Figure 6: Induction of fibrotic reaction. A & B Induction of TGFβ1 expression. BEAS-2B cells were treated with SWCNTs at 20 μg/ml or IL-1β at 5 ng/ml for 24 h. Real-time PCR for TGFβ mRNA (A) or immunoblotting for TGFβ protein (B) was performed. (C) Paracrine induction of αSMA. Macrophages were treated with SWCNTs (20 μg/ml, 16 h) or LPS (1 μg/ml, 5 h). The culture medium free of cells and SWCNTs were collected as the SWCNT-conditioned medium. WI38-VA13 cells were then cultured in the SWCNT-conditioned medium in 8-well chamber slides for 24 h. Expression of αSMA was detected by immunofluorescent staining of the cells with an anti-αSMA antibody followed by an alexia-488 conjugated second antibody. The nucleus was stained with DAPI. Cell images were taken under a Zeiss LSM510 confocal microscope. (D) Quantitative data of C from 5 separate fields.