Plasma levels of tumor necrosis factor-alpha in patients with visceral leishmaniasis (Kala-Azar). Association with activity of the disease and clinical remisson following antimonial therapy

Salomão, Reinaldo; Castelo Filho, Adauto; Medeiros, Iara Marques de; Sicolo, Miguel Angel

doi:10.1590/S0036-46651996000200005

Abstracts

Evaluation of TNF-alpha in patients with Kala-azar has drawn increasing interest due to its regulatory role on the immune system, in addition to its cachetizing activity. The objective of this study was to examine the association between plasma levels of TNF-alpha, measured by immunore-activity (ELISA) and bioactivity (cytotoxicity assay with L-929 cells), and clinical manifestations of visceral leishmaniasis. Plasma samples from 19 patients with Kala-azar were obtained before, during and at the end of antimonial therapy. TNF-alpha determinations was done by using the cytotoxicity assay (all patients) and the enzyme-linked immunoassay (ELISA - 14 patients). A discrepancy between results obtained by ELISA and cytotoxicity assay was observed. Levels of circulating TNF-alpha, assessed by ELISA, were higher in patients than in healthy controls, and declined significantly with improvement in clinical and laboratory parameters. Plasma levels before treatment were 124.7 ± 93.3 pg/ml (mean ± SD) and were higher than at the end of therapy 13.9 ± 25.1 pg/ml (mean ± SD) (p = 0.001). In contrast, plasma levels of TNF-alpha evaluated by cytotoxicity assay did not follow a predicted course during follow-up. Lysis, in this case, might be not totally attributed to TNF-alpha. The discrepancy might be attributed to the presence of factor(s) known to influence the release and activity of TNF-alpha.

Cytokines; TNF-alpha; Visceral Leishmaniasis (Kala-Azar)

Avaliação de TNF-alfa em pacientes com calazar tem despertado grande interesse devido ao seu papel no sistema imunológico e à sua atividade caquetizante. O objetivo deste estudo foi examinar a associação entre os níveis plasmáticos de TNF-alfa, medidos através de sua imunorreatividade (ELISA) e bioatividade (ensaio citotóxico sobre as células L-929), e as manifestações clínicas da leishmaniose visceral. Amostras de 19 pacientes foram obtidas para determinação do TNF-alfa antes, durante e após a terapia antimonial, utilizando o ensaio de citotoxicidade (todos os pacientes) e o ELISA (14 pacientes). Resultados discrepantes entre os ensaios de citotoxicidade e o ELISA foram observados. Níveis circulantes de TNF-alfa, medidos pelo ELISA, foram mais altos nos pacientes que nos controles e declinaram significantemente com a melhora clínica e laboratorial. Níveis plasmáticos antes do tratamento (média = 124,7 pg/ml; DP = 93,3) foram mais elevados que ao final da terapêutica (13,9 pg/ml; DP = 25,1; p = 0,001). Por outro lado, níveis plasmáticos de TNF-alfa, avaliados pela citotoxicidade, não seguiram um curso previsível durante a evolução. Esta discrepância pode ser devida à presença de fatores no plasma que podem influenciar a liberação e atividade do TNF-alfa. Ainda, a lise observada pode não ser totalmente atribuída ao TNF-alfa.

IMMUNOLOGY

Plasma levels of tumor necrosis factor-a in patients with visceral leishmaniasis (Kala-Azar). Association with activity of the disease and clinical remisson following antimonial therapy

Níveis plasmáticos do fator de necrose tumoral-a (TNF-a) em pacientes com leishmaniose visceral (Calazar). Associação com atividade da doença e remissão clínica com terapia antimonial

Reinaldo Salomão^I; Adauto Castelo Filho^I; Iara Marques de Medeiros^II; Miguel Angel Sicolo^II

^IDivision of Infectious Diseases. Escola Paulista de Medicina, Rua Botucatu 740, 04023-062 São Paulo, SP, Brasil

^IIDivision of Infectious Diseases. Universidade Federal do Rio Grande do Norte, Rua Cônego Monte s/n, 59000 Natal, RN, Brasil

^{Correspondence to} Correspondence to: Reinaldo Salomão Division of Infectious Diseases, Escola Paulista de Medicina Rua Botucatu 740 04023-062 São Paulo, SP, Brasil

SUMMARY

Evaluation of TNF-alpha in patients with Kala-azar has drawn increasing interest due to its regulatory role on the immune system, in addition to its cachetizing activity. The objective of this study was to examine the association between plasma levels of TNF-alpha, measured by immunore-activity (ELISA) and bioactivity (cytotoxicity assay with L-929 cells), and clinical manifestations of visceral leishmaniasis. Plasma samples from 19 patients with Kala-azar were obtained before, during and at the end of antimonial therapy. TNF-alpha determinations was done by using the cytotoxicity assay (all patients) and the enzyme-linked immunoassay (ELISA - 14 patients). A discrepancy between results obtained by ELISA and cytotoxicity assay was observed. Levels of circulating TNF-alpha, assessed by ELISA, were higher in patients than in healthy controls, and declined significantly with improvement in clinical and laboratory parameters. Plasma levels before treatment were 124.7 ± 93.3 pg/ml (mean ± SD) and were higher than at the end of therapy 13.9 ± 25.1 pg/ml (mean ± SD) (p = 0.001). In contrast, plasma levels of TNF-alpha evaluated by cytotoxicity assay did not follow a predicted course during follow-up. Lysis, in this case, might be not totally attributed to TNF-alpha. The discrepancy might be attributed to the presence of factor(s) known to influence the release and activity of TNF-alpha.

Keywords: Cytokines; TNF-a; Visceral Leishmaniasis (Kala-Azar)

RESUMO

Avaliação de TNF-a em pacientes com calazar tem despertado grande interesse devido ao seu papel no sistema imunológico e à sua atividade caquetizante. O objetivo deste estudo foi examinar a associação entre os níveis plasmáticos de TNF-a, medidos através de sua imunorreatividade (ELISA) e bioatividade (ensaio citotóxico sobre as células L-929), e as manifestações clínicas da leishmaniose visceral. Amostras de 19 pacientes foram obtidas para determinação do TNF-a antes, durante e após a terapia antimonial, utilizando o ensaio de citotoxicidade (todos os pacientes) e o ELISA (14 pacientes). Resultados discrepantes entre os ensaios de citotoxicidade e o ELISA foram observados. Níveis circulantes de TNF-a, medidos pelo ELISA, foram mais altos nos pacientes que nos controles e declinaram significantemente com a melhora clínica e laboratorial. Níveis plasmáticos antes do tratamento (média = 124,7 pg/ml; DP = 93,3) foram mais elevados que ao final da terapêutica (13,9 pg/ml; DP = 25,1; p = 0,001). Por outro lado, níveis plasmáticos de TNF-a, avaliados pela citotoxicidade, não seguiram um curso previsível durante a evolução. Esta discrepância pode ser devida à presença de fatores no plasma que podem influenciar a liberação e atividade do TNF-a. Ainda, a lise observada pode não ser totalmente atribuída ao TNF-a.

Full text available only in PDF format.

Texto completo disponível apenas em PDF.

Recebido para publicação em 09/06/1995

Aceito para publicação em 15/03/1996

This study was, in part, supported by grant 520768/93-0 from Conselho Nacional de Desenvolvimento Científico e Tecnológico (National Council for Scientific and Technologic Development) - CNPq.

Iara Marques de Medeiros was supported by a grant from Fundação Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES).

Informed Consent was obtained from patients or their parents or guardians before enrollment in this investigation.

1. AGGARWAL, B.B.; KHOR, W.J. & HASS, P.E. - Human tumor necrosis factor, production, purification, and characterization. J. biol. Chem., 260: 2345-2354, 1985.
2. AUKRUST, P.; LIABAKK, N.B.; MÜLLER, F. et al. - Serum levels of tumor necrosis factor-alpha (TNF-alpha) and soluble TNF receptors in human immunodeficiency virus type 1 infection. Correlations to clinical, immunologic, and virologic parametrs. J. infec. Dis., 169: 420-424, 1994.
3. BADARÓ, R.; JONES, T.C.; LOURENÇO, R. et al. - A prospective study of visceral leishmaniasis in an endemic area of Brazil. J. infect. Dis., 154: 639-649, 1986.
4. BARRAL, N.; BADARÓ, R.; BARRAL, A. et al. - Tumor necrosis factor (Cachectin) in human visceral leishmaniasis. J. infect. Dis., 163: 853-857, 1991.
5. BEUTLER, B.A.; MILSARK, I.W. & CERAMI, A. - Cachectin/tumor necrosis factor: production, distribution and metabolic fate in vivo. J. Immunol., 135: 3972-3977, 1985.
6. BEUTLER, B.; MILSARK, I. & CERAMI, A. - Passive immunization against cachectin/tumor necrosis factor protects mice from lethal effects ofendotoxin. Science, 229: 869-871, 1985.
7. CARVALHO, E. M.; BARRAL, A.; SAMPAIO, D. P. et al. - Immunologic markers of clinical evolution in children recently infected with Leishmania donovani chagasi. J. infec. Dis., 165: 535-540, 1992.
8. CERAMI, A.P. & BEUTLER, B. - The role of cachectin/TNF in endotoxic shock and cachexia. Immunol. today, 9: 28-31, 1988.
9. ENGELBERTS, I.; STEPHENS, S.; FRANCOT, G.J.M.; VANDER LINDEN, C.J. & BUURMAN, W.A. - Evidence for different effects of soluble TNF-receptors on various TNF measurements in human biological fluids. Lancet, 2: 515-516, 1991.
10. FOMSGAARD, A.; FREUDENBERG, K.; BENDTZEN, K. & GALANOS, C. - Quantitation and biological activities of native tumour necrosis factor from LPS-stimulated human monocytes. APMIS, 98: 529-534, 1990.
11. GALANOS, C.; FRENDENBERG, M.A.; KATSCHINSKI, T. et al. - Tumor necrosis factor and response to endotoxin. In: RYAN, J.L. Bacterial endotoxic lipopolysaccharides. Immunopharmacotogy and pathophysiology. London, CRC Press, 1992. v. 2, p. 75-100.
12. GRAU, G.E.; FAJARDO, L.F.; PIQUET, P.F. et al. - Tumor necrosis factor (cachetin) is an essential mediator in murine cerebral malaria. Science, 237: 1210-1212, 1987.
13. HO, J.L.; BADARÓ, R.; SCHWARTZ, A. et al. - Diminished in vitro production of interleukin-1 and tumor necrosis factor-alpha during acute visceral leishmaniasis and recovery after therapy. J. infect. Dis., 165: 1094-1102, 1992.
14. KINDLER, V; SAPPINO, A.P.; GRAU, G.E.; PIGUET, P.F. & VASSALLI, P. - The inducing role of tumor necrosis factor in the development of bactericidal granulomas during BCG infection. Cell, 56: 731-740, 1989.
15. LIEW, F.Y.; PARKINSON, C.; MILLOTT, S.; SEVERN, A. & CARRIER, M. - Tumor necrosis factor (TNF-alpha) in leishmaniasis. TNF-alpha mediates host protection against cutaneous leishmaniasis. Immunology, 69: 570-573, 1989.
16. NAKANE, A.; MINAGAWA, T.; KOHANAWA, M. et al. - Interactions between endogenous gamma-interferon and tumor necrosis factor in host-response against primary and secondary Listeria monocytogenes infections. Infect. Immun., 57: 3331-3336, 1989.
17. OMS - Comité de expertos en la lucha contra las leishmaniasis. Lucha contra las leishmaniasis: informe de um Comité de Expertos de la OMS. Org. Mund. Salud Ser. Inf. Técn., (793): 12, 1990.
18. PEARSON, R.D. & SOUSA, A.Q. - Leishmania species: visceral (Kala-Azar), cutaneous, and mucosal leishmaniasis. In: MANDELL, G.L.; DOUGLAS, R.G. & BENNET, J.B., ed. - Principles and practice of infectious diseases. 3. ed. New York, Churchill Livingstone, 1990. p. 2066-2077.
19. PETERSEN, C.M. & MOLLER, B.K. - Immunological reactivity and bioactivity of tumor necrosis factor. Lancet, 1: 1939-1940, 1988.
20. PISA, P.; GENNENE, M.; SODER, O. et al. - Serum tumor necrosis factor levels and disease dissemination in leprosy and leishmaniasis. J. infect. Dis., 161: 988-991, 1990.
21. SCUDERI, P.; LAM, K.S. & RYAN, K.J. - Raised serum levels of tumour necrosis factor in parasitic infections. Lancet, 2: 1364-1365, 1986.
22. TITUS, R.G.; SHERRY, B. & CERAMI, A. - Tumor necrosis factor plays a protective role in experimental murine cutaneous leishmaniasis. J. exp. Med., 170: 2079-2104, 1987.
23. TRACEY, K.J.; FONG, Y.; HESSE, D.G. et al. - Anti-cachetin/TNF monoclonal antibodies prevent septic shock during lethal bacteremia. Nature, 330: 662-664, 1987.
24. TRACEY, K.J.; WEI, H. & MANOGUE, K.R. - Cachetin/tumor necrosis factor induces cachexia, anemia, and inflammation. J. exp. Med., 167: 1211-1227, 1988.
25. TUMANG, M.C.T.; KEAGH, C.; MOLDAWER, L.L. et al. - Role and effect of TNF-alpha in experimental visceral leishmaniasis. J. Immunol., 153: 768-775, 1994.
26. VASSALLI, P. - The pathophysiology of tumor necrosis factor. Ann. Rev. Immunol., 10: 411-452, 1992.
27. WAAGE, A.; HALSTENSEN, A. & ESPEVIK, T. - Association between tumor necrosis factor in serum and fatal outcome in patients with meningococcal disease. Lancet, 1: 355-357, 1987.
28. WILLIAMS, D.M.; MAGEE, D.M.; BONEWALD, L.F. et al. - A role in vivo for tumor necrosis factor alpha in host defense against Chlamydia trachomatis. Infect. Immun., 58: 1572-1576, 1987.

Correspondence to:

Reinaldo Salomão

Division of Infectious Diseases, Escola Paulista de Medicina

Rua Botucatu 740

04023-062 São Paulo, SP, Brasil

Publication Dates

Publication in this collection
22 Sept 2006
Date of issue
Apr 1996

History

Accepted
15 Mar 1996
Received
09 June 1995

This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

[1] 1. AGGARWAL, B.B.; KHOR, W.J. & HASS, P.E. - Human tumor necrosis factor, production, purification, and characterization. J. biol. Chem., 260: 2345-2354, 1985.

[2] 2. AUKRUST, P.; LIABAKK, N.B.; MÜLLER, F. et al. - Serum levels of tumor necrosis factor-alpha (TNF-alpha) and soluble TNF receptors in human immunodeficiency virus type 1 infection. Correlations to clinical, immunologic, and virologic parametrs. J. infec. Dis., 169: 420-424, 1994.

[3] 3. BADARÓ, R.; JONES, T.C.; LOURENÇO, R. et al. - A prospective study of visceral leishmaniasis in an endemic area of Brazil. J. infect. Dis., 154: 639-649, 1986.

[4] 4. BARRAL, N.; BADARÓ, R.; BARRAL, A. et al. - Tumor necrosis factor (Cachectin) in human visceral leishmaniasis. J. infect. Dis., 163: 853-857, 1991.

[5] 5. BEUTLER, B.A.; MILSARK, I.W. & CERAMI, A. - Cachectin/tumor necrosis factor: production, distribution and metabolic fate in vivo. J. Immunol., 135: 3972-3977, 1985.

[6] 6. BEUTLER, B.; MILSARK, I. & CERAMI, A. - Passive immunization against cachectin/tumor necrosis factor protects mice from lethal effects ofendotoxin. Science, 229: 869-871, 1985.

[7] 7. CARVALHO, E. M.; BARRAL, A.; SAMPAIO, D. P. et al. - Immunologic markers of clinical evolution in children recently infected with Leishmania donovani chagasi. J. infec. Dis., 165: 535-540, 1992.

[8] 8. CERAMI, A.P. & BEUTLER, B. - The role of cachectin/TNF in endotoxic shock and cachexia. Immunol. today, 9: 28-31, 1988.

[9] 9. ENGELBERTS, I.; STEPHENS, S.; FRANCOT, G.J.M.; VANDER LINDEN, C.J. & BUURMAN, W.A. - Evidence for different effects of soluble TNF-receptors on various TNF measurements in human biological fluids. Lancet, 2: 515-516, 1991.

[10] 10. FOMSGAARD, A.; FREUDENBERG, K.; BENDTZEN, K. & GALANOS, C. - Quantitation and biological activities of native tumour necrosis factor from LPS-stimulated human monocytes. APMIS, 98: 529-534, 1990.

[11] 11. GALANOS, C.; FRENDENBERG, M.A.; KATSCHINSKI, T. et al. - Tumor necrosis factor and response to endotoxin. In: RYAN, J.L. Bacterial endotoxic lipopolysaccharides. Immunopharmacotogy and pathophysiology. London, CRC Press, 1992. v. 2, p. 75-100.

[12] 12. GRAU, G.E.; FAJARDO, L.F.; PIQUET, P.F. et al. - Tumor necrosis factor (cachetin) is an essential mediator in murine cerebral malaria. Science, 237: 1210-1212, 1987.

[13] 13. HO, J.L.; BADARÓ, R.; SCHWARTZ, A. et al. - Diminished in vitro production of interleukin-1 and tumor necrosis factor-alpha during acute visceral leishmaniasis and recovery after therapy. J. infect. Dis., 165: 1094-1102, 1992.

[14] 14. KINDLER, V; SAPPINO, A.P.; GRAU, G.E.; PIGUET, P.F. & VASSALLI, P. - The inducing role of tumor necrosis factor in the development of bactericidal granulomas during BCG infection. Cell, 56: 731-740, 1989.

[15] 15. LIEW, F.Y.; PARKINSON, C.; MILLOTT, S.; SEVERN, A. & CARRIER, M. - Tumor necrosis factor (TNF-alpha) in leishmaniasis. TNF-alpha mediates host protection against cutaneous leishmaniasis. Immunology, 69: 570-573, 1989.

[16] 16. NAKANE, A.; MINAGAWA, T.; KOHANAWA, M. et al. - Interactions between endogenous gamma-interferon and tumor necrosis factor in host-response against primary and secondary Listeria monocytogenes infections. Infect. Immun., 57: 3331-3336, 1989.

[17] 17. OMS - Comité de expertos en la lucha contra las leishmaniasis. Lucha contra las leishmaniasis: informe de um Comité de Expertos de la OMS. Org. Mund. Salud Ser. Inf. Técn., (793): 12, 1990.

[18] 18. PEARSON, R.D. & SOUSA, A.Q. - Leishmania species: visceral (Kala-Azar), cutaneous, and mucosal leishmaniasis. In: MANDELL, G.L.; DOUGLAS, R.G. & BENNET, J.B., ed. - Principles and practice of infectious diseases. 3. ed. New York, Churchill Livingstone, 1990. p. 2066-2077.

[19] 19. PETERSEN, C.M. & MOLLER, B.K. - Immunological reactivity and bioactivity of tumor necrosis factor. Lancet, 1: 1939-1940, 1988.

[20] 20. PISA, P.; GENNENE, M.; SODER, O. et al. - Serum tumor necrosis factor levels and disease dissemination in leprosy and leishmaniasis. J. infect. Dis., 161: 988-991, 1990.

[21] 21. SCUDERI, P.; LAM, K.S. & RYAN, K.J. - Raised serum levels of tumour necrosis factor in parasitic infections. Lancet, 2: 1364-1365, 1986.

[22] 22. TITUS, R.G.; SHERRY, B. & CERAMI, A. - Tumor necrosis factor plays a protective role in experimental murine cutaneous leishmaniasis. J. exp. Med., 170: 2079-2104, 1987.

[23] 23. TRACEY, K.J.; FONG, Y.; HESSE, D.G. et al. - Anti-cachetin/TNF monoclonal antibodies prevent septic shock during lethal bacteremia. Nature, 330: 662-664, 1987.

[24] 24. TRACEY, K.J.; WEI, H. & MANOGUE, K.R. - Cachetin/tumor necrosis factor induces cachexia, anemia, and inflammation. J. exp. Med., 167: 1211-1227, 1988.

[25] 25. TUMANG, M.C.T.; KEAGH, C.; MOLDAWER, L.L. et al. - Role and effect of TNF-alpha in experimental visceral leishmaniasis. J. Immunol., 153: 768-775, 1994.

[26] 26. VASSALLI, P. - The pathophysiology of tumor necrosis factor. Ann. Rev. Immunol., 10: 411-452, 1992.

[27] 27. WAAGE, A.; HALSTENSEN, A. & ESPEVIK, T. - Association between tumor necrosis factor in serum and fatal outcome in patients with meningococcal disease. Lancet, 1: 355-357, 1987.

[28] 28. WILLIAMS, D.M.; MAGEE, D.M.; BONEWALD, L.F. et al. - A role in vivo for tumor necrosis factor alpha in host defense against Chlamydia trachomatis. Infect. Immun., 58: 1572-1576, 1987.

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