Figure 7.

Effect of clusterin deficiency on apoptosis. Lung fibroblasts were transduced with clusterin shRNA (shCLU, open circles) and mock shRNA vectors (grey circles) or remained untransfected (black circles). Lung fibroblasts were seeded and treated with FasL (3 nM – 6 nM) and/or exogenous clusterin (CLU, 125 nM) for 19 h or remained untreated. Apoptotic cells (Annexin V+ and Annexin V+/ DAPI+ cells) were assessed via FACS analysis post staining of apoptotic cells with annexin V – Alexa647 and DAPI (mean ± SEM, n = 5). (A) shRNA-induced clusterin deficiency sensitized fibroblasts to basal and FasL-induced apoptosis, and this could be overcome by addition of exogenous clusterin (B). (C) Basal apoptosis in representative control (9.37 ± 0.63%) and fibrotic (11.6 ± 0.69%) lung fibroblast isolates was not significantly different. Fibrotic lung fibroblasts were more resistant to FasL-induced apoptosis compared with controls (C) and exogenous clusterin tends to reduce basal and FasL-induced apoptotic levels further (D). Data representative of at least two individual experiments with fibroblasts derived from 1 donor per group. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.