Figure 2

SubAB-increased CHOP and C/EBPG regulate SubAB-induced LCN2 expression. (A) HeLa cells were co-cultured for 24 h with the indicated STEC O113:H21 strains as shown in Fig. 1D. The chop mRNA levels were measured by RT-qPCR as described in “Methods”. GAPDH was used as an internal control. Data are mean ± SD (n = 3). *P < 0.05, versus mt SubAB-treated control cells. (B) Control (NC) or CHOP siRNA-transfected cells were incubated for 24 h with mt SubAB or SubAB (400 ng ml⁻¹). The lcn2 mRNA levels were measured by RT-qPCR as described in “Methods”. GAPDH was used as an internal control. Data are mean ± SD (n = 3). *P < 0.05, versus mt SubAB-treated control cells. (C) Control (NC) or CHOP siRNA-transfected cells were incubated for 24 h with mt or wt SubAB (400 ng ml⁻¹). Cell lysates were subjected to immunoblotting with the indicated antibodies. GAPDH served as a loading control. (D) Quantification of cPARP or LCN2 in the transfected cells with mt or wt SubAB was performed by densitometry. Data are presented as mean ± SD of values from three independent experiments and significance is *P < 0.05. (E) Cells were incubated for 24 h with mt or wt SubAB (400 ng ml⁻¹). The mRNA levels of cebpA, cebpB, or cebpG were measured by RT-qPCR as described in “Methods”. GAPDH was used as an internal control. Data are mean ± SD (n = 3). * P < 0.05, versus mt SubAB-treated control cells. (F) The indicated siRNA-transfected cells were incubated for 24 h with mt or wt SubAB (400 ng ml⁻¹). The lcn2 mRNA levels were measured by RT-qPCR as described in “Methods”. GAPDH was used as an internal control. Data are mean ± SD (n = 3). *P < 0.05, versus mt SubAB-treated control cells. (G) Control (NC), C/EBPB, or C/EBPG siRNA-transfected cells were incubated for 24 h with mt or wt SubAB (400 ng ml⁻¹). Cell lysates were subjected to immunoblotting with anti-LCN2 antibodies. GAPDH served as a loading control.