Natural killer activity in a medium-term multi-organ bioassay for carcinogenesis

Natural killer (NK) cell activity was evaluated after the initiation and promotion steps in a medium-term multi-organ bioassay for carcinogenesis. NK cell activity was assessed in vitro by Cr-51 release assay at the 4th and 30th weeks of the experiment. Male Wistar rats were sequentially initiated with N-diethylnitrosamine (DEN i.p.), N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN drinking water), N-methyl-N-nitrosourea (MNU i.p.), dihydroxy-di-N-propylnitrosamine (DHPN drinking water) and N,N'-dimethylhydrazine (DMH s.c.) at subcarcinogenic doses for 4 weeks (DMBDD initiation). One group was evaluated at the 4th week and the other was maintained without any further treatment until the 30th week. Two initiated groups were exposed through the diet to 2-acetylaminofluorene (2-AAF) or phenobarbital (PB), from the 6th until the 30th week, Five additional groups were studied to evaluate the effects of each initiator on NK activity. All groups submitted to initiation only, initiation plus promotion, or promotion only, developed significantly more preneoplastic lesions than the untreated control group. The main target organs for tumor development in the initiated animals n ere the liver and the colon, irrespective of treatment with 2-AAF or PB. NK cell activity was not affected bal exposure to genotoxic carcinogens after initiation, at the 4th week. Treatments only with PB or 2-AAF did not change NK cell activity, However, decreased NK cell activity was registered in the group only initiated with DMBDD and in the group given DMBDD+2-AAF. This late depression of NK cell activity at the 30th week could be related to the production of suppressing molecules by the tumor cells.

Enhanced natural killer activity and production of pro-inflammatory cytokines in mice selected for high acute inflammatory response (AIRmax)

Strains of mice with maximal and minimal acute inflammatory responsiveness (AIRmax and AIRmin, respectively) were developed through selective breeding based on their high- or low-acute inflammatory responsiveness. Previous reports have shown that AIRmax mice are more resistant to the development of a variety of tumours than AIRmin mice, including spontaneous metastasis of murine melanoma. Natural killer activity is involved in immunosurveillance against tumour development, so we analysed the number and activity of natural killer cells (CD49b(+)), T-lymphocyte subsets and in vitro cytokine production by spleen cells of normal AIRmax and AIRmin mice. Analysis of lymphocyte subsets by flow cytometry showed that AIRmax mice had a higher relative number of CD49b(+) cells than AIRmin mice, as well as cytolytic activity against Yac.1 target cells. The number of CD3(+) CD8(+) cells was also higher in AIRmax mice. These findings were associated with the ability of spleen cells from AIRmax mice in vitro to produce higher levels of the pro-inflammatory cytokines tumour necrosis factor-alpha, interleukin-12p40 and interferon-gamma but not the anti-inflammatory interleukin-10. Taken together, our data suggest that the selective breeding to achieve the AIRmax and AIRmin strains was able to polarize the genes associated with cytotoxic activity, which can be responsible for the antitumour resistance observed in AIRmax mice.

Natural killer activity in mice infected with rabies virus and submitted to P-acnes (Propionibacterium acnes) as immunomodulator

The natural killer (NK) activity and lethality were evaluated in swiss mice experimentally infected with street rabies virus and submitted to immunomodulation by P. acnes (formerly Corynebacterium parvum). The infected animals were sacrificed at different times and spleen non-adherent cells were obtained through ficoll-hypaque gradient and depletion of glass-adherent cells. Immunosuppression was observed in rabies virus infected mice correlated with lower NK activity in clinically ill animals. Higher NK activity and percentual of survival were observed in the group submitted to P. acnes. The increased survival correlated with higher NK activity induced by P. acnes suggests a protective role of this natural barrier against rabies virus infection in mice. (C) 2000 Elsevier B.V. Ltd. All rights reserved.

Natural killer activity in the experimental privational rickets

To study the 'in vivo' importance of vitamin D on the natural killer (NK) activity, rats were submitted to privational rickets induced by a diet deficient in vitamin D and phosphorus (D-P-). Thirty days after the beginning of treatment the animals showed low body weight, changes in the bone development, and decreased levels of 25-hydroxyvitamin D-3 (25-OH D-3). NK activity, evaluated using a cytotoxicity assay against Cr-51-labeled Yac.1 target cells, was not modified by the rickets-inducing treatment during the first 30 days. Following a long-term treatment (60 days) the rachitic rats (D-P-) exhibited higher NK activity than control animals (D + P +) (P < 0.05). on the other hand, D - P + animals showed higher cytotoxic activity than D - P - and D + P + groups. Feed replacement to the rachitic rats by a complete diet (D - P - /D + P +) led to a partial recuperation of growth, bone development, and 25-OH D-3 scrum. levels. The NK activity was also influenced by vitamin D intake, decreasing after treatment. (C) 2002 Elsevier B.V. B.V. All rights reserved.

Increased natural killer activity does not prevent progression of experimental kala-azar

Kala-azar is the visceral form of leishmaniasis and it is caused by intracellular parasites from the complex Leishmania donovani. Golden hamster (Mesocricetus auratus) infected with Leishmania donovani develop a disease very similar to human Kala-azar. There is conspicuous hipergammaglobulinaemia and their T cells do not respond to stimulation with parasite antigens. We used this experimental model to evaluate the natural killer (NK) activity during the initial phase of the disease. Outbred hamsters infected by intravenous route with 5.106 amastigotes of L. donovani 1S showed a concurrent increase in the spleen weight and in the spleen cell number. Using the single cell assay we detected a significant increase in the percentage of NK effector cells on the 4th day of infection. Imprints from spleen and liver showed at days 14 and 28 a significant increase in the parasite burden. These results show that the increased NK activity in the beginning of the infection was not able to restrain the progression of the disease in this experimental model.

Natural killer activity in a medium-term multi-organ bioassay for carcinogenesis

Natural killer (NK) cell activity was evaluated after the initiation and promotion steps in a medium-term multi-organ bioassay for carcinogenesis. NK cell activity was assessed in vitro by Cr51 release assay at the 4th and 30th weeks of the experiment. Male Wistar rats were sequentially initiated with N-diethylnitrosamine (DEN i.p.), N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN drinking water), N-methyl-N-nitrosourea (MNU i.p.), dihydroxy-di-N-propylnitrosamine (DHPN drinking water) and N,N'-dimethylhydrazine (DMH s.c.) at subcarcinogenic doses for 4 weeks (DMBDD initiation). One group was evaluated at the 4th week and the other was maintained without any further treatment until the 30th week. Two initiated groups were exposed through the diet to 2-acetylaminofluorene (2-AAF) or phenobarbital (PB), from the 6th until the 30th week. Five additional groups were studied to evaluate the effects of each initiator on NK activity. All groups submitted to initiation only, initiation plus promotion, or promotion only, developed significantly more preneoplastic lesions than the untreated control group. The main target organs for tumor development in the initiated animals were the liver and the colon, irrespective of treatment with 2-AAF or PB. NK cell activity was not affected by exposure to genotoxic carcinogens after initiation, at the 4th week. Treatments only with PB or 2-AAF did not change NK cell activity. However, decreased NK cell activity was registered in the group only initiated with DMBDD and in the group given DMBDD+2-AAF. This late depression of NK cell activity at the 30th week could be related to the production of suppressing molecules by the tumor cells.

Lack of Chemopreventive Activity of Agaricus blazei Mushroom on the Development of 1,2-Dimethylhydrazine-Induced Colonic Aberrant Crypt Foci in Rats

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

Effects of extracts from Brazilian sun-mushroom (Agaricus blazei) on the NK activity and lymphoproliferative responsiveness of Ehrlich tumor-bearing mice

Agaricus blazei Murrill, is an edible and medicinal mushroom which is popularly consumed due to its antitumoral properties. The immunomodulatory effects of methanol (METH), dichloromethane (DM) and n-hexane (HEX) extracts of this mushroom were evaluated in Ehrlich tumor-bearing mice. Subcutaneous inoculation of Ehrlich tumor cells inhibited the natural killer (NK) activity of spleen cells (specific lysis = 6.18 +/- 2.56%) compared with normal mice (17.59 +/- 7.77%). Treatment of tumor-bearing mice with the extracts for 10 days restored the natural killer activity against Yac-1 target cells and the best results were observed by treatment with the HEX extract (21.48 +/- 15.26%). Treatment of the animals with the HEX extract for 10 days was also able to stimulate the mitogen-induced lymphoproliferative activity of spleen cells. Thirty days after the treatment, all groups presented low proliferative activity. Specific antibody production was observed to be higher in the groups treated with the DM or METH extract 30 days after the treatment. Analysis of the 3 extracts by gas chromatography mass spectrum (GCMS) and magnetic nuclear resonance (MNR) showed that the HEX extract contains mainly sugar and fatty acids and that the METH extract also contains sugar and possibly amino acids. (C) 2004 Elsevier Ltd. All rights reserved.

Purification of Candida guilliermondii and Pichia ohmeri killer toxin as an active agent against Penicillium expansum

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Occurrence of killer yeasts in leaf-cutting ant nests

Killer activity was screened in 99 yeast strains isolated from the nests of the leaf-cutting ant Atta sexdens against 6 standard sensitive strains, as well as against each other. Among this yeast community killer activity was widespread since 77 strains (78 %) were able to kill or inhibit the growth of at least one standard strain or nest strain. Toxin production was observed in representatives of all the studied genera including Aureobasidium, Rhodotorula, Tremella and Trichosporon, whose killer activity has not yet been described.

Evaluation of the biological control by the yeast Torulaspora globosa against Colletotrichum sublineolum in sorghum

The yeasts are microorganisms with great potential for biotechnological applications in diverse areas. The biological control of phytopathogens by yeasts has showed satisfactory results under laboratory conditions, and it has already produced commercial formulations. With this as focus, this work aims to perform in vitro and in vivo evaluations of the action of a Torulaspora globosa yeast strain (1S112), isolated from sugarcane rhizosphere, against the phytopathogenic mold Colletotrichum sublineolum, the causative agent of anthracnose in sorghum. In vitro experiments included the antagonism test in Petri dishes with morphological hyphal evaluation; yeast killer activity; siderophore, volatile compound and hydrolytic enzyme production. In vivo experiments were conducted in greenhouse conditions with a sorghum variety susceptible to C. sublineolum by evaluating the anthracnose disease for 6 weeks. The results indicated that the yeast strain significantly controlled the fungal growth, either in vitro or in vivo. The strain of T. globosa exhibited killer activity against two sensitive strains, which is a novel capacity for this species. The yeast did not produce siderophores, volatile compounds or hydrolytic enzymes, although it has reduced the mycelial growth, resulting in hyphal deformities but not cell death. The yeast controlled the anthracnose disease in sorghum, either inoculated before or after the fungal spores, suggesting that the competition for space and nutrients to dominate the mold and killer toxin production, altering the hyphal morphology, are mechanisms utilized by the yeast in the biocontrol.

Bioprospection of yeasts as biocontrol agents against phytopathogenic molds

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Immunomodulatory activities associated with beta-glucan derived from Saccharomyces cerevisiae

In this study we investigated the effect of beta-glucan derived from Saccharomyces cerevisiae on fungicidal activity, cytokine production and natural killer activity. Spleen and peritoneal cells from female C57BL/6 mice, previously injected (24 or 48 h) with 20 or 100 mu g of glucan by i.p. route, were assayed. In vivo mu-glucan administration primed spleen cells for a higher production of IL-12 and TNF-alpha when S. aureus was used as a stimulus. In addition, beta-glucan increased NK spleen cells activity against YAC target cells. Some immunomodulatory activities not yet described for beta-glucan were observed in this work.

Immunomodulatory activities associated with β-glucan derived from Saccharomyces cerevisiae

In this study we investigated the effect of β-glucan derived from Saccharomyces cerevisiae on fungicidal activity, cytokine production and natural killer activity. Spleen and peritoneal cells from female C57BL/6 mice, previously injected (24 or 48 h) with 20 or 100 μg of glucan by i.p. route, were assayed. In vivo β-glucan administration primed spleen cells for a higher production of IL-12 and TNF-α when S. aureus was used as a stimulus. In addition, β-glucan increased NK spleen cells activity against YAC target cells. Some immunomodulatory activities not yet described for β-glucan were observed in this work. © 2005 Institute of Physiology, Academy of Sciences of the Czech Republic.

Estudo dos mecanismos de ação de leveduras envolvidos no biocontrole de doenças de pós-colheita em citros

Pós-graduação em Microbiologia Agropecuária - FCAV