High Lactate Dehydrogenase Activity Research Articles

Computational fluid dynamics simulations of mitral paravalvular leaks in the human heart

Abstract Funding Acknowledgements Type of funding sources: Public Institution(s). Main funding source(s): Warsaw University of Technology - BIOTECHMED-3 project under the program Excellence Initiative: Research University. Background Presence of paravalvular leaks (PVL) can lead to hemolysis and high shear stress that affects erythrocytes passing through PVL channels may be the main reason behind this pathology. In vitro, shear stress forces above 300 Pa cause erythrocyte destruction. Distribution of shear stress can currently be precisely determined with computational fluid dynamics (CFD). CFD is a numerical modelling method based on solving equations of differential balance of mass, momentum and energy for a small section of fluid known as a computational cell. Purpose To investigate the magnitude of shear stress forces that affect erythrocytes in PVL channels and define properties of PVL channels that increase the likelihood of hemolysis. Methods Models of left-sided heart chambers with artificial mitral valves and PVLs were created based on cardiac computed tomography data from subjects with PVLs. Computational fluid dynamics principles were applied to create patterns of blood flow through PVL channels. 4 channels with cross-sectional areas (CSA) of 26.2 mm2 (model I), 15.7 mm2 (model 2), 20.4 mm2 (model 3) and 32 mm2 (model 4) were examined. Length of PVL channels was 6.5, 9.5, 7.3 and 3.25 mm respectively. In models I,III and IV the PVL channel was located anteriorly around the mitral annulus, whereas in model II a posterior location was described. Blood flow was simulated using a commercial CFD software (ANSYS Fluent 2020R1). Simulations were performed for heart rate of 60 beats per minute. The following variables were examined regarding blood flow through PVL channels: wall shear stress, shear stress in fluid, volume of PVL channels in which shear stress exceeded 300 Pa and duration of exposure of red blood cells to shear stress above 300 Pa. Obtained values were compared with lactate dehydrogenase (LDH) activities in subjects whose cardiac CTs were used to create PVL models. One of the models of the left ventricle is presented in picture 1. Studied PVL channels are presented in picture 2. Results In all models shear stress value of 300 Pa was exceeded. Maximal wall shear stress and maximal shear stress in fluid values were not related to CSA of the PVL channel. Lowest values of all examined variables were obtained for model 2, which had the longest PVL channel and a posterior location. The lowest LDH activity was noted in model with the lowest volume of the PVL channel with shear stress above 300 Pa (model 2) and the highest LDH activity in model with both the highest volume of the PVL channel with shear stress above 300 Pa and longest exposure to shear stress exceeding 300 Pa (model III). Conclusions Presence of PVLs is always accompanied by some degree of hemolysis. Shear stress values in PVL channels are not related to CSA of these channels. Volume of PVL channels in which shear stress exceeds 300 Pa and duration of exposure of erythrocytes to shear stress above 300 Pa may influence the severity of hemolysis.

Abstract 2177: Roles of lactate dehydrogenase (LDH) in melanoma: an underestimated prognostic biomarker

Abstract Introduction: High LDH activity is associated with a poor prognosis in many cancer types, especially in melanoma where it is the strongest indicator of poor outcome even with the most potent anticancer immunotherapies or targeted therapies. Although high LDH blood activity is often associated with high tumor burden, this is not always the case and there is no definitive demonstration that blood LDH activity is directly derived from tumors cells. Our objectives were 1) to explore the biological impact of LDHA and LDHB independently of their metabolic function 2) to explore the respective prognostic values of the 5 LDH isoforms as well as the relationship between tumor and blood LDH isoforms repartition in melanoma patients. Material, Patients, Methods: A375 melanoma cells cultured in glucose or galactose (to evaluate glycolytic independent effects) were silenced for LDHA or LDHB expression using si-RNAs. RNAseq analysis of total RNAs and polysomal RNAs was performed to explore the transcriptional and translational impacts of LDHA and LDHB expression. In melanoma patients treated with immunotherapy, the enzymatic activities of each of the 5 LDH isoforms (encoded by LDHA and LDHB) were measured in tumors and blood samples. The prognostic values of the distinct isoforms as well as the correlation between the isoforms distribution in blood and tumors were evaluated. Results: In vitro, independently of their metabolic effect, both LDHA and LDHB had distinct transcriptional and translational impacts on the control of several key oncogenic pathways (adhesion, survival, proliferation, immunity) with a more important effect of LDHA vs LDHB silencing. In vivo, among 64 melanoma patients including 25% with high LDH, high LDH1 (B4) activity was significantly associated with response and overall survival (OS) whereas high LDH4 (A3B1) was inversely associated with response and OS. LDH1 and LDH4 were associated with survival even among patients with normal LDH levels. For patients with paired (tumor and blood) isoforms analysis, there was no correlation between isoforms repartition suggesting that blood LDH activity is not directly related to tumor LDH. Conclusion: In addition to their metabolic activity, LDH enzymes, particularly LDHA, control several key oncogenic pathways at the transcriptional level but also at the translational level for a set of genes involved in cancer biology. LDH1 and LDH4 isoforms blood levels are differentially correlated with response to immunotherapy and survival. They are more robust prognostic biomarkers than total blood LDH level used in our daily practice. Thus, LDH is much more than just a marker of tumor burden in melanoma and the origins, the significance and the roles of blood LDH isoforms need to be revisited. These results highlight the need to develop anticancer drugs targeting LDH, and especially LDHA. Citation Format: Laura Soumoy, Virginie Quidville, Lisa Fredeau, Giuseppina Claps, Caroline Pradon, Ludovic Lacroix, Emilie Routier, Djaouida Belkadi, Séverine Roy, Feras Chehade, Stephan Vagner, Caroline Robert. Roles of lactate dehydrogenase (LDH) in melanoma: an underestimated prognostic biomarker [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2177.

Study of the Arginase Activity and Other Biochemical Parameters in Patients with Coronary Artery Disease in Baghdad Governorate-Iraq

The most prevalent form of heart disease and the main cause of death in both developed and developing nations is CAD. It happens when "plaque," or cholesterol or other fatty deposits that accumulate on the inner wall of the artery, narrows or blocks the arteries that deliver blood to the heart. Over time, chest pain might develop as a result of the reduction in blood flow to the heart caused by this plaque accumulation. The study was designed to find if Arginase acts as a biomarker for diagnosing Coronary Artery Disease (CAD). A total of 90 individual samples were included in the present study, the control group consist of 40 healthy individual samples, while the CAD patients were 50 individual samples. Some biochemical parameters such as fasting blood glucose (FBG), troponin I(TnI), high sensitivity C-reactive protein (hs-CRP), lipid profile, lactate dehydrogenase (LDH), and Arginase activity were analyzed. The results of the current study showed no significant differences in the average age of patients (67.00±6.78) when compared with the control group (61.10±6.46), P>0.05. A significant increase Was found in the FBI level, cholesterol, triglycerides, very low-density lipoprotein (VLDL), low-density lipoprotein (LDL), TnI, hs-CRP, LDH, and Arginase activity in the patient's group when compared with the control group. While significant decrease (P<0.05) was revealed in the high-density lipoprotein (HDL) level in CAD patients in comparison to the control group. Also, there was a positive significant correlation between Arginase activity with each age and FBG. As for the ROC operator curve for Arginase, it was found that the area under the curve (AUC) was 0.953 with a sensitivity of 90%, and specificity of 95%. The results in the present study indicate a possible use of Arginase as a diagnostic marker for CAD.

Enhanced Activity of NLRP3 Inflammasome in the Lung of Patients with Anti-Synthetase Syndrome.

Anti-synthetase syndrome (ASSD) is an autoimmune disorder characterized by inflammatory interstitial lung disease (ILD). The main objective of this work was to quantify the concentrations of cytokines and molecules associated with inflammasome activation in bronchoalveolar lavage (BAL) of patients with ASSD and a comparison group of systemic sclerosis (SSc) patients. Cytokines and lactate dehydrogenase (LDH) were determined using the concentrated BAL protein. The activity of caspase-1 and concentration of NLRP3 with the protein purified from the cell pellet in each group of patients. We found higher caspase-1 levels in ASSD vs. SSc, 1.25 RFU vs. 0.75 RFU p = 0.003, and LDH levels at 0.15 OD vs. 0.09 OD p < 0.001. A significant difference was observed in molecules associated with inflammasome activation, IL-18: 1.42 pg/mL vs. 0.87 pg/mL p = 0.02 and IFN-γ: 0.9 pg/mL vs. 0.86 pg/mL, p = 0.01. A positive correlation was found between caspase-1 and LDH in the patients with ASSD Rho 0.58 (p = 0.008) but not in the SSc group. In patients with ASSD, greater caspase-1 and higher LDH activity were observed in BAL, suggesting cell death due to pyroptosis and activation of the inflammasome pathway.

Tonsil microbiocenosis and salivary lactate dehydrogenase activity in children with infectious mononucleosis

The article analyzes clinical data, blood test results, tonsil mucus bacteriological testing, and salivary lactate dehydrogenase activity in children with acute infectious mononucleosis caused by an Epstein-Barr viral infection. Research involves 280 children aged from 7 months to 19 years 8 months with infectious mononucleosis, who were hospitalized in the Lviv Infectious Cli-nical Hospital. Group 1 included 234 children infectious mononucleosis caused by Epstein-Barr virus, group 2 — 46 patients with infectious mononucleosis caused by the combination of Epstein-Barr virus and cytomegalovirus. Severe hepatosplenomegaly, lymphadenitis, nasal speech and snoring, hyperthermic syndrome, acute tonsillitis were found in patients with infectious mononucleo-sis. A bacteriological study identified a number of pathogens that can be classified as pathogenic or opportunistic flora. Staphylococcus aureus, Streptococcus pneumoniae, Pseudomonas aeruginosa, Candida fungi were identified in 43.8 % of children with an isolated form of mononucleosis and in 51.7 % of children with mixed infection. In children with infectious mononucleosis caused by mixed infection, we observed the more severe course of the disease and a significantly higher lactate dehydrogenase activity. A significantly increased activity of lactate dehydrogenase in the saliva of patients with infectious mononucleosis in the presence of pathogenic microflora on the mucous membranes indicates destructive processes in the tissues of the tonsils.

Tailored upconversion nanomaterial: A hybrid nano fluorescent sensor for evaluating efficacy of lactate dehydrogenase inhibitors as anticancer drugs

High Lactate Dehydrogenase (LDH) activity, a distinct feature of cancer cells, is a possible causative factor for cancer cells to metastasize, infiltrate, invade, and deteriorate. The exploration of upconversion hybrid nano fluorescent sensor (HNFS) with the capability of evaluation the LDH inhibitors efficacy will be useful to accelerate the development of anti-cancer drugs from LDH inhibitors. Herein, we design an HNFS that can evaluate the inhibitors efficacy in vitro from the changes of the upconversion luminescence in the red-light region. HNFS is composed of two components: a Nile Blue A derivative (CPNB) that detects LDH activity, and tailored manganese doped upconverting nanoparticles that produce bright red emission which absorbs by CPNB via Förster resonance energy transfer. The cytotoxicity of HNFS in living cells (HeLa, A549 and Ges-1) was investigated by using the CCK-8 assay, which showed that HNFS possesses excellent biocompatibility. After incubating HNFS with three types of cells (A549, HeLa, and Ges-1), the mean fluorescence intensities of confocal images in vitro can illustrate the efficacy of LDH inhibitors to different cancer cells and distinguish healthy cells (Ges-1) from cancer cells (A549 and HeLa). Therefore, the systematic study of HNFS in vitro does encourage the potential applications of evaluating the LDH inhibitors efficacy in cancer cells.

Mitochondrial physiology and responses to elevated hydrogen sulphide in two isogenic lineages of an amphibious mangrove fish

Hydrogen sulphide (H2S) is toxic and can act as a selective pressure on aquatic organisms, facilitating a wide range of adaptations for life in sulphidic environments. Mangrove rivulus (Kryptolebias marmoratus) inhabit mangrove swamps and have developed high tolerance to environmental H2S. They are hermaphroditic and can self-fertilize, producing distinct isogenic lineages with different sensitivity to H2S. Here, we tested the hypothesis that observed differences in responses to H2S are the result of differences in mitochondrial functions. For this purpose, we performed two experimental series, testing (1) the overall mitochondrial oxidizing capacities and (2) the kinetics of apparent H2S mitochondrial oxidation and inhibition in two distinct lineages of mangrove rivulus, originally collected from Belize and Honduras. We used permeabilized livers from both lineages, measured mitochondrial oxidation, and monitored changes during gradual increases of sulphide. Ultimately, we determined that each lineage has a distinct strategy for coping with elevated H2S, indicating divergences in mitochondrial function and metabolism. The Honduras lineage has higher anaerobic capacity substantiated by higher lactate dehydrogenase activity and higher apparent H2S oxidation rates, likely enabling them to tolerate H2S by escaping aquatic H2S in a terrestrial environment. However, Belize fish have increased cytochrome c oxidase and citrate synthase activities as well as increased succinate contribution to mitochondrial respiration, allowing them to tolerate higher levels of aquatic H2S without inhibition of mitochondrial oxygen consumption. Our study reveals distinct physiological strategies in genetic lineages of a single species, indicating possible genetic and/or functional adaptations to sulphidic environments at the mitochondrial level.

Biochemical evaluation of processed ascites in patients undergoing cell-free and concentrated ascites reinfusion therapy.

The biochemical composition of processed ascites is not well researched and may differ among institutions. This prospective study was conducted to evaluate the biochemical characteristics of processed ascites of 11 patients with liver cirrhosis and carcinoma who underwent cell-free and concentrated ascites reinfusion therapy. The ascites due to carcinoma were more acidic and had higher lactate dehydrogenase activity than those due to liver cirrhosis. The ascites due to liver cirrhosis contained a higher amount of immunoglobulin than those due to carcinoma. Immunoglobulin preparations were approximately 2.95% IgG in liver cirrhosis ascites and 2.25% IgG in carcinoma ascites. Moreover, the concern about IgA infusion in the patient with IgA deficiency made it important to identify the source of the ascites. The present study provided fundamental information regarding the safety of cell-free and concentrated ascites reinfusion therapy.

Subclinical Mastitis Survey on Milk Composition in Dairy Sheep in Kurdistan Region of Iraq

Mastitis is an inflammatory condition of the mammary gland, characterized by the changes in the physical characteristics of the udder or milk. A cross sectional study was carried out in June, 2017 to December, 2018 to estimate the effect of subclinical mastitis (SCM) on milk composition in dairy sheeps in Kafri city of Kurdistan region of Iraq. Milk samples were gathered from residences of 295 sheeps with subclinical mastitis (California mastitis test (CMT) positive and somatic cell counts (SCC) &gt;600,000 cells/ml in individual quarter foremilk), as well as from 50 healthy controls. Contrasted to the levels watched in milk from healthy quarters, milk from quarters with subclinical mastitis exhibited raised high chloride (&gt;0.12 in contrast with &lt;0.12 g/dl), pH (5.65 in comparison to 5.57), sodium (86.87 vs 47.81 mg/dl), albumin (4.52 in contrast with 1.75 g/dl), immunoglobulins (24.66% in comparison to 5.73%) and lactate dehydrogenase (LDH) activity (1344.14 vs 449.84 IU/L). In compare, reduced values were discovered for potassium (147.47 in comparison to 161.34 mg/dl), inorganic phosphorous (19.42 in comparison to 26.48 mg/dl), calcium (86.35 vs 121.12 mg/dl), β-lactoglobulin (30.22% in comparison to 52.18%) and α- lactalbumin (19.15% vs 24.52%). In this study, no changes were seen in blood serum LDH activity. Moreover, an increase in positive response to CMT was found to be accompanied by an almost proportionate increase in immunoglobulin values to 44.32% and reduce of α-lactalbumin levels in milk serum (p&lt;0.01). These alterations in LDH activity, pH, mineral concentrations and protein fractions in milk of quarters display the presence of tissue injury provoked by SCM. Therefore, these parameters can be used in the diagnosis of mastitis. The current study revealed that changes of the foremilk chemical composition are connected to the subclinical mastitis; and that mastitis progression of quarters (CMT scores) influenced protein fractions in milk.

Altered glucose catabolism in the presynaptic and perisynaptic compartments of SOD1G93A mouse spinal cord and motor cortex indicates that mitochondria are the site of bioenergetic imbalance in ALS.

Amyotrophic lateral sclerosis is an adult-onset neurodegenerative disease that develops because of motor neuron death. Several mechanisms occur supporting neurodegeneration, including mitochondrial dysfunction. Recently, we demonstrated that the synaptosomes from the spinal cord of SOD1G93A mice, an in vitro model of presynapses, displayed impaired mitochondrial metabolism at early pre-symptomatic stages of the disease, whereas perisynaptic astrocyte particles, or gliosomes, were characterized by mild energy impairment only at symptomatic stages. This work aimed to understand whether mitochondrial impairment is a consequence of upstream metabolic damage. We analyzed the critical pathways involved in glucose catabolism at presynaptic and perisynaptic compartments. Spinal cord and motor cortex synaptosomes from SOD1G93A mice displayed high activity of hexokinase and phosphofructokinase, key glycolysis enzymes, and of citrate synthase and malate dehydrogenase, key Krebs cycle enzymes, but did not display high lactate dehydrogenase activity, the key enzyme in lactate fermentation. This enhancement was evident in the spinal cord from the early stages of the disease and in the motor cortex at only symptomatic stages. Conversely, an increase in glycolysis and lactate fermentation activity, but not Krebs cycle activity, was observed in gliosomes from the spinal cord and motor cortex of SOD1G93A mice although only at the symptomatic stages of the disease. The cited enzymatic activities were enhanced in spinal cord and motor cortex homogenates, paralleling the time-course of the effect observed in synaptosomes and gliosomes. The observed metabolic modifications might be considered an attempt to restore altered energetic balance and indicate that mitochondria represent the ultimate site of bioenergetic impairment.

Abstract P2-02-08: Slit2 induced anti-tumor activity may be mediated through metabolism driven immunomodulation

Abstract Metabolism of immune cells plays an important role in regulating tumor growth by modulating the anti-tumor M1 phenotype or pro-tumor M2 phenotype in macrophages. However, the role of bone marrow derived macrophages (BMDM) and their metabolic profile in promoting tumor growth is unknown. Slit2 is an anti-tumor molecule that is suppressed in breast cancer; however, the mechanism by which Slit2 mediates its function is not fully elucidated. We hypothesize that Slit2 mediated metabolic reprogramming of BMDMs favors anti-tumor M1 phenotype in these macrophages, which in turn reduces tumor growth. Here we assessed cellular metabolism in BMDMs from the MMTV-PyMT mouse, a model showing spontaneous tumor development. Shortly, age matched female mice with palpable tumors and control mice without tumors were treated with recombinant Slit2 (Slit2) or PBS intraperitoneally every third for 2 weeks (n=4 mice per group). Tumor volume was measured in each mouse before and at the end of treatment. Next, mice were euthanized and bone marrow was flushed from both the tibia and femor for culturing in vitro in the presence of macrophage chemotactic factor rich conditioned media. Rate of glycolysis was described based on extracellular acidification rate (ECAR) as measured by the Seahorse Bioanalyzer® under glycostress conditions. Lactate dehydrogenase (LDH) activity, which is linked with breast cancer progression and pro-tumor macrophage phenotype in experimental models, was also assayed using a commercially available kit. Finally, to elucidate potential pathways involved in Slit2 induced metabolic change, we assessed the expression of several proteins and factors involved in cellular metabolism. Firstly, we observed that PyMT mice treated with Slit2 showed lower tumor volume compared to mice treated with PBS confirming Slit2 anti-tumor activity. Hematoxylin &amp; Eosin staining of tumor sections from these mice also showed better tissue structure, with a higher cytoplasm to nuclear ratio in Slit2 treated PyMT mice compared to PBS treated mice. Furthermore, BMDMs from PyMT mice show lower aerobic glycolysis with higher lactate dehydrogenase activity (LDH) compared to control mice. Moreover, treatment with Slit2 appeared to lower LDH activity and trended to increase glycolysis in the BMDMs isolated from Slit2 treated PyMT compared to PBS treated PyMT. Expression analyses using quantitative PCR showed a 2-4 fold decrease in PGC-1α and CPT-2 in Slit2 treated BMDMs, indicating a reduction in fatty acid oxidation in these cells. This coupled with a 3 fold increase in IL-6 expression, and 2-3 fold decrease in arginase and IL-10 expression in tumor tissue suggest a potential shift from pro-tumor M2 to anti-tumor M1 phenotype. In spite of these preliminary trends, changes in metabolism and associated signals may be clearer in isolated, enriched populations of macrophages alone. Nevertheless, our findings suggest that Slit2 reduces tumor growth by affecting immune cell metabolism. Furthermore, these studies provide novel evidence of the potential immunomodulatory effects of Slit2 on macrophages. This may lead to development of Slit2 as a novel non-invasive therapeutic strategy against highly aggressive and metastatic cancers associated with high mortality and low quality of life. Citation Format: Kaul K, Ahirwar DK, Benej M, Denko N, Ganju R. Slit2 induced anti-tumor activity may be mediated through metabolism driven immunomodulation [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P2-02-08.

Primary and Secondary Central Nervous System Involvement in Acute Myeloid Leukemia

Introduction: Central Nervous System involvement (CNSi) in patients with Acute Myeloid Leukemia (AML) rarely occurs. It is not well characterized clinically and lacks standardized treatments. Patients and methods: A retrospective analysis of 77 consecutive AML patients with primary and secondary CNSi during 2004-2016 was performed in eight Polish haematological centres. Results: 77 patients (38 with primary CNSi-AML) were included. Median age was 44 years in both groups. Elevated lactate dehydrogenase activity was found in the majority of subjects. Patients in primary CNSi-AML group more often had myelomonocytic and monoblastic AML subtypes (68.4%) compared to secondary CNSi AML (43.5%) (p=0.039). There were no differences between both groups in the number of leukocytes or the proportion of blast cells rates in peripheral blood or bone marrow at AML diagnosis. There were also no statistically significant differences between both groups in the incidence of cytogenetic or molecular abnormalities. In both groups, CNSi was most frequently found in the cerebrospinal fluid and the most common neurological symptom was headache. The following manifestations were more frequent in secondary than in primary CNSi-AML: lower extremity weakness (38.46% vs. 13.16%; p=0.023), paraesthesia (38.46% vs. 13.16%; p=0.023), motor deficits (31.58% vs. 10.53%; p=0.047), and asymmetry of reflexes (26.32% vs. 2.7%; p=0.007). Median pleocytosis was also significantly higher in secondary than in primary CNSi-AML: 27 (IQR 2-146) vs. 2 (IQR: 1-12; p=0.004). Both groups had rather short Overall Survival (OS), with a median of 16.6 months (9.9-NA) for patients with primary CNSi-AML and 15.4 months (10.1-21.1) for patients with secondary CNSi. Conclusion: Patients with CNSi AML were relatively young, having high lactate dehydrogenase activity and high rates of the myelomonocytic and monoblastic/monocytic AML subtypes. The advisability of undertaking CNS examination and prophylaxis in patients with such characteristics thus merits further reassessment.

Hyperbaric oxygen rescues lung cancer cells from chemical hypoxia-induced low differentiation and apoptosis resistance

Hypoxia induces vigorous growth and a higher malignant phenotype in solid tumors. Hyperoxic treatment using hyperbaric oxygen (HBO) has previously been shown as a highly effective method to attenuate hypoxia. We aimed to investigate the effect of HBO on hypoxia-induced malignancy of lung cancer cells. Cobalt chloride (CoCl2) was used to induce chemical hypoxia in lung cancer cell line A549. Hypoxic inducible factor-1α (HIF-1α) expression, lactate dehydrogenase (LDH) activity, migration and invasion capacity, expression profiles of epithelial-mesenchymal transition (EMT) markers and apoptotic markers were assessed in CoCl2-treated A549 cells, with or without HBO treatment. Chemical hypoxia caused by CoCl2 resulted in high LDH activity, increased migration and invasion, decreased E-cadherin/N-cadherin ratio, enhanced EMT phenotype, higher Bcl-2/Bax ratio and elevated GRP78 expression. HBO treatment could significantly attenuate hypoxia-induced LDH activity, migration and invasion, restore hypoxia-reduced E-cadherin/N-cadherin ratio and EMT phenotype, as well as hypoxia-induced Bcl-2/Bax ratio, and repress GRP78 expression. HBO could serve as a reliable adjuvant treatment targeting the hypoxia microenvironment in solid tumors.

Structure and significance of cytogenetic abnormalities in adult patients with Ph-negative acute lymphoblastic leukemia.

To evaluate occurrence, variety, structural peculiarities and prognostic meaning of cytogenetic abnormalities in adult patients with Ph-negative acute lymphoblastic leukemia (ALL) receiving therapy according to ALL-2009 protocol. The study included 115 adult patients with firstly diagnosed Ph-negative ALL: 58 male and 57 female aged from 15 to 61 years (mean age 26.5 years), who underwent treatment from September 2009 to September 2015 in National Medical Research Center for Hematology MH RF (n=101) and in hematology departments of regional hospitals (n=14). All patients received therapy of ALL-2009 protocol (ClinicalTrials.gov, NCT01193933). The median follow-up was 24.5 months (0.2-94.4 months). As a part of the study results of a standard cytogenetic assay (SCA) were analyzed and fluorescence hybridization in situ (FISH) with the use of DNA-probes was performed on archived biological material for structural changes in gene locuses MLL/t(11q23), с-MYC/t(8q24), TP53/ deletion 17p13, CDKN2A/ deletion 9p21, translocation t(1;19)/E2A-PBX1 и t(12;21)/ETV6-RUNX1; iAMP21 identification. Karyotype was defined using SCA in 86% of patients. Normal karyotype was found in 48.5% of them, chromosome aberrations in 51.5% (structural changes were found in 19.2%, hyperploidy in 27.2%, and hypoploidy in 5.1%). In 17.2% of patients complex karyotype abnormalities were found. With the use of FISH technique aberrations were found in 67% of patients: 9p21/CDKN2A deletion in 24.3%, MLL/t(11q23) gene abnormalities in 7.8%, 17p13/TP53 deletion in 5.2%, abnormalities of c-MYC/t(8q24) in 1.7%, t(1;19)/E2A-PBX1 in 0.8%, and iAMP21 in 0.8%, other abnormalities (additional signals/absence of signals from gene locuses) in 26.4%, t(12;21)/ETV6-RUNX1 was not found. FISH technique use in addition to SCA allows to increase aberrant karyotype location from 51.5 to 67%. A statistically significant correlation of 9p21/CDKN2A deletion with high serum lactate dehydrogenase activity (p=0.02); MLL/t(11q23) gene abnormalities - with leucocytosis and high blast cells level in blood (p=0.0016), hyperploidy - with normal leukocyte count (p=0.02) was shown. In groups with different cytogenetic abnormalities no statistically significant differences of treatment with ALL-2009 protocol were found (in terms of complete remission, early mortality and treatment resistance). When connection of cytogenetic abnormalities and their combinations with long-term results were analyzed according to ALL-2009 protocol, only two characteristics - MLL/t(11q23) and c MYC/t(8q24) gene abnormalities had a statistically significant influence on disease-free survival (HR - 176.9; p&lt;0.0001) and chance of recurrence (HR - 6.4; p=0.02). Adverse prognostic factors in terms of therapeutic management provided in ALL-2009 protocol were MLL/t(11q23) and с-MYC/t(8q24) genes abnormalities. CDKN2A/9p21 and TP53/17p13 genes deletions, quantative and complex karyotype abnormalities were not prognostic factors in adult patients with Ph-negative ALL in ALL-2009 protocol use.

Mapping Metabolism: Monitoring Lactate Dehydrogenase Activity Directly in Tissue.

Mapping enzymatic activity in space and time is critical for understanding the molecular basis of cell behavior in normal tissue and disease. In situ metabolic activity assays can provide information about the spatial distribution of metabolic activity within a tissue. We provide here a detailed protocol for monitoring the activity of the enzyme lactate dehydrogenase directly in tissue samples. Lactate dehydrogenase is an important determinant of whether consumed glucose will be converted to energy via aerobic or anaerobic glycolysis. A solution containing lactate and NAD is provided to a frozen tissue section. Cells with high lactate dehydrogenase activity will convert the provided lactate to pyruvate, while simultaneously converting provided nicotinamide adenine dinucleotide (NAD) to NADH and a proton, which can be detected based on the reduction of nitrotetrazolium blue to formazan, which is visualized as a blue precipitate. We describe a detailed protocol for monitoring lactate dehydrogenase activity in mouse skin. Applying this protocol, we found that lactate dehydrogenase activity is high in the quiescent hair follicle stem cells within the skin. Applying the protocol to cultured mouse embryonic stem cells revealed higher staining in cultured embryonic stem cells than mouse embryonic fibroblasts. Analysis of freshly isolated mouse aorta revealed staining in smooth muscle cells perpendicular to the aorta. The methodology provided can be used to spatially map the activity of enzymes that generate a proton in frozen or fresh tissue.

Metabolic and oxidative stress responses of the jellyfish Cassiopea to pollution in the Gulf of Aqaba, Jordan

Physiological responses of jellyfish to pollution are virtually overlooked. We measured the activity of two glycolytic enzymes (pyruvate kinase (PK) and lactate dehydrogenase (LDH)), lipid peroxidation (LPO), protein and chlorophyll a content in the jellyfish Cassiopea sp. from polluted and reference sites along the Gulf of Aqaba, Jordan. In jellyfish from polluted sites, low PK/LDH ratios and high LDH activity clarify their reliance on anaerobic metabolism. PK and LDH were positively correlated in the jellyfish. While medusae from polluted sites showed no signs of oxidative stress damage, protein content was significantly lower. This might suggest protein utilization for energy production needed for maintenance. Unchanged LPO in polluted sites indicates the ability of jellyfish to keep reactive oxygen species under control. Overall these results suggest that the jellyfish seems to tolerate the current levels of pollution at the studied sites and they might be anaerobically poised to live at such habitats.

Biochemical and ultrasonographic evaluation of exertional rhabdomyolysis and myositis in draft horses

The purpose of this study was to evaluate the biochemical alterations in horses with exertional rhabdomyolysis (ER) and those with myositis and to characterize the progressive changes of both diseases by using ultrasonography (US). For this reason, 40 horses with various symptoms of musculoskeletal disorders were studied. Based on the competent case history, clinical symptoms, and in combination with clinicopathological findings as well as ultrasonographic alterations, the investigated horses were allocated into two main groups: ER-group (n = 7) and myositis group (n = 33). Blood samples were drawn from all investigated horses for laboratory assessment. The obtained findings revealed that horses with mild and severe forms of ER had a statistically significant (P < 0.05) high serum enzymatic activity of creatine phosphokinase (CPK), lactate dehydrogenase (LDH), and aspartate aminotransferase (AST), with significant (P < 0.05) low values of serum calcium compared with those of myositis as well as a control group. Horses with severe ER also demonstrated a statistically significant (P < 0.05) high value of blood urea nitrogen, serum magnesium, phosphorus, and potassium, with significant (P < 0.05) low values of serum chloride and sodium compared with other groups. Ultrasonically, it appeared that horses with severe ER have had hypoechoic loss of muscle texture, while mildly affected cases showed small hypoechoic cavities replacing the affected muscles. Small hypoechoic cavities separating muscle fibers were also observed in mild cases of myositis, but fibrinous hyperechoic dots prevailed in severe cases. From the results obtained, it can be concluded that the sole use of US for the diagnosis of ER and myositis in investigated horses was not really conclusive, but a combination of serum biochemical analyses, clinical examination findings, and US could facilitate early diagnosis of the studied diseases.

High resolution hyperpolarized 13 C MRSI using SPICE at 9.4T.

To test the feasibility of using the SPICE (SPectroscopic Imaging by exploiting spatiospectral CorrElation) technique, which uses the partial separability of spectroscopic data, for high resolution hyperpolarized (HP) 13 C spectroscopic imaging. Numerical simulations were performed to investigate the impact of transient HP signals on SPICE reconstruction. Furthermore, spectroscopic imaging exams from SPICE and conventional EPSI (echo-planar spectroscopic imaging) were simulated for comparison. For in vivo experiments, HP 13 C SPICE was performed in a mouse kidney by means of the injection of HP [1-13 C] pyruvate at 9.4T. The variation of lactate/pyruvate from the simulated SPICE was less than 4% under various factors that affect the transient HP signal, suggesting that the impact is negligible. We found that while HP 13 C EPSI was limited to the low signal-to-noise ratio (SNR) of lactate, these limitations were mitigated through HP 13 C SPICE, facilitating the improved SNR of lactate and the distinction of tissues. Acquisition of a high resolution HP 13 C spectroscopic image was possible for the in vivo experiments. With the fine structural information, the acquired image showed higher signal of pyruvate and lactate in the renal cortices than in the medullas, which is known to be attributed to higher activity of lactate dehydrogenase. The feasibility of HP 13 C SPICE was investigated. Simulation studies were conducted and in vivo experiments were performed in the mouse kidney at 9.4T. Results confirmed that a high resolution HP 13 C spectroscopic image with adequate spectral resolution can be obtained. Magn Reson Med 80:703-710, 2018. © 2018 International Society for Magnetic Resonance in Medicine.

Study of the influence of medicinal preparations on the indicators of mixed saliva in patients with essential hypertension

Aim. To identify the effect of drugs on the functional state of the salivary glands in patients with essential hypertension. Methods. A total of 38 hypertensive patients were examined. Depending on the prescribed drug therapy, all patients were divided into four groups: group 1 received an angiotensin-converting enzyme inhibitor; group 2 received a beta-blocker; group 3 received a slow calcium channel blocker, group 4 received a statin. For the analysis of salivary glands, the salivation rate was calculated, pH of the mixed saliva and activity of lactate dehydrogenase, alkaline phosphatase, aspartate aminotransferase, and alanine aminotransferase were measured. Results. In the mixed saliva of hypertensive patients, the salivation rate decreased in patients taking statins. The pH values of saliva in all groups were within neutral values, from 6.80 to 7.04. The amount of total protein was increased in patients taking angiotensin-converting enzyme inhibitors and statins, and was reduced in patients taking beta-adrenoblockers and slow calcium channel blockers. High activity of lactate dehydrogenase and aspartate aminotransferase was detected in patients taking statins, which indicates activation of anaerobic bacteria and inflammatory processes in the oral cavity. The increased activity of alkaline phosphatase, noted in patients taking beta-adrenoblockers, suggests possible violation of the processes of enamel mineralization. High activity of alkaline phosphatase and aspartate aminotransferase in the mixed saliva of observed patients coincides with the increase of papillary marginal alveolar index up to 27±4.6%, which indicates the development of pathology in the periodontal tissues. Conclusion. In patients receiving statins salivation function was suppressed; increased activity of lactate dehydrogenase and aspartate aminotransferase in the mixed saliva indicates activation of anaerobic microflora and tissue protein breakdown in the oral cavity.

Assessing Prostate Cancer Aggressiveness with Hyperpolarized Dual-Agent 3D Dynamic Imaging of Metabolism and Perfusion.

New magnetic resonance (MR) molecular imaging techniques offer the potential for noninvasive, simultaneous quantification of metabolic and perfusion parameters in tumors. This study applied a three-dimensional dynamic dual-agent hyperpolarized 13C magnetic resonance spectroscopic imaging approach with 13C-pyruvate and 13C-urea to investigate differences in perfusion and metabolism between low- and high-grade tumors in the transgenic adenocarcinoma of mouse prostate (TRAMP) transgenic mouse model of prostate cancer. Dynamic MR data were corrected for T1 relaxation and RF excitation and modeled to provide quantitative measures of pyruvate to lactate flux (kPL ) and urea perfusion (urea AUC) that correlated with TRAMP tumor histologic grade. kPL values were relatively higher for high-grade TRAMP tumors. The increase in kPL flux correlated significantly with higher lactate dehydrogenase activity and mRNA expression of Ldha, Mct1, and Mct4 as well as with more proliferative disease. There was a significant reduction in perfusion in high-grade tumors that associated with increased hypoxia and mRNA expression of Hif1α and Vegf and increased ktrans , attributed to increased blood vessel permeability. In 90% of the high-grade TRAMP tumors, a mismatch in perfusion and metabolism measurements was observed, with low perfusion being associated with increased kPL This perfusion-metabolism mismatch was also associated with metastasis. The molecular imaging approach we developed could be translated to investigate these imaging biomarkers for their diagnostic and prognostic power in future prostate cancer clinical trials. Cancer Res; 77(12); 3207-16. ©2017 AACR.