PROGRAMMED IRRIGATION ASPIRATION SANATION IN COMPLEX TREATMENT OF PATIENTS WITH CHRONIC POSTTRAUMATIC OSTEOMYELITIS OF THE LONG BONES.

The analysis of treatment results of 128 patients aged from 21 to 62 years old with chronic posttraumatic osteomyelitis of the long bones was made at the period from 2006 to 2013. The main group included 67 patients and the method of programmed irrigation aspiration sanation was applied for them. The comparison group consisted of 61 patients and drainage was performed for these patients using the conventional ways. The authors noted good immediate results in the main group in 56 (83,58%) out of 67 patients and in the comparison group - in 43 (70,49%) out of 61 patients. The long-term results were analyzed in 116 (90,6%) out of 128 patients in terms from two to five years after treatment. The rate of recurrences such as formation of purulent fistula were twice less in patients of the main group, than in the comparison group. According to the results of questionnaire SF-36, there was noticed, that patients of the main group got better mean indices of quality of life on all 8 scales compared with the other group.

[The role of telomerase activity in non-invasive diagnostics of bladder cancer].

PURPOSE: To evaluate the potentials of determining the telomerase activity (TA) in the cellular material of the urine for noninvasive diagnosis of bladder cancer (BC). MATERIALS AND METHODS: Evaluation of TA was performed in the urine of 48 patients with bladder cancer (study group) before and after transurethral resection of the bladder wall (n=38), an open resection of the bladder (n=4), and cystectomy (n=6). TA was also evaluated in 48 tumor tissue samples obtained from these patients during removal of the bladder tumor. Each sample of the tumor tissue was separated into two parts, one of which was subjected to histological examination, and the latter was used to determine the telomerase activity. In all cases, the diagnosis of bladder cancer was confirmed morphologically. Determination of TA in the samples was performed by the modified TRAP-method (telomerase repeat amplification protocol), RT-PCR, PCR, and electrophoresis. As a control, cell material of the urine and tissue in 12 patients with chronic cystitis was investigated. RESULTS: TA before surgery was found in 45 (93.75%) of 48 samples of cellular material of the urine from patients with suspected bladder cancer. BC was histologically verified in all patients in this group. In the postoperative period, TA was not observed in the 48 samples of cellular material of the urine from patients with BC. In the control group of patients with histologically verified cystitis, weak TA was determined only in one sample of cellular material of the urine. The analysis indicates statistically significant predominance of patients with bladder cancer in case of TA in the urine (P=0.001). TA was detected in all samples of tumor tissue. We also analyzed the dependence of TA levels in urine and tissue on the degree of BC differentiation. In patients with highly differentiated BC, mean AT in the cellular materials of the urine was 0,61% (n=15), in patients with moderately differentiated BC - 0.95% (n=23), in patients with low-grade bladder cancer - 1.33% (n=10); in other words, increase in the TA levels with decreasing the degree of differentiation was observed. This finding can be used in the prognosis of the course of disease based on determining the TA level in these patients. CONCLUSIONS: Preliminary data indicate the possibility of use of determining the TA in cellular material of the urine for the diagnosis and monitoring of bladder cancer recurrence.

Variation of microsatellite loci in Vincetoxicum Wolf. species in the south-east of Ukraine.

The genetic diversity of 13 Vincetoxicum Wolf species distributed in Ukraine was investigated using 4 of the 8 nuclear microsatellite markers, previously developed for Vincetoxicum atratum from Japan. The number of alleles ranged from 8 to 25. The expected heterozygosities were 0,690­0,938 and observed ones ranged from 0,205 to 0,806. In general, the level of genetic variation in studied representatives of the genus Vincetoxicum from Ukraine proved to be comparable with that of Vincetoxicum atratum. The microsatellite loci Vinc5, Vinc104, Vinc123, Vinc124 can be successfully used to assess intra- and interspecific polymorphisms of species of the genus Vincetoxicum Wolf in Ukraine.

[A New Simple Technique for Producing Labeled Monoclonal Antibodies for Antibody Pair Screening in Sandwich-ELISA].

A simple and fast method for obtaining biotin-labeled monoclonal antibodies was developed usingcontent of hybridoma culture supernatant sufficient to select antibody pairs in sandwich ELISA. The method consists in chemical biotinylation of antigen-bound antibodies in a well of ELISA plate. Using as an example target Vaccinia virus A27L protein it was shown that the yield of biotinylated reactant is enough to set comprehensive sandwich ELISA for a moderate size panel of up to 25 monoclonal antibodies with an aim to determine candidate pairs. The technique is a cheap and effective solution since it avoids obtaining preparative amounts of antibodies.

[Development of noninvasive bladder cancer diagnosis on basis of telomerase and it's subunits hTERT and hTR detection].

Telomerase activity (TA) and expression of genes coding it's subunits (hTERT and hTR) have been examined in tumor tissue and urine sediment samples taken from patients with bladder cancer (BC) using the modified TRAP assay (in the case of telomerase detection) and RT-PCR (in the case of hTERT and hTR expression). Results obtained in this study demonstrate possibility of noninvasive diagnosis of BC with sensitivity of 96% and specificity of 100% in the case of telomerase detection and with sensitivity of 80% and specificity of 100% in the case of hTERT detection in urine sediment samples.

Telomerase as a tumor marker in diagnosis of prostatic intraepithelial neoplasia and prostate cancer.

BACKGROUND: Early diagnosis of prostate cancer (CaP) can be addressed by studying prostatic intraepithelial neoplasia (PIN) as precancer (high-grade PIN or HGPIN). This article attempts to analyze the diagnostic role of telomerase as an early marker of carcinogenesis. METHODS: Complex urological patient evaluation and assessment of telomerase activity. RESULTS: Out of 92 patients 44% were diagnosed with CaP, 49% with low-grade PIN (LGPIN) in association with benign prostatic hyperplasia (BPH), and 7% with HGPIN in association with BPH. Active telomerase (AT) in prostate biopsy specimens was detected in 98% of patients with CaP, in 33% of patients with HGPIN, and in 20% of patients with LGPIN. In the event of simultaneous detection of AT and PIN in initial prostate biopsy specimens, further monitoring for 0.5-4.0 years revealed CaP development in 50-56% of cases. Further follow-up of patients with PIN and absent telomerase activity in initial biopsy specimens did not demonstrate the development of CaP. The PSA level was significantly higher in patients with active telomerase in the prostate tissue than in telomerase negative patients. CONCLUSIONS: Telomerase activity in the prostate tissue increases the risk of CaP development in patients with PIN. Detection of telomerase activity in prostate biopsy specimens from patients with PIN enables selection of a group of patients with high risk of CaP development and reduction of the number of prostate biopsies performed in other patients.

[Electrophysiological mechanisms of heart rate stability of the hibernating mammals during hypothermia].

Despite more than 100 years of study, the mechanisms of natural resistance of the hibernator heart to cardiac arrhythmias during hypothermia has remained unknown. Renewed optimism in this area of research comes with recent methodological advances which enable to shed light on the hidden secrets of the hibernator's heart. This review discusses basic mechanisms of hypothermic ventricular arrhythmias and highlights some recent findings from the hibernator's heart electrophysiology, which may have an antiarrhythmic potential for the human heart as well.

[Development of a set of diagnostic test systems for analysis of expression of C-myc, mad1, max, p53, and E2F1 gene oncomarkers by the reverse transcription reaction-polymerase chain reaction alignment technique].

Based on the performed studies, the authors developed test systems to analyze the expression of mRNA of the p53, C-myc, mad1, max, and E2F1 genes. These test systems could reveal a statistically significant difference between follicular adenoma and carcinoma of the thyroid in their expression of p53 mRNA. It should be noted that the use of our developed test systems is promising when searching for the diagnostic and prognostic markers of cancer, analyzing, and creating the genetic networks characterizing this or that cancer.

[Detection of telomerase activity in gastric cancer].

Telomerase activity (TA) was examined in gastric adenocarcinomas and gastric lymphoma using a modified TRAP assay. TA was present in 16 of 18 (89%) gastric adenocarcinomas and in gastric lymphoma, whereas no TA was detected in normal tissue. Almost all samples had "high" and "very high" TA levels. Telomerase is undoubtedly associated with the process of malignant transformation and therefore can be an important marker for diagnostics of gastric cancer.