Claudia de Mello Bertoncheli

A method for isolation of rat lymphocyte-rich mononuclear cells from lung tissue useful for determination of nucleoside triphosphate diphosphohydrolase activity

Methods for the isolation of peripheral blood mononuclear cells (PBMCs) and human lung mononuclear cells (LMCs) have been proposed previously. This study describes a method that allows the separation of lymphocyte-rich LMCs from rats. Trypan blue was applied to determine cell viability. White blood cell and differential cell counts were also performed. Relationships between nucleoside triphosphate diphosphohydrolase (NTPDase, EC 3.6.1.5) activities expressed in milligrams of protein, millions of cells, and millions of viable cells were examined as linear correlations. The lung tissue yielded 82.46% lymphocytes, 8.6% macrophages, 2.20% monocytes, and 1.27% polymorphonuclear cells (PMNs). In LMCs, a very strong correlation was observed as follows: between NTPDase activity, as determined using ATP or ADP as a substrate, expressed in milligrams of protein and that expressed in millions of cells (r ≥ 0.91), between that expressed in milligrams of protein and that expressed in millions of viable cells (r ≥ 0.91), and between that expressed in millions of cells and that expressed in millions of viable cells (r ≥ 0.98). Based on our results, we affirm that NTPDase activity could be expressed in millions of viable cells, millions of cells, or milligrams of protein.

Lactating and nonlactating rats differ to renal toxicity induced by mercuric chloride: the preventive effect of zinc chloride.

This study evaluated the effects of HgCl2 on renal parameters in nonlactating and lactating rats and their pups, as well as the preventive role of ZnCl2. Rats received 27 mg kg−1 ZnCl2 for five consecutive days and 5 mg kg−1 HgCl2 for five subsequent days (s.c.). A decrease in δ‐aminolevulinic acid dehydratase (δ‐ALA‐D) activity in the blood and an increase in urine protein content in renal weight as well as in blood and urine Hg levels were observed in lactating and nonlactating rats from Sal―Hg and Zn―Hg groups. ZnCl2 prevented partially the δ‐ALA‐D inhibition and the proteinuria in nonlactating rats. Renal Hg levels were increased in all HgCl2 groups, and the ZnCl2 exposure potentiated this effect in lactating rats. Nonlactating rats exposed to HgCl2 exhibited an increase in plasma urea and creatinine levels, δ‐ALA‐D activity inhibition and histopathological alterations (necrosis, atrophic tubules and collagen deposition) in the kidneys. ZnCl2 exposure prevented the biochemical alterations. Hg‐exposed pups showed lower body and renal weight and an increase in the renal Hg levels. In conclusion, mercury‐induced nephrotoxicity differs considerably between lactating and nonlactating rats. Moreover, prior exposure with ZnCl2 may provide protection to individuals who get exposed to mercury occupationally or accidentally. Copyright

Triazenos e atividade antibacteriana

Fifteen triazenes compounds were studied concerning their antibacterial activity by broth microdilution method. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) was determined with E. coli ATCC 25922, Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 27853, Acinetobacter baumannii, Acinetobacter lwoffii, Ralstonia pickettii, Bordetella bronchiseptica, Micrococcus sp., Enterococcus sp., Staphylococcus epidermidis, Staphylococcus saprophyticus, Streptococcus agalactiae, Bacillus cereus, Corynebacterium sp., Rhodococcus sp., Salmonella sp., Serratia marcescens, Morganella morganii, Enterobacter cloacae, Shigella flexneri, Shigella sonnei, Shigella sp., Klebsiella pneumoniae, ESBL Klebsiella oxytoca 14, ESBL Klebsiella pneumoniae 23, ESBL Klebsiella pneumoniae 24, ESBL Klebsiella pneumoniae 25, ESBL Escherichia coli 26, ESBL Klebsiella pneumoniae 27, ESBL Klebsiella pneumoniae 31, ESBL Escherichia coli 32, ESBL Klebsiella pneumoniae 37 e ESBL Escherichia coli 38. The highest effect was evidenced by the compound 1-methyl-3-(p-carboxyphenyl)triazene 1-oxide (2) against Streptococcus agalactiae (MIC = 16 µg/mL and MBC = 32 µg/mL). The compounds 1-phenyl-3-(4-nitrophenyl)triazene-1-oxide (9), 1-(4-nitrophenyl)-3-(4-carboxyphenyl)triazene (10) e 1-(4-acethyl amine phenyl)-3-(4-carboxyphenyl)triazene (11) presented MIC between 32 and 64 µg/mL against S. edipermidis, S. saprophyticus, Rhodococcus sp. and E. cloacae. The compounds 1-methyl-3-phenyltriazene-1-oxide (1) , bis-1,3-(4-acethyl oxime)triazene (3), bis-1,3 (4-acethyl phenyl)triazene (4), 1-(2-fluorophenyl)-3-(4-acethylphenyl)triazene (5), 1,3-(3-hydroxy-diphenyltriazenide)(piridil)(bis-oxo-vanadium) (12), 1-(3-nitrophenyl)-3-phenyltriazene (14), 1-(4-nitrophenyl)-3-benzyltriazene (15) presented MIC of the 128 µg/mL against S. aureus ATCC 25923, P. aeruginosa ATCC 27853, A. lwoffii, Micrococcus sp., S. epidermidis, S. saprophyticus, Corynebacterirum sp., E. cloacae, S. flenneri e S. sonnei. The compounds 1-phenyl-3-(4-acethylphenyl)triazene (6), 1,3-bis-(4-ethoxycarbonylphenyl) triazene (7), and 3-(4-carboxylatephenyl)-1-methyltriazene 1-oxide of potassium tetrahydrate (13) presented MICs equals or highest than 128 µg/mL. The results have demonstrated the potential biological activity of these compounds against Gram-positive and Gram-negative bacteria.

Uma revisão sobre metalo-β-lactamases

A emergencia e disseminacao da resistencia aos antimicrobianos sao problemas de grande importância mundial, particularmente entre patogenos nosocomiais de importância clinica como os bacilos Gram negativos nao fermentadores e membros da familia Enterobacteriaceae. O principal mecanismo de resistencia desses patogenos e a producao de β-lactamases, que sao enzimas que hidrolisam o anel β-lactâmico impedindo assim a acao dos antimicrobianos β-lactâmicos. As β-lactamases foram dividas em quatro classes de acordo com a sua estrutura primaria e podem tambem ser classificadas dentro de dois grupos com base no seu mecanismo catalitico, isto e, serina-β-lactamases (Classes A, C e D) e metalo-β-lactamases (Classe B). As metalo-β-lactamases (MβL) utilizam ions divalentes, comumente zinco, como co-fator para sua atividade catalitica e sao atualmente uma das classes que mais merece destaque, devido a sua capacidade de hidrolisar todos os antimicrobianos β-lactâmicos, incluindo os carbapenens. Estes antimicrobianos sao utilizados no tratamento de infeccoes causadas por bacterias Gram-negativas multirresistentes e conseguem se manter estaveis frente as serina-β-lactamases. A deteccao de microrganismos produtores de MβL tem por finalidade auxiliar a Comissao de Controle de Infeccao Hospitalar (CCIH) na prevencao da disseminacao desse mecanismo de resistencia no ambiente hospitalar e impedir que ele chegue ate a comunidade, bem como enfatizar o uso racional dos antimicrobianos disponiveis para uso clinico, pois, atualmente,ha poucos investimentos da industria farmaceutica na pesquisa de novos agentes antimicrobianos.

Increased NTPDase activity in lymphocytes during experimental sepsis.

We investigated in rats induced to sepsis the activity of ectonucleoside triphosphate diphosphohydrolase (NTPDase; CD39; E.C. 3.6.1.5), an enzyme involved in the modulation of immune responses. After 12 hours of surgery, lymphocytes were isolated from blood and NTPDase activity was determined. It was also performed the histology of kidney, liver, and lung. The results demonstrated an increase in the hydrolysis of adenosine-5′-triphosphate (ATP) (P < 0.01), but no changes regarding adenosine-5′-monophosphate (ADP) hydrolysis (P > 0.05). Histological analysis showed several morphological changes in the septic group, such as vascular congestion, necrosis, and infiltration of mononuclear cells. It is known that the intracellular milieu contains much more ATP nucleotides than the extracellular. In this context, the increased ATPasic activity was probably induced as a dynamic response to clean up the elevated ATP levels resulting from cellular death.

An experimental model of contact dermatitis: Evaluation of the oxidative profile of Wistar rats treated with free and nanoencapsulated clobetasol

Abstract Objective An experimental animal model of contact dermatitis (CD) was used to investigate the effects of free and nanoencapsulated clobetasol propionate on the skin and on the oxidative profile of liver tissue. Methods Female Wistar rats were divided into six groups, each containing eight rats. The first group, control (C), was sensitized with solid vaseline. Group 2, (CD), was sensitized with 5% NiSO4. Groups 3 and 4 were sensitized with 5% NiSO4 and treated with free (FC) and nanoencapsulated (NC) clobetasol (0.42 mg/g), respectively, daily for 5 days. Group 5 was treated with nanoencapsulated clobetasol (0.42 mg/g) on days 1, 3, and 5 (C135) and group 6 received a hydrogel containing empty nanoparticles (NP) daily for 5 days. Thiobarbituric acid reactive substances (TBARS), carbonyl levels, non-protein sulfhydryl groups (NPSH) and catalase activity were measured in liver homogenates. Results A significant increase was observed in the levels of TBARS, NPSH, and catalase activity for the groups CD and NP. Discussion Our results suggest that both NiSO4 sensitization and NP administration induced oxidation of cellular lipids and activated the antioxidant enzyme catalase to protect from this damage. These results also indicated that daily treatment with the free and nanoencapsulated clobetasol, as well as treatment with the nanoencapsulated clobetasol every other day, were able to prevent these redox alterations and protect against histological damage.

Clinical signs and histopathology of brain, spinal cord and muscle of the pelvic limb of rats experimentally infected with Trypanosoma evansi

The aim of this study was to investigate whether rats infected with Trypanosoma evansi had neurological and locomotor signs, as well as histological lesions in the central nervous system (CNS) and pelvic muscles. To carry out this study, 52 rats were used and divided into two groups. The animals in Group A (n=40) were infected with T. evansi, and the rats in Group B (n=12) were used as negative controls (non-infected). Neurological examination was performed at Days 5, 15, 30 and 150 post-infection (PI) with eventual euthanasia of the rats. Samples of brain, spinal cord and skeletal muscle (biceps femoris and gastrocnemius muscles) were collected. The neurological tests evaluated motor capacity, balance and pain sensitivity. At Day 5 PI in Subgroup A1, the rats showed high parasitemia, became apathetic and presented with slow movements and signs of disorientation. After Day 15 PI in Subgroup A2 and Day 30 PI in Subgroup A3, no more clinical abnormalities were observed. Histologically, there was no damage to the CNS in these three subgroups, but within Subgroup A3, mononuclear infiltration of the muscle was observed. Rats chronically infected (Subgroup A4 - Day 150 PI) showed muscle atrophy, walking dysfunction and paralysis of the hind limbs. Mild mononuclear inflammatory infiltrates and perivascular cuffs were observed in the CNS of some of the animals in Subgroup A4. In these rats, severe muscle damage was observed in the skeletal muscle which included atrophy and loss of muscle fibers, multinucleated giant muscle cells, mononuclear myositis, Wallerian degeneration of the innervating fibers and mononuclear inflammatory infiltrate in the perineurium and adipose tissue. Based upon these findings, we conclude that infection by T. evansi in rats leads to muscle damage, which is probably the cause of the paralysis of hind limbs.

Oxidative Stress in the Heart of Rats Infected with Trypanosoma evansi.

This study was conducted to investigate the occurrence of oxidative stress in the heart tissue of rats infected with Trypanosoma evansi. Rats were divided into 2 groups (A and B) with 12 animals each, and further subdivided into 4 subgroups (A1 and A2, 6 animals/each; and B1 and B2, 6 animals/each). Animals in the groups B1 and B2 were subcutaneously inoculated with T. evansi. Thiobarbituric acid reactive substances (TBARS), superoxide dismutase activity (SOD), glutathione S-transferase activity (GST), reduced glutathione activity (GSH), and non-protein thiols (NPSH) in the heart tissue were evaluated. At day 5 and 15 post-infection (PI), an increase in the TBARS levels and a decrease in the SOD activity (P<0.05) were observed. GSH and GST activities were decreased in infected animals at day 15 PI (P<0.05). Considering the proper functioning of the heart, it is possible that the changes in the activity of these enzymes involved in the oxidative stress may be related, at least in part, in the pathophysiology of rats infected with T. evansi.

Trypanosoma evansi: Ca2+ ATPase activity and lipid peroxidation in skeletal muscle from rats experimentally infected

The aim of this study was to evaluate Ca(2+) ATPase activity and the lipid peroxidation in muscles from rats experimentally infected by Trypanosoma evansi and its roles in the muscle pathogenesis in trypanosomosis. Thirty-six rats were divided in two groups. Group A was infected with an isolate from T. evansi and group B was used as a negative control. Group A was divided into three subgroups (A1, A2 and A3), three animals each group, as well as group B (B1, B2 and B3). The collection of samples were performed at days 5 (A1 and B1), 15 (A2 and B2) and 30 (A3 and B3) post-infection (PI) with the purpose of comparison between healthy and infected rats in the course of the disease. The Ca(2+) ATPase enzyme activity was determined in skeletal muscle samples. Muscle tissue lipid peroxidation was determined by TBARS levels, and histopathologically it was investigated a possible damage to the muscle tissue of rats infected with T. evansi. It was observed a significant decrease of Ca(2+) ATPase activity in infected rats compared to not-infected. This enzymatic inhibition was observed at days 5, 15 and 30 PI. A significant increase was observed for TBARS levels in the muscles of infected rats at days 5, 15 and 30 PI. It was not identified any histological alterations for gastrocnemius in rats infected by T. evansi at days 5 and 15 PI. Nevertheless, at day 30 PI it was verified inflammatory infiltrate with mononuclear cells between muscle fibers in three infected rats (50%). T. evansi infections in rats showed a negative correlation between Ca(2+) ATPase and TBARS levels. Based on these results we suggest that the leg weakness and muscle injuries common in infected animals with T. evansi may be related to a reduced activity of Ca(2+) ATPase and oxidative stress.

Perfil de suscetibilidade dos microorganismos isolados de uma unidade de terapia intensiva neonatal

O presente estudo descreve o perfil epidemiologico das infeccoes na unidade de terapia intensiva neonatal (UTIN) no Hospital Universitario de Santa Maria, RS, Brasil, e o perfil de resistencia aos antimicrobianos, no periodo compreendido entre junho de 2004 a julho de 2005. Os dados foram coletados retrospectivamente de 1411 amostras cultivadas nesse periodo. Staphylococcus coagulose negativa (SCN) doi o patogeno mais frequentemente isolado, seguido por Escherichia coli e especies de Candida .