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Advances in Environmental Biology, 5(8): 2117-2121, 2011
ISSN 1995-0756
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ORIGINAL ARTICLE
The Level of Ige and Iga Isotypes in Leishamania Infantum Resistant and Susceptible
Mice
1
Eilyad Issabeagloo, 2Parviz Kermanizadeh, 3Farhad Ahmadpoor, 4Jamshid Ghiasi Ghalahkandi
1
Department of Pharmacology, Medical Sciences Faculty, Tabriz branch, Islamic Azad University, Tabriz, Iran.
Department of Parasitology, Medical Sciences Faculty, Tabriz branch, Islamic Azad University, Tabriz, Iran.
3
Department of Internal Medicine, Medical Sciences Faculty, Tabriz branch, Islamic Azad University, Tabriz,
Iran.
4
Department of animal Sciences, Veterinary Faculty, shabestar branch, Islamic Azad University, shabestar,
Iran.
2
Eilyad Issabeagloo, Parviz Kermanizadeh, Farhad Ahmadpoor, Jamshid Ghiasi Ghalahkandi: The Level
of Ige and Iga Isotypes in Leishamania Infantum Resistant and Susceptible Mice
ABSTRACT
IgE and IgA isotopes in serum were considered in susceptible BALB/C and resistant C57BL/6 mice during
36 weeks leishmania infection. Association of the IgA level in both mice with early phase of the disease may
discriminate the acute and chronic infection. And correlation of IgE level with intensity of the disease may
be used as a tool to estimate the effects of the drug in the treatment.
Key words: leishmania, IgA, IgE, C57BL/6, BALB/C.
Therefore studying the IgE during visceral
leishmaniasis was of great interest.
Introduction
Leishmaniases refers to a number of diseases
caused by the intracellular parasite Leishmania [1].
Leishmania presents different clinical expressions
depending on both species of parasite and host
immune responses [2,3].
BALB/c mice are susceptible to parasite and
develop large and non healing lesions in the case of
infection, other mouse strains, such as C3H, CBA/J,
and C57BL/6 are resistant and develop small lesions
that heal easily. Resistivity is related to Th1-type
immune responses and Th1 derived cytokines.
Susceptibility on the other hand depends on the Th2
immune response and its characteristic mediators
[4,5]. Depressed cell mediated immunity and elevated
humoral immune responses are associated with the
pathogenesis of visceral leishmaniais. The major role
for IgE and IgA antibodies has been demonstrated
[6,7,8,910] on gastrointestinal dwelling parasites.
Material and method
A total of 210 female BALB/C and C57BL/6, 8
to 10 weeks old mice were infected with L infantum,
by injecting of 2 X 106 promastigotes into the
peritoneum. Four infected & two control BALB/C
and C57BL/6 mice were anesthetized each every ten
days and blood was obtained through their heart and
Serum was frozen until examined for Ab detection.
Leishmania antigen isolated from promastigote of L.
infantum which was grown in culture medium [11].
The amount of protein of antigen was tested by the
method of Lowry [12].
The amount of IgE and IgA antibodies was
measured by the Enzyme-linked immunosorbent assay
(ELISA) method, which has been described already
[13,14,15].
Corresponding Author
Eilyad Issabeagloo, Department of Pharmacology, Medical Sciences Faculty, Tabriz branch,
Islamic Azad University, Tabriz, Iran.
Tel: +989144079927
E-mail: [email protected]
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Adv. Environ. Biol., 5(8): 2117-2121, 2011
Immunosorbent assay-standard micro-well and
Peroxidase conjugated antibodies specific to mice
IgA, IgE and ELISA reader was employed. The
results of the reaction were read at 492nm and
expressed as optical density (O.D.). Serums were
examined throughout 36 weeks of infection.
-
-
Results:
-
-
Infected serum was diluted 1:10, 1:50, 1:100 and
1:200. The best response was observed with the
50 fold dilution in preliminary examination;
therefore all the tests were carried on with 1:50
dilution during the experiments. Concentration of
anti-leishmanial IgE and IgA was explained by
means of optical density reading. An increase
in the IgA level was observed in infected
BALB/C mice after 10 days of infection and
reached a maximum at 21 weeks and gradually
decreased later (Fig.1).
The IgE level in infected BALB/C mice was
also increased 10 days after infection and stayed
increasing until the endof the experiments
(Fig.2).
-
-
The IgE level in infected C57BL/6 mice was
increased 20 days after infection and reached a
maximum at 20 weeks and was gradually
reduced until the end of the experiments (Fig.3).
The IgA level in infected C57BL/6 mice was
increased 40 days after infection and reached a
maximum at 24 weeks and was reduced sharply
by week 27 post infection (Fig.4).
At the beginning the level of IgE in infected
BALB/C and C57BL/6 mice was gradually
increased and reached a maximum at week 20.
In comparison to infected BALB/C the level of
IgE in infected C57BL/6 mice gradually
decreased by week 22 post infections (Fig.5).
The level of IgA in infected BALB/C and
C57BL/6 mice was increased and reached a
maximum at week 22 post infection. The level
of IgA in infected BALB/C was always higher.
After week 22 the level of IgA in both strains
decreased gradually and was reduced to the same
level as the control animals at 28 weeks post
infection (Fig.6).
Fig. 1: Mean IgA level in Leishmania infantum infected and control BALB/C mice.
Fig. 2: Mean IgE level in leishmania infantum infected & controls BALB/C mice .
Adv. Environ. Biol., 5(8): 2117-2121, 2011
Fig. 3: Mean IgE level in leishmania infantum infected and controls C57BL/6 mice
Fig. 4: Mean IgA Level in leishmania infantum infected and controls C57 BL/6 mice.
Fig. 5: Mean IgE level in Leishmania infantum infected BALBE/C & C57 BL/6 mice.
Fig. 6: Mean IgA level in Leishmania infantum infected BALBC& C57 BL/6 mice.
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Adv. Environ. Biol., 5(8): 2117-2121, 2011
Discussion:
Successful host defence and pathogenicity in
leishmaniasis depends on T-cell polarization.
Resistance to the Leishmania infection in the murine
model is based on the activation of the cellular
immune responses organized by the Th1 cells,
making special cytokins including IFN-γ that recruits
CMI. Leishmania resistant strain of mice such as
C57Bl/6 genetically produces Th1 immune responses
and shows only a local reaction that heals easly
[16,17]. On the other hand infected BALB/C mice
generally activate Th2 cells and regulate humoral
immune responses which are associated with severe
systemic diseases [18]. The effect of humoral
response in acute and chronic phase of parasitic
diseases have been studied [19,20]. Systemic parasitic
infections are associated with high level antigenspecific antibodies [21,22] IgE production is
dependent on IL-4, a cytokine involved in
differentiation of CD4+ Th2 cells [23]. Th2 cells
may also increase IgA production, but it is not fully
investigated in the Leishmaniasis. IgE can be used
to detect the occurrence of some Th2 depentent
pathological conditions such as autoimmune diseases,
parasitic, and viral infections [24,25,26]. IgE is also
associated with allergic reactions [27] and has been
used in diagnosis of parasitic diseases [28]. A high
level of IgA has been found in schistosomiasis
against schistosoma antigen at acute phase of
infection [29,30]. IgA might protect patients against
schistosomiasis [32,33].
In this experiment we assayed specific IgE and
IgA antibodies to L.infantum antigens by ELISA
method during 36 weeks. In comparison to control
animals we observed high levels of IgE and IgA
isotypes in the leishmania-infected BALB/C and
C57BL/6 mice Fig.1, 2, 3, 4. Continuously increasing
the level of IgE in leishmania infected BALB/C mice
explains the susceptibility of the animals to infection
and the fact that parasites still exsist in the body.
Decreasing the level of IgE in C57BL/6 mice after
week 21 may be an evidence for improving the CMI
in these mice and clearance or reducing of the
parasites in the body. Increasing the level of IgA
until week 28 and reducing the antibody to a very
low level later on in both BALB/C and C57BL/6
mice may explain a role for IgA in early phase of
the infection. The same role that has been found in
schistosomiasis [29,30]. During the experience the
level of IgA in BALB/C mice was always higher
than C57BL/6. This makes it hard to speculate any
protective role for IgA antibody during the
leishmaniasis. The following ideas can be
hypothesized from the results of this experiment:
1.
A high level of IgA against leishmania antigen
may be useful to differentiation of acute and
chronic phase of leishmania infection.
2.
3.
A sufficient quantity of IgA in acute phase of
leishmaniasis may play a preliminary supportive
role in providing the protection against the
parasite.
Detection of high level of IgM against
leishmania antigens may be considerd as a serum
marker for evaluation of the disease activity and
proper treatments.
References
1.
Peter, W., Killick-Kendrick., 1987. The
leishmaniasis in biology and Medicine, 1 and 2.
Academic press, London, pp: 941.
2. Carvalho, E.M., A. Barral, J.M.L. Costa, A.
Bittencourt and P. Marsden., 1994. Clinical and
imunopathological aspects of disseminated
cutaneous leishmaniasis. Acta Trop., 56: 315325.
3. Neva, F.A., 1992. Leishmaniasis, pp: 1982-1987.
In J. B. Wyngaarden, L. H. Smith, Jr., and J. C.
Bennett (ed.), Cecil textbook of medicine, 19th
ed. Saunders Company, London, United
Kingdom.
4. Heinzel, F.P., M.D. Sadick, S.S. Mutha, and R.
M. Locksley., 1991. Production of interferongamma, interleukin-2, interleukin-4 and
interleukin-10 by CD4+ lymphocytes in vivo
during healing and progressive murine
leishmaniasis. Proc. Natl. Acad. Sci. USA 88:
7011-7015.
5. Hunter, C.A. and S.L. Reiner., 2000. Cytokines
and T cells in host defense. Curr. Opin.
Immunol., 12: 413-418.
6. Miller, H.R.P., 1987. Immunopathology of
nematode infection and expulsion, In
Immunopathology of the Small Intestine (ed.
Marsh, M.N.), pp: 177-208, John Wiley press,
New York,
7. Omata, Y., K. Terada, A. Taka, T. Isamida, M.
Kanda, A. Saito, 1997. Positive evidence that
anti-Toxoplasma gondii IgA antibody exists in
the intestinal tract of infected cats and exerts
protective activity against the infection,
Veterinary parasitology 73(1): 15: 1-11(11).
8. Else, K.J. and D. Wakelin, 1990. Geneticallydetermined influences on the ability of poor
responder mice to respond to immunization
against Trichuris muris, Parasitology 100: 479489.
9. Roach, T., K.J. Else, D. Wakelin, D.J. McLaren
and R.K. Grencis, 1991. Trichuris muris:
Antigen recognition and transfer of immunity in
mice by IgA monoclonal antibodies, Parasite
Immunol., 13: 1-12.
10. Inaba, H., Sato, H. Kamiya and Monoclonal,
2003. IgA antibody-mediated expulsion of
Trichinella from the intestine of mice,
Parasitology, 126: 591-598.
2121
Adv. Environ. Biol., 5(8): 2117-2121, 2011
11. Afrin, F. and N. Ali., 1997. Adjuvanticity and
protective immunity elicited by Leishmania
donovani antigen encapsulated in positively
charged liposomes. Infect. Immun., 65: 23712377.
12. Lowry, O.H., N.J. Rosebrough, A.L. Farr and R.
J. Randall., 1951. Protein measurement with the
Folin phenol reagent. J. Biol. Chem., 193: 265275.
13. Susan Johnson, Dorothee Bourges, Odilia
Wijburg, Richard A. Strugnell and Andrew M.
Lew. Milk, 2006. IgA responses are augmented
by antigen delivery to the mucosal addressin
cellular adhesion molecule1. Vaccine 24: 55525558.
14. Rabello, A.L.T., M.M.A. Garcia, Dias E. Neto,
R.S. Rocha, N. Katz, 1993. Dot-dye-immuno
assay and dor-ELISA for the serological
differentiation of acute and chronic
schistosomiasis using keyhole limpet
haemocyanin as antigen. Trans R Soc Trop Med
Hyg., 87: 279-281.
15. Rabello, A.L.T., M.M.A. Garcia, Pinto da R.
Silva, R.S. Rocha, P. Chaves, N. Katz, 1995.
Humoral immune responses in acute
schistosomiasis mansoni: Relation to morbidity.
Clin Infec Dis., 21: 608-615.
16. Carvalho, E.M., R.S. Teixeira and W.D.
Johnson., 1981. Cell-mediated immunity in
American visceral leishmaniasis: reversible
immunosuppression during acute infection.
Infect. Immun., 33: 498-502.
17. Carvalho, E.M., R. Badaró, S.G. Reed, T.C.
Jones and W.D. Johnson., 1985. Absence of
gamma-IFN and interleukin-2 production during
active visceral leishmaniasis. J. Clin. Investig.
76: 2066-2069.
18. Heinzel, F.P., M.D. Sadick, B.J. Holaday, R.L.
Coffman and R.M. Locksley, 1989. Reciprocal
expression of interferon γ or interleukin 4 during
the resolution or progression of murine
leishmaniasis, J. Exp. Med., 169: 59-72.
19. Gazzinelli, G., J.R. Lambertucci, N. Katz, R.S.
Rocha, M.S. Lima, D.G. Colley, 1985. Immune
reponses during human schistosomiasis mansoni.
XI. Immunologic status of patients with acute
infection and after treatment. J Immunol., 135:
2121-2127.
20. Simpson, A.J.G., Y. Xinyuan, Lillywhite, 1990.
Dissociation of antibody responses during human
schistosomiasis and evidence for enhancement of
granuloma size by anti-carbohydrate IgM. Trans
R. Soc Trop Med Hyg., 84: 808-814.
21. Reis, A.B., A. Teixeira-Carvalho, A.M. Vale,
M.J. Marques, R.C. Giunchetti, W. Mayrink,
L.L. Guerra, R.A. Andrade, R. Correa-Oliviera
and O.A. Martins-Filho, 2006. Isotype patterns
of immunoglobulins: hallmarks for clinical status
22.
23.
24.
25.
26.
27.
28.
29.
30.
31.
32.
33.
and tissue parasite density in Brazilian dogs
naturally infected by Leishmania (Leishmania)
chagasi, Vet. Immunol. Immunopathol., 112:
102-116.
Inestia, M. Gallego and M. Portus, G.
Immunoglobulin and E 2005. Responses in
various stages of canine leishmaniosis, Vet.
Immunol. Immunopathol., 103: 77-81.
Coffman., R.L., K. Varkila, P. Scott and R.
Chatelain, 1991. Role of cytokines in the
differentiation of CD4+ T cell subsets in vivo.
Immunological Reviews, 123: 1-19.
Nagpal, S., P. Sriramarao, P.R. Krishnaswamy,
D.D. Metcalfe and Subba P.V. Rao, 1990.
Demonstration of IgE antibodies to nucleic acid
antigens in patients with SLE. Autoimmunity, 8:
59-64.
King, C.L. and T.B. Nutman, 1993. IgE and IgG
subclass regulation by IL-4 and IFN-gamma in
human helminth infections. Assessment by B cell
precursor frequencies. Journal of Immunology,
151: 458-465
Hagan, P., 1993. IgE and protective immunity to
helminth infections. Parasite Immunology, 15: 14.
Wong, S.Y., M.P. Hajdu, R. Ramirez, P.
Thulliez, R. McLeod and J.S. Remington, 1993.
Role of specific immunoglobulin E in diagnosis
of acute Toxoplasma infection and
toxoplasmosis. Journal of Clinical Microbiology,
31: 2952-2959.
Ishizaka, T., 1982. IgE and mechanisms of IgEmediated hypersensitivity. Annals of Allergy, 48:
313-319.
Kanamura, H.Y., S. Hoshino-Shimizu, M.E.
Camargo, da L.C. Silva, 1979. Class specific
antibodies and fluorescent staining pafferns in
acute and chronic schistosomiasis mansoni. Am
J Trop Med Hyg., 28: 242-248.
Kanamura, H.Y., R.M. Silva, A.L.T. Rabello,
R.S. Rocha, N. Katz, 1991. Anticorpo serico IgA
no diagnostico da fase aguda da esquistossomose
mansoni. Rev Inst Adolfo Lutz, 52: 101-104.
Rabello, A.L.T., M.M.A. Garcia, Pinto da R.
Silva, R.S. Rocha, P. Chaves, N. Katz, 1995.
Humoral immune responses in acute
schistosomiasis mansoni: Relation to morbidity.
Clin Infec Dis., 21: 608-615.
Grzych, J.M., D. Grezel, Bo C. X.u, 1993. IgA
antibodies to a protective antigen in human
schistosomiasis mansoni. J Immunol., 150: 527535.
Boulanger, D., G.D.F. Reid, R.F. Sturrock, 1991.
Immunization of mice and baboons with the
recombinant Sm 28 GST affects both viability
and fecundity after experimental infection with
Schistosoma mansoni. Parasite Immunol., 13:
473-490.