Age-related changes and effects of regular low-intensity exercise on gait, balance, and oxidative biomarkers in the spinal cord of Wistar rats.

The present study aimed to analyze age-related changes to motor coordination, balance, spinal cord oxidative biomarkers in 3-, 6-, 18-, 24-, and 30-month-old rats. The effects of low-intensity exercise on these parameters were also analyzed in 6-, 18-, and 24-month-old rats. Body weight, blood glucose, total cholesterol, and high-density lipoprotein (HDL) cholesterol were assessed for all rats. The soleus muscle weight/body weight ratio was used to estimate skeletal muscle mass loss. Body weight increased until 24 months; only 30-month-old rats exhibited decreased blood glucose and increased total cholesterol and HDL cholesterol. The soleus muscle weight/body weight ratio increased until 18 months, followed by a small decrease in old rats. Exercise did not change any of these parameters. Stride length and step length increased from adult to middle age, but decreased at old age. Stride width increased while the sciatic functional index decreased in old rats. Performance in the balance beam test declined with age. While gait did not change, balance improved after exercise. Aging increased superoxide anion generation, hydrogen peroxide levels, total antioxidant capacity, and superoxide dismutase activity while total thiol decreased and lipid hydroperoxides did not change. Exercise did not significantly change this scenario. Thus, aging increased oxidative stress in the spinal cord, which may be associated with age-induced changes in gait and balance. Regular low-intensity exercise is a good alternative for improving age-induced changes in balance, while beneficial effects on gait and spinal cord oxidative biomarkers cannot be ruled out because of the small number of rats investigated (n=5 or 6/group).

Age-related changes and effects of regular low-intensity exercise on gait, balance, and oxidative biomarkers in the spinal cord of Wistar rats

The present study aimed to analyze age-related changes to motor coordination, balance, spinal cord oxidative biomarkers in 3-, 6-, 18-, 24-, and 30-month-old rats. The effects of low-intensity exercise on these parameters were also analyzed in 6-, 18-, and 24-month-old rats. Body weight, blood glucose, total cholesterol, and high-density lipoprotein (HDL) cholesterol were assessed for all rats. The soleus muscle weight/body weight ratio was used to estimate skeletal muscle mass loss. Body weight increased until 24 months; only 30-month-old rats exhibited decreased blood glucose and increased total cholesterol and HDL cholesterol. The soleus muscle weight/body weight ratio increased until 18 months, followed by a small decrease in old rats. Exercise did not change any of these parameters. Stride length and step length increased from adult to middle age, but decreased at old age. Stride width increased while the sciatic functional index decreased in old rats. Performance in the balance beam test declined with age. While gait did not change, balance improved after exercise. Aging increased superoxide anion generation, hydrogen peroxide levels, total antioxidant capacity, and superoxide dismutase activity while total thiol decreased and lipid hydroperoxides did not change. Exercise did not significantly change this scenario. Thus, aging increased oxidative stress in the spinal cord, which may be associated with age-induced changes in gait and balance. Regular low-intensity exercise is a good alternative for improving age-induced changes in balance, while beneficial effects on gait and spinal cord oxidative biomarkers cannot be ruled out because of the small number of rats investigated (n=5 or 6/group).

Treatment with ascorbic acid and α-tocopherol modulates oxidative-stress markers in the spinal cord of rats with neuropathic pain.

Vitamin E (vit. E) and vitamin C (vit. C) are antioxidants that inhibit nociception. The effect of these vitamins on oxidative-stress markers in the spinal cord of rats with chronic constriction injury (CCI) of the sciatic nerve is unknown. This study investigated the effect of intraperitoneal administration of vit. E (15 mg·kg-1·day-1) and vit. C (30 mg·kg-1·day-1), given alone or in combination, on spinal cord oxidative-stress markers in CCI rats. Adult male Wistar rats weighing 200-250 g were divided equally into the following groups: Naive (rats did not undergo surgical manipulation); Sham (rats in which all surgical procedures involved in CCI were used except the ligature), and CCI (rats in which four ligatures were tied loosely around the right common sciatic nerve), which received injections of vitamins or vehicle (saline containing 1% Tween 80) for 3 or 10 days (n=6/each group). The vitamins prevented the reduction in total thiol content and the increase in superoxide-anion generation that were found in vehicle-treated CCI rats. While nitric-oxide metabolites increased in vehicle-treated CCI rats 3 days after surgery, these metabolites did not show significant changes in vitamin-treated CCI rats. In all rats, total antioxidant capacity and hydrogen-peroxide levels did not change significantly. Lipid hydroperoxides increased 25% only in vehicle-treated CCI rats. These changes may contribute to vit. C- and vit. E-induced antinociception, because scavenging reactive oxygen species seems to help normalize the spinal cord oxidative status altered by pain.

Effects of N-acetylcysteine on spinal cord oxidative stress biomarkers in rats with neuropathic pain.

N-acetylcysteine (NAC) inhibits nociceptive transmission. This effect has been associated partly with its antioxidant properties. However, the effect of NAC on the levels of lipid hydroperoxides (a pro-oxidant marker), content of ascorbic acid (a key antioxidant molecule of nervous tissue) and total antioxidant capacity (TAC) is unknown. Thus, our study assessed these parameters in the lumbosacral spinal cord of rats with chronic constriction injury (CCI) of the sciatic nerve, one of the most commonly employed animal models of neuropathic pain. Thirty-six male Wistar rats weighing 200-300 g were equally divided into the following groups: Naive (rats did not undergo surgical manipulation); Sham (rats in which all surgical procedures involved in CCI were used except the ligature), and CCI (rats in which four ligatures were tied loosely around the right common sciatic nerve). All rats received intraperitoneal injections of NAC (150 mg·kg-1·day-1) or saline for 1, 3, or 7 days. Rats were killed 1, 3, and 7 days after surgery. NAC treatment prevented the CCI-induced increase in lipid hydroperoxide levels only at day 1, although the amount was higher than that found in naive rats. NAC treatment also prevented the CCI-induced increase in ascorbic acid content, which occurred at days 1, 3, and 7. No significant change was found in TAC with NAC treatment. The changes observed here may be related to the antinociceptive effect of NAC because modulation of oxidative-stress parameters seemed to help normalize the spinal cord oxidative status altered by pain.

N-acetylcysteine downregulates phosphorylated p-38 expression but does not reverse the increased superoxide anion levels in the spinal cord of rats with neuropathic pain.

We determined the effect of N-acetylcysteine (NAC) on the expression of the phosphorylated p38 (p-p38) protein and superoxide anion generation (SAG), two important players in the processing of neuropathic pain, in the lumbosacral spinal cord of rats with chronic constriction injury (CCI)-induced neuropathic pain. The sciatic functional index (SFI) was also measured to assess the functional recovery post-nerve lesion. Thirty-six male Wistar rats were divided equally into the following groups: Naive (rats did not undergo surgical manipulation); Sham (rats in which all surgical procedures involved in CCI were used except the ligature), and CCI (rats in which four ligatures were tied loosely around the right common sciatic nerve), which received 2, 4, or 8 intraperitoneal injections of NAC (150 mg·kg-1·day-1) or saline beginning 4 h after CCI. Rats were sacrificed 1, 3, and 7 days after CCI. The SFI was measured on these days and the lumbosacral spinal cord was used for analysis of p-p38 expression and SAG. CCI induced a decrease in SFI as well as an increase in p-p38 expression and SAG in the spinal cord. The SFI showed a partial recovery at day 7 in saline-treated CCI rats, but recovery was improved in NAC-treated CCI rats. NAC induced a downregulation in p-p38 expression at all time-points evaluated, but did not reverse the increased SAG induced by CCI. Since p-p38 is a mediator in neuropathic pain and/or nerve regeneration, modulation of this protein may play a role in NAC-induced effects in CCI rats.

Moderate hypoxia is able to minimize the manganese-induced toxicity in tissues of silver catfish (Rhamdia quelen).

The aim of this study was to compare the effects of manganese (Mn) on silver catfish exposed to different levels of dissolved oxygen. Silver catfish (Rhamdia quelen) were exposed to increasing concentrations of Mn (4.2, 8.4 or 16.2mgL(-1)) under either normoxia (100 percent saturation) or moderate hypoxia (51.87 percent saturation) for 15 days. Under normoxia, Mn exposure increased lipid peroxidation (LP) in brain and kidney; it increased gluthatione (GSH) levels in brain and decreased catalase (CAT) activity in both tissues. Moderate hypoxia was able to prevent Mn-induced LP in brain and to reduce this oxidative parameter in kidney; GSH level was increased in brain, while CAT activity was reduced in both tissues. Activity of isolated mitochondria of liver and gills was reduced by Mn exposure under both levels of dissolved oxygen, but this effect was more prominent in normoxia. As expected, liver, kidney and gills showed an increase of Mn accumulation according to waterborne levels, and these parameters presented positive relationship. The highest waterborne Mn (8.4 and 16.2mgL(-1)) resulted in greater accumulation under normoxia, indicating that moderate hypoxia can stimulate mechanisms capable of reducing Mn accumulation in tissues (though not in blood). Moderate hypoxia can be considered a stress factor and Mn an aquatic anthropogenic contaminant. Therefore we hypothesized that these two conditions together are able to invoke defense mechanisms in juvenile silver catfish, acting in a compensatory form, which may be related to adaptation and/or hormesis.

Immersion anaesthesia with tricaine methanesulphonate or propofol on different sizes and strains of silver catfish Rhamdia quelen.

The efficacy of immersion anaesthesia with tricaine methanesulphonate (MS222) or propofol on albino and grey silver catfish Rhamdia quelen was assessed through induction and recovery times and observation of mortality. Besides reporting a novel, efficient and practical use of propofol as an immersion anaesthetic, the study shows that it is essential to consider size and strain when anaesthetizing R. quelen with MS222 or propofol bath solution in order to minimize physiological impact.

Redox profile in liver of Leporinus macrocephalus exposed to different dissolved oxygen levels.

Aquatic organisms are continuously exposed to environmental variations, which can lead to physiological and biochemical alterations. Leporinus macrocephalus, known as piavuçu, is a migratory species that may be exposed to variations in dissolved oxygen levels. Studies evaluating oxidative changes undergone by this species in these conditions are scarce. Therefore, this investigation aimed at evaluating oxidative alterations in L. macrocephalus exposed to different oxygen levels for 96 h: 6.12 ± 0.18, 3.99 ± 0.17, 3.22 ± 0.17, 2.47 ± 0.30 and 0.710 ± 0.07 mg L(-1). At the end of the experimental period, fish were euthanized and livers used to determine lipid hydroperoxides, thiobarbituric acid reactive substances, catalase, glutathione-S-transferase, superoxide dismutase and thiol groups, which are an indirect measure of reduced glutathione. Results indicated a decrease in the studied parameters in hypoxic situations, suggesting a possible metabolic depression.

Oxidative stress parameters in silver catfish (Rhamdia quelen) juveniles infected with Ichthyophthirius multifiliis and maintained at different levels of water pH.

The aim of this study was to determine oxidative stress parameters in the liver, gill and muscle of silver catfish juveniles infected with Ichthyophthirius multifiliis and maintained at pH 5.0 or 7.0 for three days. Juveniles were infected by adding one I. multifiliis-infected juvenile and water containing theronts to tanks. After the appearance of white spots on the skin, infected juveniles exposed to pH 5.0 and 7.0 showed significantly higher thiobarbituric acid reactive substances (TBARS) levels in the liver and gills compared to uninfected juveniles. Liver of infected juveniles exposed to pH 7.0 showed higher catalase (CAT) and lower glutathione-S-transferase (GST) activities, but those maintained at pH 5.0 showed significantly higher GST activity than uninfected juveniles. The gills of infected juveniles showed significantly higher CAT (day two) and GST activity at both pH 5.0 and 7.0 compared to uninfected juveniles. Muscle of infected juveniles showed significantly lower CAT and GST activity and TBARS levels (at day three) when maintained at both pH 5.0 and 7.0 compared to uninfected juveniles. In conclusion, I. multifiliis infection induces liver and gill damage via lipid peroxidation products in silver catfish, but higher antioxidant enzyme activity could indicate a greater degree of protection against this parasite.

Characterization of a protease of a feather-degrading Microbacterium species.

AIMS: To characterize a new feather-degrading bacterium. METHODS AND RESULTS: The strain kr10 producing a high keratinolytic activity when cultured on native feather broth was identified as Microbacterium sp., based on phenotypical characteristics and 16S rDNA sequence. The bacterium presented optimum growth and feather-degrading activity at pH 7.0 and 30 degrees C. Complete feather degradation was achieved during cultivation. The keratinase was partially purified by gel filtration chromatography. It was optimally active at pH 7.0 and 55 degrees C. The enzyme was inhibited by 1,10-phenanthroline, EDTA, p-chloromercuribenzoic acid, 2-mercaptoethanol and metal ions like Hg(2+), Cu(2+) and Zn(2+). SIGNIFICANCE AND IMPACT OF THE STUDY: A new Microbacterium sp. strain was characterized presenting high feather-degrading activity, which appears to be associated to a metalloprotease-type keratinase. This micro-organism has enormous potential for use in biotechnological processes involving keratin hydrolysis.

Isolation and characterization of a feather-degrading bacterium from the poultry processing industry.

A Flavobacterium sp. producing a high keratinolytic activity was isolated from a poultry industry after growth on selective feather meal agar. This bacterium grew on feather meal broth, producing keratinase, and was also capable of complete degradation of raw feathers. The proteolytic activity was assessed in the presence of specific protease inhibitors. The crude enzyme showed mainly metalloprotease character. This novel isolate would have potential biotechnological use in processes involving keratin hydrolysis.

In vitro antimicrobial activity of a new series of 1,4-naphthoquinones.

The antibacterial activity of a series of 1,4-naphthoquinones was demonstrated. Disk diffusion tests were carried out against several Gram-positive and Gram-negative bacteria. The compound 5-amino-8-hydroxy-1,4-naphthoquinone was the most effective, presenting inhibition zones measuring 20 mm against staphylococci, streptococci and bacilli at 50 microg/ml. Methicillin-resistant Staphylococcus aureus and several clinical isolates of this bacterium were also inhibited. Naphthazarin, 5-acetamido-8-hydroxy-1,4-naphthoquinone, and 2,3-diamino-1,4-naphthoquinone were the next most active compounds. The minimal inhibitory concentration of the active compounds was determined against S. aureus, ranging from 30 to 125 microg/ml. All compounds presented a minimal bactericidal concentration higher than 500 microg/ml, indicating that their effect was bacteriostatic. The EC50, defined as the drug concentration that produces 50% of maximal effect, was 8 microg/ml for 5-amino-8-hydroxy-1,4-naphthoquinone against S. aureus, S. intermedius, and S. epidermidis. These results indicate an effective in vitro activity of 5-amino-8-hydroxy-1,4-naphthoquinone and encourage further studies for its application in antibiotic therapy.

In vitro antimicrobial activity of a new series of 1, 4-naphthoquinones

The antibacterial activity of a series of 1,4-naphthoquinones was demonstrated. Disk diffusion tests were carried out against several Gram-positive and Gram-negative bacteria. The compound 5-amino-8-hydroxy-1,4-naphthoquinone was the most effective, presenting inhibition zones measuring 20 mm against staphylococci, streptococci and bacilli at 50 æg/ml. Methicillin-resistant Staphylococcus aureus and several clinical isolates of this bacterium were also inhibited. Naphthazarin, 5-acetamido-8-hydroxy-1,4-naphthoquinone, and 2,3-diamino-1,4-naphthoquinone were the next most active compounds. The minimal inhibitory concentration of the active compounds was determined against S. aureus, ranging from 30 to 125 æg/ml. All compounds presented a minimal bactericidal concentration higher than 500 æg/ml, indicating that their effect was bacteriostatic. The EC50, defined as the drug concentration that produces 50 percent of maximal effect, was 8 æg/ml for 5-amino-8-hydroxy-1,4-naphthoquinone against S. aureus, S. intermedius, and S. epidermidis. These results indicate an effective in vitro activity of 5-amino-8-hydroxy-1,4-naphthoquinone and encourage further studies for its application in antibiotic therapy

Peritoneal fluid modulates the sperm acrosomal exocytosis induced by N-acetylglucosaminyl neoglycoprotein.

The effect of peritoneal fluid (PF) on the human sperm acrosome reaction (AR) was tested. Sperm was pre-incubated with PF and the AR was induced by calcium ionophore A23187 and a neoglycoprotein bearing N-acetylglycosamine residues (NGP). The AR induced by calcium ionophore was inhibited 40% by PF from controls (PFc) and 50% by PF from the endometriosis (PFe) group, but not by PF from infertile patients without endometriosis (PFi). No significant differences were found in the spontaneous AR. When the AR was induced by NGP, pre-incubation with PFc reduced (60%) the percentage of AR, while PFe and PFi caused no significant differences. The average rates of acrosome reactions obtained in control. NGP- and ionophore-treated sperm showed that NGP-induced exocytosis differed significantly between the PFc (11%) and PFe/PFi groups (17%), and the ionophore-induced AR was higher for PFi (33%) than PFc/PFe (25%). The incidence of the NGP-induced AR was reduced in the first hour of pre-incubation with PFc and remained nearly constant throughout 4 h of incubation. The present data indicate that PF possesses a protective factor which prevents premature AR.

Peritoneal fluid modulates the sperm acrosomal exocytosis induced by N-acetylglucosaminyl neoglycoprotein

The effect of peritoneal fluid (PF) on the human sperm acrosome reaction (AR) was tested. Sperm was pre-incubated with PF and the AR was induced by calcium ionophore A23187 and a neoglycoprotein bearing N-acetylglycosamine residues (NGP). The AR induced by calcium ionophore was inhibited 40 percent by PF from controls (PFc) and 50 percent by PF from the endometriosis (PFe) group, but not by PF from infertile patients without endometriosis (PFi). No significant differences were found in the spontaneous AR. When the AR was induced by NGP, pre-incubation with PFc reduced (60 percent) the percentage of AR, while PFe and PFi caused no significant differences. The average rates of acrosome reactions obtained in control, NGP- and ionophore-treated sperm showed that NGP-induced exocytosis differed significantly between the PFc (11 percent) and PFe/PFi groups (17 percent), and the ionophore-induced AR was higher for PFi (33 percent) than PFc/PFe (25 percent). The incidence of the NGP-induced AR was reduced in the first hour of pre-incubation with PFc and remained nearly constant throughout 4 h of incubation. The present data indicate that PF possesses a protective factor which prevents premature AR

Purification and characterization of beta-N-acetylhexosaminidase from the phytopathogenic fungus Bipolaris sorokiniana.

N-acetylhexosaminidase (HEX) from the phytopathogenic fungus Bipolaris sorokiniana was isolated and characterized. The production of HEX by B. sorokiniana was not altered by growing on different carbon sources. Enzyme purification was carried out by sequential liquid chromatography on Sephacryl S-200 HR, and p-aminobenzyl-2-acetamido-2-deoxy-beta-D-thioglucopyranoside agarose. The purification was about 70-fold, with a yield of 41%, determined with p-nitrophenyl-N-acetylglucosaminide as substrate. The enzyme had pH and temperature optima of 4.5 and 55 degrees C, respectively. The molecular weight of non-denatured enzyme was estimated as 120,000 Da by gel filtration chromatography, and about 55,000 Da by SDS-PAGE. The fungal HEX had glycosylated residues as evidenced by binding to Concanavalin-A. Bipolaris sorokiniana enzyme was also active with p-nitrophenyl-chitobioside and p-nitrophenyl-N-acetylgalactosaminide as substrates.