Differential decline in Leishmania membrane antigen-specific immunoglobulin G (IgG), IgM, IgE, and IgG subclass antibodies in Indian kala-azar patients after chemotherapy

Abstract

Pathogenesis in kala-azar is associated with depressed cellular immunity and significant elevation of antileishmanial antibodies. Since these antibodies are present even after cure, analysis of the parasite-specific isotypes and immunoglobulin G (IgG) subclasses in kala-azar patients may shed new light on the immune responses during progression and resolution of infection. Using leishmanial membrane antigenic extracts, we investigated the relative levels of specific IgG, IgM, IgA, IgE, and IgG subclasses in Indian kala-azar patient sera during disease, drug resistance, and cure. Acute-phase sera showed strong stimulation of IgG, followed by IgE and IgM and lastly by IgA antibodies. IgG subclass analysis revealed expression of all of the subclasses, with a predominance of IgG1 during disease. Following sodium stibogluconate (SAG) resistance, the levels of IgG, IgM, IgE, and IgG4 remained constant, while there was a decrease in the titers of IgG2 and IgG3. In contrast, a significant (2.2-fold) increase in IgG1 was observed in these individuals. Cure, in both SAG-responsive and unresponsive patients, correlated with a decline in the levels of IgG, IgM, IgE, and all of the IgG subclasses. The stimulation of IgG1 and the persistence, most importantly, of IgE and IgG4 following drug resistance, along with a decline in IgE, IgG4, and IgG1 with cure, demonstrate the potential of these isotypes as possible markers for monitoring effective treatment in kala-azar.

FIG. 1

Levels of LAg-specific IgG (A), IgM (B), IgA (C), and IgE (D) in the sera of Indian kala-azar patients before treatment and following SAG unresponsiveness and cure. Symbols represent individual patients, and the same symbols indicating the same patients are used in Fig. 2. The heavy line represents the mean (± standard error) of the Ig isotype level for five individuals.

FIG. 2

Specific IgG subclass levels in kala-azar patient sera before treatment and following SAG resistance and cure. The symbols and the heavy line are as described for Fig. 1.

FIG. 3

LAg-specific IgG (A), IgM (B), IgA (C), and IgE (D) serum antibodies of kala-azar patients before and after SAG therapy. Symbols represent individual patients, and the same symbols depict the same patients in Fig. 4. The heavy line represents the mean (± standard error) of the Ig isotype level for 10 samples.

FIG. 4

IgG subclass antibody titers of Indian kala-azar patients before and after treatment with SAG. The symbols and the heavy line are as described for Fig. 3.

FIG. 5

Immunoblots of LAg with sera from untreated and cured kala-azar patients. Lanes 1 to 3, reactivity with untreated, SAG-unresponsive, and amphotericin B-cured sera of one kala-azar patient at 0, 45, and 75 days posttreatment, respectively. Lanes 4 and 5, reactivity with untreated and SAG-cured sera of another kala-azar patient at 0 and 50 days posttreatment, respectively. The reactivities with the major bands are indicated on the right.