Increased Lactate Dehydrogenase Activity Research Articles

A comparison of seasonal flexibility in pectoralis muscle fiber type and enzyme activity in migratory and resident sparrow species

The pectoralis muscle in birds is important for flight and thermogenesis. In migratory songbirds this muscle exhibits seasonal flexibility in size, but whether this flexibility reflects changes in muscle fiber type has not been well documented. We investigated how seasonal changes in photoperiod affected pectoralis muscle fiber type and metabolic enzymes, comparing among three closely-related sparrow species: two seasonal migrants and one year-round, temperate climate resident. We quantified fast oxidative glycolytic (FOG) and fast glycolytic (FG) fibers histologically, and measured activities of citrate synthase (CS) and lactate dehydrogenase (LDH) in the pectoralis muscle of the three species that were acclimated to long- or short-daylight length conditions. In all species FOG was the predominant fiber type, but song sparrows had FG fibers regardless of daylight length. By contrast, Lincoln's sparrows incorporated FG fibers only under short-daylight length onditions, and house sparrows did not significantly express FG fibers, regardless of conditions. Both migratory species increased LDH activity in short-daylight conditions, but did not alter CS activity. In contrast, resident house sparrows did not alter CS or LDH activity with changes in daylight length. Our findings suggest that the presence of FG fibers is important for seasonal flexibility in LDH activity. Additionally, migratory species exhibited seasonal flexibility in muscle fiber type and enzyme activity, presumably to support migratory flight, while the resident species did not exhibit such seasonal flexibility, suggesting that this consistent phenotype is important year-round, despite changing thermogenic requirements.

Furan impairs cell function by inducing oxidative stress, DNA damage and apoptosis in mouse Sertoli cells in vitro.

Research on heat-induced food contaminants, such as furan, has shown its harmful effects on various systems. However, the impact of furan on Sertoli cells, a crucial male reproductive system cell, has not been studied. The investigation involved the treatment of furan to TM4 Sertoli cells at various concentrations (750, 1500, and 3000 µM) over a period of 24 h. This in vitro study determined that furan causes a decrease in Sertoli cell viability and an increase in lactate dehydrogenase activity, leading to cytotoxicity. Additionally, we observed an increase in MDA, one of the oxidative stress markers, in Sertoli cells, indicating that furan exposure leads to lipid peroxidation. It was determined that enzyme activities in the antioxidant defense system in Sertoli cells decreased after furan exposure. The findings indicate that furan induces oxidative damage in Sertoli cells by impairing the activity of antioxidant enzymes and promoting the production of ROS. This study discovered that furan triggers apoptosis in Sertoli cells by damaging DNA and altering the expression levels of apoptotic genes. Moreover, results suggest that furan causes cellular toxicity and apoptosis, leading to damage to Sertoli cells and thus causing male infertility.

Citreoviridin induces apoptosis through oxidative damage and inflammatory response in PC-12 cells.

Citreoviridin (CIT) is a mycotoxin produced by various fungi. Although CIT has been reported to cause neurotoxicity, the molecular mechanism is poorly understood. Therefore, the aim of this study was to investigate the effects and molecular mechanisms of CIT in neurotoxicity. Different concentrations of CIT were treated to rat pheochromocytoma (PC-12 cells), and oxidative stress parameters, cytokine levels, and cell apoptosis were evaluated. CIT treatment (5 and 10μM) significantly induced PC-12 cell apoptosis and increased lactate dehydrogenase activity. Additionally, CIT treatment induced oxidative stress, as evidenced by a significant increase in intracellular levels of reactive oxygen species, malondialdehyde, and superoxide dismutase and a decrease in glutathione activity. Moreover, CIT treatment induced an inflammatory response, as evidenced by a significant increase in the intracellular levels of the proinflammatory cytokines tumor necrosis factor-alpha and interleukin-1-beta in PC-12 cells. Furthermore, quantitative PCR and western blotting showed that CIT treatment increased both the protein and mRNA expression of GADD45α and p21 in PC-12 cells, suggesting that CIT may induce apoptosis by inhibiting cell cycle, blocking cell growth, and damaging DNA. Conclusively, this study contributes the understanding the toxicity mechanisms of CIT to nerve cells.

Regulatory mechanism of antioxidant enzymes on microbial metabolism and NADH in anaerobic fermentation of food waste for hydrogen production

Anaerobic fermentation is frequently hampered by toxicity arising from oxidative stress, and antioxidant enzymes play a crucial role in combating oxidative stress. In this study, the mechanism of microbial consortium and metabolic pathways regulated by antioxidant enzyme genes in anaerobic fermentation with different pH values was revealed. The results showed that antioxidant enzyme genes, such as glutathione peroxidase, peroxidase, and superoxide dismutase, were 5 times, 3 times, and 2 times higher at pH 7 than at pH 5, respectively. This allowed Clostridium to effectively resist oxidative stress, with substrate metabolism dominated by glycolysis, leading to an increase in the NADH/NAD+ ratio. Furthermore, the relative abundance of hydrogenase and electron transfer efficiency increased by 4.5 times and 2.71 times, respectively, resulting in a hydrogen production of 21.48 L/L. When antioxidant enzyme genes were inhibited at pH 5, the substrate mainly produced biomolecules for combating oxidative stress through the pentose phosphate and glycerophospholipid pathways, and led to the transformation of dominant genus into Lactobacillus. With an increased lactate dehydrogenase activity of 5.34 times, the final lactate production reached 65.17 g/L, which was 19.69 times higher than at pH 7. These results elucidate the regulatory mechanism of pH mediated antioxidant enzyme genes on hydrogen production and improve the controllability of anaerobic fermentation.

Hypoxia-acclimation adjusts skeletal muscle anaerobic metabolism and burst swim performance in a marine fish

Red drum, Sciaenops ocellatus, are a marine teleost native to the Gulf of Mexico that routinely experiences periods of low oxygen (hypoxia). Recent work has demonstrated this species has the capacity to improve aerobic performance in hypoxia through respiratory acclimation. However, it remains unknown how hypoxia acclimation impacts anaerobic metabolism in red drum, and the consequences of exhaustive exercise and recovery. Juvenile fish were acclimated to normoxia (n = 15, DO 90.4 ± 6.42 %) or hypoxia (n = 15, DO 33.6 ± 7.2 %) for 8 days then sampled at three time points: at rest, after exercise, and after a 3 h recovery period. The resting time point was used to characterize the acclimated phenotype, while the remaining time points demonstrate how this phenotype responds to exhaustive exercise. Whole blood, red muscle, white muscle, and heart tissues were sampled for metabolites and enzyme activity. The resting phenotype was characterized by lower pHe and changes to skeletal muscle ATP. Exhaustive exercise increased muscle lactate, and decreased phosphocreatine and ATP with no effect of acclimation. Interestingly, hypoxia-acclimated fish had higher pHe and pHi than control in all exercise time points. Red muscle ATP was lower in hypoxia-acclimated fish versus control at each sample period. Moreover, acclimated fish increased lactate dehydrogenase activity in the red muscle. Hypoxia acclimation increased white muscle ATP and hexokinase activity, a glycolytic enzyme. In a gait-transition swim test, hypoxia-acclimated fish recruited anaerobic-powered burst swimming at lower speeds in normoxia compared to control fish. These data suggest that acclimation increases reliance on anaerobic metabolism, and does not benefit recovery from exhaustive exercise.

A case of infectious mononucleosis caused by Epstein-Barr virus complicated by autoimmune hemolytic anemia

We represent a rare clinical case of infectious mononucleosis with hepatitis, caused by Epstein-Barr virus, complicated by rare secondary autoimmune hemolytic anemia associated with warm agglutinins in a 16-year-old patient. The disease was long-lasting, with a two-wave course of febrile fever. Signs of hepatitis with the development of anemia of moderate severity appeared during the second wave of febrile fever. The etiology of the disease was confirmed by the detection of Epstein-Barr virus DNA in blood and the detection of anti-VCA IgM antibodies to Epstein-Barr virus in the absence of anti-EBNA IgG antibodies. Hemolytic anemia was verified using markers of hemolysis (increased lactate dehydrogenase activity, decreased haptoglobin concentration), and positive results of a direct Coombs test. After successful initiation of prednisone therapy and clinical improvement, the patient was discharged from the hospital on the 24th day of disease to continue treatment on an outpatient basis (prednisone was replaced by methylprednisolone).This case demonstrates the development of a rare autoimmune complication in infectious mononucleosis caused by the Epstein-Barr virus.

MiR-224-5p overexpression inhibits oxidative stress by regulating the PI3K/Akt/FoxO1 axis to attenuate hypoxia/reoxygenation-induced cardiomyocyte injury

To investigate the regulatory role of miRNA-224-5p in hypoxia/reoxygenation (H/R) -induced H9c2 cardiomyocyte injury. Plasma samples were collected from 160 patients with acute myocardial infarction and 80 healthy controls(HC) to measure miRNA-224-5p levels and other biochemical parameters. In cultured H9c2 cells with H/R injury, the effects of transfection with miR-224-5p mimics or a negative control sequence on cell viability, malondialdehyde (MDA) content, and superoxide dismutase 2 (SOD2) and lactate dehydrogenase (LDH) activities were tested. Dual luciferase reporter gene assay was performed to verify the targeting relationship between miR-224-5p and PTEN. Bioinformatics methods were used to analyze the potential mechanisms of the target genes. The expression of miRNA-224-5p in the treated cells was detected with qRT-PCR, the protein expressions of PTEN, Bcl-2, Bax, cleaved caspase-3, SOD2, p-PI3K/PI3K, p-Akt/Ak and p-FoxO1/FoxO1 were determined using Western blotting, and cell apoptosis was analysed with flow cytometry. The levels of blood glucose, C-reactive protein, CK, CK-MB and cTnI were significantly higher in the AMI group compared with the HC group (P < 0.05). The expression level of miR-224-5p was significantly lowered in patients with STEMI and NSTEMI and in H9c2 cells with H/R injury. The viability of H9c2 cells decreased time-dependently following H/R injury. PTEN was a target gene of miR-224-5p, and the PI3K/Akt pathway was the most significantly enriched pathway. H9c2 cells with H/R injury showed significantly decreased SOD2 activity, increased LDH activity and MDA content, increased cell apoptosis, decreased protein expression levels of p-PI3K, p-Akt, p-FoxO1, SOD2, and Bcl-2, and increased expressions of PTEN, Bax, and cleaved caspase-3. These changes were obviously attenuated by trasnfection of the cells with miR-224-5p mimics prior to H/R exposure. MiR-224-5p overexpression upregulates the expression of the antioxidant gene SOD2 through the PI3K/Akt/FoxO1 axis to relieve H/R-induced oxidative stress and reduce apoptosis of H9c2 cells.

All-cause mortality and related factors in patients with varying degrees of peripheral blood eosinophilia.

The aim of this study was to investigate all-cause mortality rates and related factors in patients with different levels of eosinophilia. This retrospective cohort study was conducted between January 2020 and December 2022 in the Internal Medicine Department of Dr Sadi Konuk Training and Research Hospital, Istanbul, Turkiye. A total of 161 patients with eosinophilia (at least 3 times) were included and divided into groups with absolute eosinophil counts of 500-999/µL (mild), 1000-1500/µL (moderate), and >1500/µL (severe). The mean age of patients was 65.67 ± 16.64 years at the time of admission, and 45 patients (57.8%) were male. The rates of mortality, oncological disease, and organ involvement were significantly higher in the severe group (P < .05). Increased serum total immunoglobulin E and vitamin B12, hematocrit value, eosinophil-to-lymphocyte ratio, and leukocyte were observed in eosinophilic patients. Decreased lymphocyte count, hemoglobin and hematocrit values were higher in deceased patients than in survivors (P < .05). Increased eosinophil-to-lymphocyte ratio, C-reactive protein, vitamin B12, and lactate dehydrogenase (LDH) activity were observed in participants who died compared to those who survived (P < .05). Multivariable logistic regression revealed that advanced age and higher LDH activity were independently associated with greater mortality risk while receiving non-steroid anti-inflammatory drugs or proton-pump inhibitors were associated with reduced mortality risk (P < .05). Advanced age and increased LDH activity were independently associated with greater risk for mortality, whereas absolute eosinophil counts was not. Considering the literature on this topic, our results show the need for further clinical and fundamental research to understand the role of eosinophils in human disease.

Mechanism of tetramethylpyrazine attenuates inflammatory injury in endothelial cells by activating the SIRT1 signaling pathway

To study the effects and mechanisms of tetramethylpyrazine (TMP) on tumor necrosis factor-α (TNF-α)-induced inflammatory injury in human coronary artery endothelial cells (HCAEC). HCAEC were randomly divided into four groups: the control group (no treatment), the model group (treated with TNF-α, 50 ng/mL for 24 hours), the TMP group (pre-treated with TMP, 80 μg/mL for 12 hours followed by TNF-α treatment for 24 hours), and the SIRT1 inhibitor group (pre-treated with TMP and the specific SIRT1 inhibitor EX527 for 12 hours followed by TNF-α treatment for 24 hours). Cell viability was assessed using the CCK-8 method, lactate dehydrogenase (LDH) activity was measured using an LDH assay kit, reactive oxygen species (ROS) levels were observed using DCFH-DA staining, expression of pyroptosis-related proteins was detected by Western blot, and SIRT1 expression was analyzed using immunofluorescence staining. Compared to the control group, the model group showed decreased cell viability, increased LDH activity, ROS level and expression of pyroptosis-related proteins, and decreased SIRT1 expression (P<0.05). Compared to the model group, the TMP group exhibited increased cell viability, decreased LDH activity, ROS level and expression of pyroptosis-related proteins, and increased SIRT1 expression (P<0.05). In comparison to the TMP group, the SIRT1 inhibitor group showed decreased cell viability, increased LDH activity, ROS level and expression of pyroptosis-related proteins, and decreased SIRT1 expression (P<0.05). TMP may attenuate TNF-α-induced inflammatory injury in HCAEC, which is associated with the inhibition of pyroptosis and activation of the SIRT1 signaling pathway.

Atividade da lactato desidrogenase e proteína total como marcadores diagnósticos de efusões cavitárias e identificação de efusões neoplásicas em cães e gatos

ABSTRACT: The analysis and classification of cavitary effusions play a crucial role in determining a patient’s diagnosis and prognosis. This prospective study assed ed the activity of Lactate Dehydrogenase (LDH) and the concentration of Total Protein (TP) in serum, pleural, and peritoneal effusions in dogs and cats. Effusions were categorized into three groups: GI (low protein transudate), GII (high protein transudate), and GIII (exudate) following conventional classification. These groups, irrespective of species, were further divided into NPG (neoplastic) and NNG (non-neoplastic) groups. In dogs, significant differences were observed among groups GI, GII, and GIII in terms of effusion LDH activity/serum LDH (LDHR), effusion LDH activity, serum TP/effusion TP (TPR), and effusion TP. Increased LDH activity in effusion was associated with the presence of neoplasia. The groups organized based on the etiopathogenesis of the effusion exhibited varying values of serum TP, TPR, LDHR, effusion TP, and LDH activity in the effusion. Conversely, statistical analysis of the data from cats showed no differences between the groups for the parameters evaluated in this study. Compared to conventional classification, Light’s criteria demonstrated greater sensitivity in distinguishing between transudates and exudates and higher specificity in identifying transudates. We propose the use of established biochemical analyses to discern the mechanism of cavitary effusion formation and advocate for LDH activity measurement in effusions as a complementary diagnostic tool for cavitary neoplasms, especially in cases where cytological analysis of the effusion yields inconclusive results.

A New Role for Lactate in Breast Cancer

Lactate dehydrogenase (LDH) is an enzyme that plays a key role in cellular metabolism by converting pyruvate to lactate during anaerobic glycolysis. LDH has been implicated in various cancer types, including breast cancer, where increased LDH activity is associated with tumor growth, invasion, and resistance to therapy. LDH inhibitors are being investigated as potential therapeutic agents in cancer treatment, including breast cancer. Khajah and colleagues investigated the role of LDH inhibitors in both estrogen receptor (ER)+ and ER‑ breast cancer cell lines and in normal breast epithelial cells. A significant reduction in extracellular lactate level, cell proliferation, motility, and invasion was observed after treatment with LDH inhibitors. Subsequently, changes in the levels of vimentin, E‑cadherin, p38 MAPK, ERK1/2, and AKT were also observed. Cancer cell proliferation was preferentially blocked compared with normal epithelial cell inhibition by LDH inhibitors such as quercetin and lonidamine. These results further highlight the role of lactate in breast cancer progression and indicate the practical use of various commercially available LDH inhibitors as promising therapeutic agents to oppose the processes leading to cancer progression. While LDH inhibitors show promise, challenges include achieving selectivity for cancer cells and minimizing toxicity to normal tissues. The development of specific LDH inhibitors with favorable pharmacokinetic properties is an ongoing area of research. Mol Med Rep. 2024 Jan;29(1):12. doi: 10.3892/mmr.2023.1313.

Therapeutic efficacy of S-adenosylmethionine in chronic uranium intoxication in rats

Introduction. Chronic exposure to uranium compounds causes adverse cytotoxic and immunopathological effects in the body. The aim of the work was to evaluate the therapeutic efficacy of S-adenosylmethionine when administered intragastrically with respect to hepato-, nephro- and immunotoxic effects in chronic intoxicationwith uranyl acetate dihydrate (UAD). Materials and methods. Chronic intoxication was modeled by injecting 85 male rats with UAD solution (5.0 mg/kg/day by element) for one month. S-adenosylmethionine was administered to animals at a dose of 72.7 mg/kg for 21 days. Results. In chronic poisoning with UAD, 65% lethality, decrease of liver mass ratios (MR) and increase of renal MR were observed. Decrease in alkaline phosphatase activity, glucose level, lactic acid, number of CD4+ T-lymphocytes, increase in lactate dehydrogenase activity and creatinine level, number of CD8+ T-lymphocytes and apoptotic cell death, formation of catabolic pool of low and medium molecular weight substances (LMSMS) of blood plasma were registered. The results of urine analysis revealed the development of hyposthenuria, signs of glucosuria, hematosuria, proteinuria and leukocyturia, appearance of bilirubin in the urine. S-adenosylmethionine contributed to the decrease in the degree of target organ damage (reduction of fibroplastic and dystrophic changes in the liver and kidneys), normalization of immune system parameters (increase in CD4+ T-cells, decrease in CD8+ T-cells and frequency of apoptotic death of lymphocytes in immunocompromised animals) and endotoxicosis (decrease in the level of LMSMS from the catabolic pool area). Discussion. According to the totality of all revealed pathological changes, UAD poisoning led to the development of tubulointerstitial nephritis, metabolic disorders of detoxification functions of the liver. The causes of animal death during the first week of the experiment were acute kidney damage or confluent pneumonia. The appearance of bilirubin in the urine was associated with porphyrin metabolism disorders. In the remote period of intoxication an imbalance of the T-cell link of the immune system developed, as well as, probably, a decrease in nonspecific resistance, which led to the development of pneumonia in rats. Conclusion. S-adenosylmethionine therapy of hepato-, nephro- and immunotoxic effects in chronic UAD intoxication contributed to a decrease in the degree of damage to target organs, normalization of immune system parameters and endogenous intoxication.

Potential mechanisms of lung injury and repair after hexavalent chromium [Cr(VI)] aerosol whole-body dynamic exposure

Hexavalent chromium [Cr(VI)], known as "Top Hazardous Substances", poses a significant threat to the respiratory system. Nevertheless, the potential mechanisms of toxicity and the lung's repair ability after injury remain incompletely understood. In this study, Cr(VI) aerosol whole-body dynamic exposure system simulating real exposure scenarios of chromate workers was constructed to evaluate the lung injury and repair effects. Subsequently, miRNA sequencing, mRNA sequencing and metabolomics analyses on lung tissue were performed to explore the underlying mechanisms. Our results revealed that Cr(VI) exposure led to an increase in lactic dehydrogenase activity and a time-dependent decline in lung function. Notably, after 13 w of Cr(VI) exposure, alveolar hemorrhage, thickening of alveolar walls, emphysema-like changes, mitochondrial damage of alveolar epithelial cells and macrophage polarization changes were observed. Remarkably, a two-week repair intervention effectively ameliorated lung function decline and pulmonary injury. Furthermore, significant disruptions in the expressions of miRNAs and mRNAs involved in oxidative phosphorylation, glycerophospholipid metabolism and inflammatory signaling pathways were found. The two-week repair period resulted in the reversal of expression of oxidative phosphorylation related genes, and inhibited the inflammatory signaling pathways. This study concluded that the inhibition of the mitochondrial oxidative phosphorylation pathway and the subsequent enhancement of inflammatory response might be key mechanisms underlying Cr(VI) pulmonary toxicity, and timely cessation of exposure could effectively alleviate the pulmonary injury. These findings shed light on the potential mechanisms of Cr(VI) toxicity and provide crucial insights into the health protection for occupational populations exposed to Cr(VI).

Oxidative Stress Accompanies HIF1-Dependent Impairment of Glucose Metabolism in the Hippocampus of Adult Rats That Survived Prenatal Severe Hypoxia

Introduction: Many socially significant diseases are associated with prenatal developmental disorders. Previously, we showed the pathological role of hypoxia-inducible factor-1 (HIF1) in post-hypoxic reoxygenation. This study aimed to investigate the effect of prenatal severe hypoxia (PSH) on HIF1α protein expression as well as on HIF1-dependent activity of the pentose phosphate pathway (PPP) and anaerobic glycolysis in the hippocampus (HPC) of offspring that reached adulthood. Methods: PSH was induced during the critical period of fetal hippocampal formation on gestation days 14–16 in a hypobaric chamber (180 Torr, 5% oxygen, 3 h). Subsequent studies were conducted on both the HPC of adult control and PSH rats under normal conditions, as well as in response to severe hypoxia (SH) or psycho-emotional stress (“learned helplessness” [LH] model). We evaluated HIF1α protein levels using both immunohistochemistry and Western blotting techniques. The amount of glucose-6-phosphate dehydrogenase (G6PD) was also determined by Western blotting. Colorimetric enzymatic assays were employed to analyze enzymatic activity of lactate dehydrogenase (LDH), the concentration of lactate, NADPH, reduced glutathione (GSHred), and malonic dialdehyde (MDA). Results: We showed that PSH caused a stable increase in the content of HIF1α protein in the HPC, which was accompanied by an increase in the efficiency of anaerobic glycolysis. This was confirmed by increased LDH activity and lactate concentration. At the same time, the amounts of G6PD, NADPH, and GSHred decreased in the HPC of PSH rats, whereas the concentration of MDA, an oxidative stress marker, exceeded the control values. In a series of experiments using the LH or SH stress, it was shown that in the HPC of control rats, there was an increase in the amount of HIF1α in response to stress, which was also accompanied by more efficient anaerobic glycolysis and decrease of PPP-dependent NADPH production, similar to the intact PSH rats. In PSH rats, emotional stress resulted in higher HIF1α levels without affecting glycolysis or PPP. Conclusion: Therefore, the increased content and activity of the transcription factor HIF1α in the HPC of adult rats exposed to prenatal hypoxia leads to an imbalance between glycolysis and PPP, which is accompanied by oxidative stress.

Antioxidants As a Novel Treatment to Revert Impaired Angiogenic Potential Driven By Metabolic Alterations Observed in Shwachman-Diamond Syndrome Derived Mesenchymal Stromal Cells

Introduction Shwachman-Diamond Syndrome (SDS) is a rare bone marrow (BM) failure disorder. SDS BM biopsies show tortuous and collapsed vessels, highlighting angiogenic abnormalities. Our group previously demonstrated that SDS BM mesenchymal stromal cells (MSCs) have in vitro and in vivo impaired angiogenic potential compared to healthy donors (HD)-MSCs. Interestingly, recent evidence underlines that pathological angiogenesis could be accompanied by altered metabolism and that antioxidants could represent a novel strategy to target angiogenic defects. Aims We aimed to characterize the metabolic status of SDS-MSCs and to investigate if the treatment with antioxidants may restore their impaired angiogenic potential. Methods BM derived HD- and SDS-MSCs were stimulated with DMEM ± N-Acetylcysteine (NAC, 1mM) or dimethylsulfoxide (DMSO, 0.05% v/v) for 48h. Oxidative phosphorylation (OxPhos) was assayed by oximetry and bioluminescent ATP synthesis assay. Spectrophotometric analyses were performed to evaluate ATP/AMP ratio, malondialdehyde (MDA) level, and Complex IV and lactate dehydrogenase (LDH) activity. The amount of reactive oxygen species (ROS) was quantified by flow cytometry. The angiogenic capability was evaluated by performing in vitro Matrigel angiogenesis assay. Results Firstly, we evaluated OxPhos oxygen consumption and ATP production (n=6). Concerning the I-III-IV mitochondrial complexes pathway, SDS-MSCs consumed 57% less oxygen (p=0.004) and produced 64% less ATP compared to HD-MSCs (p=0.002). Accordingly, the analysis of the II-III-IV complexes pathway demonstrated that the oxygen consumption was reduced by 62% (p=0.002) and the ATP synthesis was 67% lower than HD-MSCs (p=0.002). Furthermore, the P/O ratio, index of OxPhos efficiency, was significantly reduced in SDS-MSCs in both the electron transport chain pathways (p=0.002 for both). Therefore, we demonstrated that Complex IV activity was 61% lower in SDS- vs HD-MSCs (p=0.002), highlighting its role in the SDS OxPhos defect. As for the energetic status, SDS-MSCs showed a low intracellular ATP/AMP ratio (mean=1.1, range=0.8-1.6 vs mean=3.6, range=3.1-4.0, in HDs; p=0.002) that was accompanied by 30% increase of LDH activity in SDS-MSCs (p=0.002), an expression of increased anaerobic glycolysis to compensate the mitochondrial defect. The amount of ROS in SDS-MSCs was increased by 27% compared to HD-MSCs (n=5) and also the SDS-MSCs lipid peroxidation level was significantly higher over HDs (mean=12.7 µM of MDA/mg, range=10.5-14.1µM/mg vs mean=6.6µM/mg, range=5.9-7.4µM/mg; p=0.002). Then we treated SDS-MSCs for 48h with NAC, a broad-range antioxidant (n=5), or with DMSO, which acts at very low concentrations as scavenger, specifically on lipid peroxidation products (n=6). Interestingly, NAC and DMSO stimulated SDS-MSCs increased by 75% the oxygen consumption and by 70% the ATP synthesis in both the electron transport chain pathways, thus resulting in levels comparable to HD-MSCs. The SDS OxPhos restoration was respectively associated to a 53% and 60% increase of Complex IV activity after NAC and DMSO stimulation (p&amp;lt;0.0001 for both). Moreover, antioxidants corrected the SDS energetic defect by restoring ATP/AMP ratio and LDH activity to the levels of HDs. Importantly, SDS-MSCs lipid peroxidation level was drastically reduced after NAC and DMSO treatments (mean=12.7µM/mg, range=10.5-14.1µM/mg vs NAC mean=5.1µM/mg, range=4.3-5.4µM/mg and DMSO mean=6.2µM/mg, range=5.8-6.5µM/mg; p&amp;lt;0.0001 for both). Finally, we approached SDS-MSCs angiogenic defect and showed that SDS-MSCs defective capability to recreate a defined capillary-like network under angiogenic stimuli was completely restored after antioxidant stimulations (n≥5). As shown by ImageJ Angiogenesis Analyzer several angiogenic elements, including branches and segments, were significantly increased in NAC and DMSO stimulated SDS-MSCs that become comparable to HD-MSCs. Conclusions We demonstrated that the altered OxPhos metabolism of SDS-MSCs significantly contributes to their angiogenic defect and, importantly, that antioxidants restored the metabolic alterations and the angiogenic potential in SDS-MSCs, paving the way for new therapeutic strategies.

Peculiarities of the relationship between the terminal site of glycolysis and the initial segment of gluconeogenesis in the myocardium and skeletal muscles of animals irradiated at different doses

Existing types of muscles differ not only structurally, but also metabolically (direction of energy exchange, choice of biosubstrates, dependence on the action of mediators, hormones, etc.). A number of studies have been devoted to the study of the impact of ionizing radiation on metabolism in muscle tissue. After irradiation, thickened muscle cells with enlarged nuclei appear in the heart muscle, the number of which does not decrease for 6 months, which indicates a violation of nuclear-cytoplasmic relations in myocardial cells. Phosphoenolpyruvate carboxykinase which ensures the utilization of cytoplasmic oxaloacetate and its transformation into phosphoenolpyruvate, completes the initial stage of gluconeogenesis and can limit the rate of gluconeogenesis from lactate. It should be emphasized that phosphoenolpyruvate carboxykinase is more active in skeletal muscles, where the activity of pyruvate kinase and lactate dehydrogenase is increased. The purpose of the work is to investigate the peculiarities of the relationship between the terminal site of glycolysis and the initial segment of gluconeogenesis in the myocardium and skeletal muscles of animals irradiated at different doses. The authors proved that in animals irradiated at a dose of 0.5 Gy, the activity of pyruvate kinase in the myocardium and skeletal muscle increases compared to intact animals. In the blood, on the contrary, there is a decrease in the activity of this enzyme compared to intact animals. When animals are irradiated at a dose of 1.0 Gy, diametrically opposite changes are observed. When animals are irradiated at a dose of 0.5 Gy, there is a slight decrease in the activity of lactate dehydrogenase in the myocardium and blood against the background of an increase in the activity of this enzyme in skeletal muscle. Irradiation of animals at a dose of 1.0 Gy leads to an acute increase in lactate dehydrogenase activity in the myocardium and skeletal muscle. The data obtained revealed that in animals irradiated at a dose of 0.5 Gy, the activity of phosphoenolpyruvate carboxykinase in skeletal muscle increases, while in cardiac muscle, on the contrary, its activity decreases as well as in blood. With an increase in the radiation dose to 1.0 Gy, diametrically opposite changes in the activity of phosphoenolpyruvate carboxykinase are observed. The authors concluded that outlined changes in the waу of intramuscular glycolysis and gluconeogenesis reactions show the pathophysiologcal mechanisms of biochemical supply restructuring as the result of ionizing irradiation influence. From the fundamental point of view these results show the direction of pathophysiologically oriented pharmacological correction of radiation-provoked muscle disturbances.

Incidence and Role of Recipient-Specific Antibodies in Allogeneic Hematopoietic Cell Transplantation from Mismatched Related Donors

High titer of donor-specific antibodies (DSAs) increases the risk of graft rejection after mismatched related hematopoietic cell transplantation (HCT). There are no data regarding the incidence of anti-HLA recipient-specific antibodies (RSAs) and their role after transplantation. Here we aimed to identify the incidence of RSAs in a mismatched related hematopoietic cell donor population and their possible impact on immune-mediated complications, such as acute graft-versus-host disease (aGVHD), and complications resulting from endothelial injury, such as transplantation-associated thrombotic microangiopathy (TA-TMA) and veno-occlusive disease (VOD). We prospectively analyzed the incidence of anti-HLA antibodies in 28 mismatched related pairs of recipients and their donors who underwent HCT at our center between 2020 and 2022. In positive samples screened for anti-HLA class I and/or II antibodies, the specificity of the HLA antibodies was analyzed. All recipients had a hematologic malignancy and received a myeloablative conditioning regimen and immunosuppression consisting of post-transplantation cyclophosphamide, tacrolimus, and mycophenolate mofetil. Patients were tested for TA-TMA and aGVHD development during routine post-transplantation visits up to 100 days post-transplantation. We used modified Jodele criteria for TA-TMA diagnosis, and based aGVHD grading on the MAGIC criteria. VOD was assessed using the European Society for Blood and Marrow Transplantation. Anti-HLA antibodies were detected in 12 donors (43%) and in 9 recipients (32%). There were no significant differences between donors and recipients according to age (median, 42 years [range, 17 to 69 years] versus 39 years [range, 8 to 68 years]), sex, or pregnancy history. No transfusion history was noted in the donor group (P < .05). RSA antibodies were present more often than DSAs and were detected in 9 out of 12 (75%) anti-HLA-positive donors and in only 2 out of 9 (22%) recipients, respectively (P < .05). During the follow-up, 11 patients (39%) developed aGVHD, including grade I-II in 9 (32%) and grade III-IV in 2 (7%). Twelve patients (43%) met the criteria for TA-TMA, and only 1 patient (3.5%) was diagnosed with VOD by day 100 post-HCT. RSAs were detected significantly more often in the TA-TMA group; among 12 patients diagnosed with TA-TMA, 7 (58%) had RSAs (P < .05). We did not find a correlation between RSAs and aGVHD. The patient with VOD did not have an RSA-positive donor. There was no difference in membrane attack complex (MAC) concentration in the RSA-positive group on day 30 and day 60 post-HCT; however, there was a trend toward higher MAC concentration in the RSA-positive group on day 100 (median, 912 ng/mL [range, 788 to 1120 ng/mL] versus 616 ng/mL [range, 352 to 1244 ng/mL]; P=.055). Patients with RSA suffered more often from platelet and red blood cell decreases or transfusion refractoriness, and increased lactate dehydrogenase activity was observed in all RSA-positive cases. The donor immune status and the presence of RSA may be associated with higher rates of TA-TMA in mismatched HCT recipients. Antibody-mediated complement activation might be an additional factor influencing TA-TMA occurrence.

Hesperetin Alleviates Doxorubicin-Induced Cytotoxicity in H9c2 Cells by Activating SIRT1/NRF2 Signaling

To investigate whether hesperetin (Hes) alleviates doxorubicin (DOX)-induced cardiomyocytotoxicity by reducing oxidative stress via regulating silent information regulator 1 (SIRT1)/nuclear transcription factor E2-related factor 2 (NRF2) signaling in H9c2 cells. H9c2 cells were treated with DOX to establish the cardiotoxicity model and were randomly assigned to four groups, a control group (Control) and three treatment groups, receiving respectively DOX (the DOX group), Hes+DOX (the DOX+Hes group), and Hes+SIRT1 inhibitor EX527+DOX (the DOX+Hes+EX527 group). Cellular morphology was observed by the light microscope. Cell viability was evaluated by CCK-8. DOX-induced apoptosis in H9c2 cells was examined by flow cytometry. The levels of reactive oxygen species (ROS) in the H9c2 cells of the four groups were determied with 2'-7'-dichlorodihydrofluorescein diacetate (DCFH-DA) staining. The activities of lactate dehydrogenase (LDH), superoxide dismutase (SOD), catalase (CAT), and SIRT1 as well as the malondialdehyde (MDA) content were measured using ELISA kits. The expressions of cleaved caspase-3, cytochrome c, SIRT1, Ac-FOXO1, NRF2, and heme oxygenase 1 (HO-1) were determined by Western blot. Compared with the Control group, the DOX group showed swollen cellular morphology, decreased cell density and viability, and increased LDH activity in the medium ( P<0.01); both apoptosis and the expression of cleaved caspase-3 and cytochrome c increased ( P<0.01); the activities of CAT and SOD decreased while the contents of MDA and ROS increased ( P<0.01); the expression of SIRT1, NRF2, and HO-1 decreased, the activity of SIRT1 decreased, and the expression of Ac-FOXO1 increased ( P<0.01). Compared with the DOX group, the DOX+Hes group showed improved cellular morphology, increased cell density and viability, and decreased LDH activity in the medium ( P<0.01); the apoptosis and the expression of cleaved caspase-3 and cytochrome c decreased ( P<0.01); the activities of CAT and SOD increased while the levels of MDA and ROS decreased ( P<0.01); the expression of SIRT1, NRF2, and HO-1 increased, the activity of SIRT1 increased, and the expression of Ac-FOXO1 decreased ( P<0.01). Comparison of the findings for the DOX+Hes group and the DOX+Hes+EX527 group showed that EX527 could block the protective effects of Hes against DOX-induced cell injury, oxidative stress, and SIRT1/NRF2 signaling. Hes inhibits oxidative stress and apoptosis via regulating SIRT1/NRF2 signaling, thereby reducing DOX-induced cardiotoxicity in H9c2 cells.

Effects of Non-Alcoholic Beer after Running in Three Consecutive Days on Antioxidant Enzyme Activity and Muscle Damage Biomarkers

Running recovery is challenging for several body systems and can be improved by nutritional focus. Non-alcoholic beer is a widely used post-exercise beverage for its antioxidant and energetic properties. After three consecutive days of 1 h submaximal running (80% HRmax), antioxidant enzyme activity (glutathione peroxidase [GPx], glutathione reductase [GR], catalase), lactate dehydrogenase (LDH) activity as a muscle damage blood marker, and lower limb thermographic values were determined in order to observe possible changes in 20 subjects divided into two groups: control (n = 10) and NAB (n = 10). NAB drank 10 mL/kg of non-alcoholic beer post-exercise (both groups drank water ad libitum). Non-alcoholic beer did not show statistically significant changes compared to water. Regarding the effect size, the NAB group had a medium increase in thermography values (15′Post-15′Pre) on days 1 and 2 compared to the control group; a large increase in LDH activity (both 60′Post-0′Post and 60′Post-Pre) on day 2, and a medium increase (60′Post-0′Post) on day 3; a medium decrease in GR (60′Post-Pre) on days 1 and 3; and a large (60′Post-0′Post) and medium (60′Post-Pre) decrease in GPx on day 3. These findings support the idea that non-alcoholic beer is not an appropriate recovery beverage after 1 h running for three consecutive days.

BHTCM Protects Müller Cells from Diabetic Retinopathy by Reducing Abnormal Changes of Kir4.1 and AQP4, Suppressing VEGF and IL-1β, and Enhancing PEDF Production

Background: In the diabetic condition, damage to the Müller cells contributes to the pathogenesis of diabetic retinopathy. Aims: This study aimed to investigate the protective effect of Bushen Huoxue, Traditional Chinese Medicine (BHTCM), on Müller in diabetic retinopathological conditions. Methods: Primary rat retinal Müller cells (RRMC) were isolated and cultured under high glucose (50 nmol/L). The advanced glycation end products (AGEs) and sodium dithionite were applied to treat highglucose administrated RRMC to mimic diabetic retinopathological conditions. The effects of BHTCM on diabetic retinopathological RRMC were evaluated. The expressions of aquaporin-4 (AQP4) and Kir4.1 were determined by double-labeling immunofluorescence and ELISA. Levels of vascular endothelial growth factor (VEGF), interleukin-1β (IL-1β) and pigment epithelium-derived factor (PEDF) were examined with ELISA. Lactate dehydrogenase (LDH) activity was also evaluated. Results: Retinal Müller cells were successfully isolated and identified. RRMC treated with AGEs and sodium dithionite resulted in the increase of AQP4 and decrease of Kir4.1 in RRMC, increase of VEGF and IL-1β secretion, increase of LDH activity, decrease of PEDF secretion in culture medium, all of which, in a dose-dependent or time-dependent manner. Post treating RRMC with AGEs and dithionite, BHTCM reversed changes in expression of AQP4 and Kir4.1 in RRMC, and reversed VEGF levels, PEDF and IL-1β secretion in the culture medium. Moreover, BHTCM reversed the decrease of RRMC cell membrane integrity after AGEs and dithionite treatment. Conclusions: BHTCM protected Müller cells from diabetic damage by reducing abnormal changes of Kir4.1 and AQP4, inhibiting VEGF and IL-1β, increasing PEDF production, and maintaining cell membrane integrity. Therefore, BHTCM is a potential drug for the treatment of diabetic retinopathy, which can correct the function of Müller cells.