FIG 3.

MK2206 alters Akt signaling and does not allow development of cellular responses to pH1N1 infection. (A) NCI-H1666 cells were nontreated or MK2206 treated (10 μM) and mock or A/Helsinki/P14/2009 infected (MOI, 3), cells were collected after 0.5 h, and phosphorylation levels of kinases and their substrates were profiled using a human phosphokinase array. The relative intensities of spots were calculated in ImageJ and plotted. (B and C) NCI-H1666 cells were treated as for panel A, cells were collected, and total RNA was extracted at 8 h, labeled, and used for whole-genome gene expression analysis. A heat map of 70 significantly changed genes is shown. The heat map represents normalized expression data on the logarithmic scale (log fold change of >2 and <−2), where genes are ordered by means of hierarchical clustering. Total RNA was isolated at 0, 2, 4, 8, and 16 h and subjected to RT-qPCRs detecting IFN-β and IL-29 mRNA levels. The data points are mean values, the numbers of observations used to derive the values are 2 and 3, respectively, and error bars represent the SD. (D) NCI-H1666 cells were treated as for panel A, cell culture supernatants were collected at 24 h postinfection, and cytokine levels were determined using human cytokine array panel A. A heat map is also shown.