Kinetics of Recovery of Naïve and Memory T Cells in Acute Leukemia Patients after Allogeneic Stem Cell Transplantation Depending on Different GVHD Prophylaxis Regimens.

Background: Memory T cells are a heterogeneous population of immune cells that provide adaptive immunity. Its full recovery seems essential for graft-versus-tumor reactions that provide an opportunity for biological cure in patients with acute leukemia. The use of mismatched or haploidentical donors has increased, which has become possible because of modifications in graft versus host disease (GVHD) prophylaxis. Materials and Methods: Sixty-five leukemia patients (acute myeloid leukemia - 40, acute lymphoblastic leukemia - 25), median age 33 (17-61) years, underwent allo-HSCT from 2016 to 2019 in the National Research Centre for Hematology. Patients were divided into three groups based on the impact of GVHD prophylaxis on T cell recovery: horse antithymocyte globulin (ATG)-based regimen (n=32), horse ATG combined with posttransplant cyclophosphamide (PT-Cy) (n=18), and ex vivo T cell depletion (n=15). Results: The early period after transplantation (before day +100) was characterized by significantly lower absolute numbers of T naïve, memory stem and T central memory cells in peripheral blood in patients after ATG+PT-Cy-regimen or ex vivo T cell depletion than after ATG-based prophylaxis (p<0.05). Moreover, strong depletion of naïve T and memory stem cells prevents the development of GVHD, and determining the absolute number of CD8+ naïve T and memory stem cells with a cutoff of 1.31 cells per microliter seems to be a perspective in assessing the risks of developing acute GVHD (p=0.008). The dynamics of T cell recovery showed the involvement of either circulating or bone marrow resident T effector cells shortly after allogeneic transplantation in all patients, but the use of manipulated grafts with ex vivo T cell depletion requires the involvement of naïve and memory stem cells. There was no significant effect of T cell recovery on leukemia relapse after allogeneic transplantation. Conclusion: These experimental outcomes contribute to providing the best understanding of immunological events that occur early after transplantation and help in the rational choice of GVHD prophylaxis in patients who will undergo allogeneic transplantation. Our study demonstrated the comparable immunological effects of posttransplant cyclophosphamide and ex vivo T cell depletion and immunological inefficiency of horse ATG for GVHD prevention.

Morphological parameters of hepatocytes in the European mole (Talpa europaea) and herb field mouse (Sylvaemus uralensis) under industrial pollution: Qualitative and quantitative assessment.

Morphological alterations of cells and tissues usually occur in biological organisms exposed to environmental contaminants, there by acting as a biomarker of environmental pollution, thus, making this study highly pertinent. The effect of industrial pollution on the qualitative and quantitative morphological parameters of hepatocytes (through histological analysis and cytomorphometry) was studied in two contrasting species of small mammals (Talpa europaea and Sylvaemus uralensis), taking into account the animal age (young and adult groups) and liver concentrations of heavy metals (Cu, Zn, Cd, Pb). Studies were performed in the regions exposed to emissions from two currently operating copper smelters: Middle Ural Copper Smelter (Middle Urals, T. europaea catching area) and Karabash Copper Smelter (Southern Urals, S. uralensis catching area). Seven morphometric parameters of hepatocytes were measured, of which two key parameters were selected by the method of principal components-the cell packing density and nuclear-cytoplasmic ratio (N/C). It was found that cell packing density in T. europaea from the impact zone decreased relative to the background area in young animals. At the same time, the differences in this parameter between the age groups from the background zone were leveled in the impact area of catching. The N/C ratio in T. europaea hepatocytes showed no correlation with either animal age or site of capture (background or impact area). In S. uralensis, both parameters, even taking into account the age, were found to be insensitive to indicate an effect of industrial pollution. Dystrophic changes (tested through histological analysis) in the liver tissue were revealed in all animal groups, but their frequency did not depend on any of the factors (age, zone) as well as the level of accumulation of toxic heavy metals (Cd, Pb). Morphometric parameters of hepatocytes have proved to be more reliable indicators of pollution, compared to the frequency of liver histopathology, due to lower subjectivity in their evaluation.

T Cell and Cytokine Dynamics in the Blood of Patients after Hematopoietic Stem Cell Transplantation and Multipotent Mesenchymal Stromal Cell Administration.

Multipotent mesenchymal stromal cells (MSCs) are currently under intensive investigation for the treatment and prevention of graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT), owing to their substantial immunomodulatory properties. The responses of recipients to MSC infusion following allo-HSCT are not yet well understood. T cells are central to the adaptive immune system, protecting the organism from infection and malignant cells. Memory T cells with different phenotypes, gene expression profiles, and functional properties are critical for immune processes regulation. The aim of this study was to study the dynamics of memory T cell subpopulations and cytokines in the blood of allo-HSCT recipients after MSC administration. In clinical trial NCT01941394, patients after allo-HSCT were randomized into 2 groups, one receiving standard GVHD prophylaxis and the other also receiving MSC infusion on the day of leukocyte recovery to 1000 cells/µL (engraftment, day E0). Blood samples of patients from both groups were analyzed on days E0, E+3, and E+30. T cell subpopulations were studied by flow cytometry, and cytokine concentrations were evaluated by the Bio-Plex Pro Human Cytokine Panel. Administration of MSCs to patients on day E0 did not affect the overall dynamics of restoration of absolute numbers and proportions of T and B lymphocytes after 3 and 30 days. At 3 days after MSC injection, only the numbers of CD8+ effector cells (CD8+TE, CD8+TM, and CD8+EM) were found to increase significantly. A significant increase in the number of CD4+ cells after 30 days compared to day E0 was observed only in patients who received MSCs, indicating faster recovery of the CD4+ cell population following MSC injection. An increase in CD8+ cell number by day E+30 was significant regardless of MSC administration. To characterize the immune status of patients following allo-HSCT in more detail, changes in the cytokine concentration in the peripheral blood of patients on days E0, E+3, and E+30 after MSC administration were investigated. On day E+30, significant increases in the numbers of CD4+CM and activated CD4+CD25+ cells were observed. The concentrations of proinflammatory and anti-inflammatory cytokines IL-6, IL-8, IL-17, TNF-α, and IFN-γ were increased significantly in patients injected with MSCs. Analysis of growth factor levels showed that in the group of patients who received MSCs, the concentrations of G-CSF, GM-CSF, PDGFbb, FGFb, and IL-5 increased by day E+30. Among the cytokines involved in regulation of the immune response, concentrations of IL-9, eotaxin, IP-10, MCP-1, and MIP-1a were increased after 30 days irrespective of MSC administration. The administration of MSCs exerts a positive effect on the restoration of T cell subpopulations and immune system recovery in patients after allo-HSCT.

Structural-functional aberrations of erythrocytes in the northern red-backed vole (Clethrionomys rutilus Pallas, 1779) that inhabits the zone of influence of the copper smelter (the Middle Ural).

Red blood cell parameters were assessed in a natural population of the northern red-backed vole (Clethrionomys rutilus Pallas, 1779) in the zone of influence of the Kirovgrad Copper Smelter along a gradient of pollution by heavy metals (Cu, Zn, Cd, and Pb) at three catching sites (polluted [Imp] and controls [Bg-1, and Bg-2]). The difference of the smelter area (Imp group of voles) from both background groups (Bg-1 and Bg-2) was proven by means of a set of 13 parameters in univariate and multivariate analyses. Among the detected erythrocyte disturbances, we noted the following: a decrease in activities of Na+,K+-ATPase and antioxidant enzymes (SOD, GSH-Px, and CAT); an increase in the concentration of lipid peroxidation products, in osmotic fragility, and in intravascular hemolysis; interruption of carbohydrate metabolism; and lowered oxygen-carrying capacity. A higher load of Cd (p = 0.0009) and possibly Pb (p = 0.054) in the Imp animals was confirmed by quantitation of heavy metals in the liver. Most erythrocyte parameters (11 out of 13) covaried with individual Cd load by obeying a semilogarithmic dependence; such a relation was not found for Cu, Zn, and Pb. A decrease in the growth rate of structural and functional erythrocyte aberrations ("resistance improvement") with increasing cadmium load is probably due to compensatory enhancement of the synthesis of metallothioneins in the liver and kidneys and hence a greater proportion of Cd bound to metallothioneins. Problems of differences/similarities in Cd-associated reactivity among the animals are discussed too, taking into account the catching sites (polluted [Imp] and controls [Bg-1, and Bg-2]) and reproductive-age (i.e., immature underyearlings, mature underyearlings, and individuals that overwintered). The persistence of differences in erythrocyte status observed by us between the Imp and background groups after normalization to Cd load may be due to the action of other (unexamined) adverse factors and calls for further ecotoxicological studies.

Management of percutaneous treatment of aorta coarctation diagnosed during pregnancy.

Management of coarctation of the aorta (CoA) during pregnancy is complicated by increased procedural risks to the pregnant woman and her fetus. The aim of this research was to analyze 10-years of experience of CoA treatment diagnosed during pregnancy. During 2010-2020 we performed percutaneous stents implantations (SI) in 4 women during 15-23 weeks of pregnancy and in 6 women 48 hours - 5 years after delivery. In all presented cases, successful CoA repair was achieved. There was a significant decrease of peak-to-peak invasive systolic pressure gradient across the CoA (60.0±31.2 and 11.8±7.3 mmHg, p=0.001) and mean noninvasive systolic arterial pressure (163.0±46.2 and 120.5±9.2 mmHg, p=0.01) after SI. All percutaneously treated women during pregnancy (n=4) delivered healthy full-term babies. At follow-up (from 2 months to 10 years), all 10 women are alive without significant Doppler gradient across CoA with no signs of aortic aneurysm formation. To the best of our knowledge, we presented the largest published cohort of CoA percutaneous treatment during pregnancy. We categorized our experience in managing aortic coarctation diagnosed during pregnancy in one algorithm. Our experience demonstrates that excellent maternal and neonatal pregnancy outcomes can be obtained in women after CoA percutaneous repair, diagnosed during pregnancy. An aortic stent implantation is effective and safe for both mother and fetus.

Small mammals of background areas in the vicinity of the Karabash copper smelter (Southern Urals, Russia).

BACKGROUND: The dataset contains records of small mammals (Eulipotyphla and Rodentia) collected in the background (unpolluted) areas in the vicinity of Karabash copper smelter (Southern Urals, Russia) and the territory of the Sultanovskoye deposit of copper-pyrite ores before the start of its development. Data were collected during the snowless periods in 2007 (18 sampling plots), 2008-2010 (13 plots annually), 2011 (30 plots) and 2012-2014 (19 plots annually). The capture of animals was carried out in different types of forests (pine, birch, mixed and floodplain), sparse birch stands, reed swamps, marshy and dry meadows, border areas, a household waste dump, areas of ruderal vegetation and a temporary camp. Our study of small mammals was conducted using trap lines (snap and live traps). During the study period, 709 specimens of small mammals were caught, which belonged to five species of shrews and 13 species of rodents. The dataset may be highly useful for studying regional fauna and the distribution of species in different habitats and could also be used as reference values for environmental monitoring and conservation activities. NEW INFORMATION: Our dataset contains new information on occurrences of small mammals. It includes the peculiarities of their habitat distribution in the background areas in the vicinity of the large copper smelter and the deposit of copper-pyrite ores before the start of its development (Chelyabinsk Oblast, Russia). All occurrence records of 18 mammal species with georeferencing have been published in GBIF.

Immunophenotype of Measurable Residual Blast Cells as an Additional Prognostic Factor in Adults with B-Cell Acute Lymphoblastic Leukemia.

Measurable residual disease (MRD) is a well-known independent prognostic factor in acute leukemias, and multicolor flow cytometry (MFC) is widely used to detect MRD. MFC is able not only to enumerate MRD accurately but also to describe an antigen expression profile of residual blast cells. However, the relationship between MRD immunophenotype and patient survival probability has not yet been studied. We determined the prognostic impact of MRD immunophenotype in adults with B-cell acute lymphoblastic leukemia (B-ALL). In a multicenter study RALL-2016 (NCT03462095), 267 patients were enrolled from 2016 to 2022. MRD was assessed at the end of induction (day 70) in 94 patients with B-ALL by six- or 10-color flow cytometry in the bone marrow specimens. The 4 year relapse-free survival (RFS) was lower in MRD-positive B-ALL patients [37% vs. 78% (p < 0.0001)]. The absence of CD10, positive expression of CD38, and high expression of CD58 on MRD cells worsened the 4 year RFS [19% vs. 51% (p = 0.004), 0% vs. 51% (p < 0.0001), and 21% vs. 40% (p = 0.02), respectively]. The MRD immunophenotype is associated with RFS and could be an additional prognostic factor for B-ALL patients.

Severe vancomycin-induced thrombocytopenia in a patient with meningitis.

Vancomycin is a widely used antibiotic and rarely can cause drug-induced thrombocytopenia. A patient with hospital-acquired meningitis after neurosurgery was treated with systemic and intrathecal vancomycin. On 9th day of antibiotic treatment, the patient's platelets dropped to 0.68×109/L. Multiple platelet transfusions had minimal influence on platelet count. After cessation of vancomycin therapy, platelets returned to normal values without any additional interventions. Diagnosis of vancomycin-induced thrombocytopenia was confirmed by detection of drug-dependent antiplatelet IgG antibodies.

Humoral Effect of a B-Cell Tumor on the Bone Marrow Multipotent Mesenchymal Stromal Cells.

The properties of bone marrow (BM)-derived multipotent mesenchymal stromal cells (MSCs) are altered in the patients with the diffuse large B cell lymphoma (DLBCL) without BM involvement. It was suggested that plasma from the patients contains soluble factors that affect MSCs. Plasma and BM-derived MSCs from the DLBCL patients at the onset of the disease and one month after the end of treatment were studied. Concentration of the plasma cytokines and gene expression in the MSCs were evaluated by the Bio-Plex Pro Human Cytokine Panel kit to measure 27 analytes and real-time PCR. Plasma and MSCs from the healthy donors were used as controls. Analysis of cytokines in the plasma from healthy donors and patients before and one month after the end of treatment revealed significant differences in the concentration of 14 out of 27 cytokines. Correlations between the levels of secreted cytokines were altered in the plasma from patients indicating that the immune response regulation was disturbed. Cultivation of the MSCs from the healthy donors in the medium supplemented with the plasma from patients led to the changes in the MSC properties, similar to those observed in the MSCs from patients. The BM-derived MSCs were shown to participate in the humoral changes occurring in the DLBCL patients. For the first time, it was shown that the precursors of the stromal microenvironment - multipotent mesenchymal stromal cells - are altered in the patients with DLBCL without bone marrow involvement due to the humoral effect of the tumor and the response of organism to it. Comprehensive analysis of the results shows that, when remission is achieved in the patients with DLBCL, composition of the plasma cytokines normalizes, but does not reach the level observed in the healthy donors. The discovery of a new aspect of the effect of the tumor B-cells on the organism could help to reveal general regularities of the humoral effect of various tumors on the bone marrow stromal cells.

Rapid chemically selective 3D imaging in the mid-infrared.

The emerging technique of mid-infrared optical coherence tomography (MIR-OCT) takes advantage of the reduced scattering of MIR light in various materials and devices, enabling tomographic imaging at deeper penetration depths. Because of challenges in MIR detection technology, the image acquisition time is, however, significantly longer than for tomographic imaging methods in the visible/near-infrared. Here we demonstrate an alternative approach to MIR tomography with high-speed imaging capabilities. Through femtosecond nondegenerate two-photon absorption of MIR light in a conventional Si-based CCD camera, we achieve wide-field, high-definition tomographic imaging with chemical selectivity of structured materials and biological samples in mere seconds.

High expression of granzyme B in conventional CD4+ T cells is associated with increased relapses after allogeneic stem cells transplantation in patients with hematological malignancies.

Granzyme B is known to be a serine protease contained in granules of cytotoxic T cells. We have previously reported an influence of granzyme B expression in T regulatory cells (Tregs) on the risk of acute graft versus host disease (GVHD) onset. However, it is still unknown if conventional T cells (Tcon) use the granzyme B pathway as a mechanism of alloimmunity. We hypothesized that granzyme B in Tcon may affect recurrence within the first 6 months after allogeneic transplantation (allo-HSCT). A total of 65 patients with different hematological malignancies were included in this study. Blood samples were collected on day +30 after allo-HSCT. The percentage of granzyme B positive conventional T cells in patients who developed relapse in the first 6 months after allo-HSCT was 11.3 (4.5-35.3) compared to the others in continuous complete remission-1.3 (3.65-9.7), р = 0.011. The risk of relapse after allo-HSCT was in 3.9 times higher in patients with an increased percentage of granzyme B positive conventional T cells. The findings demonstrated that the percentage of granzyme B positive conventional T cells on day +30 after allo-HSCT could be a predictable marker of relapse within the first 6 months after allo-HSCT.

Diagnostic significance of flow cytometry scales in diagnostics of myelodysplastic syndromes.

BACKGROUND: Myelodysplastic syndromes (MDS) can present a challenge for clinicians. Multicolor flow cytometry (MFC) can aid in establishing a diagnosis. The aim of this study was to determine the optimal MFC approach for MDS. METHODS: The study included 102 MDS (39 low-grade MDS), 83 cytopenic patients without myeloid neoplastic disorders (control group), and 35 healthy donors. Bone marrow was analyzed using a six-color MFC. Analysis was conducted according to the "Ogata score," "Wells score," and the integrated flow cytometry (iFC) score. RESULTS: The respective sensitivity and specificity values were 77.5% and 90.4% for the Ogata score, 79.4% and 81.9% for the Wells score, and 87.3% and 87.6% for the iFC score. Specificity was not 100% due to deviations of MFC parameters in the control group. Patients with paroxysmal nocturnal hemoglobinuria (PNH) had higher levels of CD34+ CD7+ myeloid cells than donors. Aplastic anemia and PNH were characterized by a high proportion of CD56+ cells among CD34+ precursors and neutrophils. The proportion of MDS-related features increased with the progression of MDS. The highest number of CD34+ blasts was found in MDS with excess blasts. MDS with isolated del(5q) was characterized by a high proportion of CD34+ CD7+ cells and low granularity of neutrophils. In 39 low-grade MDS, the sensitivities were 53.8%, 61.5%, and 71.8% for Ogata score, Wells score, and iFC, respectively. CONCLUSION: The results support iFC as a useful diagnostic tool in MDS.

Immunophenotypic characteristics of multipotent mesenchymal stromal cells that affect the efficacy of their use in the prevention of acute graft vs host disease.

BACKGROUND: Multipotent mesenchymal stromal cells (MSCs) are widely used in the clinic due to their unique properties, namely, their ability to differentiate in all mesenchymal directions and their immunomodulatory activity. Healthy donor MSCs were used to prevent the development of acute graft vs host disease (GVHD) after allogeneic bone marrow transplantation (allo-BMT). The administration of MSCs to patients was not always effective. The MSCs obtained from different donors have individual characteristics. The differences between MSC samples may affect their clinical efficacy. AIM: To study the differences between effective and ineffective MSCs. METHODS: MSCs derived from the bone marrow of a hematopoietic stem cells donor were injected intravenously into allo-BMT recipients for GVHD prophylaxis at the moment of blood cell reconstitution. Aliquots of 52 MSC samples that were administered to patients were examined, and the same cells were cultured in the presence of peripheral blood mononuclear cells (PBMCs) from a third-party donor or treated with the pro-inflammatory cytokines IL-1ß, IFN and TNF. Flow cytometry revealed the immunophenotype of the nontreated MSCs, the MSCs cocultured with PBMCs for 4 d and the MSCs exposed to cytokines. The proportions of CD25-, CD146-, CD69-, HLA-DR- and PD-1-positive CD4+ and CD8+ cells and the distribution of various effector and memory cell subpopulations in the PBMCs cocultured with the MSCs were also determined. RESULTS: Differences in the immunophenotypes of effective and ineffective MSCs were observed. In the effective samples, the mean fluorescence intensity (MFI) of HLA-ABC, HLA-DR, CD105, and CD146 was significantly higher. After MSCs were treated with IFN or cocultured with PBMCs, the HLA-ABC, HLA-DR, CD90 and CD54 MFI showed a stronger increase in the effective MSCs, which indicated an increase in the immunomodulatory activity of these cells. When PBMCs were cocultured with effective MSCs, the proportions of CD4+ and CD8+central memory cells significantly decreased, and the proportion of CD8+CD146+ lymphocytes increased more than in the subpopulations of lymphocytes cocultured with MSC samples that were ineffective in the prevention of GVHD; in addition, the proportion of CD8+effector memory lymphocytes decreased in the PBMCs cocultured with the effective MSC samples but increased in the PBMCs cocultured with the ineffective MSC samples. The proportion of CD4+CD146+ lymphocytes increased only when cocultured with the inefficient samples. CONCLUSION: For the first time, differences were observed between MSC samples that were effective for GVHD prophylaxis and those that were ineffective. Thus, it was shown that the immunomodulatory activity of MSCs depends on the individual characteristics of the MSC population.

Structural-functional modifications of the liver to chronic radioactive exposure in pygmy wood mouse (Apodemus uralensis) within the East-Urals Radioactive Trace.

The hepatic parameters (contents of glycogen, total lipids, nuclear and cytoplasmic proteins, DNA and RNA, fructose-6-phosphate, water, lipid peroxidation products, as well as activities of succinate dehydrogenase and glucose phosphate isomerase), radiometric data, and the relative population abundance of the pygmy wood mouse (Apodemus uralensis Pall., 1811) inhabiting natural (Middle Urals, Southern Urals, and Trans-Urals) areas and radioactivity territory (the EURT zone after of the Kyshtym accident in the South Urals in 1957) were analysed. Structural-functional modifications of the liver in A. uralensis from the EURT area are presented, taking into account irradiation power by dose-forming radionuclides (external and internal exposure to 137Cs and 90Sr), population size, and reproductive status (sexually immature and sexually mature yearlings, representing different ontogenetic patterns). The sexually immature mice from the EURT area can be considered to be the more sensitive (reactive) intrapopulation group to synergistic factors, such as radiation burden and population overabundance. The extent of structural-functional hepatic modification under current conditions of radionuclide exposure, in addition to the 60 year long effect of radioactive contamination in the EURT, can exceed the level of natural (geographic) variation observed in this species in the Urals region, which points to a long term evolutionary-ecological process.