Effect of the novel influenza A (H1N1) virus in the human immune system

Abstract

Background: The pandemic by the novel H1N1 virus has created the need to study any probable effects of that infection in the immune system of the host.

Methodology/principal findings: Blood was sampled within the first two days of the presentation of signs of infection from 10 healthy volunteers; from 18 cases of flu-like syndrome; and from 31 cases of infection by H1N1 confirmed by reverse RT-PCR. Absolute counts of subtypes of monocytes and of lymphocytes were determined after staining with monoclonal antibodies and analysis by flow cytometry. Peripheral blood mononuclear cells (PBMCs) were isolated from patients and stimulated with various bacterial stimuli. Concentrations of tumour necrosis factor-alpha, interleukin (IL)-1beta, IL-6, IL-18, interferon (FN)-alpha and of IFN-gamma were estimated in supernatants by an enzyme immunoassay. Infection by H1N1 was accompanied by an increase of monocytes. PBMCs of patients evoked strong cytokine production after stimulation with most of bacterial stimuli. Defective cytokine responses were shown in response to stimulation with phytohemagglutin and with heat-killed Streptococcus pneumoniae. Adaptive immune responses of H1N1-infected patients were characterized by decreases of CD4-lymphocytes and of B-lymphocytes and by increase of T-regulatory lymphocytes (Tregs).

Conclusions/significance: Infection by the H1N1 virus is accompanied by a characteristic impairment of the innate immune responses characterized by defective cytokine responses to S.pneumoniae. Alterations of the adaptive immune responses are predominated by increase of Tregs. These findings signify a predisposition for pneumococcal infections after infection by H1N1 influenza.

Figure 1. Absolute counts of monocytes, of natural killer T-cell (NKT) and of natural killer (NK) cells.

Results refer to 10 healthy volunteers, 18 patients with flu-like syndrome and 31 patients infected by the H1N1 virus. Expression of HLA-DR on monocytes is also shown. a: denotes statistically significant differences compared with healthy volunteers; b: denotes statistically significant differences compared with patients with a flu-like syndrome.

Figure 2. Production of tumour necrosis factor-alpha (TNFα).

Peripheral blood mononuclear cells of 10 healthy volunteers, of 18 patients with flu-like syndrome and of 31 patients infected by the H1N1 virus were stimulated with endotoxins (LPS), phytohemagglutin (PHA), and heat-killed isolates of Candida albicans, of Pseudomonas aeruginosa, of methicillin-resistant Staphylococcus aureus producing Panton-Valentine leukocidin (MRSA-PVL) and of penicillin-susceptible Streptococcus pneumoniae. Superscript “a” denotes statistically significant differences compared with healthy volunteers; superscript “b” denotes statistically significant differences compared with patients with flu-like syndrome.

Figure 3. Production of interleukin-1beta (IL-1β).

Peripheral blood mononuclear cells of 10 healthy volunteers, of 18 patients with flu-like syndrome and of 31 patients infected by the H1N1 virus were stimulated with endotoxins (LPS), phytohemagglutin (PHA), and heat-killed isolates of Candida albicans, of Pseudomonas aeruginosa, of methicillin-resistant Staphylococcus aureus producing Panton-Valentine leukocidin (MRSA-PVL) and of penicillin-susceptible Streptococcus pneumoniae.

Figure 4. Cytokine production by peripheral blood mononuclear cells of healthy volunteers, of patients with flu-like syndrome and of patients infected by the H1N1.

a) interleukin-6 (IL-6) after stimulation with one heat-killed isolate of penicillin-susceptible Streptococcus pneumoniae; b) IL-18 after stimulation with one heat-killed isolate of penicillin-susceptible Streptococcus pneumoniae; c) interferon-alpha (IFNα) after stimulation with one heat-killed isolate of penicillin-susceptible Streptococcus pneumoniae; d) interferon-gamma (IFNγ) after stimulation with phytohemagglutin (PHA); and e) IFNγ after stimulation with one heat-killed isolate of penicillin-susceptible Streptococcus pneumoniae. Superscript “a” denotes significant differences compared with healthy volunteers.

Figure 5. Absolute counts of CD4-lymphocytes, of CD8-lymphocytes, of T-regulatory cells and of B-lymphocytes.

Results refer to 10 healthy volunteers, 18 patients with flu-like syndrome and 31 patients infected by the H1N1 virus. a: denotes statistically significant differences compared with healthy volunteers; b: denotes statistically significant differences compared with patients with flu-like syndrome.

Figure 6. Absolute counts of T-regulatory cells of patients infected with the H1N1 virus.

Results are given separately for 25 patients without pneumonia and for six patients with pneumonia (p<0.0001).

Figure 7. Serum levels of tumour necrosis factor-alpha (TNFα), of interleukin-1beta (IL-1β) and of IL-6.

Results refer to 10 healthy volunteers, 18 patients with flu-like syndrome and 31 patients infected by the H1N1 virus enrolled in the study. Superscript “a” denotes significant differences compared with healthy volunteers.

Figure 8. Absolute counts of monocytes and B-lymphocytes.

Results refer to 31 patients infected by the H1N1 virus at baseline and after 48 hours. P values denote statistically significant differences between the two time points.