Inhibition Of Lactate Dehydrogenase Activity Research Articles

Inhibition of Lactate Dehydrogenase-A by Singlet Oxygen and Hypochlorous Acid via Cysteine Oxidation and Irreversible Conformational Changes

Muscle lactate dehydrogenase (LDH-A) catalyzes the reduction of pyruvate to lactate, the end product of anaerobic glycolysis. LDH-A is overexpressed in many cancers prior to and even when tumors receive adequate oxygen, and lactate has multiple cellular roles. We assessed the effect of singlet oxygen and hypochlorous acid (HOCl) on mammalian LDH-A. Oxidants induced distinct patterns of protein crosslinks observed by SDS-PAGE under reducing conditions. LDH-A cysteines were detected using fluorescein-modified maleimide to assess their oxidation and accessibility. Singlet oxygen initially increased cysteine exposure, but higher doses resulted in their oxidation in addition to non-reducible covalent crosslinks. LDH-A cysteines were oxidized by micromolar HOCl (1–10 equivalents over enzyme) but were resistant to millimolar H2O2, chloramines and Angeli’s salt. HOCl oxidation inhibited LDH-A activity and yielded inter-chain disulfides observed by nonreducing SDS-PAGE. Disulfide reduction did not restore LDH-A activity that was lost due to HOCl oxidation. An irreversible conformational change induced by HOCl was detected by native gel electrophoresis and tryptophan fluorescence. In the absence of pyruvate, LDH-A enhanced NADH oxidation resulting in H2O2 formation. Singlet oxygen, but not HOCl, initiated this superoxide-dependent chain reaction. Once damaged by both singlet oxygen or HOCl, LDH-A had decreased NADH oxidation activity.

Microcapsules of a cinnamon, peppermint, and lemon essential oil mix by spray drying: Preparation, characterization and antibacterial functions

Plant essential oils (EO) are increasingly used as food preservatives due to their antibacterial and antioxidant activities. For such applications, EO are commonly microencapsulated to overcome their poor stability, low solubility and dominant aromas. But, typically, individual EO act against only a narrow range of bacterial species. Therefore, in this study, we aimed to develop a microencapsulated preservative using a combination of three EO and to determine its anti-bacterial action against three common food contaminants, viz. Escherichia coli, Salmonella and Staphylococcus aureus, and one, usually, beneficial species, viz. Lactoplantibacillus plantarum. The EO of cinnamon (Cinnamomum cassia), peppermint (Mentha haplocalyx) and lemon (Citrus limon) were combined in the proportions, 1.0 : 1.8: 2.4, respectively and were encapsulated in gelatin-chitosan by spray drying. The particle size, zeta potential, moisture content, and encapsulation rate of the essential oil microcapsules (EOM) were measured. With increasing EOM exposure time, the concentrations of E. coli, Salmonella, S. aureus and L. plantarum decreased by 3, 3, 4, and 5 orders of magnitude, respectively. Cell apoptosis accompanied loss of membrane integrity and permeability, inhibition of lactate dehydrogenase activity and disruption of organic acid metabolism. Gram-positive bacteria (S. aureus and L. plantarum) were more susceptible to those effects of EOM than Gram-negative bacteria (E. coli and Salmonella). It also demonstrates that in applications as preservatives for probiotic foods requiring live L. plantarum, EOM comprising cinnamon, peppermint or lemon EO should be avoided.

ROCK2 knockdown alleviates LPS-induced inflammatory injury and apoptosis of renal tubular epithelial cells via the NF-κB/NLRP3 signaling pathway.

Rho-associated protein kinase 2 (ROCK2) is an important regulator of the inflammatory response and has been reported to serve a role in sepsis. The present study aimed to investigate whether ROCK2 served a role in sepsis-associated acute kidney injury (S-AKI). HK-2 cells were stimulated with lipopolysaccharide (LPS) to simulate S-AKI in vitro. Subsequently, the change in ROCK2 expression levels were determined. ROCK2 in LPS-induced HK-2 cells was knocked down using short hairpin RNA-ROCK2, in the absence or presence of phorbol 12-myristate 13-acetate (PMA), an activator of NF-κB. Cell viability, cytotoxicity, inflammation and apoptosis were assessed using MTT, lactate dehydrogenase (LDH) release, reverse transcription-quantitative PCR, ELISA, TUNEL and western blotting assays. The protein expression levels of proteins involved in the NF-κB/NLR family pyrin domain containing 3 (NLRP3) signaling pathway were also assessed using western blotting. The results demonstrated that ROCK2 was upregulated in HK-2 cells upon LPS treatment. LPS also reduced cell viability, promoted LDH activity and increased TNF-α, IL-6 and IL-1β mRNA expression levels and concentrations. Apoptosis was also induced by LPS as indicated by an increase in the proportion of TUNEL-positive cells, decreased Bcl-2 protein expression levels and increased cleaved caspase-3 and cleaved poly (ADP-ribose) polymerase protein expression levels. However, ROCK2 knockdown in LPS-induced HK-2 cells reversed cell viability damage and inhibited LDH activity, the generation of pro-inflammatory cytokines and apoptosis caused by LPS. Furthermore, ROCK2 knockdown inhibited the LPS-induced expression of phosphorylated-NF-κB p65, NLRP3, apoptosis-associated speck-like protein containing a CARD and caspase-1 p20. PMA treatment reversed all the aforementioned effects of ROCK2 knockdown on LPS-treated HK-2 cells. Therefore, ROCK2 knockdown may alleviate LPS-induced HK-2 cell injury via the inactivation of the NF-κB/NLRP3 signaling pathway.

First-in-class topical therapeutic omilancor ameliorates disease severity and inflammation through activation of LANCL2 pathway in psoriasis

Psoriasis (PsO) is a complex immune-mediated disease that afflicts 100 million people. Omilancor is a locally-acting, small molecule that selectively activates the Lanthionine Synthetase C-like 2 (LANCL2) pathway, resulting in immunoregulatory effects at the intersection of immunity and metabolism. Topical omilancor treatment in an imiquimod-induced mouse model of PsO ameliorates disease severity, epidermal hyperplasia and acanthosis. Further, pharmacological activation of LANCL2 results in significant downregulation of proinflammatory markers including local reduction of IL17, and infiltration of proinflammatory cell subsets. These therapeutic effects were further validated in an IL-23 PsO model. This model reported increased preservation of homeostatic skin structure, accompanied by a decreased infiltration of proinflammatory T cell subsets. In CD4+ T cells and Th17 cells, the LANCL2 pathway regulates proinflammatory cytokine production, proliferation and glucose metabolism. Metabolically, the loss of Lancl2 resulted in increased glycolytic rates, lactate production and upregulated enzymatic activity of hexokinase and lactate dehydrogenase (LDH). Inhibition of LDH activity abrogated the increased proliferation rate in Lancl2−/− CD4+ T cells. Additionally, topical omilancor treatment decreased the metabolic upregulation in keratinocytes, keratinocyte hyperproliferation and expression of inflammatory markers. Omilancor is a promising topical, LANCL2-targeting therapeutic candidate for the treatment of PsO and other dermatology indications.

Ameliorative effect of piracetam on emamectin benzoate induced perturbations in the activity of lactate dehydrogenase in murine system

Emamectin benzoate (4´-deoxy-4´-epimethylaminoavermectin B1) is a macrocyclic lactone developed for the control of insect pests in edible plant crops. The current study was taken to assess the possible ameliorative effect of piracetam (P) on emamectin benzoate (EMB) induced alterations in energy metabolism in liver, kidney, testis, heart, cerebral cortex, cerebellum and muscle of rats. Rats of normal control group (N) received water and normal diet ad libitum and (EMB as well as EMB+P) group received 5 mg/kg bw emamectin benzoate via drinking water for 4 weeks. After four weeks of EMB treatment, rats of (EMB+P) received piracetam (500 mg/kg bw, iv) for 15 days. The results demonstrate that emamectin benzoate caused a significant increase in lactate dehydrogenase activity and the level of lactate indicating organ damage in treated animals. Piracetam, the so-called ‘nootropic’ drug is mainly used to treat cognitive impairment in aging, brain injuries, as well as dementia. In the present study piracetam treatment caused a significant inhibition of lactate dehydrogenase activity in liver, kidney, testis, heart, cerebral cortex, cerebellum and muscle of rats. The amelioration of toxic effects by piracetam treatment in EMB treated rats indicate that this drug has the potential to mitigate emamectin benzoate induced toxicity in mammalian systems.

Targeting Cancer Metabolism Breaks Radioresistance by Impairing the Stress Response.

Simple SummaryIonizing radiation is a major pillar in the therapy of solid tumors. However, normal tissue toxicities and radioresistance of tumor cells can limit the therapeutic success of radiotherapy. In this study, we investigated the coregulation of the cancer metabolism and the heat shock response with respect to radioresistance. Our results indicate that an inhibition of lactate dehydrogenase, either pharmacologically or by gene knockout of LDHA and LDHB, significantly increases the radiosensitivity in tumor cells by global impairing of the stress response. Therefore, inhibition of the lactate metabolism might provide a promising strategy in the future to improve the clinical outcome of patients with highly aggressive, therapy-resistant tumors.The heightened energetic demand increases lactate dehydrogenase (LDH) activity, the corresponding oncometabolite lactate, expression of heat shock proteins (HSPs) and thereby promotes therapy resistance in many malignant tumor cell types. Therefore, we assessed the coregulation of LDH and the heat shock response with respect to radiation resistance in different tumor cells (B16F10 murine melanoma and LS174T human colorectal adenocarcinoma). The inhibition of LDH activity by oxamate or GNE-140, glucose deprivation and LDHA/B double knockout (LDH−/−) in B16F10 and LS174T cells significantly diminish tumor growth; ROS production and the cytosolic expression of different HSPs, including Hsp90, Hsp70 and Hsp27 concomitant with a reduction of heat shock factor 1 (HSF1)/pHSF1. An altered lipid metabolism mediated by a LDHA/B double knockout results in a decreased presence of the Hsp70-anchoring glycosphingolipid Gb3 on the cell surface of tumor cells, which, in turn, reduces the membrane Hsp70 density and increases the extracellular Hsp70 levels. Vice versa, elevated extracellular lactate/pyruvate concentrations increase the membrane Hsp70 expression in wildtype tumor cells. Functionally, an inhibition of LDH causes a generalized reduction of cytosolic and membrane-bound HSPs in tumor cells and significantly increases the radiosensitivity, which is associated with a G2/M arrest. We demonstrate that targeting of the lactate/pyruvate metabolism breaks the radioresistance by impairing the stress response.

Synthesis and initial screening of lactate dehydrogenase inhibitor activity of 1,3-benzodioxole derivatives

Cancer is one of the main causes of mortality in the world. Many cancer cells produce ATP through high-level lactic acid fermentation catalyzed by lactate dehydrogenase (LDH), which converts pyruvic acid to lactic acid. LDH plays a dominant role in the Warburg effect, wherein aerobic glycolysis is favored over oxidative phosphorylation. Due to the high lactic acid production level in cancer cells, LDH-targeting could be a potential cancer treatment strategy. A few approaches, such as drug treatment, reportedly inhibited LDH activity. In this study, we describe new 1,3-benzodioxole derivatives that might be potential small molecule candidates for LDHA inhibition. The synthesis was carried out by trans-esterification between aryl ester and alcohol groups from piperonyl alcohol. Compounds 2 and 10 exhibited a selective LDHA IC50 value of 13.63 µM and 47.2 µM, respectively. Whereas only compound 10 showed significant cytotoxicity in several lines of cancer cells, especially in human pancreatic cancer PANC-1 cells. These synthesized compounds possess 2 aromatic rings and –CF3 moiety, which expectedly contributes to LDHA inhibition. The presented products have the potential to become a promising LDHA inhibitor drug candidate.

Melatonin protects against myocardial ischemia-reperfusion injury by inhibiting contracture in isolated rat hearts

To investigate the protective effect of melatonin against myocardial ischemia reperfusion (IR) injury in isolated rat hearts and explore the underlying mechanisms. The isolated hearts from 40 male SD rats were randomly divided into 4 groups (n=10): the control group, where the hearts were perfused with KH solution for 175 min; IR group, where the hearts were subjected to global ischemia for 45 min followed by reperfusion for 120 min; IR+melatonin (Mel+IR) group, where melatonin (5 μmol/L) was administered to the hearts 1 min before ischemia and during the first 5 min of reperfusion, followed by 115 min of reperfusion; and IR+2, 3-butanedione monoxime (IR+BDM) group, where the hearts were treated with BDM (20 mmol/L) in the same manner as melatonin treatment. Myocardial injury in the isolated hearts was assessed based on myocardial injury area, caspase-3 activity, and expressions of cytochrome C and cleaved caspase-3 proteins. Cardiac contracture was assessed using HE staining and by detecting lactate dehydrogenase (LDH) activity and the content of cardiac troponin I (cTnI) in the coronary outflow, measurement of left ventricular end-diastolic pressure (LVEDP) and electron microscopy. The content of ATP in the cardiac tissue was also determined. Compared with those in the control group, the isolated hearts in IR group showed significantly larger myocardial injury area and higher caspase-3 activity and the protein expressions of cytochrome C and cleaved caspase-3 with significantly increased LDH activity and cTnI content in the coronary outflow and elevated LVEDP at the end of reperfusion; HE staining showed obvious fractures of the myocardial fibers and the content of ATP was significantly decreased in the cardiac tissue; electron microscopy revealed the development of contraction bands. In the isolated hearts with IR, treatment with Mel or BDM significantly reduced the myocardial injury area, caspase-3 activity, and protein expressions of cytochrome C and cleaved caspase-3, obviously inhibited LDH activity, lowered the content of cTnI and LVEDP, reduced myocardial fiber fracture, and increased ATP content in the cardiac tissue. Both Mel and BDM inhibited the formation of contraction bands in the isolated hearts with IR injury. Mel can alleviate myocardial IR injury in isolated rat hearts by inhibiting cardiac contracture, the mechanism of which may involve the upregulation of ATP in the cardiac myocytes to lessen the tear of membrane and reduce cell content leakage.

Integrated multi-techniques to probe the binding mechanism between amlodipine and lactate dehydrogenase

The interaction mechanism of amlodipine (AD) and lactate dehydrogenase (LDH) was examined using different spectroscopic and molecular docking methods. Spectroscopy results demonstrated that AD could bind with LDH, thus inhibiting its activity in a time-dependent and dose-dependent manner. Based on the thermodynamic parameters, it was concluded that the reaction between LDH and AD occurred spontaneously, where H-bonding and van der Waals force occupied vital roles. The association constant (K) was the order of magnitude of 105, the number of binding site (n) was approximately 1, and binding distance between AD and LDH was estimated as 3.5 nm. Synchronous fluorescence, circular dichroism, and FT-IR spectra indicated that AD led to conformational variations in LDH. The results from molecular docking found that AD was inserted into the hydrophobic cavity of LDH and interacted with residue His192, resulting in the inhibition of LDH activity. The results of Discovery Studio Visualizer highlighted the role of H-bonding in the binding process, in agreement with the solution experiments. This work would help in understanding the binding mode of LDH-AD system and would provide new insights into the effect of AD on human health.

Suppression of tumor growth and metastasis by ethanol extract of Angelica dahurica Radix in murine melanoma B16F10 cells.

The roots of Angelica dahurica have long been used as a traditional medicine in Korea to treat various diseases such as toothache and cold. In this study, we investigated the effect of ethanol extract from the roots of this plant on metastatic melanoma, a highly aggressive skin cancer, in B16F10 melanoma cells and B16F10 cell inoculated-C57BL/6 mice. Our results showed that the ethanol extracts of Angelicae dahuricae Radix (EEAD) suppressed cell growth and induced apoptotic cell death in B16F10 cells. EEAD also activated the mitochondria-mediated intrinsic apoptosis pathway, with decreased mitochondrial membrane potential, and increased production of intracellular reactive oxygen species and ration of Bax/Bcl-2 expression. Furthermore, EEAD reduced the migration, invasion, and colony formation of B16F10 cells through the reduced expression and activity of matrix metalloproteinase (MMP)-2 and -9. In addition, in vivo results demonstrated that oral administration of EEAD inhibited lactate dehydrogenase activity, hepatotoxicity, and nephrotoxicity without weight loss in B16F10 cell inoculated-mice. Importantly, EEAD was able to markedly suppress lung hypertrophy, the incidence of B16F10 cells lung metastasis, and the expression of tumor necrosis factor-alpha in lung tissue. Taken together, our findings suggest that EEAD may be useful for managing metastasis and growth of malignant cancers, including melanoma.

Enzymatic and thermodynamic profiles of a heterotetramer lactate dehydrogenase isozyme in swine

Lactate dehydrogenase (LDH) is a glycolytic enzyme that catalyzes the final step of glycolysis and produces NAD+. In somatic cells, LDH forms homotetramers and heterotetramers that are encoded by two different genes: LDHA (skeletal muscle type, M) and LDHB (heart type, H). Analysis of LDH isozymes is important for understanding the physiological role of homotetramers and heterotetramers and for optimizing inhibition of their enzymatic activity as it may result in distinct effects. Previously, we reported that hydroxychloroquine (HCQ) inhibited LDH activity, but we did not examine isozyme specificity. In the present study, we isolated heterotetrameric LDH (H2M2) from swine brain, determined its kinetic and thermodynamic properties, and examined the effect of HCQ on its activity compared to homotetrameric LDH isozymes. We show that: (1) the Km values for H2M2–mediated catalysis of pyruvate or lactate were intermediate compared to those for the homotetrameric isozymes, M4 and H4 whereas the Vmax values were similar; (2) the Km and Vmax values for H2M2–mediated catalysis of NADH were not significantly different among LDH isozymes; (3) the values for activation energy and van't Hoff enthalpy changes for pyruvate reduction of H2M2 were intermediate compared to those for the homotetrameric isozymes; (4) the temperature for half residual activity of H2M2 was closer to that for M4 than for H4. We also show that HCQ had different affinities for various LDH isozymes.

Myristica fragrans Suppresses Tumor Growth and Metabolism by Inhibiting Lactate Dehydrogenase A.

Most cancer cells predominantly produce ATP by maintaining a high rate of lactate fermentation, rather than by maintaining a comparatively low rate of tricarboxylic acidcycle, i.e., Warburg's effect. In the pathway, the pyruvate produced by glycolysis is converted to lactic acid by lactate dehydrogenase (LDH). Here, we demonstrated that water extracts from the seeds of Myristica fragrans Houtt. (MF) inhibit the in vitro enzymatic activity of LDH. MF effectively suppressed cell growth and the overall Warburg effect in HT29 human colon cancer cells. Although the expression of LDH-A was not changed by MF, both lactate production and LDH activity were decreased in MF-treated cells under both normoxic and hypoxic conditions. In addition, intracellular ATP levels were also decreased by MF treatment, and the uptake of glucose was also reduced by MF treatment. Furthermore, the experiment on tumor growth in the in vivo mice model revealed that MF effectively reduced the growth of allotransplanted Lewis lung carcinoma cells. Taken together, these results suggest that MF effectively inhibits cancer growth and metabolism by inhibiting the activity of LDH, a major enzyme responsible for regulating cancer metabolism. These results implicate MF as a potential candidate for development into a novel drug against cancer through inhibition of LDH activity.

Inhibition of key markers linked with spermatogenesis and cellular ATP by subchronic exposure to leachate in a rat model.

Few or no studies have measured the toxic effects of subchronic exposure to leachate using key markers linked with spermatogenesis and cellular adenosine triphosphate (ATP) in an experimental rat model. This study was undertaken to evaluate the toxic effects of leachate obtained from the Elewi Odo municipal battery-recycling site (EOMABRL) on male reproductive function using testicular hormones and biomarker of cellular ATP. EOMABRL was administered at 0, 20, 40, 60, 80, and 100% concentrations to adult male rats for 60 days. After exposure, serum was collected for hormonal biochemistry assays, and testes were collected to determine the activity of xanthine oxidase (XO) and lactate dehydrogenase (LDH). Exposure of animals to EOMABRL resulted in a 519.7, 285.7, 569.1, 606.1, and 1,793.2% increase in XO activity with a sequential decrease in LDH activity (marker of cellular ATP) by 44.1, 55.9, 61.4, 69.3, and 89.7%, respectively, compared with the control. Furthermore, EOMABRL caused a significant inhibitory effect on serum luteinizing hormone, follicle-stimulating hormone, and testosterone levels. We conclude that some possible mechanisms by which EOMABRL elicits toxicity in male rat testes could be through inhibition of LDH activity and depletion of serum hormone levels.

Inhibition of Panton-Valentine Leukocidin Toxin Induced Neutrophil Cell Lysis by Vancoplus in Methicillin-Resistant <i>Staphylococcus Aureus</i> Infections

Pantone Valentine Leukocidin (PVL) is a cytotoxin associated with virulence of Methicillin-Resistant Staphylococcus aureus (MRSA). This study was undertaken to analyze the prevalence of PVL among MRSA strains and to study the effect of adjuvants (L-lysine, L-arginine, Ethylenediamine Tetraacetic Acid (EDTA) as well as drugs on PVL induced neutrophil cell lysis. Further, in vitro activity of Vancoplus, vancomycin, linezolid, teicoplanin and daptomycin against PVL positive MRSA strains was analyzed. A total of 67 Staphylococcus aureus strains collected were screened for the prevalence of methicillin resistance and PVL by Polymerase Chain Reaction (PCR). Effects of various adjuvants and drugs on PVL induced neutrophil cell lysis were studied by Lactate Dehydrogenase (LDH) assay. Antibiotic susceptibility was conducted using Clinical and Laboratory Standards Institute (CLSI) guidelines. Our results showed that of 67 clinical isolates, 71.6% (48/67) isolates were confirmed to be MRSA. Of these MRSA strains, 75% (36/48) isolates were observed to be PVL positive. Pus swab (86.3%; 19/22) had more proportion of PVL than urine (75%; 6/8); sputum (70%; 7/10) and blood (50%; 4/4). When diverse adjuvants alone (L-lysine, L-argine and EDTA) or in combinations were tested on PVL induced neutrophil cell lysis, 10 mM L-lysine alone and combinations of 10 mM L-lysine +10 mM EDTA and 100 mM L-arginine +10 mM EDTA produced the highest 78.7±7.1, 92.6±8.8 and 93.4±6.3%, respectively, inhibition of LDH activity (known marker of cell lysis). Further, when Vancoplus reconstituted with solvent produced 93.1±6.4% inhibition of LDH activity whereas other comparator drugs produced only 12.7±2.4 to 30.5±3.1% inhibition of LDH activity. Vancoplus appeared to be the most efficacious against 94.4% of the MRSA strains with MIC values 0.25 to 4 µg mL-1. The susceptibility of other drugs vancomycin, linezolid, teicoplanin and daptomycin against MRSA varied from 22.3 to 77.8% of isolates with MICs 0.5 to 32 µg mL-1. This study provides new insight into the prevalence of PVL toxins among MRSA. Results of this study showed that Vancoplus is the most active against MRSA and it additionally protects the neutrophil cells from PVL toxins and prevents from sequels of MRSA infections caused by these cytotoxins.

Inhibition of Lactate Dehydrogenase Activity as an Approach to Cancer Therapy

In the attempt of developing innovative anticancer treatments, growing interest has recently focused on the peculiar metabolic properties of cancer cells. In this context, LDH, which converts pyruvate to lactate at the end of glycolysis, is emerging as one of the most interesting molecular targets for the development of new inhibitors. In fact, because LDH activity is not needed for pyruvate metabolism through the TCA cycle, inhibitors of this enzyme should spare glucose metabolism of normal non-proliferating cells, which usually completely degrade the glucose molecule to CO2. This review is aimed at summarizing the available data on LDH biology in normal and neoplastic cells, which support the anticancer therapeutic approach based on LDH inhibition. These data encouraged pharmaceutical industries and academic institutions in the search of small-molecule inhibitors and promising candidates have recently been identified. The availability of inhibitors with drug-like properties will allow the evaluation in the near future of the real potential of LDH inhibition in anticancer treatment, also making the identification of the most responsive neoplastic conditions possible.

Synergistic anti-cancer effect of phenformin and oxamate.

Phenformin (phenethylbiguanide; an anti-diabetic agent) plus oxamate [lactate dehydrogenase (LDH) inhibitor] was tested as a potential anti-cancer therapeutic combination. In in vitro studies, phenformin was more potent than metformin, another biguanide, recently recognized to have anti-cancer effects, in promoting cancer cell death in the range of 25 times to 15 million times in various cancer cell lines. The anti-cancer effect of phenformin was related to complex I inhibition in the mitochondria and subsequent overproduction of reactive oxygen species (ROS). Addition of oxamate inhibited LDH activity and lactate production by cells, which is a major side effect of biguanides, and induced more rapid cancer cell death by decreasing ATP production and accelerating ROS production. Phenformin plus oxamate was more effective than phenformin combined with LDH knockdown. In a syngeneic mouse model, phenformin with oxamate increased tumor apoptosis, reduced tumor size and 18F-fluorodeoxyglucose (FDG) uptake on positron emission tomography/computed tomography compared to control. We conclude that phenformin is more cytotoxic towards cancer cells than metformin. Furthermore, phenformin and oxamate have synergistic anti-cancer effects through simultaneous inhibition of complex I in the mitochondria and LDH in the cytosol, respectively.

From sub cellular to community level: Toxicity of glutaraldehyde to several aquatic organisms

The biocide glutaraldehyde (GA) is widely used as a disinfectant and sterilizing agent against bacteria and virus in hospital and veterinary facilities. GA or its metabolites may reach aquatic ecosystems due to incomplete or inadequate treatment of wastewaters. Data about GA effects at lethal and a sub lethal level to non-target organisms is needed so that a risk assessment to aquatic ecosystems can be done. Thus, in this work a battery of toxicity tests with primary producers, primary consumers and secondary consumers were performed and a species sensitive distribution (SSD) for GA was built. Moreover, effects on biomarkers (catalase, lactate dehydrogenase, glutathione-S-transferase, and cholinesterase) were measured in Danio rerio embryos and adults. Primary consumers (Thamnocephalus platyurus 24h — EC50=3.6mg/l; Daphnia magna 48h — EC50=6.6mg/l) and D. rerio adults (96h — LC50=5.5mg/l) were slightly more sensitive to GA than D. rerio embryos (96h — LC50=22.2mg/l) and primary producers (Lemna minor 168h — EC50=73.8mg/l; Pseudokirchneriella subcapitata 72h — EC50=12.3mg/l; Chlamydomonas reinhardtii 72h — EC50=14.6mg/l; Chlorella vulgaris 72h — EC50=31.3mg/l). However, no significant differences between the trophic levels were found and general HC5 and HC50 values of 0.6 and 11.4mg/l were respectively estimated. Despite the low GA toxicity to D. rerio embryos, hatching delay and malformations were found (96h — EC50=11.9mg/l). For biomarkers, an inhibition of lactate dehydrogenase activity was observed in embryos whereas an inhibition in catalase, lactate dehydrogenase and glutathione-S-transferase activities was observed in adults. Thus, GA is moderately toxic (doses>1mg/l) to aquatic organisms, independently of the trophic level. However, considering the varied range of effects depending on the life stage and organism tested and relatively low HC5 value of 0.6mg/l, mesocosm and chronic toxicity tests seem to be the next step in direction of more realistic scenarios of GA risk assessment in aquatic ecosystems.

The protective effect of picroside II against hypoxia/reoxygenation injury in neonatal rat cardiomyocytes

Context: Picroside II, an iridoid glucoside found in the root of Picrorhiza scrophulariiflora Pennell (Scrophulariaceae), has been demonstrated to possess potent antioxidant activity. However, whether picroside II has a protective effect against hypoxia/reoxygenation (H/R)-induced cardiomyocyte injury is poorly understood.Objective: To explore the cardioprotective role of picroside II against oxidative stress induced by H/R injury in neonatal rat cardiacmyocytes.Materials and methods: The viability and cellular damage of cardiomyocytes were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolim bromide (MTT) and lactate dehydrogenase (LDH) assays, respectively. The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), the levels of reduced (GSH) and oxidized glutathione (GSSG), and the contents of malondialdehyde (MDA) were determined by a colorimetric method. The levels of intracellular reactive oxygen species (ROS) and calcium were evaluated by flow cytometric analysis.Results: We analyzed the effective half-maximal concentration for protection from the dose-response curves and obtained the concentration of 50 µg/mL as EC50. Pretreated cardiomyocytes with picroside II (50–200 µg/mL), prior to H/R exposure, inhibited LDH activity in culture media and increased cell viability in a dose-dependent manner. This protective effect was accompanied by significantly increasing reduced GSH contents and the activities of SOD and GSH-Px and attenuating MDA and GSSG contents in response to H/R injury. Furthermore, treatment with picroside II also inhibited ROS production and calcium accumulation in cardiomyocytes.Discussion and conclusion: The present study demonstrates that picroside II protects cardiomyocytes against oxidative-stress injury induced by H/R through reduction of ROS production and calcium accumulation and enhancement of the activity of antioxidant defense.

Effects of the antibacterial monomer 12-methacryloyloxydodecylpyridinium bromide (MDPB) on bacterial viability and metabolism

The antibacterial monomer 12-methacryloyloxydodecylpyridinium bromide (MDPB) is a strong bactericide when unpolymerized and has the potential to be utilized in various resinous biomaterials. To analyze the antibacterial characteristics of this monomer in detail, the ability of high concentrations of unpolymerized MDPB to kill Streptococcus mutans in planktonic or biofilm forms within a short time-period of contact, and the inhibitory effects of low concentrations of MDPB on the metabolic function of S. mutans, were examined. High concentrations of MDPB showed effective killing of planktonic and biofilm S. mutans cells within 60 s, and complete killing was obtained by contact with 1,000 μg ml(-1) of MDPB for 60 s. At a concentration of 4-8 μg ml(-1) , MDPB demonstrated growth inhibition, inducing elongation of the lag phase and of the doubling time, when the bacterial number was low. Inhibition of the production of acid from S. mutans by 8 μg ml(-1) of MDPB may have been caused by the inhibition of lactate dehydrogenase activity. At high concentrations, MDPB is lethal to both planktonic and biofilm forms of S. mutans in a short time-period, and at low concentrations, MDPB inhibits metabolic enzymatic activity.

Electrochemical behavior of lactate dehydrogenase immobilized on “silica sol–gel/nanometre-sized tridecameric aluminium polycation” modified gold electrode and its application

This paper reports the electrochemical behavior of lactate dehydrogenase (LDH) immobilized in the silica sol-gel film on gold electrode after adding nanometre-sized tridecameric aluminium polycation (nano-Al13, also called nanopolynuclear Al13) as a promoter. A pair of surface controlled quasi-reversible cyclic voltammetry peaks with the formal potential (E0') of 154 mV (vs. SCE) was found in the presence of nano-Al13. A potential application of the nano-Al13-LDH electrode for the determination of resorcinol and p-xylene was also investigated. The experimental results showed that both resorcinol and p-xylene inhibited LDH activity, and the calibration ranges were 5.0 x 10(-6)-3.0 x 10(-4) mol L(-1) for resorcinol and 1.0 x 10(-6)-1.0 x 10(-5) mol L(-1) for p-xylene, respectively. The nano-Al13-LDH electrode can be anticipated to be applied to environmental toxic assessments.