Involvement of T cells in enhanced resistance to Klebsiella pneumoniae septicemia in mice treated with liposome-encapsulated muramyl tripeptide phosphatidylethanolamine or gamma interferon

Abstract

We have previously shown that prophylactic administration of the liposome-encapsulated immunomodulating agents muramyl tripeptide phosphatidylethanolamine (MTPPE) and gamma interferon (IFN-gamma) results in strongly increased survival of mice from a normally lethal septicemia with Klebsiella pneumoniae. It was anticipated that the treatment acts on macrophages and nonspecifically augments host resistance to various infections. In the present study, we provide evidence for a key role for T cells in host defense potentiation by the liposomal immunomodulators toward K. pneumoniae septicemia. It is shown that both CD4 and CD8 cells are important in immunomodulation, most likely due to production of IFN-gamma. Depletion of circulating IFN-gamma resulted in strong reduction of the antimicrobial host defense activation. Administration of interleukin-10 resulted in decreased antimicrobial host defense activation by liposomal immunomodulators. Moreover, administration of liposomal immunomodulators was shown to induce predominantly T-helper type 1 (Th1) cell populations in the spleen. These findings indicate that immunomodulation with liposomal MTPPE and IFN-gamma favors Th1 and NK cell activation.

FIG. 1

Survival of T-cell-depleted mice infected by i.p. inoculation of 10³ CFU of K. pneumoniae. T cells were depleted by injection of CD3-specific IgG2b MAb. Mice were treated before infection with LE-MTPPE/IFN-γ (——–), LE-MTPPE (––––), LE-IFN-γ (—––), or PL (———). Control mice were injected with rat IgG2b anti-β-galactosidase MAb and treated with LE-MTPPE/IFN-γ (——). Each group contained 20 mice.

FIG. 2

Survival of CD4⁺ cell-depleted mice infected by i.p. inoculation of 10³ CFU of K. pneumoniae. CD4⁺ cells were depleted by injection of CD4-specific IgG2b MAb. Mice were treated before infection with LE-MTPPE/IFN-γ (——–), LE-MTPPE (––––), LE-IFN-γ (—––), or PL (——). Control mice were injected with rat IgG2b anti-β-galactosidase MAb and treated with LE-MTPPE/IFN-γ (——). Each group contained 20 mice.

FIG. 3

Survival of CD8⁺ cell-depleted mice infected by i.p. inoculation of 10³ CFU of K. pneumoniae. CD8⁺ cells were depleted by injection of CD8-specific IgG2b MAb. Mice were treated before infection with LE-MTPPE/IFN-γ (—–), LE-MTPPE (––––), LE-IFN-γ (——––), or PL (——). Control mice were injected with rat IgG2a anti-β-galactosidase MAb and treated with LE-MTPPE/IFN-γ (——). Each group contained 20 mice.

FIG. 4

Survival of mice depleted of endogenous IFN-γ and infected by i.p. inoculation of 10³ CFU of K. pneumoniae. IFN-γ was depleted by injection of IFN-γ-specific IgG2a MAb. Mice were treated before infection with LE-MTPPE/IFN-γ (——–), LE-MTPPE (––––), LE-IFN-γ (——––), or PL (——). Control mice were injected with rat IgG2a anti-β-galactosidase MAb and treated with LE-MTPPE/IFN-γ (——). Each group contained 20 mice.

FIG. 5

Survival of mice injected i.p. with IL-10-producing cells in alginate and infected by i.p. inoculation of 10³ CFU of K. pneumoniae. Mice were treated before infection with LE-MTPPE/IFN-γ (——–), LE-MTPPE (––––), LE-IFN-γ (——–), or PL (——). Control mice were injected with mock-transfected alginate-encapsulated cells and treated with LE-MTPPE/IFN-γ (——). Each group contained 20 mice.

FIG. 6

Survival of mice depleted of endogenous TNF-α and infected by i.p. inoculation of 10³ CFU of K. pneumoniae. TNF-α was depleted by injection of TNF-α-specific polyclonal rabbit IgG. Mice were treated before infection with LE-MTPPE/IFN-γ (——–) or PL (——). Control mice were injected with an equal amount of rabbit IgG and treated with LE-MTPPE/IFN-γ (——). Each group contained 20 mice.

FIG. 7

Ia antigen expression on spleen cells measured by flow cytometry in mice treated with LE-MTPPE/IFN-γ, PL, or PBS. One group of mice was also infected by i.p. inoculation of 10³ CFU of K. pneumoniae (LE-MTPPE/IFN-γ day 4 inf) 12 h after the end of treatment. Spleen cells were examined 12 h and 4 days (day 4) after the end of treatment. Each bar represents the average + standard deviation of three mice. FITC, fluorescein isothiocyanate.

FIG. 8

Presence of Th1 and Th2 cells in spleens of mice treated with LE-MTPPE/IFN-γ, measured by production of IFN-γ and IL-4. Three mice were used per treatment group, and cytokine production was determined in quadruplicate. T cells were isolated 12 h (A and C and 4 days (B and D after the end of treatment. Cells were cultured in anti-CD3-coated plates in the presence of IL-2 (IL-2), IL-2, IFN-γ, and anti-IL-4 (Th1), IL-2, IL-4, and anti-IFN-γ (Th2), or medium alone (-). Horizontal lines represent averages of the determinations. ○, PL-treated controls; •, LE-MTPPE/IFN-γ-treated mice.