Fig. 3.

The cytokine and chemokine responses induced by influenza virus infection of mouse lung slices. (A, B) The analysis of the mRNA levels of selected cytokines and chemokines. Following infection by 200 μl of 10⁵ PFU/ml of PR8 (H1N1) (A) and HUBEI (H3N2) (B), lung slices were collected at indicated times post infection and were lysed to quantify the mRNA levels of cytokines and chemokines by real time RT-PCR. Each data point was from three lung slices and virus diluent was used as a negative control. The gene expression normalised to β-actin mRNA was shown as fold expression values over negative control. The data were analysed using the 2^−ΔΔCt method. The data are expressed as the means ± standard error of the means (SEMs). (C, D) The analysis of the protein levels of selected cytokines and chemokines. Following exposure to 200 μl of 10⁵ PFU/ml of PR8 (H1N1) virus, the supernatants were sampled at 24 h (C) and 48 h (D) post infection and were used to determine the protein levels of cytokines and chemokines by ELISA. To show the induced changes, cytokines and chemokines were placed in separate panels according to scales of the protein levels. Each data point was from three lung slices, and virus diluent was used as a negative control. Comparisons between the infected group and the negative control were performed by the unpaired t-test (*, 0.01 < p < 0.05; **, 0.001 < p < 0.01). The data are expressed as the means ± SEM. ND, not detected (for real-time PCR, no amplification was detected during the 40 cycles. For ELISA, the signal was below the detection limit of the commercial kit).