(A) Constitutive levels of HAS1 (2A-i), HAS2 (2A-ii), HAS3 (2A-iii), HA (2A-iv) and CD44 and RHAMM (2A-v) in immortalized BEAS-2B bronchial cells and NSCLC cell lines. (B) Modulation of levels of HAS2 (2B-i), HAS3 (2B-ii), HA (2B-iii), CD44 and RHAMM (2B-iv; 2B-v) in NSCLC cells treated with triptolide (25 nM) or DMSO. Cells were treated with 25 nM of triptolide for 72 h with the exception of the results shown in Figure 2B-iv in which cell were exposed to 0, 25 or 50 nM of triptolide. (C) Exogenous HA attenuated triptolide-induced cytotoxicity and modulation of cell proliferation and survival-related proteins. C-i, A549 cells grown in RPMI media supplemented with 2.5% FBS were treated with DMSO, triptolide (25 nM), HA (2.5 mg/mL), or triptolide + HA for 72 h and cell viability determined by MTT assay. C-ii, images of A549 cells exposed to DMSO, triptolide (25 nM), HA, or triptolide + HA for 72 h. C-iii, Western immunoblotting assays showing attenuation by HA of triptolide-induced modulation in the expression of cell proliferation and survival-related proteins. *P < 0.05. Assays were performed in triplicate and repeated three times on different days. C, Control; T, triptolide.