Experimental peritonitis in horses: peritoneal fluid composition

Dezesseis eqüinos adultos foram aleatoriamente divididos em quatro grupos de quatro animais que receberam inoculação intraperitoneal das seguintes suspenções: grupo I, 100×10(7) unidades formadoras de colônias (CFU) de E. coli diluídas em 500ml de solução salina a 0,9%; grupo II, 100×10(7) CFU de Bacteroides fragilis em 500ml de solução salina a 0,9%; grupo III, 100×10(7) CFU de E. coli combinados com 100×10(7) CFU de B. fragilis em 500ml de solução salina a 0,9%; grupo IV, 500ml de solução salina a 0,9%. Observou-se aumento significativo do número de leucócitos no líquido peritoneal quatro horas após as inoculações dos animais dos grupos I e II, e oito horas após as inoculações dos animais do grupo III. A contagem mais elevada foi de 516×10³ leucócitos/mm³. Aumentos significativos nas concentrações de fibrinogênio (1g/dl) e proteína total (9,1%) foram também observados. Eqüinos inoculados com culturas puras, tanto de E. coli quanto de B. fragilis, apresentaram peritonites mais brandas e autolimitantes, enquanto que eqüinos inoculados com associação das duas bactérias apresentaram alterações laboratoriais com maior intensidade e duração.

Endotoxemia por lipopolissacarídeo de Escherichia coli, em eqüinos: efeitos de antiinflamatórios nas concentrações sérica e peritoneal do fator de necrose tumoral alfa (TNF-alfa)

Avaliou-se a inibição da produção do fator de necrose tumoral alfa (TNF-alfa) devido ao pré-tratamento com antiinflamatório esteroidal (dexametasona) e não esteroidal (diclofenaco sódico) em eqüinos com endotoxemia induzida experimentalmente. Foram utilizados 15 cavalos machos não castrados, distribuídos em três grupos de cinco animais: controle (C), diclofenaco sódico (DS) e dexametasona (DM). A endotoxemia subletal foi induzida pela infusão intravenosa (IV) de 0,1mg/kg/pv de lipopolissacarídeo (LPS) de Escherichia coli 055:B5, administrado em 250ml de solução estéril de cloreto de sódio a 0,9%, durante 15min. Os cavalos do grupo-controle foram tratados com solução de cloreto de sódio a 9% IV. Nos animais do grupo DS, administraram-se, por via oral, 2,2mg/kg de diclofenaco sódico e, nos do grupo DM, 1,1mg/kg de dexametasona IV, respectivamente, 60 e 30min antes da infusão da endotoxina. Mensurou-se, por meio de ensaio de toxicidade com células da linhagem L929, a concentração de TNF-alfa no soro e no líquido peritoneal às 0, 1¼, 3 e 6 horas após injeção do LPS. No grupo-controle, observou-se aumento significativo de TNF-alfa sérico, em relação ao valor basal e aos grupos DS e DM, 1,15 horas após a indução da endotoxemia. No líquido peritoneal, as concentrações observadas estavam abaixo daquelas da curva padrão de TNF-alfa, não havendo diferença entre os grupos (P>0,05).

Morphological alteration of peritoneal mast cells and macrophages in the mouse peritoneal cavity during the early phases of an allergic inflammatory reaction

We investigated the presence of mast cell granules in macrophages following an in vivo model of an allergic reaction. Injection of ovalbumin (100 mug) into the peritoneal cavity of sensitised mice produced a rapid (within 2 h) influx of neutrophils followed by a slower (after >4 h) eosinophil migration. Ovalbumin treatment induced a high incidence (similar to 50%) of mast cell degranulation compared to control phosphated-buffered saline-treated mice. The majority (similar to 90%) of peritoneal macrophages contained mast cell granules as early as 2 It post-ovalbumin, with lower values at later time-points, as determined by staining with Toluidine blue and Berberine sulphate. This was confirmed by electron microscopy which enabled us to identify the complex mast cell granule sub-structural components in macrophage phagosomes. In conclusion, we used histochemical and ultrastructural analyses to show that mast cell granules become internalised with macrophages during the early stages of an experimental allergic reaction. (C) 2001 Academic Press.

Exame do fluido peritoneal e hemograma de eqüinos submetidos à laparotomia e infusão intraperitoneal de carboximetilcelulose

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Características do líquido peritoneal de eqüinos normais após punção cecal percutânea

Foram avaliados 10 eqüinos da raça Manga Larga, machos, inteiros clinicamente sadios, submetidos à punção cecal percutânea. Analisou-se a resposta clínica, celular, bioquímica e microbiológica do líquido peritoneal por um período de 24 horas após a punção cecal, nos tempos T0, T6, T12 e T24. Foi observada elevação na freqüência respiratória em T6 e na temperatura retal entre T6 e T12. Decorridas 24 horas da punção cecal, ocorreu aumento na concentração de proteínas totais do líquido peritoneal e na atividade da fosfatase alcalina. Tanto a atividade da ALT quanto os níveis de hemoglobina apresentaram diminuição em T6. Não foram registradas alterações na celularidade do plasma ou do líquido peritoneal e obteve-se resultado negativo para a cultura microbiológica do líquido. Considerando a inexistência de efeitos adversos, além das poucas alterações descritas, conclui-se que a punção cecal percutânea é um procedimento seguro e factível, se praticada criteriosamente.

Immunocytochemical localization of heparin in secretory granules of rat peritoneal mast cells using a monoclonal anti-heparin antibody (ST-1)

We performed immunogold labeling with an ST-1 monoclonal antibody (IgM), specific for intact heparin, to define the subcellular localization of heparin in mast cells. Rat peritoneal mast cells were fixed by a modified Karnovsky method and embedded in Araldite. Ultrathin sections were first treated with sodium periodate and then sequentially incubated with MAb ST-1, rabbit anti-mouse IgM, and protein A-gold. By transmission electron microscopy, gold particles were localized inside cytoplasmic granules of peritoneal mast cells. In contrast, with the same procedure, no labeling was observed in mast cells from rat intestinal mucosa. Control sections of rat peritoneal or intestinal mucosa mast Mast cells cells treated with an irrelevant MAb (IgM) did not show any labeling. Treatment with nitrous Heparin acid abolished the reactivity of MAb ST-1 with peritoneal mast cells. These results Granules show that different mast cells can be identified regarding their heparin content by immunochemical procedures using MAb ST-1.

Evaluation of nanoparticles loaded with benzopsoralen in rat peritoneal exudate cells

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Inhibition of hydrogen peroxide, nitric oxide and TNF-alpha production in peritoneal macrophages by ethyl acetate fraction from Alchornea glandulosa

The effects of Alchornea glandulosa ethyl acetate fraction (AGF) on hydrogen peroxide (H2O2), nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) production in peritoneal macrophages activated with lipopolysaccharide (LPS) or phorbol myristate acetate (PMA) were investigated. Analysis by thin layer chromatography (TLC) of AGF showed several constituents, including flavonoids, which may have anti-inflammatory activity. Inhibitory effects of the fraction in H2O2 and NO production ranged from 8.59 +/- 7.84% to 70.56 +/- 4.16% and from 16.06 +/- 3.65% to 38.73 +/- 3.90%, respectively. The TNF-alpha production was only partially inhibited in the tested concentrations (12.21 +/- 6.23%-15.16 +/- 0.96%). According to these results, it is suggested that AGF has anti-inflammatory activity. This medicinal plant may have therapeutic potential in the control of inflammatory disorders.

Phagocytosis of PLGA microparticles in rat peritoneal exudate cells: A time-dependent study

With the purpose of enhancing the efficacy of microparticle-encapsulated therapeutic agents, in this study we evaluated the phagocytic ability of rat peritoneal exudate cells and the preferential location of poly(D,L-lactide-co-glycolic acid) (PLGA) microparticles inside these cells. The microparticles used were produced by a solvent evaporation method and were characterized by dynamic light scattering (DLS), transmission electron microscopy (TEM), and scanning electron microscopy (SEM). Size distribution analysis using DLS and SEM showed that the particles were spherical, with diameters falling between 0.5 and 1.5 mu m. Results from cell adhesion by SEM assay, indicated that the PLGA microparticles are not toxic to cells and do not cause any distinct damage to them as confirmed by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay. Among the large variety of cell populations found in the peritoneal exudates (neutrophils, eosinophils, monocytes, and macrophages), TEM showed that only the latter phagocytosed PLGA microparticles, in a time-dependent manner. The results obtained indicate that the microparticles studied show merits as possible carriers of drugs for intracellular delivery.

Evaluation of nanoparticles loaded with benzopsoralen in rat peritoneal exudate cells

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

EVALUATION OF PERITONEAL-FLUID IN HORSES WITH EXPERIMENTAL ENDOTOXEMIA

Clinical parameters and biochemical and cellular changes in the plasma and peritoneal fluid were evaluated in horses after i.v. injection of a sub-lethal dose (50 ng/kg) of E. coli endotoxin. A significant decrease in the number of neutrophils and lymphocytes occurred in the blood 1h 15 min and 3 hours after injection of endotoxin; body temperature was increased significantly at the 3rd hour. No changes were detected in the total number of white blood cells in the peritoneal fluid. No significant differences in biochemical values were detected in either plasma or peritoneal fluid. Endotoxemia caused an alteration in blood cellularity, without effecting the peritoneal cellular population.

Determination of trace elements in serum samples by isotope dilution inductively coupled plasma mass spectrometry using on-line dialysis

An on-line dialysis flow system coupled to inductively coupled plasma mass spectrometry to determine trace elements in serum samples by isotope dilution is presented. Isotope dilution was performed on samples incubated with enriched Cu-65, Zn-66, Se-77 and Pb-206 for 24 h at 36degreesC prior to dialysis to quantified total element concentrations. The sample and acceptor solutions flowed through the dialysis unit with cellophane membrane placed in between the compartments. The serum sample (1 mL) was left to recycle in a closed path while the acceptor solution was continuously pumped along the dialyzer channel and through a cationic AG50W X-8 resin column. After 10 min, around 70% of Na, K and Cl migrate from the sample. Three replicate injections of 0.1 mL were performed for the clean sample after each separation step. The on-line coupling of the dialyzer to ICP-MS allowed isotope dilution for total element determination either in the cleaned sample or by eluting the cations retained in the resin to be carried out. Results demonstrated no matrix effects from alkaline elements or spectral interference from ArNa+ on Cu-63, ArCl+ on Se-77 and (SO2+)-S-34 on Zn-66. The precision of isotope ratio measurements for Cu and Zn was around 1% and for Se and Pb was around 2.5%. The values found for the reference serum sample IMEP-17 were in good agreement with the certified values for Cu, Zn and Se.

Different inflammatory stimuli in the footpad of mice influence the kinetics of resident peritoneal cells

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Peritoneal effusion in a dog secondary to visceral mast cell tumor: A case report

BACKGROUND: Mast cell tumor, one of the most common skin tumors in dogs, may also be found in visceral sites (mainly spleen and liver). When a visceral mast cell tumor is present, neoplastic mast cells may be found in any effusion secondary to the tumor. Therefore, the diagnosis may be made by cytologic analysis of the effusion. CASE: An 8-year-old, spayed, female Siberian husky presented with a peritoneal effusion secondary to a visceral mast cell tumor. Seven months earlier, the dog had presented with a cutaneous nodule diagnosed as a well-differentiated mast cell tumor. The peritoneal fluid was classified as a transudate. Numerous neoplastic mast cells were found in the effusion. Although the mast cell tumor presented with characteristics of the well-differentiated tumor, its biologic behavior was that of a malignant tumor. CONCLUSION: Care should be taken to evaluate the prognosis of mast cell tumors in dogs since their biologic behavior is extremely variable.