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. 2013 Jun 6;9(6):e1003407. doi: 10.1371/journal.ppat.1003407

Figure 6. Control of Mtb infection by co-cultured epithelial cells is dependent on epithelial IL1R1 and DEFB4 expression.

Figure 6

(A) CFU of macrophages (Mφ) infected with H37Rv at an MOI of 5 for 4 hours and treated with vehicle control or 100 nM 1,25D (+D), with and without the additional presence of SAECs in transwell co-culture (CC). Data are from three experimental replicates (mean and SD) and representative of three independent experiments using different donors of primary cells. Statistical significance was determined by one-way ANOVA. (*P<0.05). (B) Validation of siRNA-mediated knockdown of IL1R1 expression in SAECs. RNA was extracted from SAECs cells 3 days after the initiation of co-culture and 4.5 days after transfection of control siRNA (siCTL) or siIL1R1. Data are from three experimental replicates (mean and SD), **P<0.01 as determined by student's T-test relative to respective siCTL controls. (C) CFU quantification of Mtb in THP-1 cells infected at an MOI of 5 for 4 hours after 72 hours of co-culture with SAEC cells transfected with control siRNA (siCTL) or siRNA specific to IL1R1. 1,25D (D) was added as indicated. Data are from three experimental replicates (mean and SD) and representative of two independent experiments. Statistical significance was determined by one-way ANOVA. (**P<0.01). (D) Validation of siRNA-mediated knockdown of DEFB4 expression in SAECs. RNA was extracted from SAECs cells 3 days after the initiation of co-culture and 4.5 days after transfection of control siRNA (siCTL) or siDEFB4. Data are from three experimental replicates (mean and SD) and representative to two independent replicates. **P<0.01 as determined by student's T-test relative to siCTL control. (E) CFU quantification of Mtb in THP-1 cells infected at an MOI of 5 for 4 hours after 72 hours of co-culture with SAEC cells transfected with control siRNA (siCTL) or siRNA specific to IL1R1 or DEFB4. Data are from three experimental replicates (mean and SD) and representative of two independent experiments using separate donors of SAECs. Statistical significance was determined by one-way ANOVA. (*P<0.05).