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. Author manuscript; available in PMC: 2014 Jan 5.
Published in final edited form as: J Immunol. 2012 Feb 27;188(7):10.4049/jimmunol.1100573. doi: 10.4049/jimmunol.1100573

Figure 5.

Figure 5

SHP-1 regulates M. pneumoniae-induced IL-8 production through PI3K/Akt in airway epithelial cells from nonasthmatic and asthmatic subjects. (A) Fold change in IL-8 production in nonasthmatic and asthmatic airway epithelial cells with or without pretreatment with NF-κB inhibitor II (30 μM), LY294002 (25 μM), or Akt inhibitor (50 nM) for 30 min, followed by mycoplasma infection for 48 h. IL-8 levels are expressed as fold change from cells in the same group without mycoplasma infection. *p < 0.01, #p < 0.05, compared with mycoplasma-infected cells without inhibitor pretreatment in the same group. The data are presented as mean ± SEM (n = 10 for nonasthma; n = 12 for asthma). (B) Immunoblot analysis of Akt phosphorylation (P-Akt) in total cell lysates of nonasthmatic and asthmatic cells with or without mycoplasma infection for 2 h (left panel). The levels of P-Akt 2 h after M. pneumoniae infection in asthmatic and nonasthmatic cells, with or without SHP-1 knockdown or overexpression, are expressed as fold change of densitometry from cells in the same group transduced with control rAAV only (right panel). *p < 0.01, compared with control rAAV-transduced cells treated with mycoplasma in the same group; ¥,#p < 0.01, compared with control rAAV-transduced nonasthmatic cells treated with mycoplasma only; †p < 0.01, compared with SHP-1 knockdown cells with mycoplasma infection only in the same group. Data are presented as mean ± SEM (n = 5). Mycoplasma-induced phosphorylation of Akt in asthmatic cells, which was significantly higher than that in nonasthmatic cells, was reversed by pretreatment with LY294002 and attenuated by SHP-1 overexpression. SHP-1 knockdown dramatically increased phosphorylation of Akt in nonasthmatic cells, but not in asthmatic cells, which was reversed by pretreatment with LY294002. (C) IL-8 fold change in SHP-1 knocked down nonasthmatic and asthmatic airway epithelial cells that were pretreated or not with NF-κB inhibitor II (30 μM), LY294002 (25 μM), or Akt inhibitor (50 nM) for 30 min and then infected with mycoplasma for 48 h. SHP-1 knockdown dramatically accentuated IL-8 production in nonasthmatic cells, but not in asthmatic cells, which were all significantly inhibited by these three inhibitors. *p < 0.01 compared with mycoplasma-infected “S−” cells in the same group, without pretreatment of inhibitors; #p < 0.01, compared with “Ctr” cells of nonasthma, without pretreatment of inhibitors. The data are presented as mean ± SEM (n = 10 for nonasthma; n = 12 for asthma). Ctr, control rAAV-transduced airway epithelial cells; S−, airway epithelial cells with SHP-1 knockdown; S+, airway epithelial cells with SHP-1 overexpression.