Resistance to and recovery from lethal influenza virus infection in B lymphocyte-deficient mice

Abstract

In the adaptive immune response to most viruses, both the cellular and humoral arms of the immune system play complementary roles in eliminating virus and virus-infected cells and in promoting recovery. To evaluate the relative contribution of CD4+ and CD8+ effector T lymphocytes in virus clearance and recovery, we have examined the host response to lethal type A influenza virus infection in B lymphocyte-deficient mice with a targeted disruption in the immunoglobulin mu heavy chain. Our results indicate that naive B cell-deficient mice have a 50- 100-fold greater susceptibility to lethal type A influenza virus infection than do wild type mice. However, after priming with sublethal doses of influenza, immune B cell-deficient animals show an enhanced resistance to lethal virus infection. This finding indicates that an antibody-independent immune-mediated antiviral mechanism accounts for the increased resistance to lethal virus challenge. To assess the contribution of influenza-specific CD4+ and CD8+ effector T cells in this process, defined clonal populations of influenza-specific CD4+ and CD8+ effector T cells were adoptively transferred into lethally infected B cell-deficient mice. Cloned CD8+ effectors efficiently promoted recovery from lethal infection, whereas cloned CD4+ T cells conferred only partial protection. These results suggest that memory T lymphocytes can act independently of a humoral immune response in order to confer resistance to influenza infection in immune individuals. The potential implications of these results for vaccination against human influenza infection are discussed.

Figure 1

B cell–deficient mice are more susceptible to influenza viral challenge. μKO (a) and C57Bl/6 (b) mice were intranasally inoculated with varying doses (♦, 10⁻³ dilution; ▪, 10⁻⁴ dilution; •, 10⁻⁵ dilution; ▴, 10⁻⁶ dilution; and ✖, 10⁻⁷ dilution) of mouse adapted influenza virus and watched for 21 d for morbidity and mortality.

Figure 2

μKO mice can initiate and maintain an influenza-specific CTL response after challenge with influenza virus. (a) Lungs were removed from μKO (shaded bars) and C57BL/6 (open bars) mice on day 12 after intranasal viral challenge with a sublethal dose of A/JAPAN/57 (attenuated strain). Cell suspensions from the lungs were obtained by processing through a sieve, and were Ficoll purified and plated for a final effector/targer (E/T) ratio of 50:1. Assay time was 6 h and spontaneous release for all targets was <20%. Results of two separate experiments with two mice per experimental group are shown. (b) Influenza specific bulks from four individual μKO (shaded bars) and from C57BL/6 (open bars) mice were tested for their ability to lyse uninfected and A/JAPAN/57 infected class II negative (EL4) and class I and II positive (LB15.13) target cells in a standard chromium release assay. Assay time = 6 h; E/T = 10:1. Spontaneous release for all targets was <15%. Experiment is representative of three separate experiments.

Figure 3

B cell–deficient mice are more susceptible to rechallenge with influenza. Groups of 7–12 age-matched μKO (open symbols) and C57Bl/6 (closed symbols) mice were intranasally infected with attenuated A/JAPAN/57. 28 d later the animals were rechallenged intranasally with the dilutions of mouse adapted A/JAPAN/57: ▵, 10⁻¹ dilution; ○, 10⁻² dilution; ⋄, 10⁻³ dilution; □, 10⁻⁴ dilution. Animals were followed for 21 d for mortality. Data is representative of two experiments.

Figure 4

CD8⁺, but not CD4⁺, T cell clones promote full recovery in B cell–deficient mice. B cell–deficient (μKO) mice were infected intranasally with a 10 LD₅₀ dose of mouse-adapted A/JAPAN/57 virus followed by an intravenous injection of 10⁷ cells. The cells transferred included 4D7 (⋄, a CD4⁺ clone), 11E4 (▴, a CD8⁺ clone), and B1.11 (•, a CD8⁺ clone). Mice receiving intranasal influenza, but no cell transfer, are denoted as □. C57Bl/6 mice receiving intranasal influenza without cell transfer are also shown (✖). Each group represents 5–7 animals. Adoptive transfer of these clones and media control into lethally challenged C57Bl/ 6 mice was done simultaneously as a control.