Specific Activity Of Lactate Dehydrogenase Research Articles

Toxicological assessment of crude oil vapour and the modulating effect of vitamin E on the heart of albino rats

This study investigates the effect of vapour of various concentrations of crude oil and oral co-administration of vitamin E on the heart of albino rats. Thirty (30) albino rats were randomly grouped into six (6) of five (5) rats each and designated A to E. Rats neither exposed to crude oil vapour nor oral administration of vitamin E, rats exposed to oral administration of 15 mg/kg bw vitamin E but not exposed to vapour of crude oil were designated groups A and B. Rats exposed to oral co-administration of vitamin E and vapour of 25%, 50%, 75% and 100% crude oil were designated groups C, D, E and F respectively. Rats’ treatment was over a period of 28 days, after which the rats were euthanized for biochemical analyses. Serum total protein (TP), total cholesterol (CHOL) and triglycerides (TRIG) levels were determined. Standard enzyme assay was conducted for alanine transaminase (ALT), aspartate transaminase (AST), gamma glutamyl transferase and lactate dehydrogenase (LDH) of the heart. Generally, the concentration of TP of serum of other groups of rats, except group B, was significantly higher relative to the Control (p≤0.05). Serum CHOL and TRIG levels increased as the concentration of crude oil increased. Heart AST activities of rats in groups A, B and C were not significantly different (p≤0.05), similarly the AST activities of heart of rats in groups D, E and F were not significantly different (p>0.05). There was no significant difference between the gamma glutamyl transferase (GGT) activity of Control rats not exposed to crude oil vapour and not given oral administration of vitamin E (A) and the Control in Group B (given oral administration of vitamin E but not exposed to crude oil vapour). The specific activity of lactate dehydrogenase (LDH) of heart of both control rats and those of rats exposed to the vapour of various concentrations of crude oil were not significantly different (p>0.05). Experimental evidence from this study revealed that vapour of crude oil can induce biochemical changes in the heart of rats while oral co-administration of vitamin E could reduce the risk posed by exposure to vapour of crude oil by 50%.

Increasing of LDH Specific Activity and PEPCK Level Play a Role on Activation of Gluconeogenesis Pathway in Early Onset Pre-Eeclampsia Placenta.

Recent advancement on experiment concluded that etiology of pre-eclampsia (PE) could be explained by the "two-stage" theory. The theory of which explained that pre-eclampsia occurs due to abnormalities in spiral arteries development and release of inflammatory response. Failure of spiral arteries development, the lesion of damage could be due to ischemia-reperfusion or hypoxia-reoxygenation. Hypoxia in pre-eclamptic placenta leads to metabolic change to anaerobic in glycolysis. Lactate dehydrogenase (LDH) has important role in anaerobic glycolysis that catalyzes the conversion of lactate to pyruvate during hypoxia. On the other hand, phosphoenolpyruvate carboxy kinase (PEPCK) is merely an enzyme of gluconeogenesis. This research conduct to reveal that in early onset pre-eclampsia the placenta still hypoxic and undergoes gluconeogenesis even after delivery, through metabolic enzyme of LDH and PEPCK level. This cross-sectional study compared early onset PE (< 34 weeks) with normal term placenta. We measured LDH enzyme activity using colorimetric assay and PEPCK protein using ELISA method. Result show that placental LDH specific activity was increased significantly in PE with median 2.750 (0.030 - 5.680) U/mg compared to normal term placenta 0.255 (0.032 - 1.194) U/mg (Mann-Whitney, p< 0.001). PEPCK level was significantly increased in PE 8.998 (1.737-44.914) ng/mg compared to normal term placenta 1.552 (0.741-8.832) ng/mg (Mann-Whitney, p< 0.001). We conclude that anaerobic glycolysis and gluconeogenesis pathway are increased in early onset PE placenta as adaptation mechanism to hypoxic condition.

Cytoprotective effect of taurine against sodium chlorate-induced oxidative damage in human red blood cells: an ex vivo study.

Sodium chlorate (NaClO3) is a common non-selective herbicide that is also used in paper and pulp mills and is produced as a by-product during drinking water disinfection by chlorine dioxide. Here, we report the effect of dietary antioxidant taurine on NaClO3-induced cytotoxicity in human red blood cells (RBC). RBC were treated with 5mM NaClO3, either alone or in presence of 1, 2.5 and 5.0mM taurine. Incubation of RBC with NaClO3 alone caused hemolysis, increased oxidation of lipids and proteins, methemogobin level and decreased total sulfhydryl and glutathione content. It lowered the activities of antioxidant enzymes thioredoxin reductase, glutathione peroxidase, catalase and glutathione reductase, while Cu-Zn superoxide dismutase activity was increased. The antioxidant capacity of RBC was impaired. This strongly suggests that NaClO3 causes the induction of oxidative stress condition in RBC. The specific activities of lactate dehydrogenase, glucose 6-phosphate dehydrogenase and plasma membrane bound enzymes, were also greatly altered. However, prior treatment of RBC with taurine conferred significant protection against NaClO3-induced oxidative damage and also improved the antioxidant defence system of cells. These results were supported by electron microscopy images of RBC. Treatment with NaClO3 alone converted the normal biconcave discoidal RBC to acanthocytes and echinocytes but this transformation was greatly prevented in the presence of taurine. Thus, taurine mitigates the cytotoxicity of NaClO3 in human RBC and can function as an effective chemoprotectant.

Metabolic Regulation of Organic Acid Biosynthesis in Actinobacillus succinogenes.

Actinobacillus succinogenes is one of the most promising strains for succinic acid production; however, the lack of efficient genetic tools for strain modification development hinders its further application. In this study, a markerless knockout method for A. succinogenes using in-frame deletion was first developed. The resulting ΔpflA (encode pyruvate formate lyase 1-activating protein) strain displayed distinctive organic acid synthesis capacity under different cultivation modes. Additional acetate accumulation was observed in the ΔpflA strain relative to that of the wild type under aerobic conditions, indicating that acetate biosynthetic pathway was activated. Importantly, pyruvate was completely converted to lactate under anaerobic fermentation. The transcription analysis and enzyme assay revealed that the expression level and specific activity of lactate dehydrogenase (LDH) were significantly increased. In addition, the mRNA expression level of ldh was nearly increased 85-fold compared to that of the wild-type strain during aerobic–anaerobic dual-phase fermentation, resulting in 43.05 g/L lactate. These results demonstrate that pflA plays an important role in the regulation of C3 flux distribution. The deletion of pflA leads to the improvement of acetic acid production under aerobic conditions and activates lactic acid biosynthesis under anaerobic conditions. This study will help elaborate the mechanism governing organic acid biosynthesis in A. succinogenes.

Identification and application of novel low pH-inducible promoters for lactic acid production in the tolerant yeast Candida glycerinogenes

Bioproduction of organic acids under low pH condition without adding inducer and neutralizer is an economical craft to decrease downstream cost. Candida glycerinogenes had higher tolerances to low pH and lactic acid than Saccharomyces cerevisiae. QRT-PCR analysis showed that four of fifteen candidates functioned as potentially acid-inducible promoters in C. glycerinogenes. In particular, PCggmt1 showed the strongest induction ability at pH 2.5. Fluorescence analysis indicated that the induction ability of PCggmt1 gradually increased as the pH decreased. In addition, PCggmt1 had the binding sites of Msn2p/4p, Azf1p, and Nrg1p. PCggmt1 was further used to control the expression of lactate dehydrogenase gene from Rhizopus oryzae in C.glycerinogenes. Compared with pH 5.5, the specific activity of lactate dehydrogenase and lactic acid titer at pH 2.5 were increased by 229% and 218%, reaching 13.8mU/mg and 12.3g/L, respectively. These results presented here showed a potential to produce organic acid economically at low pH by the stress tolerant C.glycerinogenes and the novel low-pH inducible promoter PCggmt1.

Expression of Lactate Dehydrogenase in Aspergillus niger for L-Lactic Acid Production.

Different engineered organisms have been used to produce L-lactate. Poor yields of lactate at low pH and expensive downstream processing remain as bottlenecks. Aspergillus niger is a prolific citrate producer and a remarkably acid tolerant fungus. Neither a functional lactate dehydrogenase (LDH) from nor lactate production by A. niger is reported. Its genome was also investigated for the presence of a functional ldh. The endogenous A. niger citrate synthase promoter relevant to A. niger acidogenic metabolism was employed to drive constitutive expression of mouse lactate dehydrogenase (mldhA). An appraisal of different branches of the A. niger pyruvate node guided the choice of mldhA for heterologous expression. A high copy number transformant C12 strain, displaying highest LDH specific activity, was analyzed under different growth conditions. The C12 strain produced 7.7 g/l of extracellular L-lactate from 60 g/l of glucose, in non-neutralizing minimal media. Significantly, lactate and citrate accumulated under two different growth conditions. Already an established acidogenic platform, A. niger now promises to be a valuable host for lactate production.

Cognitive-enhancing and antioxidant activities of inhaled coriander volatile oil in amyloid β(1–42) rat model of Alzheimer's disease

Coriandrum sativum L., commonly known as coriander and belonging to the Apiaceae family is cultivated throughout the world for its nutritional value. In traditional medicine, coriander is recommended for the relief of pain, anxiety, flatulence, loss of appetite and convulsions. In the present study, the effects of inhaled coriander volatile oil (1% and 3%, daily, for 21days) extracted from C. sativum var. microcarpum on spatial memory performance were assessed in an Aβ(1–42) rat model of Alzheimer's disease. The Aβ(1–42)-treated rats exhibited the following: decrease of spontaneous alternations percentage within Y-maze task and increase of working memory errors, reference memory errors and time taken to consume all five baits within radial arm maze task. Exposure to coriander volatile oil significantly improved these parameters, suggesting positive effects on spatial memory formation. Assessments of oxidative stress markers in the hippocampal tissue of Aβ(1–42)-treated rats showed a significant increase of superoxide dismutase (SOD), lactate dehydrogenase (LDH) and a decrease of glutathione peroxidase (GPX) specific activities along with an elevation of malondialdehyde (MDA) level. Coriander volatile oil significantly decreased SOD and LDH specific activities, increased GPX specific activity and attenuated the increased MDA level. Also, DNA cleavage patterns were absent in the coriander rats, thus suggesting antiapoptotic activity of the volatile oil. Therefore, our results suggest that exposure to coriander volatile oil ameliorates Aβ(1–42)-induced spatial memory impairment by attenuation of the oxidative stress in the rat hippocampus.

Production of 2,3-butanediol by a low-acid producing Klebsiella oxytoca NBRF4

2,3-Butanediol (2,3-BDO) is a value-added chemical with great potential for the industrial production of synthetic rubber, plastic and solvent. For microbial production of 2,3-BDO, in this study, Klebsiella oxytoca NBRF4 was constructed by chemical mutation and screening against NaBr, NaBrO(3) and fluoroacetate. Among metabolic enzymes involved in the production of lactate, acetate and 2,3-BDO, K. oxytoca NBRF4 possessed 1.2 times lower specific activities of lactate dehydrogenase and phosphotransacetylase, and 22% higher specific acetoin reductase activity than the K. oxytoca ATCC43863 control strain. A series of batch fermentations in a defined medium and application of a statistical tool of response surface method led to the determination of optimal culture conditions: 10% dissolved oxygen level, pH 4.3 and 38°C. The actual results of batch fermentation at the optimal conditions using 44 g/L glucose were coincident with the predetermined values: 14.4 g/L 2,3-BDO concentration, 0.32 g/g yield. To increase 2,3-BDO titer, fed-batch fermentation of K. oxytoca NBRF4 was performed by an intermittent feeding of 800 g/L glucose to control its concentration around 5-20 g/L in the culture broth. Finally, 34.2g/L 2,3-BDO concentration and 0.35 g/g yield were obtained without organic acid production in 70 hours of the fed-batch culture, which were 2.4 and 1.2 times higher than those of the batch fermentation using 44 g/L glucose.

Analysis of lactate dehydrogenase activities and isoenzyme patterns in colorectal cancer tissues.

To investigate the relationship between lactate dehydrogenase (LDH) activity or LDH isoenzyme patterns and the pathogenesis of colorectal cancer. Activities of tissue LDH and LDH isoenzyme patterns in 16 patients with colorectal cancer were assayed using spectrophotometric procedures and agarose gel electrophoresis, respectively. The total and specific activities of LDH were significantly higher in colorectal cancer tissues than those in adjacent noncancerous tissues (P < 0.001). The LDH isoenzyme pattern was also different from that in the control. The percentage of LDH5 doubled and the ratio of LDH4 + LDH5/LDH1 + LDH2 was 3.6 ± 1.4 in cancer tissue, significantly greater than in the control. The increased LDH activity in colorectal cancer tissues resulted mainly from the increased LDH5, suggesting that the alteration of LDH activity and isoenzyme patterns were related to the pathogenesis of colorectal cancer.

Ecophysiological category based toxicological responses in metabolism of earthworms: Impact of a pyrethroidal insecticide

Effects of alphamethrin (0.04 ppm, 0.08 ppm, 0.16 ppm, 0.32 ppm) on some metabolic dehydrogenases and proteins for 1, 2, 3, 4, 8 and 16 days using three ecologically different earthworm species ( Perionyx sansibaricus, Lampito mauritii and Metaphire posthuma) were studied. The first significant effect was on 2nd/3rd day and maximum response was achieved on 16th day of exposure of alphamethrin at all concentrations. Similarly, maximum effect was obtained at 0.32 ppm alphamethrin at different exposure periods. It showed a dose- and duration-dependent inhibitory effects of a pyrethroid on enzymes and proteins of earthworms. There were approximately 47%, 43% and 41% declines in specific activities of cytoplasmic malate dehydrogenase (cMDH) of P. sansibaricus, L. mauritii and M. posthuma, respectively, in response to 0.32 ppm exposure of alphamethrin for 16 days. In case of similar treatment, specific activities of mitochondrial malate dehydrogenase (mMDH) showed 57%, 51% and 45% reductions in epigeic ( P. sansibaricus), anecic ( L. mauritii) and endogeic ( M. posthuma) earthworms, respectively. Exposure of alphamethrin (0.32 ppm) also decreased the specific activities of lactate dehydrogenase (LDH) by 47% ( P. sansibaricus), 35% ( L. mauritii) and 39% ( M. posthuma) in different types of earthworms. The reductions in specific activities of a Krebs cycle enzyme (mMDH) were greater than that of a glycolytic enzyme (LDH). The protein contents declined significantly in the earthworms exposed to alphamethrin. The decreases of 41%, 36% and 30% were obtained in cytoplasmic proteins of P. sansibaricus, L. mauritii and M. posthuma, respectively. However, the mitochondrial proteins showed reductions of 45% in P. sansibaricus, 46% in L. mauritii and 38% in M. posthuma in response to alphamethrin (0.32 ppm) intoxication for 16 days. The decrease in the protein content reflected an inhibitory effect of alphamethrin on protein turnover. The most pronounced effect of alphamethrin was on metabolic enzymes and proteins of P. sansibaricus and least effect was found on M. posthuma. This clearly showed that surface dwelling species of earthworms (epigeic) are more vulnerable to toxic chemical than deep burrowing species (endogeic). Thus a possible ecophysiological link exists between the toxicological responses and ecological categories at metabolic level in tropical earthworms.

Abstract A64: Chemopreventive potential of Bauhinia variegata against DMBA-induced skin papillomagenesis in Swiss albino mice

Abstract The present study is an attempt to evaluate the cancer chemopreventive potential of Bauhinia variegata bark against DMBA-induced skin papillomagenesis in mice. We investigated chemopreventive effect of three different doses (2%, 4% and 6% w/w) of test diets of bark of B. variegata during DMBA-induced skin papillomagenesis in mice and tumor cell proliferation and apoptosis. Its effect was also studied in a short-term animal protocol on carcinogen metabolizing enzymes in liver. DMBA and croton oil was used to induce and promote skin papillomagenesis, respectively. Skin papillomagenesis study was done by using three different doses of B. variegata before, during and after the DMBA treatment to mice till termination of the experiment (120 days). In addition, the expression of proliferation [proliferating cell nuclear antigen (PCNA), glutathione-S-transferse (GST-π) and apoptosis were analyzed in the skin tumors by immunohistochemistry. For biochemical studies, mice were given with the same doses of B. variegata in the test diet for two weeks. The specific activities/levels of phase-I and phase-II enzymes, antioxidant enzymes, lactate dehydrogenase, glutathione content, and peroxidative damage were determined in the liver of the mice. Results showed a significant reduction in the skin tumors in B. variegata treated animals, as compared to the control group of animals. The tumor incidence in case of control group of animals was 100% whereas it was 90%, 70% and 99% in groups treated with 2%, 4% and 6% test diets, respectively. Tumor multiplicity in control group was 12±3.9 whereas it was reduced to 8±3.7, 3±2.9 and 4±2.4 by the lower, intermediate and higher dose of B. variegata treatment. The dietary administration of three different doses of B. variegata significantly decreased the expression of PCNA, GST-π, Bcl-2 and VEGF and on the other hand significantly increased the expression of Bax and total Caspase 3 as compared to DMBA and croton oil treated group of animals. The effect of 2 weeks of dietary feeding of B. variegata inhibited phase I, activated phase II and antioxidant enzymes in the mouse liver. The content of reduced glutathione was significantly elevated whereas the peroxidative damage and specific activity of lactate dehydrogenase exhibited a significant reduction with all the doses of test diets, respectively. All these findings were indicative of that Bauhinia variegata has chemopreventive potential against DMBA-induced skin carcinogenesis. It would be of interest to investigate for active cancer chemopreventive agents in Bauhinia variegata for its future implication in cancer prevention and control. Citation Information: Cancer Prev Res 2010;3(1 Suppl):A64.

Prevention of chemically induced mammary tumorigenesis by daidzein in pre-pubertal rats: the role of peroxidative damage and antioxidative enzymes

Isoflavones are biologically active plant derived compounds that have several health promoting effects. In the present study hitherto unknown effects of one of the well known isoflavonoids, daidzein, has been evaluated on its chemo-preventive action against breast cancers in pre-pubertal rats. Either daidzein (500 mug/g bwt) or vehicle, dimethyl sulphoxide (DMSO), was administered at 16th, 18th, and 20th day post-partum and the chemopreventive efficacy was evaluated in dimethylbenz[a]nthracene (DMBA) induced Sprague-Dawley rats, at 50th day. To elucidate the mechanism of action, the antioxidative status was also examined in the liver and mammary gland of prebubertal rats using two different doses of daidzein (0.5 mg/kg bwt and 50 mg/kg bwt, p.o.) for 10 days. The specific activity of antioxidant enzymes as well as reduced glutathione (GSH) level and peroxidative damage were evaluated spectrophotometrically, both in liver as well as in mammary gland. Animals treated with daidzein pre-pubertally, showed a significant reduction in the tumorigenesis of mammary gland up to 37.4% as compared to animals induced for tumors with DMBA. In animals treated with 50 mg/kg of daidzein, a significant increase in the specific activities of the antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione transferase (GST), DT-diaphorase (DTD), and in GSH content were observed in both liver and mammary gland. Expectedly, the specific activity of lactate dehydrogenase (LDH) and level of peroxidative damage was decreased, as compared to that of control group of animals. Our results suggest that, daidzein can be considered as a potent chemopreventive agent against mammary carcinogenesis in pre-pubertal animals, with modulation of antioxidant enzymes being one of its mechanisms of actions.

Chemoprevention of mammary tumorigenesis and chemomodulation of the antioxidative enzymes and peroxidative damage in prepubertal Sprague Dawley rats by Biochanin A

Although chemopreventive action of Biochanin A against various cancers including that of prostate, breast, colon, and fore-stomach has been reported earlier, none of the studies was made in prepubertal subjects. The present study appears to be the first one on prepubertal rats that indicates the efficacy of the test compound in the prevention of tumorigenesis. The antioxidative status and xenobiotic metabolism were also evaluated to understand the mechanism of Biochanin A induced prevention of cancer. For the tumorigenesis study 500 microg/g bwt of Biochanin A or vehicle dimethyl sulfoxide (DMSO) s.c, was injected at 16th, 18th, and 20th days post-partum followed by the administration of dimethylbenz[a]nthracene (DMBA) (80 microg/g bwt) at 50th day. In another set of experiments, to study the involvement of peroxidative process in the mechanism of action of test compound, different antioxidant parameters were studied following the administration of two different doses of Biochanin A (0.5 and 50 mg/kg bwt, through oral gavage for 10 days) in the prepubertal rats from day 16 post-partum. Results showed a significant reduction in the mammary tumors (more than 40%) in Biochanin A treated animals, as compared to animals treated with DMBA only. Spectrophotometric enzyme estimations revealed that the specific activities of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione transferase (GST), DT-diaphorase (DTD), and reduced glutathione (GSH) levels were increased, whereas specific activities of lactate dehydrogenase (LDH) and lipid peroxidation (LPO) were decreased significantly, both in liver as well as in mammary gland, in animals treated with Biochanin A prepubertally. These results reveal the possible involvement of the antioxidative and metabolic enzymes in the suppression of cancer burden and incidence in a prepubertal rat model suggesting that the intake of this phytoestrogen at an early stage may help in lowering the risk of mammary tumor.

Improvement of L(+)-Lactic Acid Production of Rhizopus Oryzae by Low-Energy Ions and Analysis of Its Mechanism

The wild type strain Rhizopus oryzae PW352 was mutated by means of nitrogen ion implantation (15 keV, 7.8 × 1014 ∼ 2.08 × 1015 ions/cm2) to find an industrial strain with a higher L(+)-lactic acid yield, and two mutants RE3303 and RF9052 were isolated. In order to discuss the mechanism primarily, Lactate Dehydrogenase of Rhizopus oryzae was studied. While the two mutants produced L(+)-lactic acid by 75% more than the wild strain did, their specific activity of Lactate Dehydrogenase was found to be higher than that in the wild strain. The optimum temperature of Lactate Dehydrogenase in Rhizopus oryzae RF9052 was higher. Compared to the wild strain, the Michaelis constant (Km) value of Lactate Dehydrogenase in the mutants was changed. All these changes show that L(+)-lactic acid production has a correlation with the specific activity of Lactate Dehydrogenase. The low-energy ions, implanted into the strain, may improve the specific activity of Lactate Dehydrogenase by influencing its gene structure and protein structure.

Skeletal muscle expression of LDH and monocarboxylate transporters in growing rats submitted to protein malnutrition

In different circumstances such as infant malnutrition, old age or chronic disease, decline of muscular strength, particularly anaerobic power, is shown. In this context, our laboratory, has demonstrated a decrease in anaerobic glycolytic power in pre-pubertal Bolivian children living at low and high altitude and suffering from marginal protein malnutrition. To bring molecular support to the relationship between protein malnutrition and anaerobic glycolytic metabolism, we studied the impact of prolonged protein malnutrition on lactate metabolism in different muscles of growing rats. Lactate dehydrogenase (LDH), monocarboxylate transporters (MCT1, MCT4) and membrane protein CD147 were chosen as specific markers of anaerobic glycolytic metabolism. Two groups of 10 weaning male rats were fed for 10 weeks either ad libitum with a well-balanced diet containing 18% protein or an isocaloric-diet containing 8% protein. LDH activity and mRNA amounts of LDH isoforms, MCT, CD147 were measured. Protein deprivation during rat growth induced a decrease of LDH specific activity in skeletal muscles (mean value of -41%), accompanied by isoform distribution modifications in soleus, but not in glycolytic muscles (extensor digitorum longus (EDL) or plantaris). A reduction in mRNA amounts encoding the LDH A and B subunits was observed in EDL. A decrease in LDH B mRNA amounts was monitored in plantaris, whereas no modification in both LDH isoform mRNA quantities was observed in soleus. MCT1 mRNA quantities were decreased in EDL but MCT4 mRNA quantities remained stable. CD147 mRNA amounts were unchanged except for EDL with a 42% increase. The global decreases of LDH activity, LDH and MCT gene expressions in growing rat skeletal muscles support the observed alterations of lactate metabolism associated with lowered muscular anaerobic performances in protein malnutrition.

Differences in some properties of lactate dehydrogenase from muscles of the carp Cyprinus carpio and trout Salmo gairdneri

Some catalytic and kinetic properties of lactate dehydrogenase (LDH) isolated from trout and carp skeletal muscles were compared. The specific activity of LDH in the carp muscle was lower by about one third than the activity in the trout muscle. Temperature and pH optima for LDH isolated from the carp muscle were higher than those for the trout muscle LDH. Moreover, in direct reaction, the carp muscle LDH had a higher affinity both for pyruvate and for NADH, i.e., it had lower KM values. Instead, the trout muscle LDH showed the positive kinetic cooperativity (the Hill coefficient > 1) of the substrate and coenzyme binding sites. Thus, the carp LDH seems to function more effectively under anaerobic conditions and at higher temperatures.

PPARα agonists clofibrate and gemfibrozil inhibit cell growth, down-regulate hCG and up-regulate progesterone secretions in immortalized human trophoblast cells

We studied effects of PPARα agonists clofibric acid and gemfibrozil on cell growth and functions of immortalized human extravillous trophoblast cells. Levels of DNA and protein gradually increased during incubation for 4 days. Gemfibrozil (>0.25 mM) and clofibric acid (2.5 mM) suppressed the rate of increase in DNA and protein. Specific activities of fatty acyl-CoA oxidase and catalase were increased to about 1.2–2.0 times the control value by 0.05 mM gemfibrozil and 1.0 and 2.5 mM clofibric acid after incubation for 3 days. Acid phosphatase activity showed a small increase in response to both agents, but esterase activity changed little. The secretion of progesterone from the cells into the medium was increased by 0.25 mM gemfibrozil and 1.0 and 2.5 mM clofibric acid after incubation for 3 days, but that of human chorionic gonadotropin (hCG) was decreased by 0.35 mM gemfibrozil and 2.5 mM clofibric acid. The specific activity of lactate dehydrogenase in the cells was hardly changed at all after incubation for 3 days. These results suggest that gemfibrozil and clofibric acid inhibit the proliferation of trophoblast cells. Cell metabolism is probably affected by both agents. The two agents may down-regulate hCG and up-regulate progesterone secretions.

Eugenol as An In Vivo Radioprotective Agent

In the present work, an attempt has been made to evaluate the possible in vivo radioprotection by eugenol. Swiss albino mice were administered different doses of eugenol (75,150 and 300 mg/kg) before exposure to 1.5 Gy of gamma radiation. The micronucleus test was carried out to determine the genetic damage in bone marrow. Our results demonstrated significant reduction in the frequencies of micronucleated polychromatic erythrocytes (MnPCEs) with all three eugenol doses. Eugenol (150 mg/kg) was also tested against different doses of radiation (0.5, 1, 1.5, and 2 Gy) and was found to afford significant radioprotection. Reduction in the incidence of MnPCEs could be noticed up to 72 h postirradiation (1.5 Gy). Moreover, the level of peroxidative damage and the specific activities of lactate dehydrogenase (LDH) and methylglyoxalase I (Gly I) were observed in the liver of mice treated with eugenol for seven days in comparison to untreated mice. The results revealed that eugenol exerted significant protection against oxidative stress. This possibility was further supported by the enhanced response of Gly I and the lowered activity of LDH. The present findings suggested that eugenol has a radioprotective potential.

Lactic acid production by Rhizopus oryzae transformants with modified lactate dehydrogenase activity.

Rhizopus oryzae is capable of producing high levels of lactic acid by the fermentation of glucose. Yields typically vary over 60-80%, with the remaining glucose diverted primarily into ethanol fermentation. The goal of this work was to increase lactate dehydrogenase (LDH) activity, so lactic acid fermentation could more effectively compete for available pyruvate. Three different constructs, pLdhA71X, pLdhA48XI, and pLdhA89VII, containing various lengths of the ldhA gene fragment, were transformed into R. oryzae. This fungus rarely integrates DNA used for transformation, but instead relies on extra-chromosomal replication in a high-copy number. Plasmid pLdhA48XI was linearized prior to transformation in order to facilitate integration into the pyrG gene used for selection. Isolates transformed with ldhA containing plasmid were compared with both the wild-type parent strain and the auxotrophic recipient strain containing vector only. All isolates transformed with pLdhA71X or pLdhA48XI had multiple copies of the ldhA gene that resulted in ldhA transcript accumulation, LDH specific activity, and lactic acid production higher than the controls. Integration of plasmid pLdhA48XI increased the stability of the strain, but did not seem to offer any benefit for increasing lactic acid production. Since lactic acid fermentation competes with ethanol and fumaric acid production, it was not unexpected that increased lactic acid production was always concomitant with decreased ethanol and fumaric acid. Plasmid pLdhA71X, containing a large ldhA fragment (6.1 kb), routinely yielded higher levels of lactic acid than the smaller region (3.3 kb) used to construct plasmid pLdhA48XI. The greatest levels of ldhA transcript and enzyme production occurred with isolates transformed with plasmid pLdhA89VII. However, these transformants always produced less lactic acid and higher amounts of ethanol, fumaric, and glycerol compared with the control.

Expression of lactate dehydrogenase A and B genes in different tissues of rats adapted to chronic hypobaric hypoxia.

Lactate dehydrogenase (LDH) is a tetramer made up of two different subunits A and B. In cellular models, severe hypoxia increases LDH A gene expression whereas LDH B gene does not exhibit any regulation. The aim of our work was to characterise LDH expression in different tissues of rats bred at high altitude. For this purpose, we chose a Sprague-Dawley rat strain adapted to chronic hypoxia in La Paz (3700 m), Bolivia. Two normoxic control groups were bred at low altitude in Clermont-Ferrand (350 m), France, one group was ad libitum with free access to food and water as was the hypoxic one, and the second normoxic group was nourished with the food intakes measured for the animals from La Paz. We measured total LDH specific activity, isoform distribution and LDH A and B mRNA amounts in three skeletal muscles (soleus, extensor digitorum longus (EDL), plantaris), heart and brain. Our study demonstrates that, unlike what has been shown in cellular models under severe hypoxia, LDH A gene is not systematically up-regulated in tissues of rats living at high altitude. Furthermore, chronic hypoxia limits LDH B gene transcription or its mRNA stability in both soleus and EDL. These regulations occur at various molecular levels like gene transcription, mRNA stabilisation or translation and protein stability, depending on the tissue studied, and are partly attributed to caloric restriction provoked by high altitude. These data provide insight into LDH gene expression underlying the diverse and complex tissue-specific response to chronic hypoxia.