INTERDEPENDENCE BETWEEN THE PHENOTYPE OF DENDRITIC CELLS AND AMOUNTS OF BLOOD PROINFLAMMATORY MONOCYTES IN PATIENTS WITH KIDNEY CANCER

The aim of the study was to investigate an interdependence between the phenotype of dendritic cells (DC) differentiated from monocytes and the number of pro-inflammatory monocytes in peripheral blood of patients with kidney cancer (KC). The study involved 28 patients at the age of 40-55 years suffering with KC (Т3N0М0, clear cell type) before surgical treatment. The diagnosis was verified histologically. 31 healthy agematched persons were examined as a control group. Mononuclear cells were isolated from heparinized venous blood by centrifugation in a Histopaque®-1077 density gradient followed by plastic adsorption in RPMI 1640 medium supplied with 10% autologous serum. Immature DCs (iDCs) were generated from blood monocytes by culturing for 5 days with GM-CSF and IFNα. Activation of DCs (mDCs) was induced by incubation with the tumor cell lysate and TNFα, followed by incubation for 48 hours. A tumor fragment was used to prepare the lysate of autologous tumor cells. Phenotyping of blood monocytes and DC at various maturation stages was performed by flow cytometry. The numbers of CD14+CD16+ monocytes in peripheral blood of KC patients were decreased (up to 42% of the total monocyte level) against the control ranges. In this regard, the analysis of the dependence between the phenotype of DCs differentiated from monocytes and the number of pro-inflammatory blood monocytes was carried out by comparing the groups with a high content of pro-inflammatory monocytes in the blood in KC patients (> 42%, near-control range) and low content (resp., < 42%). We have found that the contents of tolerogenic iDC in cell culture are increased in KC patients with low amounts of pro-inflammatory monocytes in blood (< 42%). A relatively increased expression of antigen-presenting and co-stimulatory molecules proved to be the specific feature of iDC phenotype in patients with high contents (> 42%) of proinflammatory monocytes in blood. The phenotype of dendritic cells in KC patients with different content of proinflammatory monocytes during maturation/activation showed more differences. In the patients with low levels of pro-inflammatory monocytes, the cell pool of in vitro maturing DCs was characterized by low level of CD86 and HLA-DR receptor expression, thus reflecting a weak co-stimulating and antigen-presenting activity. In the patients with high levels of pro-inflammatory monocytes in blood, the in vitro activated DCs showed higher level of functional activity using the above markers. The revealed differences in the DC phenotype and interrelations with amounts of blood monocyte subpopulations in KC patients may presume the programmed cell differentiation mechanisms depending on the microenvironment, under pathogenic conditions (i.e., in presence of malignant tumor growth)

Dependence of phenotype and chemiluminescent activity of monocytes on the Tregulatory cells content in patients with kidney cancer

The aim of this work was to reveal the interrelations between the number of T regulatory cells (Tregs) in patients with kidney cancer (KC) and phenotype of peripheral blood monocytes and their capacities to produce ROS. Patients with KC (T3N0M0, clear cell type) were examined prior to surgical treatment. Tregs phenotype and blood monocytes were identified by flow cytometry. ROS production of purified monocytes was carried out through the determination of lucigenin- and luminol-dependent spontaneous and zymosan-induced chemiluminescence activity. It has been found that the relative number of Tregs within total lymphocyte subset in KC patients was increased if compared to control values (in KC patients — Me = 6.3%). Then the patients were divided into two groups according to the median of Tregs number (less and more than 6.3%). The most pronounced changes in the phenotype of monocytes and their chemiluminescent activity were found in KC patients with the Tregs count of less than 6.3%. Our findings suggest that low frequency of Tregs in the periphery was associated with increased relative numbers of “intermediate” and “non-classical” (“pro-inflammatory”) monocytes as it was shown on the samples from patients with KC with a low level of Tregs. According to our data, both groups of KC patients had low levels of HLA-DR expression when comparing to control group. Furthermore, both groups of patients had decreased rates of HLA-DR and CD64 co-expressing cells. Changes in the phenotype of monocytes in patients with KC were closely linked with imbalance in ROS production. Thus, the monocytes spontaneous superoxide radical (primary ROS) synthesis in KC patients with a low Treg numbers were characterized by redused NADPH-oxidase activation time and increased level of its activity if compared to patients with a high Treg rates in peripheral blood. Next, the activation index for lucigenin-dependent chemiluminescence in KC patients was reduced, as well as it was independent of circulating Tregs rates and was determined apparently by the insufficiency of metabolic reserves. Similarly, spontaneous secondary ROS production by the monocytes in KC patients was lower then in healthy controls and was also independent of circulating Tregs rates. Finally, the induced secondary ROS synthesis and activation index for their synthesis in monocytes were reduced only in patients with KC with a low number of Tregs in the blood. In general, the characteristics of the chemiluminescent reaction of monocytes in patients with KC determined the imbalance in peripheral blood monocytes primary and secondary ROS production. Monocytes in patients with KC with a low number of Tregs in the blood were characterized by more pro-inflammatory activity due to the rapid activation and intensity of the synthesis of primary ROS

Immunophenotype and metabolism are linked in peripheral blood neutrophils from patients with kidney cancer

The aim of the present study was to analyze the relationships between expression of activation and adhesion receptors on peripheral blood neutrophils, and intracellular activity of some neutrophil enzymes in patients with kidney cancer (KC). Patients and methods: the KC patients (n = 72) (T3N0M0, clear-cell type) were examined prior to surgical treatment at the Krasnoyarsk Regional Oncology Center. The diagnosis was verified histologically for all KC patients. The phenotype of blood neutrophils was studied using flow cytometry. The surface receptor expression levels of the neutrophils were evaluated by mean fluorescence intensity. NAD and NADP-dependent dehydrogenases activities in purified peripheral blood neutrophils were measured by bioluminescent method. Results: we have found that the phenotypic alterations in circulating KC patients’ neutrophils appeared along with inhibition of main intracellular metabolic processes and were closely linked with them. The features of the phenotypic imbalance in the neutrophils from KC patients were associated with a decrease in blood cells expressing adhesive (CD11b and CD62L) and functional (CD64 and HLA-DR) receptors. Moreover, the patient’s neutrophils expressed CD11b, CD16 and HLA-DR on their cell surface more intensively, than neutrophilic leukocytes from control group. These phenotypic changes in KC patients’ blood neutrophils occurred in parallel with pronounced decrease in immature cells numbers. The metabolic changes of neutrophil cytoplasmic compartment in KC patients were determined by a decrease in Glu6PDH activity (a key and initializing enzyme of the pentose phosphate cycle) and NADH-LDH (anaerobic glycolysis). Mitochondrial metabolism in neutrophils of KC patients was characterized by multidirectional changes in the activity of NAD- and NADP-dependent glutamate dehydrogenases (decreased activity of NAD-dependent and increased activity of NADP-dependent) and a decrease in NADH-MDH activity. The established features in mitochondrial enzymes activities suggest some disturbances of NAD-dependent processes that could lead to down-regulation of aerobic energy processes. We guess that the decreased activity of plastic and energy processes in blood neutrophils of KC patients could affect the receptor expression levels. By means of correlation analysis, we have found that the relationships in KC patients were determined by negative effects of NADHGDH and NADH-LDH activities upon expression of activation and adhesion receptors in blood neutrophils. Of these enzymes, only glutathione reductase activity in neutrophils from KC patients was positively linked with the CD23 and HLA-DR expression. Thus, an increase in activity of energy processes (e.g., coupling the tricarboxylic acid cycle to amino acid metabolism) in blood neutrophils from the patients with kidney cancer could stimulate expression levels of activation and adhesion receptors and potentially increase antitumor activity of neutrophils

FUNCTIONAL ACTIVITY OF NEUTROPHILS AND HEMOSTASIS PATTERN IN PATIENTS WITH ACUTE DESTRUCTIVE PANCREATITIS

The aim of present study was to investigate relationships between indicators of functional activity of neutrophilic granulocytes, and hemostasis parameters in patients with acute destructive pancreatitis (ADP). The study included thirty-three patients with ADP. 35 healthy persons were examined as a control group. The phagocytosis level in neutrophils was determined by flow cytometry using FITC-labeled staphylococcal protein A. We have calculated the percentage of fluorescent neutrophils as phagocytic index, and average cell fluorescence assumed phagocytic number. The intensity of respiratory burst observed in neutrophil samples was evaluated using chemiluminescence assay. All the persons under study were also tested for blood coagulation and vascular-platelet hemostasis. It was found that the ADP patients with decreased number of phagocytic neutrophils in the blood showed a decrease in respiratory burst intensity in the neutrophils. Moreover, spontaneous and induced synthesis of the primary reactive oxygen species (ROS) in neutrophils of ADP patients proceeded faster than in healthy people, but its intensity was much lower. The maximal level of spontaneous and induced synthesis of secondary ROS in neutrophils of patients was significantly higher than in healthy individuals, but its rapid may be generally characterized by insufficient respiratory burst in these patients. A reduced neutrophil phagocytic activity and kinetic characteristics of primary and secondary ROS synthesis may be attributed to the effects produced by pancreatic enzymes entering blood flow which may alter functional activity of the blood neutrophils. Concerning hemostasis in patients with ADP, some disturbances were found only in the coagulation link which seem to depend on increase in fibrinogen, soluble fibrin monomer complexes and D-dimer in blood plasma, along with reduced antithrombin III levels. Such a change in blood coagulation indexes is typical to inflammatory processes and presumes activation of the coagulation cascade and higher risk of septic complications. In patients with ADP, we have found a significantly increased number of correlations between indicators of functional activity of neutrophils and hemostasis parameters. This analysis revealed a relationship by the patients with ADP reflect some unidirectional changes in functional activity of neutrophils (as phagocytosis and respiratory burst), and blood coagulation parameters (as blood clotting and vascular/ platelet links). The changes in functional activity of neutrophils and the state of hemostasis in the ADP patients, as well as correlations between their alterations are omvolved into the pathogenesis of this disorder, and determine potential mechanisms for evolving complications

Determination of temperature dependences of Young's modulus and internal friction of fuel cladding by resonance method

We study elastic characteristics and internal friction of fuel claddings to improve computer codes for VVER-1000 fuel rods. We analytically described elastic characteristics of cladding material and obtained coefficient of the form of the first longitudinal frequency numerically. We described new measuring module for automatic acquisition data. We’ve established temperature dependences of Young’s modulus and internal friction via high-temperature facility and developed electronic module and noted maximum of these characteristics at the temperature 1160 K. It can be explained by the destruction of the texture in the material of claddings

Association of mechanical bowel preparation with oral antibiotics and anastomotic leak following left sided colorectal resection:an international, multi-centre, prospective audit

Introduction: The optimal bowel preparation strategy to minimise the risk of anastomotic leak is yet to be determined. This study aimed to determine whether oral antibiotics combined with mechanical bowel preparation (MBP+Abx) was associated with a reduced risk of anastomotic leak when compared to mechanical bowel preparation alone (MBP) or no bowel preparation (NBP). Methods: A pre-planned analysis of the European Society of Coloproctology (ESCP) 2017 Left Sided Colorectal Resection audit was performed. Patients undergoing elective left sided colonic or rectal resection with primary anastomosis between 1 January 2017 and 15 March 2017 by any operative approach were included. The primary outcome measure was anastomotic leak. Results: Of 3676 patients across 343 centres in 47 countries, 618 (16.8%) received MBP+ABx, 1945 MBP (52.9%) and 1099 patients NBP (29.9%). Patients undergoing MBP+ABx had the lowest overall rate of anastomotic leak (6.1%, 9.2%, 8.7% respectively) in unadjusted analysis. After case-mix adjustment using a mixed-effects multivariable regression model, MBP+Abx was associated with a lower risk of anastomotic leak (OR 0.52, 0.30–0.92, P = 0.02) but MBP was not (OR 0.92, 0.63–1.36, P = 0.69) compared to NBP. Conclusion: This non-randomised study adds ‘real-world’, contemporaneous, and prospective evidence of the beneficial effects of combined mechanical bowel preparation and oral antibiotics in the prevention of anastomotic leak following left sided colorectal resection across diverse settings. We have also demonstrated limited uptake of this strategy in current international colorectal practice

Evaluating the incidence of pathological complete response in current international rectal cancer practice

The mainstay of management for locally advanced rectal cancer is chemoradiotherapy followed by surgical resection. Following chemoradiotherapy, a complete response may be detected clinically and radiologically (cCR) prior to surgery or pathologically after surgery (pCR). We aim to report the overall complete pathological response (pCR) rate and the reliability of detecting a cCR by conventional pre-operative imaging.A pre-planned analysis of the European Society of Coloproctology (ESCP) 2017 audit was performed. Patients treated by elective rectal resection were included. A pCR was defined as a ypT0 N0 EMVI negative primary tumour; a partial response represented any regression from baseline staging following chemoradiotherapy. The primary endpoint was the pCR rate. The secondary endpoint was agreement between post-treatment MRI restaging (yMRI) and final pathological staging.Of 2572 patients undergoing rectal cancer surgery in 277 participating centres across 44 countries, 673 (26.2%) underwent chemoradiotherapy and surgery. The pCR rate was 10.3% (67/649), with a partial response in 35.9% (233/649) patients. Comparison of AJCC stage determined by post-treatment yMRI with final pathology showed understaging in 13% (55/429) and overstaging in 34% (148/429). Agreement between yMRI and final pathology for T-stage, N-stage, or AJCC status were each graded as 'fair' only (n = 429, Kappa 0.25, 0.26 and 0.35 respectively).The reported pCR rate of 10% highlights the potential for non-operative management in selected cases. The limited strength of agreement between basic conventional post-chemoradiotherapy imaging assessment techniques and pathology suggest alternative markers of response should be considered, in the context of controlled clinical trials