RESUMO
BACKGROUND: Despite the use of a sterile technique and the administration of prophylactic antibiotics during surgical procedures, mesh infection continues to complicate the use of biomaterials. The purpose of this study was to compare the susceptibility to infection of prosthetic biomaterials in a live-animal model. METHODS: The following seven prosthetic mesh biomaterials were used in this study. Expanded polytetrafluoroethylene (ePTFE) with silver/chlorhexidine (DM+), ePTFE (DM), porcine intestinal submucosa (S), polypropylene (M), ePTFE/polypropylene (X), hyaluronate/carboxymethylcellulose/polypropylene (SM), and human acellular dermal matrix (A). Lewis rats (n = 108) underwent creation of a single ventral hernia; 105 of them were repaired with a different mesh (2-cm2 piece). Twelve pieces of each mesh were inoculated at the time of hernia repair with 10(8) Staphylococcus aureus (n = 84). Three pieces of each mesh were placed without bacterial inoculation (n = 21). In three animals, no mesh was placed; instead, the peritoneum of the hernia defect was inoculated (n = 3). After 5 days, the animals were killed and the mesh was explanted (peritoneum for the nonmesh control). The mesh was vortex-washed and incubated in tryptic soy broth. Bacterial counts were determined using serial dilutions and spot plates and quantified in colony-forming units (CFU) per square centimeter of mesh present in the vortex wash fluid (wash count) and the soy broth (broth count). Data are presented as the mean log(10), with analysis of variance (ANOVA) and Tukey's test used to determine significance (p < 0.05). RESULTS: The DM+ material had no detectable live bacteria in the wash or broth counts in 10 of 12 tested samples (p = 0.05). Of the samples that showed bacterial growth, the peritoneum control group had a lower wash count than A (p = 0.05) and the lowest broth count of all the materials except for DM+ (p = 0.05). In addition, SM had a significantly lower wash count than A (p = 0.05), with no broth count difference. In regard to wash and broth counts, DM, M, X, SM, S, and A were no different (p = NS). CONCLUSIONS: The DM+ material was the least susceptible to infection. Impregnation with silver/chlorhexidine killed the inoculated bacteria, preventing their proliferation on the mesh surface. Other than DM+, native peritoneal tissue appears to be the least susceptible to infection. Silver/chlorhexidine appears to be an effective bactericidal agent for use with mesh biomaterials.
Assuntos
Infecções Bacterianas/epidemiologia , Infecções Bacterianas/etiologia , Materiais Biocompatíveis , Infecções Relacionadas à Prótese/epidemiologia , Infecções Relacionadas à Prótese/etiologia , Telas Cirúrgicas/efeitos adversos , Animais , Masculino , Ratos , Ratos Endogâmicos LewRESUMO
Lumpectomy coupled with radiation therapy and/or chemotherapy is commonly used to treat breast cancer patients. We are developing an enhanced thermal IR imaging technique that has the potential to provide real-time imaging to guide tissue excision during a lumpectomy by delineating tumor margins. This enhanced thermal imaging method is a combination of IR imaging (8 to 10 µm ) and selective heating of blood (â¼0.5°C ) relative to surrounding water-rich tissue using LED sources at low powers. Postacquisition processing of these images highlights temporal changes in temperature and the presence of vascular structures. In this study, fluorescent, standard thermal, and enhanced thermal imaging modalities, as well as physical caliper measurements, were used to monitor breast cancer tumor volumes over a 30-day study period in 19 mice implanted with 4T1-RFP tumor cells. Tumor volumes calculated from fluorescent imaging follow an exponential growth curve for the first 22 days of the study. Cell necrosis affected the tumor volume estimates based on the fluorescent images after day 22. The tumor volumes estimated from enhanced thermal imaging, standard thermal imaging, and caliper measurements all show exponential growth over the entire study period. A strong correlation was found between tumor volumes estimated using fluorescent imaging, standard IR imaging, and caliper measurements with enhanced thermal imaging, indicating that enhanced thermal imaging monitors tumor growth. Further, the enhanced IR images reveal a corona of bright emission along the edges of the tumor masses associated with the tumor margin. In the future, this IR technique might be used to estimate tumor margins in real time during surgical procedures.
Assuntos
Neoplasias da Mama/patologia , Interpretação de Imagem Assistida por Computador/métodos , Raios Infravermelhos , Neovascularização Patológica/patologia , Termografia/métodos , Animais , Neoplasias da Mama/complicações , Linhagem Celular Tumoral , Feminino , Aumento da Imagem/métodos , Camundongos , Camundongos Endogâmicos BALB C , Invasividade Neoplásica , Neovascularização Patológica/complicações , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
Physical exercise and diet changes have been shown to affect immune parameters, and similar effects are also induced by the administration of a nonmetabolizable glucose analog, 2-deoxy-D-glucose (2-DG). The present study was designed to characterize the effects of glucoprivation induced by 2-DG administration on concentrations of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and IL-6 in the blood and interferon-gamma (IFN-gamma), IL-2, and IL-4 in vitro production by partially purified T splenocytes in BDF1 mice. Mice (n = 8 per group) were injected intraperitoneally one or three times with 0, 500, 750, or 1000 mg/kg of 2-DG, and blood and spleens were collected 2 h after the last injection. Partially purified T splenocytes were cultured 24 h in the presence of concanavalin A (ConA). A significant increase in the corticosterone levels with the amount of 2-DG injected was observed after one or three injections (p<0.05). The amount of 2-DG injected was associated with an increase in TNF-alpha, IL-1beta, and IL-6 concentrations in the blood of mice after one or three injections of 2-DG (p<0.05). A significant decrease in in vitro proliferation of partially purified splenocytes in the presence of ConA was associated with a decrease in IFN-gamma production in the culture supernatants and an increase in IL-1 receptor expression on the cell surface (p<0.05).
Assuntos
Citocinas/biossíntese , Desoxiglucose/farmacologia , Animais , Glicemia/metabolismo , Divisão Celular/efeitos dos fármacos , Corticosterona/sangue , Citocinas/sangue , Desoxiglucose/administração & dosagem , Feminino , Técnicas In Vitro , Interferon gama/biossíntese , Interleucina-1/sangue , Interleucina-2/biossíntese , Interleucina-4/biossíntese , Interleucina-6/sangue , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Receptores de Interleucina-1/metabolismo , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/biossínteseRESUMO
The expression of the beta 3 tubulin gene is regulated, at the transcriptional level, by the steroid hormone ecdysone, in Drosophila Kc cells. Using a transient expression assay, we show that 360 bp from the first intron of the beta 3 tubulin gene, associated with the 5' flanking sequences, are essential to confer ecdysone inducibility on a minimum promoter driving the chloramphenicol acetyl transferase (CAT) gene. The 5' flanking region contains ecdysone-independent cis-positive elements located in proximity to the promoter. Deletion analysis of the 360 bp intronic region reveals that a fragment of 57 bp is crucial for the ecdysone response of the beta 3 tubulin gene. This fragment contains 5'-TGA(A/C)C-3' motifs homologous to ecdysone responsive elements (EcRE) half sites. Band shift assays show that this 57-bp fragment is bound by three specific complexes. One of them appears to be involved in the level of the ecdysone response.
Assuntos
Drosophila melanogaster/genética , Ecdisona/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Genes de Insetos , Íntrons , Sequências Reguladoras de Ácido Nucleico , Tubulina (Proteína)/genética , Animais , Sequência de Bases , Linhagem Celular , Regulação da Expressão Gênica/genética , Dados de Sequência Molecular , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética , Transcrição Gênica/efeitos dos fármacos , Tubulina (Proteína)/biossínteseRESUMO
We have studied the transcriptional regulation of the beta 3 tubulin gene by the steroid hormone 20-hydroxyecdysone (20-E) in Drosophila Kc cells. A series of hybrid genes, with different fragments of the beta 3 tubulin gene driving the bacterial chloramphenicol acetyl transferase (CAT) gene were constructed. The promoter activity was assayed after transient expression in Kc cells, in the presence and the absence of 20-E. Constructs with 0.91 kb upstream from the transcription start site and 360 bp from the first large intron allowed the hormonal regulation, i.e. a repression in the absence of 20-E and a derepression-activation in the presence of the hormone. This 360 bp fragment contains several enhancers and silencer(s) sequences. The regulation of the expression of the beta 3 tubulin gene results from the combined activity of all the positive and negative regulatory sequences of the first intron, and a dialogue with the promoter sequences. The nucleotide sequence of this intronic regulatory-fragment has been established and we have identified several EcRE (ecdysone responsive element) consensus sequences.
Assuntos
Drosophila melanogaster/genética , Ecdisterona/farmacologia , Tubulina (Proteína)/genética , Animais , Sequência de Bases , Sequência Consenso , DNA/genética , Elementos Facilitadores Genéticos , Regulação da Expressão Gênica/efeitos dos fármacos , Íntrons , Dados de Sequência Molecular , Receptores de Esteroides/genética , Transcrição Gênica/efeitos dos fármacosRESUMO
Melanoma inhibitory activity (MIA) was originally identified as an 11 kDa protein secreted from malignant melanoma cells. We have shown that MIA is strongly expressed in melanoma and melanoma cell lines but not in melanocytes and normal skin. We also observed that MIA mRNA expression correlates with progressive malignancy of melanocytic tumors. Measuring MIA in serum or plasma by a sensitive and quantitative ELISA and investigating the potential of MIA serum levels as a novel marker for malignant melanomas showed that the protein can be used to monitor therapy and follow-up. The present study measured the variations in blood concentrations of MIA in 84 patients with stage II-IV melanoma by ELISA. Patients treated with repeated injections of a polyvalent melanoma vaccine (PMV), interferon-alpha-2b (IFN-alpha 2b) or interleukin-2 (IL-2) were followed during treatment duration. Before treatment, patients treated with PMV or IFN-alpha 2b had comparable low MIA concentrations, whereas most IL-2-treated patients had higher MIA levels. At the end of treatment, MIA concentrations were higher in patients with progressive disease (PD) than in patients with no clinical evidence of melanoma (NPD) for PMV, IFN-alpha 2b or IL-2 therapy (3.7 +/- 0.2 vs 11.5 +/- 5.4 ng/ml, 3.8 +/- 0.2 vs 8.3 +/- 1.7 ng/ml, and 2.3 +/- 0.7 vs 20.2 +/- 7.4 ng/ml, respectively, p < 0.05). In contrast to the stable MIA concentrations measured in NPD patients, significant increase in MIA levels were observed in PD patients over time regardless of treatment. For PMV- and IFN-alpha 2b-treated patients, a rise in MIA levels occurred significantly earlier than clinical diagnosis of melanoma recurrence. In conclusion, our data suggest that quantitation of MIA serum levels may be used for detection of both clinically apparent and non-apparent metastatic melanoma disease and for monitoring therapy.
Assuntos
Biomarcadores Tumorais/sangue , Melanoma/sangue , Proteínas de Neoplasias/sangue , Neoplasias Cutâneas/sangue , Animais , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos/uso terapêutico , Vacinas Anticâncer/uso terapêutico , Cricetinae , Progressão da Doença , Ensaio de Imunoadsorção Enzimática , Proteínas da Matriz Extracelular , Feminino , Seguimentos , Humanos , Interferon alfa-2 , Interferon-alfa/uso terapêutico , Interleucina-2/uso terapêutico , Masculino , Melanoma/diagnóstico , Melanoma/terapia , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/sangue , Estadiamento de Neoplasias , Valor Preditivo dos Testes , Estudos Prospectivos , Proteínas Recombinantes , Neoplasias Cutâneas/diagnóstico , Neoplasias Cutâneas/terapia , Vacinas CombinadasRESUMO
As an adjuvant therapy for patients with high risk of recurrent melanoma, high-dose interferon (IFN)-alpha2b therapy has been shown to have some efficacy. We examined 22 patients with resected melanoma who were treated with repeated injections of recombinant IFN-alpha2b during the treatment. Both angiogenic and immune parameters were measured. White blood cells (WBCs) and lymphocyte numbers, lymphocyte subpopulations, serum concentrations of IFN-alpha and anti-IFN-alpha antibodies, and the serum vascular endothelial growth factor (VEGF), interleukin (IL)-8, and basis fibroblast growth factor (bFGF) concentrations were determined over time in resected, recurrence-free patients with American Joint Committee on Cancer (AJCC) stage III melanoma with one or less (LN+ < or = 1, n = 7) or more than one (LN+ > 1, n = 8) lymph nodes involved, and AJCC stage IV resected disease (n = 7). Follow-up and recurrence-free intervals were longer in stage III (LN+ < or = 1) patients compared with stage IV patients (P < 0.05). The number of WBCs and lymphocytes decreased during the treatment for all patient groups (P < 0.001). In addition, percentages of CD8 and CD20 were higher in stage IV patients than in stage III (LN+ > 1) and stage III (LN+ < or = 1) patients at the beginning of therapy (P < 0.05). A significant increase in the percentage of CD20+ cells, mostly B lymphocytes, was observed in the stage III (LN+ > 1) and stage III (LN+ < or = 1) patients over time but not in stage IV patients (P < 0.001). Low IL-8 and bFGF concentrations at the beginning of therapy were associated with significantly longer recurrence-free survival (P < 0.05). These results warrant a larger trial to determine if the differences observed in patients before treatment can provide prognostic markers in patients receiving IFN-alpha2b therapy.
Assuntos
Antineoplásicos/uso terapêutico , Substâncias de Crescimento/sangue , Interferon-alfa/uso terapêutico , Interleucina-8/sangue , Melanoma/tratamento farmacológico , Neovascularização Patológica/imunologia , Neoplasias Cutâneas/tratamento farmacológico , Adjuvantes Imunológicos/uso terapêutico , Adulto , Idoso , Linfócitos B/imunologia , Intervalo Livre de Doença , Fatores de Crescimento Endotelial/sangue , Feminino , Fator 2 de Crescimento de Fibroblastos/sangue , Humanos , Interferon alfa-2 , Interferon-alfa/sangue , Interferon-alfa/imunologia , Contagem de Leucócitos , Linfocinas/sangue , Masculino , Melanoma/irrigação sanguínea , Melanoma/imunologia , Pessoa de Meia-Idade , Prognóstico , Proteínas Recombinantes , Neoplasias Cutâneas/irrigação sanguínea , Neoplasias Cutâneas/imunologia , Linfócitos T/imunologia , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio VascularRESUMO
Melanoma inhibitory activity protein (MIA) has been detected in patients with advanced melanoma. The present study measured the variations in blood concentrations of MIA in 84 patients with AJCC stage II to IV melanoma by ELISA. Patients treated with repeated injections of a polyvalent melanoma vaccine (PMV), interferon-alpha-2b (IFN-alpha2b), or interleukin-2 (IL-2) were followed during treatment duration. Before treatment, patients treated with PMV or IFN-alpha2b had comparable low MIA concentrations, whereas most IL-2-treated patients had higher MIA levels. At the end of treatment, MIA concentrations were higher in patients with progressive disease (PD) than in patients with no clinical evidence of melanoma (NPD) for PMV, IFN-alpha2b, or IL-2 therapy (3.7+/-0.2 vs. 11.5+/-5.4 ng/ml, 3.8+/-0.2 vs. 8.3+/-1.7 ng/ml, and 2.3+/-0.7 vs. 20.2+/-7.4 ng/ml, respectively, P<0.05). In contrast to stable MIA concentrations measured in NPD patients, significant increase in MIA levels were observed in PD patients over time regardless of treatment (P<0.05). In 20 of the 27 patients who had melanoma recurrence or progression, MIA concentrations were above 4.5 ng/ml. Finally, in these 20 patients, MIA concentrations above 4.5 ng/ml were observed prior to clinical evidence of progression (P<0.01).
Assuntos
Antineoplásicos/uso terapêutico , Biomarcadores Tumorais/sangue , Interferon-alfa/uso terapêutico , Interleucina-2/uso terapêutico , Melanoma/sangue , Proteínas de Neoplasias/sangue , Adulto , Idoso , Progressão da Doença , Proteínas da Matriz Extracelular , Feminino , Humanos , Imunoterapia , Masculino , Melanoma/patologia , Melanoma/terapia , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/sangue , Estadiamento de Neoplasias , Estudos ProspectivosRESUMO
Social conflict has been shown to affect the neuroendocrine stress response in rodents. The current study was designed to characterize the effects of social conflict on leukocyte subset distribution and function as well as in vivo bacterial growth. Male DBA/2 mice implanted or not implanted with a closed chamber containing Escherichia coli were repeatedly challenged by temporary placement in the territory of a dominant CF-1 mouse five times a day for 2 consecutive days. Nonstressed animals were similarly handled, but were not exposed to social conflict. Effects on immune responses and E. coli growth were analyzed 13 h after the last social conflict session. Social conflict alone was associated with an increase in plasma corticosterone concentration and decreases in thymocyte numbers and splenocyte ability to proliferate in vitro in the presence of lipopolysaccharide (p < 0.05). After social conflict, immature CD4+CD8+ thymocytes decreased, whereas mature T cells increased (p < 0.05). In the presence of E. coli, social conflict induced a significant increase in plasma concentration of interleukin-1beta, and a decrease in the number of thymocytes and the percentage of CD4+CD8+ T cells in the thymus (p < 0.05). In addition to the lymphocyte subpopulation changes observed with social conflict alone, the proportion of CD3+ and major histocompatibility complex (MHC) class II IAd+ cells were significantly higher in stressed mice implanted with a closed chamber containing E. coli (p < 0.05). Social conflict tended to favor E. coli growth in the closed chamber, indicating possible direct bacterial-neuroendocrine hormone interactions. Taken together, these results suggest that stress may modulate the host immune response by altering both bacterial growth and resistance to infection.
Assuntos
Nível de Alerta/fisiologia , Conflito Psicológico , Dominação-Subordinação , Escherichia coli O157/imunologia , Animais , Relação CD4-CD8 , Contagem de Colônia Microbiana , Corticosterona/sangue , Escherichia coli O157/crescimento & desenvolvimento , Tolerância Imunológica/imunologia , Interleucina-1/sangue , Masculino , Camundongos , Camundongos Endogâmicos DBA , Camundongos Endogâmicos , Técnica de Janela Cutânea , Especificidade da EspécieRESUMO
Using 2-deoxy-D-glucose (2-DG)-induced stress, our laboratory has developed studies to define stress effects on immune responses. Here, we report effects of increasing doses of 2-DG on the immune response of BALB/c, C57BL/6 and BDF(1) mice 2 h after three injections of 0 to 2000 mg/kg of 2-DG. Female 4- to 5-week-old mice were euthanized and blood and spleens were collected. A suspension of partially purified mature T splenocytes was obtained by negative selection using J11.d2 antibodies. Glucose and corticosterone levels were measured in the plasma of each mouse. Splenocyte and mature T splenocyte suspensions were tested in in vitro proliferation assays with or without concanavalin A. Splenocytes were analyzed for the following cell-surface markers: CD3, TCR alpha/beta, CD4, CD8 and major histocompatibility complex (MHC) Class II. Significant increases in blood glucose levels were observed in C57BL/6 and BALB/c strains with the highest 2-DG dose (p<0.05). Corticosterone levels were higher in BDF(1) mice and C57BL/6 mice following the administration of 1000 and 2000 mg/kg of 2-DG, respectively (p<0.01). In vitro proliferation of mature T splenocytes in the presence of concanavalin A was decreased in BDF(1) (p<0.05) but not in BALB/c and C57BL/6 mice. In addition, in BDF(1) mice the decrease was highly correlated with an increase of CD3+ and TCR alpha/beta+ cells in the spleen. These results demonstrated high variability in the response of different mouse strains to 2-DG-induced stress.
Assuntos
Desoxiglucose/imunologia , Baço/imunologia , Estresse Fisiológico/imunologia , Animais , Biomarcadores , Glicemia , Células Cultivadas , Concanavalina A/farmacologia , Corticosterona/sangue , Cruzamentos Genéticos , Desoxiglucose/administração & dosagem , Relação Dose-Resposta a Droga , Feminino , Citometria de Fluxo , Imunofenotipagem , Ativação Linfocitária/efeitos dos fármacos , Subpopulações de Linfócitos/efeitos dos fármacos , Subpopulações de Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Especificidade da Espécie , Baço/citologia , Baço/efeitos dos fármacos , Estresse Fisiológico/induzido quimicamente , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologiaRESUMO
Administration of 2-deoxy-D-glucose (2-DG) induces acute cellular glucoprivation. In the current study, we examined differences in immune parameters after 2-DG administration in both sexes. Male and female BDF1 mice were injected three times, 48 h apart, either with a saline solution (control group) or with 2-DG in saline (500 mg/kg). Two hours after the last injection, blood and spleens were collected. Plasma levels of interleukin-1beta, and interferon-gamma levels were measured. Additionally, the levels of the specific leukocyte antigens CD3, CD4, CD8, T cell receptor (TCR) alpha/beta, I-Ad, and H-2Ld/H-2Db were evaluated by flow cytometry on both blood and spleen cells. The blastogenic response of leukocytes from both tissues to mitogens was assessed. Levels of glucose, corticosterone, testosterone, progesterone, 17beta-estradiol, follicle-stimulating hormone, and luteinizing hormone were also determined. Increases in the percentage of cells bearing TCR alpha/beta and I-Ad in the blood and H-2Ld/H-2Db in the spleen were observed in the 2-DG-treated group for both sexes. In contrast, higher corticosterone and IL-1beta plasma concentrations, as well as higher percentages of splenocytes bearing TCR alpha/beta and I-Ad, and lower mitogen-induced proliferation of mature T splenocytes (79%) were observed in female but not in male mice injected with 2-DG compared with those injected with saline (p < 0.05). Taken together, these results suggest that female mice are more sensitive than male mice to immune alterations induced by 2-DG administration.
Assuntos
Adjuvantes Imunológicos/farmacologia , Antimetabólitos/farmacologia , Desoxiglucose/farmacologia , Imunidade/efeitos dos fármacos , Animais , Glicemia/metabolismo , Divisão Celular/efeitos dos fármacos , Citocinas/biossíntese , Feminino , Citometria de Fluxo , Hormônios/sangue , Ácido Láctico/sangue , Ativação Linfocitária/efeitos dos fármacos , Contagem de Linfócitos/efeitos dos fármacos , Linfócitos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBARESUMO
Calves fed soya proteins may develop severe gastrointestinal disorders. Whether these are predominantly associated with particular Ig subclasses and (or) dietary proteins remains unclear. Therefore, antibody responses to soyabean protein were analysed by dot- and blot-immunobinding in plasma and intestinal mucous secretions. One-month-old calves were fed for 2.5 months liquid diets based on skim milk powder (SMP) or a mixture (2:3, protein basis) of whey and soyabean products including a low antigenic hydrolysed soya protein isolate (HSPI) and a highly antigenic heated soya flour (HSF). Specific antibodies (Abs) of the main isotypes (IgM, IgA, IgG1, IgG2) were characterised by immunostaining of samples which had been previously incubated with nitrocellulose sheets coated with SMP, HSPI or HSF extracts. Plasma collected before feeding experimental diets showed very little specific Abs. By contrast, 2.5 months later, a three-fold increase (P < 0.05) in IgG1 and IgA titres against HSF antigens was observed in calves fed HSF compared with those fed the control or HSPI diet. IgG1 immunoblotting revealed many protein bands from soya in the molecular range of 22-32 and 38-42 kDa. Immunorecognition of specific proteins from SMP and HSPI remained low and similar among animal groups. Specific IgM, IgA and IgG1 titres against HSF, and to a lesser extent HSPI, were significantly higher (P < 0.05) in jejunal mucous secretion of calves fed HSF compared with other groups. Secretions from calves fed HSF bound to many soyabean proteins in the range of 17-23 and 26-38 kDa, with similar patterns for IgA and IgG1. By contrast, only weak bands were found for IgM and IgG2 in all groups of calves. Thus, calves fed antigenic HSF do present specific Abs including IgG1 and IgA isotypes, both systemically and locally. Therefore, IgG1 and (or) IgA rather than IgM and IgG2 Abs may be preferred for assessing the immunogenicity of soyabean products in calves. Interestingly, soyabean immunogenicity was drastically reduced by adequate proteolysis.
Assuntos
Bovinos/imunologia , Isotipos de Imunoglobulinas/análise , Secreções Intestinais/imunologia , Proteínas de Vegetais Comestíveis/imunologia , Animais , Bovinos/sangue , Proteínas Alimentares/administração & dosagem , Proteínas Alimentares/imunologia , Immunoblotting/veterinária , Imunoglobulina A/análise , Imunoglobulina A/sangue , Imunoglobulina G/análise , Imunoglobulina G/sangue , Imunoglobulina G/classificação , Isotipos de Imunoglobulinas/sangue , Imunoglobulina M/análise , Imunoglobulina M/sangue , Masculino , Proteínas de Vegetais Comestíveis/administração & dosagem , Proteínas de Soja , Glycine max/imunologiaRESUMO
Gut immune responses have been suspected in food hypersensitivity reactions such as those to soyabean proteins in early-weaned piglets. The present study examines the lymphoid cell subset distribution in piglets fed heat-treated (HTSP) or ethanol-treated soyabean proteins (ETSP). Duodenal cryosections of 4-week-old HTSP piglets (n = 10) and ETSP piglets (n = 8) were analysed for IgA, IgM, IgG1 and IgG2 positive cells, CD2, CD4, CD8, WC1 T cell positive antigens using immunohistochemical peroxidase reactions. Densities of IgM+ and IgA+ cells were three times and, IgG1+ and IgG2+ six times higher in the lamina propria of HTSP piglets compared with ETSP (P < 0.05). Increased CD2+ T cells were accounted for by a rise in both CD4+ and CD8+ T cell subsets in the lamina propria (P < 0.01) as well as in the epithelium of the duodenal mucosa of piglets fed HTSP. The density of the WC1+ T cell subset in the epithelium was significantly higher in HTSP than in ETSP piglets (P < 0.01). Immune reactions in the duodenal mucosa, involving both B and T lymphocytes may be related to atrophy of the duodenal villi in HTSP piglets.
Assuntos
Linfócitos B/imunologia , Hipersensibilidade Alimentar/veterinária , Mucosa Intestinal/imunologia , Proteínas de Vegetais Comestíveis/administração & dosagem , Doenças dos Suínos/imunologia , Linfócitos T/imunologia , Animais , Anticorpos Monoclonais , Antígenos CD/análise , Proteínas Alimentares/administração & dosagem , Proteínas Alimentares/efeitos adversos , Duodeno/imunologia , Manipulação de Alimentos , Hipersensibilidade Alimentar/etiologia , Hipersensibilidade Alimentar/imunologia , Temperatura Alta , Técnicas Imunoenzimáticas/veterinária , Isotipos de Imunoglobulinas/análise , Imunofenotipagem/veterinária , Contagem de Linfócitos/veterinária , Proteínas de Vegetais Comestíveis/efeitos adversos , Distribuição Aleatória , Proteínas de Soja , Glycine max/efeitos adversos , Suínos , Doenças dos Suínos/etiologiaRESUMO
The effect of dietary antigens on the gut morphology and density of immune cells was studied in preruminant calves fed milk substitutes containing skim milk powder (SMP), non-antigenic hydrolysed soya protein isolate (HSPI) or antigenic heated soyabean flour (HSF) as their main protein source for 3 months. The height and perimeter of proximal jejunum villi were highest in the calves fed SMP and lowest in those fed HSF (P < 0.05). In contrast, the crypt depth and perimeter were apparently not influenced by the dietary treatments studied. This morphological alteration was associated with a dramatic infiltration of the lamina propria by B and T lymphocytes in the calves fed HSF (P < 0.01). Increased B cell density was essentially accounted for by IgA-, IgG1- and IgG2-bearing cells. The density of CD2-positive T lymphocytes increased (P < 0.01) in the jejunal lamina propria of HSF calves, involving helper (CD4+) and suppressor-cytotoxic (CD8+) T cell subsets. The density of gamma-delta (WC1+) T cells also increased (P < 0.01). The major change concomitantly observed in the villus epithelium was an increased density of CD8+ cells (P < 0.05) and WC1 + cells (P < 0.01).
Assuntos
Linfócitos B/imunologia , Quimiotaxia de Leucócito/imunologia , Farinha/análise , Glycine max/imunologia , Temperatura Alta/uso terapêutico , Intestino Delgado/imunologia , Linfócitos T/imunologia , Ração Animal , Animais , Bovinos , Jejuno/imunologia , MasculinoRESUMO
The systemic and local (gut) patterns of antibodies against various proteins from soyabean were analysed in preruminant calves fed milk substitutes based on skim milk powder (SMP) or heated soyabean flour (HSF) as the main protein sources. The titres of IgM, IgA, IgG1 and IgG2 antibodies were determined against feed extracts and purified soyabean proteins by dot-blotting in plasma after three months and jejunal mucous secretions after six months of feeding the experimental diets. The calves fed HSF had higher levels of circulating IgA, IgG1 and IgG2 antibodies against raw or heated soya extracts and purified proteins including alpha-conglycinin, beta-conglycinin, Bowman-Birk protease inhibitors and lectins than the calves fed SMP. In contrast, the differences between the IgM titres of the groups were most often not significant. The patterns of specific antibodies present in the jejunum were broadly similar to those observed in the blood, although the differences between the groups of calves more often reached significance for IgG2 and IgM than for IgA and IgG1, when the purified soyabean proteins were considered.
Assuntos
Formação de Anticorpos/imunologia , Bovinos/imunologia , Hipersensibilidade Alimentar/veterinária , Glycine max/imunologia , Intestinos/imunologia , Envelhecimento/imunologia , Animais , Anticorpos Monoclonais/imunologia , Hipersensibilidade Alimentar/imunologia , Temperatura Alta , Immunoblotting/veterinária , Isotipos de Imunoglobulinas/imunologia , Masculino , Glycine max/efeitos adversos , Glycine max/química , Fatores de TempoRESUMO
The allergenicity of soya proteins was assessed by direct skin testing and by in vitro lymphoproliferation tests in calves fed milk substitutes containing skim milk powder (SMP) or an antigenic heated soya flour (HSF). During the last three weeks of treatment, the calves were injected intradermally with raw soya flour (RSF), HSF, hydrolysed soya protein isolate (HSPI), SMP or purified soya proteins, after being premedicated with anti-histamine or not. Peripheral blood leucocytes (PBL) were grown over five days with various mitogens or dietary antigens, and the incorporation of tritiated thymidine was measured. Strong skin oedema reactions to RSF, HSF and all the purified proteins were observed in the calves fed HSF at various times up to 24 hours after injection. The skin oedema was largely prevented by premedication with anti-histamine. A strong delayed skin thickening was observed in the calves fed HSF for up to five days with beta-conglycinin. PBL from the calves fed HSF proliferated in vitro with HSF, HSPI and beta-conglycinin, but not with glycinin. Thus, most proteins from soyabean were implicated in the immediate and semi-delayed immune reactions, whereas beta-conglycinin was strongly involved in a delayed type hypersensitivity in calves.
Assuntos
Alérgenos , Glycine max/imunologia , Ativação Linfocitária , Ração Animal , Animais , Animais Recém-Nascidos , Formação de Anticorpos , Bovinos , Edema/prevenção & controle , Farinha , Testes de Hemaglutinação , Técnicas In Vitro , Masculino , Leite , Proteínas de Plantas/imunologia , Pele/imunologia , Testes CutâneosRESUMO
The development of local and systemic immune responses to soybean proteins was investigated in early-weaned pigs. Pigs were given either antigenic (ASP, n = 10 pigs) or non-antigenic (NASP, n = 8 pigs) soybean products (6 g of protein/d) from d 5 to 9 of age by stomach tube. After weaning at d 21, pigs were fed diets containing the corresponding soybean products and slaughtered between d 28 to 30. Diarrhea was 2.4-fold more frequent, the size of duodenal villi was reduced by 24 to 36%, and the eosinophil density in the duodenal mucosa was 13 times greater (P < .02) in the ASP pigs compared with the NASP pigs. A larger erythema area (P = .006) was observed in the ASP group than in the NASP group 30 min after an intradermal injection of glycinin, but no significant difference could be detected with alpha- or beta-conglycinin or whole soybean extracts. No difference in skin fold thickness was apparent between groups 24 h later. Intestinal, mesenteric lymph node, and blood lymphocytes did not proliferate when cultured with soybean proteins, regardless of dietary treatment. By ELISA, no difference between groups was observed in the circulating levels of total immunoglobulins (Ig) and IgM. Immunoblotting patterns of raw soybean with sera from 28-d-old pigs showed two bands (22 and 36 kDa) recognized by IgA and IgM, respectively, in the ASP group only. Finally, the adverse effects observed with antigenic soybean flour can be overcome by the use of alcohol-treated products.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Ração Animal , Proteínas Alimentares/administração & dosagem , Glycine max , Proteínas de Vegetais Comestíveis/administração & dosagem , Suínos/imunologia , Animais , Proteínas Alimentares/imunologia , Duodeno/anatomia & histologia , Ensaio de Imunoadsorção Enzimática/veterinária , Eosinófilos/imunologia , Feminino , Immunoblotting/veterinária , Imunoglobulina M/sangue , Imunoglobulinas/sangue , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Testes Intradérmicos/veterinária , Ativação Linfocitária , Masculino , Proteínas de Vegetais Comestíveis/imunologia , Distribuição Aleatória , Proteínas de Soja , Glycine max/imunologia , Suínos/crescimento & desenvolvimento , Desmame , Aumento de PesoAssuntos
Hipersensibilidade Alimentar/imunologia , Glycine max/imunologia , Proteínas de Vegetais Comestíveis/imunologia , Animais , Formação de Anticorpos , Antígenos , Feminino , Hipersensibilidade Alimentar/etiologia , Hipersensibilidade Alimentar/patologia , Imunidade Celular , Técnicas In Vitro , Ativação Linfocitária , Masculino , Proteínas de Vegetais Comestíveis/efeitos adversos , Gravidez , Glycine max/efeitos adversos , Suínos , Aumento de PesoAssuntos
Arte , Técnicas Projetivas , Psicologia do Esquizofrênico , Tioridazina/farmacologia , Adulto , Feminino , HumanosRESUMO
OBJECTIVES: To evaluate melanoma biopsy specimens for human papilloma virus (HPV) and determine the relation between the presence of HPV, in vitro growth, and clinical progression of melanoma in the patients from whom the biopsy specimens were derived. SUMMARY BACKGROUND DATA: Ultraviolet radiation from sun exposure appears to be the primary causal agent in the development of cutaneous melanoma. However, other agents, including HPV, as observed in different epithelial carcinomas, may also play a role in melanoma development and progression. METHODS: Twelve melanoma biopsy specimens obtained from 12 patients with AJCC stage III and IV melanoma were stained with antibodies against gp-100 (HMB-45) and S-100 protein to confirm melanoma diagnosis and with a polyclonal HPV antibody. After mechanical dissociation, the melanoma specimen cells' ability to grow in vitro was assessed. Patients were evaluated for melanoma progression with physical examination, complete blood count, and liver function tests every 3 months and a chest radiograph every 6 months. RESULTS: All biopsy specimens were positive for S-100, and nine (75%) were positive for gp-100. Seven of 12 (58%) were positive for HPV by immunohistochemistry. In vitro, none of the HPV-negative tumor cells grew from the tumor biopsies, whereas five of seven (71%) of the HPV-positive melanoma tumor cells grew very well. All patients with HPV-positive tumor cells had recurrences and died of melanoma progression, whereas four of five (80%) patients with HPV-negative tumor cells remained alive and without melanoma recurrence. CONCLUSIONS: The presence of HPV was found in 58% of the biopsy specimens obtained from patients with stage III and IV melanoma and correlated with rapid melanoma progression. HPV may serve as a cofactor in the development of melanoma and may modulate a more aggressive phenotype in HPV-containing melanoma cells.