Changes of pulmonary co-signaling molecules in influenza A (H1N1) virus infected mice

Background Influenza A virus is highly infectious with a high mortality rate in severe infection cases. Co-signaling molecules are important molecules involved in immune response. Exploring the changing characteristics and correlation between pulmonary viral load and co-signaling molecules after influenza infection can provide reference for the treatment of influenza A virus infection.

Objective To explore the changing characteristics of pulmonary viral load and its association with co-signaling molecules mRNA expression in mice infected with influenza A virus A/Puerto Rico / 8/34 (H1N1) (PR8 strain).

Methods Totally 72 BALB/c mice aged 7 weeks were randomly divided into high dose virus group (10⁸TCID₅₀/mL, 50 μL/ mouse, n=32), low dose virus group (10⁴TCID₅₀/mL, 50uL/ mouse, n=32) and control group (PBS, 50uL/ mouse, n=8), 8, 8 and 2 mice were sacrificed in the three groups at each time point of 24 h, 48 h, 72 h and 96 h after infection, respectively. All mice were dissected for the left lung which used for HE staining, qRT-PCR was applied to detect the pulmonary virus load and the relative mRNA expression levels of co-signaling molecules CD28, CD226, ICOS, CTLA-4, TIGIT and PD-1. The correlation between pulmonary viral load and mRNA expression of co-signaling molecules were analyzed in all mice infected with high and low dose viruses.

Results Within 96 h after infection, the lung injury of high dose virus group was more severe than that of low dose virus group, and the mRNA expression of co-stimulatory molecules CD28, CD226 and ICOS was decreased (all P＜0.05). In the high dose virus group, the relative expression of PD-1 mRNA increased at 24 h after infection (P＜0.01), the relative expression of CTLA-4 mRNA increased (P＜0.01) and the viral replication reached to the peak at 48 h after infection. In the low dose virus group, the relative expression of PD-1 mRNA increased (P＜0.05) and the viral replication reached to the peak at 72 h after infection, the relative expression of CTLA-4 mRNA increased (P＜0.01) at 96 h after infection. The pulmonary viral load of infected mice was positively correlated with the relative expression level of PD-1 mRNA (r=0.54, P＜0.001), and negatively correlated with the relative expression level of CD226 mRNA (r=-0.34, P=0.006).

Conclusion After infected with influenza A virus, the early appearance of the increased mRNA expression of CTLA-4 and PD-1 may indicate severe infection. PD-1 and CD226 in mice lung tissue may be the co-signaling molecular targets correlated with pulmonary viral load.