Influence of disease severity on nitrite and cytokine production by peripheral blood mononuclear cells (PBMC) from patients with pulmonary tuberculosis (TB).

Earlier studies in patients with pulmonary TB have revealed a higher production of Th1 cell type cytokines in moderate TB, with predominant Th2-like responses in advanced disease. Given the influence of IL-12 in T cell differentiation, as well as the roles of transforming growth factor-beta (TGF-beta), nitric oxide and tumour necrosis factor-alpha (TNF-alpha) in the immune response against intracellular pathogens, we decided to analyse the interferon-gamma (IFN-gamma), IL-4, IL-12, TGF-beta, TNF-alpha and nitrite concentrations in culture supernatants of PBMC from TB patients showing different degrees of lung involvement. The sample population comprised 18 untreated TB patients with either moderate (n = 9) or advanced (n = 9) disease and 12 age- and sex-matched healthy controls (total population (patients and controls) 12 women, 18 men, aged 37 +/- 13 years (mean +/- s.d.)). PBMC were stimulated with whole sonicate from Mycobacterium tuberculosis and the supernatants were collected on day 4 for measurement of cytokine and nitrite levels. Antigen-stimulated IFN-gamma, TGF-beta and TNF-alpha production was found to be significantly increased in TB patients, both moderate and advanced, compared with the controls. Levels of IFN-gamma were significantly higher in moderate disease than advanced cases, whereas advanced cases showed significantly higher IL-12, TGF-beta and TNF-alpha concentrations when compared with cases of moderate TB. Nitrite levels were also increased in TB patients and the increase was statistically significant when advanced cases were compared with controls. These findings may contribute to a clearer picture of the net effect of cytokine interactions in TB, essential for a better understanding of the immunopathological mechanisms underlying the distinct clinical forms of the disease.

Mechanism of antimetastatic immunopotentiation by low-dose cyclophosphamide.

We have previously reported the antimetastatic effect of a single low-dose of cyclophosphamide (Cy) on L-TACB rat lymphoma. The phenomenon could be adoptively transferred in immunocompetent rats and is abolished in nude mice, facts for which an immunomodulatory explanation was proposed. The aim of this paper was to identify the mechanism(s) by which spleen cells from Cy-treated tumour-bearing rats could exert this antimetastatic activity. Conditioned media obtained by incubation of spleen cells from Cy-treated and non-treated tumour-bearing rats, under specific or non-specific stimulation, were assayed to evaluate their effect on lymphocyte proliferation. The production of transforming growth factor beta (TGF-beta), interleukin-10 (IL-10) and nitric oxide (NO) by conditioned media was also studied. The restoration of spleen lymphoproliferative responses to normal levels when exposed to media conditioned by splenocytes from Cy-treated tumour-bearing rats, together with a decreased production of suppressive cytokines TGF-beta, IL-10 and NO, suggest an enhancement of host antimetastatic immunity triggered by single low-dose Cy treatment.

Clinical and serological studies of tuberculosis patients in Argentina receiving immunotherapy with Mycobacterium vaccae (SRL 172).

Two small, placebo-controlled studies of immunotherapy with heat killed Mycobacterium vaccae added to routine chemotherapy for pulmonary tuberculosis, together involving 40 HIV seronegative patients, were carried out in Argentina. The immunotherapy was associated with reduced sputum smear positivity of AFB at 1 month and a greater reduction in ESR at 2 months. In the first study radiological improvement was better (P < 0.05) among immunotherapy recipients. In the second study, weight regain and time to become apyrexial were measured and were found to be improved amongst immunotherapy recipients (P < 0.05). In the first month of treatment the levels of IgG to the 65 kDa and 70 kDa heat-shock proteins showed greater falls following immunotherapy (P < 0.05 and P < 0.001, respectively). On admission serum cytokine levels of interleukins 4 and 10 (IL-4, IL-10), interferon gamma (IFN-gamma) and tumour necrosis factor alpha (TNF-alpha) were grossly raised in comparison with a matched control group (P < 0.001). After 1 month. Levels of IL-4, IL-10 and TNF-alpha fell (P < 0.001, P < 0.01 and P < 0.01, respectively) and levels of IFN-gamma rose more (P = 0.005) in immunotherapy recipients than in those receiving chemotherapy alone. The results are in accord with a switch towards a TH1 immunological status and clinical benefit for immunotherapy recipients.

In vitro synthesis of interferon-gamma, interleukin-4, transforming growth factor-beta and interleukin-1 beta by peripheral blood mononuclear cells from tuberculosis patients: relationship with the severity of pulmonary involvement.

Given the role of cell-mediated immune responses in resistance to mycobacteria, we sought to analyse whether there was a relationship between the severity of pulmonary tuberculosis (TB) and lymphocyte proliferation as well as in vitro cytokine production. To achieve this, 25 untreated TB patients showing mild (n = 5), moderate (n = 9) or advanced (n = 11) pulmonary disease, and 12 age-matched healthy controls (mean+/-SD, 37+/-14.5 years) were studied. Peripheral blood mononuclear cells were cultured for 5 days with 10 microg/ml whole, sonicated Mycobacterium tuberculosis (WSA) or 2.5 microg/ml Concanavalin A (Con A). Supernatants were collected on day 4, from cultures grown with or without WSA, for measurement of interferon-gamma (IFN-gamma), interleukin (IL)-4, IL-1beta and transforming growth factor-beta (TGF-beta). Antigen-specific proliferation was found to be reduced among patients and more profound in those with advanced disease who also displayed a depressed response to Con A. Patients with mild TB showed a preferential production of IFN-gamma over IL-4, gave the highest level of IFN-gamma synthesis upon specific antigen stimulation and showed increased levels of IL-1beta production. Findings in patients with moderate TB appeared compatible with a mixed production of IFN-gamma and IL-4 coexisting with a higher synthesis of TGF-beta, by comparison to patients with mild TB. Advanced disease showed the highest IL-4 and TGF-beta production, with IFN-gamma synthesis readily noticeable, yet decreased in comparison with the other patient groups. Differences in cytokine response according to the amount of lung involvement suggest a role for such mediators in the immunopathogenesis underlying the distinct clinical forms of pulmonary TB, that is a predominant T helper Th)1-like or Th2-like activity in mild or in progressive TB, respectively.

Circulating profile of Th1 and Th2 cytokines in tuberculosis patients with different degrees of pulmonary involvement.

To investigate whether differences in the degree of pulmonary tuberculosis lesions could be accompanied by changes in the pattern of circulating cytokines, 29 untreated tuberculosis patients showing mild (n = 10), moderate (n = 5) or advanced (n = 14) pulmonary disease, and 12 age-matched healthy controls (mean +/- S.D., 36 +/- 15 years) were studied. ELISA methods for the evaluation of interferon-gamma, interleukin-2, interleukin-4, and interleukin-10 indicated that all patients had increased serum levels of the four cytokines in relation to controls. Mean titers of interferon-gamma and interleukin-2 in mild and moderate patients appeared higher than in those with advanced disease, whereas moderate and advanced patients showed the higher levels of IL-4 in comparison to mild cases. Raised levels of interleukin-10 were more prevalent in advanced disease, and statistically different from those in mild patients. This cytokine pattern may help to explain findings wherein mild tuberculosis is characterized by preserved cellular immune responses while advanced disease is accompanied by an impairment of such parameters.

Levels of interleukin-8 in tuberculous pleurisy and the profile of immunocompetent cells in pleural and peripheral compartments.

Our study investigated the presence of IL-8 in pleural exudates from tuberculosis patients (TBP) (n = 13), and evaluated whether it was related with the profile of major immunocompetent cells present in their pleural and peripheral compartments. To allow comparisons, an additional group of patients with parapneumonic pleural effusions (PNE) (n = 7) was included. Blood peripheral immunophenotypic studies were also carried out in 12 age-matched healthy controls (Co), and 39 tuberculosis patients classified, according to the extent of pulmonary involvement, into mild (n = 9), and advanced (n = 30) cases. Patients were recruited before starting therapy, had HIV negative serology, and showed no age differences among groups (mean +/- SD., 40.7 +/- 14.7 years). IL-8 concentrations were measured by an ELISA method while immunophenotypic analysis was performed by using FITC-conjugated monoclonal antibodies reacting against the following cell surface molecules: CD3, CD4, CD8, CD25 (IL-2R+ cells), CD19, and CD68. IL-8 was detected in all pleural exudates though levels in the TB patients, 384 +/- 110 pg/ml, appeared significantly higher than the PNE group, 185 +/- 110 pg/mg, (P < 0.015, mean +/- S.D.). In turn, the former group presented values of pleural CD3+, CD4+, and CD25, which were found increased in comparison with PNE patients (P < 0.01). Unlike the pleural compartment, patients with TBP showed a marked and significant decrease in their circulating levels of cells bearing the CD3, CD4, CD19, CD25, and CD68 phenotypes not only when comparing with Co but also with PNE and mild patients. Differences between the levels of pleural and peripheral T-cells from TBP patients may be the reflection of an important influx of T-lymphocytes from the circulatory system to the pleural cavity, probably linked to the presence of chemotactic factors within the pleural fluid like IL-8.

Low occurrence of arthritic manifestations in patients with pulmonary tuberculosis. T cell subsets and humoral studies.

Given the suspected role of mycobacteria in the establishment of disorders with an autoimmune background and joint damage, a study was conducted to analyze whether rheumatic symptoms were likely to be present in tuberculosis (TB) patients. To this end, 330 patients with a bacteriologic confirmation of tuberculosis were investigated for the presence of arthritic complaints. The latter were recorded in five of them with rheumatic symptoms mostly involving interphalangeal and metacarpophalangeal joints, and preceding the clinical manifestations of the TB illness. Three out of these five patients remained arthritic by the time of the bacteriologic conversion and fulfilled the criteria for the diagnosis of rheumatoid arthritis. In the two remaining patients sputum negativization was accompanied by a disappearance of rheumatic manifestations. These patients were also assessed for their peripheral levels of major T cell subsets as well as for the presence of autoantibodies. Comparisons with a series of non-arthritic TB cases, rheumatoid arthritis patients, and controls revealed that presence of rheumatic manifestations was associated with a different profile of autoantibody formation and T cell subset changes. Evidence recorded in the present study indicates that joint affectation in TB is a rare event, being rather the exception than the rule.

Circulating immune complexes in patients with advanced tuberculosis and their association with autoantibodies and reduced CD4+ lymphocytes.

We investigated the presence of circulating immune complexes (CICs) in serum from tuberculosis (TB) patients with different degrees of pulmonary involvement. Patients were classified into four groups according to the extent of lung involvement: mild (N = 9), moderate (N = 12), moderate plus (N = 16), and severe cases (N = 10). A search for CICs by the 3.5% PEG precipitation test showed that the CIC values of patients with the moderate plus or severe form of pulmonary TB were significantly higher compared to healthy controls and to mild and moderate cases (P < 0.01 and P < 0.001, respectively). Further analysis demonstrated that increased CIC levels were associated with increased autoantibody production, since this abnormality was more prevalent in patients with advanced disease (P < 0.01), who also showed a significant reduction of CD4+ T lymphocytes. The immunoregulatory and pathogenetic implications of these findings are discussed.

Circulating immune complexes in patients with advanced tuberculosis and their association with autoantibodies and reduced CD4 + lymphocytes

We investigated the presence of circulating immune complexes (CICs) in serum from tuberculosis (TB) patients with different degrees of pulmonary involvement. Patients were classified into four groups according to the extent of lung involvement: mild (N = 9), moderate (N = 12), moderate plus (N = 16), and severe cases (N = 10). A search for CICs by the 3.5 percent PEG precipitation test showed that the CIC values of patients with the moderate plus or severe form of pulmonary TB were significantly higher compared to healthy controls and to mild and moderate cases (P<0.01 and P<0.001, respectively). Further analysis demonstrated that increased CIC levels were associated with increased autoantibody production, since this abnormality was more prevalent in patients with advanced disease (P<0.01), who also showed a significant reduction of CD4+ T. lymphocytes. The immunoregulatory and pathogenetic implications of these findings are discussed

Microorganismos enteropatógenos en niños con diarrea aguda en dos hospitales de Rosario, Argentina / Enteropathogenic microorganisms in children with acute diarrhea in 2 hospitals of Rosario, Argentina

Como parte de um estudo multicéntrico nacional se investigaron agentes enteropatógenos em 495 niños menores de 5 años con diarrea aguda, entre agosto de 1985 y diciembre de 1988. La tasa total de aislamiento fue similar en relación a la estación, edad y sexo. Estuvieron significantemente asociados a la condición de hospitalización la desnutrición, deshidratación, fiebre, leucocitos en heces y frecuencia de agentes, en especial Escherichia coli enteropatógeno (ECP) (p < 0,001). Los principales agentes enteropatógenos fueron ECEP (26,5//), E. coli enterotosigénico (ECET) (9,7//), Shigella (8,5//). Rotavirus (5,1//), Giardia (3,6//), Campylobacter (3,2//) y Salmonella (2,4//). Los serotipos de ECET 0 153:H45 y 0 128:H21 fueron los más frecuentes. El aislamiento de Shigella se relacionó claramente con la presencia de leucocitos en heces, predominó en niños internados, febriles y en mayores de 5 meses. Campylobacter se presentó en niños de 1 año y ambulatórios. Rotavirus predominó en otoño e invierno. Giardia predominó en niños internados y denutridos. En 10// de los casos hubo asociación de 2 ó mas agentes. Salmonella resultó multisensible a los antimicrobianos probados pero entre 40 y 80// de las cepas de Shigella y E. coli fueron resistentes a sulfametaxazol-trimetoprima y ampicilina (AU)

Microorganismos enteropatógenos en niños con diarrea aguda en dos hospitales de Rosario, Argentina / Enteropathogenic microorganisms in children with acute diarrhea in 2 hospitals of Rosario, Argentina

Como parte de um estudo multicéntrico nacional se investigaron agentes enteropatógenos em 495 niños menores de 5 años con diarrea aguda, entre agosto de 1985 y diciembre de 1988. La tasa total de aislamiento fue similar en relación a la estación, edad y sexo. Estuvieron significantemente asociados a la condición de hospitalización la desnutrición, deshidratación, fiebre, leucocitos en heces y frecuencia de agentes, en especial Escherichia coli enteropatógeno (ECP) (p < 0,001). Los principales agentes enteropatógenos fueron ECEP (26,5//), E. coli enterotosigénico (ECET) (9,7//), Shigella (8,5//). Rotavirus (5,1//), Giardia (3,6//), Campylobacter (3,2//) y Salmonella (2,4//). Los serotipos de ECET 0 153:H45 y 0 128:H21 fueron los más frecuentes. El aislamiento de Shigella se relacionó claramente con la presencia de leucocitos en heces, predominó en niños internados, febriles y en mayores de 5 meses. Campylobacter se presentó en niños de 1 año y ambulatórios. Rotavirus predominó en otoño e invierno. Giardia predominó en niños internados y denutridos. En 10// de los casos hubo asociación de 2 ó mas agentes. Salmonella resultó multisensible a los antimicrobianos probados pero entre 40 y 80// de las cepas de Shigella y E. coli fueron resistentes a sulfametaxazol-trimetoprima y ampicilina

[Inhibition of total and active E-rosettes mediated by serum from tubercular patients]. / Inhibición en la formación de rosetas E totales y activas mediada por el suero de pacientes tuberculosos.

AIM: To study the effect of tuberculous patients' serum upon normal lymphocytes in order to determine possible inhibition factors of immune response. PATIENTS AND METHODS: We studied total E-rosette formation (4 degrees C, RET) and active rosette formation (37 degrees C, REA) in tuberculous patients in different stages: mild (n:36), moderate (n:28), severe (n:24) and a control group of 32 patients. The same determinations were studied in normal circulating lymphocytes (LN) incubated before with serum from tuberculous patients (SP), serum of normal individuals (SN) and in Tc-199 medium. RESULTS: The number of RET and REA in tuberculous patients of different stages were significantly lower to the ones of the control group (p less than 0.01, p less than 0.05). This differences were also found when we compare the number of RET and REA after the incubation of LN with SP and SN respectively (p less than 0.01, p less than 0.05). SUMMARY: Tuberculous patients showed an impaired ability to form RET and REA. The incubation of normal lymphocytes with serum from tuberculous patients reduced significantly its capacity to form rosettes.

Evaluacion de las subpoblaciones de linfocitos T en pacientes tuberculosos empleando la modulacion con teofilina / T-lymphocyte subsets evaluation in patients with tuberculosis using modulation with theophylline

Se evaluaron las poblaciones y subpoblaciones linfocitarias en pacientes con tuberculosis pulmonar antes y durante la terapia relacionando estos valores con la incidencia y evolucion de la enfermedad. Pacientes en sus diversas manifestaciones clinicas, virgenes de tratamiento, se estudiaron por baciloscopia (BAAR), radiologia, i.d.r. Mantoux y analisis complementarios. Se cuantificaron mediante la prueba de Rosetas espontaneas (RE) linfocitos T totales y activos (RE a 4 graus Celsius y 37 graus Celsius), T colaboradores (RE Teofilina Resistentes: RETR) y supressores (RE Teofilina Sensibles: RETS). Los examenes se repitieron en los mismos sujetos iniciado el tratamiento y en testigos sanos (TS). Se demostro en los pacientes en todas sus formas clinicas un descenso significativo en los valores relativos y absolutos de celulas T y en la relacion RETR/RETS (menor de 1). Existe asociacion entre la forma clinica y el numero de linfocitos T colaboradores. Los pacientes en tratamiento con evolucion favorable, evidenciaron un incremento significativo en los linfocitos T totales, activos, colaboradores y en la relacion RETR/RETS. Los enfermos con baciloscopia altemente positiva presentaron i.d.r. Mantoux baja o negativa y marcado descenso de celulas inmunocompetentes...

[T-lymphocyte subsets evaluation in patients with tuberculosis using modulation with theophylline]. / Evaluación de las subpoblaciones de linfocitos T en pacientes tuberculosos empleando la modulación con teofilina.

T cells and T cells subsets in peripheral blood of patients with different forms of pulmonary tuberculosis were evaluated to explain some aspects of the immunocompromised state of these subjects. Diagnosis was made by baciloscopy (BAAR), chest roentgenography i.d.r. Mantoux, and other clinical analysis. Spontaneous E-Rosette test (RE) was used to quantify Total (RET 4 degrees C) and Active T cells (REA 37 degrees C) and the same test after incubation with Theophylline (The) for helper cells (The-resistant cells: RETR) and suppressor cells (The-sensitive cells: RETS). Patients were followed for at least 4 months after therapy. The data demonstrate a significant decrease of relative and absolute numbers of Total T-cells and a diminished T helper/T suppressor subset ratio (RETR/RETS) which dropped to less than 1 in untreated patients. Treated patients with a favourable evolution showed a restoration of Total active and helper T cells. RETR/RETS ratio was also significantly increased. In patients with highly positive BAAR, low on negative i.d.r. Mantoux, a decreased level of immunocompetent cells was observed. The 3 aspects were associated. Nutritional condition of the patients was also associated with the predisposition to acquire this disease.

[Inhibitory effect of Staphylococcus aureus on the growth of rat sarcoma]. / Efecto inhibidor de Staphylococcus aureus sobre el crecimiento de un sarcoma en ratas.

We have studied the effect upon the growth of a transplantable rat sarcoma, of an intraperitoneal inoculation of peritoneal exudate cell (PEC), spleen cells (SC) and non-adherent spleen cells (N-ASC) obtained from S. aureus previously inoculated rats and the same cells from normal rats (NPEC, NSC, NN-ASC). Inbred, adult rats were inoculated with 800 x 10(6) bacteria, killed by tyndalization. After 7 days, treated and normal animals were sacrificed and PEC, SC and N-ASC were obtained. Different groups of animals were inoculated with these cell populations. Simultaneously the rats received a s.c. inoculum of a transplantable sarcoma (S-E 100). Tumor size was measured on days 4, 7, 14, 21 and 28 after tumor challenge. A significant tumor growth inhibition was found with all three cell populations, but no effect was observed with normal cells. Tumor size on different days and tumor growth curves clearly demonstrate this effect. We conclude that Staphylococcus aureus inoculum as well as the cells from animals previously challenged with the bacteria induce tumor growth inhibition. The possibility that protein A of S. aureus may be involved in this phenomenon, or that the mechanism of tumor growth inhibition is mediated by an activation of different cell populations is discussed.

Efecto inhibidor de Staphylococcus aureus sobre el crecimiento de un sarcoma en ratas. / [Inhibitory effect of Staphylococcus aureus on the growth of rat sarcoma]

We have studied the effect upon the growth of a transplantable rat sarcoma, of an intraperitoneal inoculation of peritoneal exudate cell (PEC), spleen cells (SC) and non-adherent spleen cells (N-ASC) obtained from S. aureus previously inoculated rats and the same cells from normal rats (NPEC, NSC, NN-ASC). Inbred, adult rats were inoculated with 800 x 10(6) bacteria, killed by tyndalization. After 7 days, treated and normal animals were sacrificed and PEC, SC and N-ASC were obtained. Different groups of animals were inoculated with these cell populations. Simultaneously the rats received a s.c. inoculum of a transplantable sarcoma (S-E 100). Tumor size was measured on days 4, 7, 14, 21 and 28 after tumor challenge. A significant tumor growth inhibition was found with all three cell populations, but no effect was observed with normal cells. Tumor size on different days and tumor growth curves clearly demonstrate this effect. We conclude that Staphylococcus aureus inoculum as well as the cells from animals previously challenged with the bacteria induce tumor growth inhibition. The possibility that protein A of S. aureus may be involved in this phenomenon, or that the mechanism of tumor growth inhibition is mediated by an activation of different cell populations is discussed.

Efecto inhibidor de Staphylococcus aureus sobre el crecimiento de un sarcoma en ratas / Inhibitory effect of Staphylococcus aureus on the growth of rat sarcoma

Se investigó el efecto modulador de la respuesta antitumoral de S. aureus y diversas poblaciones celulares en ratas portadoras de un sarcoma transplantable (S-E 100). Se comprobó en estas experiencias la capacidad inhibitoria del crecimiento del S-E 100 en ratas endocriadas, a través de la inoculación i.p. de células de exudado peritoneal (CEP), células de bazo (C.B.) y células de bazo no-adherenrtes (CBN-A), provenientes de ratas previamente desafiadas con S. aureus. También se inocularon i.p. células de animales normales (CEPN, CBN, CBN-AN). Se midió el tamaño del S-E 100 en los días 4, 7, 14, 21 y 28 después de iniciado el tratamiento. Se determinó que S. aureus, así como las poblaciones celulares provenientes de animales previamente inoculados con el germen (CE, CB, CBN-A), inhibe significativamente el crecimiento del tumor. Las células de animales normales no inciden en tal efecto. Se discute si este fenómeno es producido por la proteína A de S. aureus o a través de la activación de las poblaciones celulares (linfocitos, macrófagos, NK) inoculadas (AU)

Efecto inhibidor de Staphylococcus aureus sobre el crecimiento de un sarcoma en ratas / Inhibitory effect of Staphylococcus aureus on the growth of rat sarcoma

Se investigó el efecto modulador de la respuesta antitumoral de S. aureus y diversas poblaciones celulares en ratas portadoras de un sarcoma transplantable (S-E 100). Se comprobó en estas experiencias la capacidad inhibitoria del crecimiento del S-E 100 en ratas endocriadas, a través de la inoculación i.p. de células de exudado peritoneal (CEP), células de bazo (C.B.) y células de bazo no-adherenrtes (CBN-A), provenientes de ratas previamente desafiadas con S. aureus. También se inocularon i.p. células de animales normales (CEPN, CBN, CBN-AN). Se midió el tamaño del S-E 100 en los días 4, 7, 14, 21 y 28 después de iniciado el tratamiento. Se determinó que S. aureus, así como las poblaciones celulares provenientes de animales previamente inoculados con el germen (CE, CB, CBN-A), inhibe significativamente el crecimiento del tumor. Las células de animales normales no inciden en tal efecto. Se discute si este fenómeno es producido por la proteína A de S. aureus o a través de la activación de las poblaciones celulares (linfocitos, macrófagos, NK) inoculadas