Enhancement of lysozyme stability and activity by polyamines

Spermine, a low molecular weight polyamine, administered orally to suckling rats induces the maturation of the small intestine. In this organ, lysozyme is an important component of the innate immunity. In this report, we analysed the binding of spermine to lysozyme and its effect on thermal inactivation of the protein by spectroscopy techniques. The activity of the enzyme was analysed in presence of spermine by lysoplate technique. We studied the effects of spermine ingestion by suckling rats on intestinal lysozyme activity and gene expression. We reported that spermine binds to lysozyme and increases in vitro the thermal stability and the activity of the protein. When administered orally to suckling rats, spermine increases the lysozyme activity in jejunum, but not in ileum. This increase is not due to a modification of the gene expression. The observed effects lead us to postulate that spermine could be used in some mammals as a promoter of the innate immunity.     

Inhibition of Na+,K+-ATPase in Penaeus indicus postlarvae by lead

The plasma membrane/mitochondrial fractions of Penaeus indicus postlarvae contain Mg2+-dependent ATPase, Na+,K+-stimulated ATPase, Na+-stimulated ATPase and K+-stimulated ATPase. The Na+,K+-activated, Mg2+-dependent ATPase was investigated further in relation to different pH and temperature conditions, and at various concentrations of protein, ouabain, ATP and ions in the incubation medium. In vitro and in vivo effects of lead were studied on the enzyme activity. In vitro lead inhibited the enzyme activity in a concentration-dependent manner with an IC50 value of 204.4 microM. In correlation with in vitro studies, in vivo investigations (both concentration and time dependent) of lead also indicated a gradual inhibition in enzyme activity. A maximum decrease of 85.3% was observed at LC50 (7.2 ppm) of lead for concentration-dependent experiments. In time-dependent studies, the decrease was maximal (81.7%) at 30 days of sublethal exposure (1.44 ppm). In addition, the substrate- and ion-dependent kinetics of Na+,K+-ATPase was studied in relation to in vitro exposure of lead; these studies suggest a non-competitive type of inhibition.     

Isocytosine-based inhibitors of xanthine oxidase: Design,synthesis, SAR,PK and in vivo efficacy in rat model of hyperuricemia

Structure–activity relationship studies were carried out for lead generation following structure-guided design approach from an isocytosine scaffold identified earlier for xanthine oxidase inhibition. A 470-fold improvement in in vitro IC₅₀ was obtained in the process. Five most potent compounds with nanomolar IC₅₀ values were selected for pharmacokinetics and in vivo experiments. The best compound showed good in vivo activity when administered intraperitoneally but was not active by oral route. The results suggest that improvement in oral exposure could improve the in vivo efficacy of this series.     

Study of Response of Thymic and Submaxillary Lymph Node Lymphocytes to Administration of Lead by Different Routes

A number of studies have reported that heavy metals are not only toxic for the organism but they may modulate immune responses. In the current study, the effect of 4-week administration of 200 ppm of PbAc₂, using different routes of administration (orally and intraperitoneal injection), on lymphatic organs was evaluated. In the thymus, the number of lymphocyte cells and the cellularity diminished significantly for both routes of treatment. Regarding the submaxillary lymph nodes, no significant variations took place. Cell-mediated immune response is commonly evaluated by cell proliferation assays. Mitogens are known to induce a vigorous proliferative response in lymphoid cells from mammals. An increase in the proliferation of T lymphocytes stimulated by concanavalin A and the proliferation of B lymphocytes stimulated with lipopolysaccharides was found in thymus for both routes of administration, whereas in the lymph nodes, there was a decrease in proliferation of T lymphocytes. Furthermore, lead administration by intraperitoneal route caused an effect on B and T lymphocyte subpopulations. Thus, there was an increase in B+ cells and a decrease in T+ cells. Regarding CD4+ and CD8+ T cells, there were only variations, concretely a drop in both subpopulations, in lymph nodes when lead was administered intraperitoneally. It is important to emphasize that an increase in apoptosis was found in this tissue. At the histological level, evident alterations were described in thymus both for the oral and for the intraperitoneal route. Therefore, it is possible to show that lead administered by both routes generated effects on an immunological level.     

Lactobacillus plantarum CCFM8661 Alleviates Lead Toxicity in Mice

Lead causes a broad range of adverse effects in humans and animals. The objective was to evaluate the potency of lactobacilli to bind lead in vitro and the protective effects of a selected Lactobacillus plantarum CCFM8661 against lead-induced toxicity in mice. Nine strains of bacteria were used to investigate their binding abilities of lead in vitro, and L. plantarum CCFM8661 was selected for animal experiments because of its excellent lead binding capacity. Both living and dead L. plantarum CCFM8661 were used to treat 90 male Kunming mice during or after the exposure to 1?g/L lead acetate in drinking water. The results showed oral administration of both living and dead L. plantarum CCFM8661 offered a significant protective effect against lead toxicity by recovering blood ??-aminolevulinic acid dehydratase activity, decreasing the lead levels in blood and tissues, and preventing alterations in the levels of glutathione, glutathione peroxidase, malondialdehyde, superoxide dismutase, and reactive oxygen species caused by lead exposure. Moreover, L. plantarum CCFM8661 was more effective when administered consistently during the entire lead exposure, not after the exposure. Our results suggest that L. plantarum CCFM8661 has the potency to provide a dietary strategy against lead toxicity.     

Regulation of postnatal beta-adrenergic receptor/adenylate cyclase development by prenatal agonist stimulation and steroids: alterations in rat kidney and lung after exposure to terbutaline or dexamethasone

Glucorticoids and adrenergic stimulation are both thought to control the development of beta-adrenergic receptors/responses. In the current study, rats were exposed to dexamethasone or terbutaline during late gestation and the development of beta-receptor binding capabilities and adenylate cyclase activity evaluated in membrane preparations from kidney and lung. Prenatal dexamethasone exposure produced postnatal adrenergic hyperreactivity of kidney adenylate cyclase; the effect resulted from increases in the enzyme itself, as both basal adenylate cyclase and forskolin-stimulation of the enzyme were also increased by dexamethasone. Similarly, prenatal terbutaline exposure evoked increases in basal, isoproterenol-stimulated and forskolin-stimulated adenylate cyclase in the kidney. In the lung, dexamethasone produced an initial postnatal deficit in basal adenylate cyclase and deficient responsiveness to isoproterenol, but the deficit resolved shortly after birth. Terbutaline selectively promoted the ability of isoproterenol to stimulate lung adenylate cyclase in the first few days after birth, without alterations in basal adenylate cyclase; this was followed by a period of prolonged subsensitivity of both basal and isoproterenol-stimulated activity. Although dexamethasone and terbutaline also caused significant changes in development of beta-receptor binding capabilities, in neither tissue could these effects account for the direction or magnitude of the changes in adenylate cyclase reactivity. Thus, glucocorticoids and beta-agonists can participate in the programming of development of postsynaptic reactivity by exerting actions upon post-receptor coupling mechanisms.     

Metabolic distribution of radioactivity in Sprague-Dawley rats and B6C3F1 mice exposed to 1,3-[2,3-14C]-butadiene by whole body exposure

The uptake of 1,3-[2,3-(14)C]-butadiene and its disposition, measured as radioactivity in urine, faeces, exhaled volatiles and CO(2) during and following 6 h whole body exposure to 20 ppm butadiene has been investigated in male Sprague-Dawley rats and B6C3F1 mice. Whilst there were similarities between the two species, the uptake and metabolic distribution of butadiene were somewhat different for rats and mice. The major differences observed were in the urinary excretion of radioactivity and in the exhalation of 14C-CO(2). After 42 h from the start of exposure, 51.1% of radioactivity was eliminated in rat urine compared with 39.5% for mouse urine. 34.9% of the recovered radioactivity was exhaled by rats as 14C-CO(2), compared with 48.7% by mice. Excretion of radioactivity in faeces was similar for both species (3.8% for rats and 3.4% for mice). The tissue concentrations of 14C-butadiene equivalents measured in liver, testes, lung and blood of exposed mice were 0.493, 0460, 0.457, and 1.626 nmol/g tissue, respectively. The values for the corresponding rat tissues were 0.869, 0.329, 0.457, and 1.626 nmol butadiene equivalents/g tissue, respectively. For rats, 6.2% of recovered radioactivity (0.288 nmol butadiene equivalents/g tissue) was retained in carcasses whereas for mice the amount was 3.6% (0.334 nmol butadiene equivalents/g tissue). There were also some significant differences between the metabolic conversion of 1,3-[2,3-(14)C]-butadiene and excretion by mice following the 20 ppm whole body exposure compared to previously reported data for nose-only exposure to 200 ppm butadiene [Richardson et al., Toxicol. Sci. 49 (1999) 186]. The main difference between the high- and low-exposure studies was in the exhalation of 14C-CO(2). At the 200 ppm exposure, 40% of the radioactivity was exhaled as 14C-CO(2) by rats whereas 6% was measured by this route for mice. The proportional conversion of butadiene to CO(2) by mice was significantly greater at the low exposure concentration compared with that reported for the higher concentration. This shift was not observed for rats. The difference between species could be caused by a saturation of metabolism in mice between 20 and 200 ppm for the pathways leading to CO(2). Restraint or error in collection of CO(2) in the 200 ppm study could also be factors.     

Effect of cadmium on pulmonary and renal microsomal drug metabolizing enzymes of the guinea-pig

1. The effect of acute cadmium (Cd) treatment on pulmonary and renal microsomal aniline 4-hydroxylase and ethylmorphine N-demethylase enzyme activities of adult male guinea-pigs were assessed 72 hr following a single dose of Cd ion (2 mg Cd2+/kg i.p.). Tissue and microsomal Cd levels were also determined. 2. There were no significant differences between either lung or kidney tissue weights, microsomal protein contents or enzyme activities of Cd treated and control animals. 3. The tissues and microsomes of Cd-treated animals were found to have significantly higher levels of Cd than those of control animals. In Cd treated animals, tissue and microsomal Cd levels of kidney were found to be higher than that of lung. 4. In vitro addition of cadmium chloride (CdCl2) to incubation mixtures produced concentration related inhibitions of microsomal drug metabolizing enzymes in each tissue. However, in vitro effect of CdCl2 was found to be stronger on drug metabolizing enzymes of kidney than those of lung. In addition, while the strength of Cd effect was more pronounced on the activity of ethylmorphine N-demethylase than that of aniline 4-hydroxylase in the lung, the opposite was observed in the kidney.     

Lead acetate: toxicity without effcts on the locomotor activity of the bluegill sunfish, Lepomis macrochirus Rafinesque

The concentration of lead acetate which caused 50% mortality of bluegill in 96 h (96 h LC50) is approximately 400 ppm. Sublethal concentrations of 0.1, 1, 10, 100, 200 and 300 ppm lead acetate did not produce any effects on the locpmotor activity of the bluegill. The observation that lead can reach concentrations which are lethal without sublethal effects on locomotor activity at lower concentrations is unusual and in sharp contrast to studies with other metals (Ellgaard et al. . 1978).     

Experimental chemotherapy of schistosomiasis mansoni. XIII. Activity of praziquantel, an isoquinoline-pyrazino derivative, on mice, hamsters and Cebus monkeys.

In mice experimentally infected with Schistosoma mansoni, praziquantel (2-cyclohexylcarbonyl-1,3,4,6,7,11b-hexahydro-2H-pyrazino[2,1-a]isoquinoline-4-one), administered orally at the levels of 100 and 50 mg/kg, for 5 consecutive days, produces oogram changes in all animals and a pronounced hepatic shift of schistosomes (97.1 and 89.1, respectively). At lowest levels (12.5 and 6.3 mg/kg), alterations in the oogram could still be detected, although hepatic shift of schistosomes was no more evident. After a single intramuscular injection, the results obtained paralleled those observed with a single-dose oral treatment. The hepatic shift was only moderate at 200 and 100 mg/kg and the percentages of worms retained in the liver, after perfusion, were particularly low. When nasal route in a 1-day regimen was used, the results obtained were slightly less evident as compared with those observed by oral route (5-day schedule). Considering the percentage of oogram changes, the degree of hepatic shift of schistosomes and the percentage of worms fixed in the liver, the antischistosomal activity of praziquantel was greater in hamsters than in mice. Actually, a daily dose as low as 12.5 mg/kg, administered for 5 consecutive days, was sufficient to shift 60.4% of the worms towards the liver and to produce alterations of the oogram in 60% of the animals. In Cebus monkeys orally treated with 10 and 20 mg/kg of praziquantel, given 3 times within a single day (total doses of 30 and 60 mg/kg, respectively), a remarkable reduction in worm burden was observed. A single oral or intramuscular dose of 100 mg/kg was found to be curative. One Cebus doses with 100 mg/kg, by nasal spray, was found to harbor only female worms at autopsy performed 69 days after treatment.     

急性铅应激诱导肝肾损伤及其分子机制初探

利用腹腔注射醋酸铅方法构建了铅染毒小鼠(Mus muscculus)模型,观察了染毒小鼠肝、肾的组织学变化,并通过免疫组织化学方法检测了染毒小鼠肝、肾组织中Caspase-3、Bcl-2和Bax蛋白的表达量.结果发现,急性铅染毒可诱导肝和肾组织学损伤,且在诱导肝细胞和肾细胞凋亡、损伤过程中,随时间的延长,Caspase-3的表达量逐渐增加,而Bcl-2与Bax两蛋白表达量的比值呈逐渐下降趋势,有一定的时效性,染毒48 h后,与对照组相比,均差异极显著,表明铅可能通过影响Caspase-3、Bcl-2和Bax的表达而诱导肝和肾细胞异常凋亡.     

Effect of chronic lead exposure on kidney function in male and female rats: determination of a lead exposure biomarker

Several cytotoxic chemical pollutants inducing peroxidative damages are liable to induce kidney failure. Among these pollutants we find heavy metals such as: lead, nickel, cadmium, vanadium and mercury. Lead is one of the most dangerous metals because it is widely spread in the environment, and because it may be a source of several nervous diseases. The aim of this study is to provide evidence concerning the effect of this metal on the renal function and to try to determine a storage corner in the organism which serves as an indicator of a lead intoxication. Lead acetate was administered by oral route in the drinking water to adult rats aged three months at the rate of 0.3% (P1) and 0.6% (P2). Reference rats received distilled water to drink under the same conditions. The treatment continued for 15, 30, 45, 60 and 90 days. The creatinemia, uremia, glycemia and creatinuria are determined by colorimetric techniques. Lead concentration in blood as well as the lead content of the tail are determined by atomic absorption after nitroperchloric mineralization at the liquid stage. The results showed an increase of creatinemia on the 30th day of the experiment for both sexes in (P1 and P2). The same happened for ureamia. The increase of these two parameters would indicate a renal deficiency which is confirmed by a decrease of creatinuria and urinary pH observed mainly on and after the 45th day of the experiment. An increase of the renal relative weight was noticed in P1 and P2 on the 30th day of the treatment. The determination of the concentration of lead in the blood shows that this factor increases among treated subjects in a constant way, independently of the dose and the duration of the treatment. Nevertheless, the rate increase of lead in the tail seems to be dose-dependent. In conclusion, lead administered by oral route causes a renal deficiency to the rat without distinction between males and females. In addition, the tail seems to be a reliable exposure biomarker that demonstrates lead intoxication. The tail seems to be a dosimeter of lead bio-accumulation. It constitutes an endogenous source of lead impregnation. The concentration of lead in the blood is only an indicator of recent exposure.     

Interaction of lead with some essential elements in rat’s kidney in relation to age

The aim of this study was to evaluate the interaction of lead given orally with Fe, Zn, and Cu in adult female rats and in their pups. Kidney was chosen for studying this interaction. Four different doses of lead (acetate) from 1500 to 7500 ppm were administered to mature female albino rats in beverages during 6 wk. The exposure lasted from mating up to 3 wk after delivery. Pb, Fe, Zn, and Cu were determined in kidneys of mothers and pups. Histopathological examinations were also performed. Results showed significantly lower concentrations of Fe, Zn, and Cu in kidneys of mothers on all lead levels. Their pups showed no change in concentrations of essential elements and even increased Fe at the highest exposure level. Concentrations of Pb and histopathological changes in the kidney were similar in mothers and offspring, although pups received only a fraction of the mothers’ doses. Our results indicate that in immature animals the interaction of Pb with essential elements in the kidney is different from that in their mothers.     

Behavioral deficits induced by lead exposure are accompanied by serotonergic and cholinergic alterations in the prefrontal cortex

The effects of long-term lead (Pb) exposure producing a blood Pb concentration of lower than 20 μg/dL, i.e. below that associated with overt neurological deficits in occupationally exposed individuals, was studied in adult rats. In order to assess gender differences, we performed parallel behavioral experiments in male and female rats. Exposure to Pb acetate (50 ppm in drinking water) for 6 months induced motor and cognitive alterations, however these effects were gender- and task-dependent. Chronic lead exposure impaired spatial learning assessed in the Morris water maze test (MWM) in both genders, whereas it only induced hyperactivity in the open field and impaired motor coordination in the rotarod test, only in male rats. Hyperactivity in male rats was accompanied by an increase in extracellular level of acetylcholine in the prefrontal cortex. Extracellular dopamine concentration in the prefrontal cortex was unaffected by lead exposure whereas serotonin concentration in the same brain area was significantly decreased in both male and female rats exposed to lead. These results unveil new molecular mechanisms underlying neuropsychiatric alterations induced by chronic lead exposure.     

Distribution of nickel,zinc, and copper in rat organs after oral administration of nickel(II) chloride

The distribution of Ni administered as NiCl₂ · 6H₂O in the drinking water (300 and 1200 ppm Ni for 90 d) was studied using male Wistar rats. Next, the effect of Ni on the concentration of zinc (Zn) and copper (Cu) in selected organs and serum was measured. The metals were analyzed in the liver, kidney, lung, spleen, brain, and serum by electrothermal (Ni) or flame (Zn, Cu) atomic absorption spectrophotometry. The results indicate that exposed rats drank less nickel solutions than the volume of water drunk by controls, but there was no mortality of animals. In comparison to control animals, a very high increase in Ni levels was found in the kidney and then lung and serum of all exposed rats. In the liver, spleen, and brain, the metal accumulation was lower. A directly proportional relation between the nickel intake and its deposition was observed in the collected organs and in the serum. The metal level did not change significantly in the course of exposure (the first analysis was after 30 d). The administration of 300 ppm Ni did not affect the zinc and copper concentration in studied organs, except the serum, where zinc content was significantly reduced. At a dose of 1200 ppm Ni, these metals were found to be depressed in the liver, kidney, serum (zinc), and copper in the kidney.     

Effects of sub-chronic low-level lead exposure on the homeostasis of copper and zinc in rat tissues

Information about the health risks or the subtle adverse health effects that might be associated with low-level lead exposure on micronutrient metabolism are scarce in the literature. The present work investigated the subtle adverse health effects of exposure to progressively low levels of lead on the metabolism of two micronutrients, copper and zinc in different tissues of the rat. Rats were exposed to 200, 300 and 400 ppm lead in their drinking water for 12 weeks. Lead, copper and zinc concentrations were determined in blood, liver, kidney, heart, spleen and brain of the animals. While the imbalance in zinc metabolism was characterized by a deposition of zinc in the kidney and to a lesser extent in the heart of the animals, imbalance in copper metabolism was characterized by a depletion of blood and splenic copper concentrations as well as renal and cardiac accumulation of copper. Hepatic and brain copper and zinc contents, together with blood zinc were not affected by the 12-week lead exposure. A linear relationship was observed between lead dose and lead accumulation in the spleen, whereas a non-linear relationship was observed between lead dose and lead accumulation in blood, liver, kidney and heart. Our findings indicate that exposure to progressively low-level lead concentrations results in imbalance in copper and zinc in the organism and this might be a factor in propensity toward behavioral disorders observed in lead exposure.     

Altered angiotensin-converting enzyme in lung and extrapulmonary tissues of hypoxia-adapted rats

The effects of exposing rats to hypoxia (10% O2) at normal atmospheric pressure for periods of 14 or 28 days on angiotensin-converting enzyme (ACE) activity and stores of angiotensin I (ANG I) and angiotensin II (ANG II) in lung, kidney, brain, and testis were examined. ACE activity was measured by spectrophotometric assay, and active sites of ACE were estimated by measuring the binding of 125I-351A [N-(1-carbonyl-3-phenyl-propyl)-L-lysyl-L-proline], a highly specific active site-directed inhibitor of ACE, to tissue homogenates and perfused lungs. Hypoxia exposure produced progressive reductions in ACE activity in lung homogenates and in ACE inhibitor binding to perfused lungs. ANG II levels in lungs from hypoxia-adapted animals were significantly less than air controls, suggesting that the reduction in intrapulmonary ACE activity was associated with reduced local generation of ANG II. ACE activity was increased in kidney and unchanged in brain and testis of hypoxia-adapted rats compared with air controls. Thus the effects of chronic hypoxia on catalytically active ACE and ACE active sites in the intact animal were organ specific. Adaptation to chronic hypoxia did not significantly alter plasma renin activity or ANG I or ANG II levels or serum ACE content. The hypoxia-induced alterations in lung and kidney ACE were reversible after return to a normoxic environment.     

Examination of Changes in Enzyme Activities of Erythrocyte Glucose 6‐Phosphate Dehydrogenase and 6‐Phosphogluconate Dehydrogenase in Rats Given Naringenin and Lead Acetate

In our study, controlled experimental groups were performed by giving substances Lead acetate, Naringenin and Naringenin + Lead acetate to rats in vivo conditions Changes in the glucose 6‐phosphate dehydrogenase (G6PD) and 6‐phosphogluconate dehydrogenase (6PGD) enzyme activities in erythrocytes of rats in these groups were compared to the Control group. An inhibition significant degree for G6PD enzyme activity was observed in all groups when compared to the Control group (p < 0.001). While inhibition significant degree for 6PGD enzyme activity was observed in Lead acetate groups (p < 0.001), no significant effect was observed in the Naringenin and Naringenin + Lead acetate groups (p > 0.05). In addition, lead levels in the groups of rats were determined using an inductively coupled plasma mass spectrometer (ICP‐MS) device. As a result of measurements by the ICP‐MS device, lead levels were found as an average of 42.9 ± 2.51, 36.71 ± 1.13, 172.16 ± 9.63, and 95.07 ± 5.87 ppm in the Control, Naringenin, Lead acetate and Naringenin + Lead acetate groups, respectively. Our results were shown that Naringenin has protective effects on the Lead acetate induced oxidative stress erythrocytes in rat.     

Lysozyme is an ozone-sensitive component of alveolar type II cell lamellar bodies

Exposure of rats to 3 ppm ozone for up to 8 h results in significant changes in lamellar bodies, the surfactant storing organelles of type II cells. We have previously shown that a 14 kDa lamellar body protein is decreased as early as 4 h after the onset of ozone exposure. We have isolated this ozone-sensitive protein from rat lung lamellar bodies and identified it as lysozyme by immunochemical methods, as well as by its amino acid composition, N-terminal amino acid sequence and bacteriolytic activity. Reduced lysozyme activity in isolated lamellar bodies is detected as early as 4 h after the start of ozone exposure. Following an 8 h ozone exposure, the activity does not return to control levels for at least 48 h. Lamellar body lysozyme is expected to be secreted with surfactant phospholipids, thereby contributing to the antimicrobial defense of the alveolar lining layer. The acute lysozyme deficiency seen in ozone-induced oxidant injury may reduce the resistance of the lung to infection.