Diverse functions of clusterin promote and protect against the development of pulmonary fibrosis.

Pulmonary fibrosis is a progressive scarring disorder of the lung with dismal prognosis and no curative therapy. Clusterin, an extracellular chaperone and regulator of cell functions, is reduced in bronchoalveolar lavage fluid of patients with pulmonary fibrosis. However, its distribution and role in normal and fibrotic human lung are incompletely characterized. Immunohistochemical localization of clusterin revealed strong staining associated with fibroblasts in control lung and morphologically normal areas of fibrotic lung but weak or undetectable staining in fibrotic regions and particularly fibroblastic foci. Clusterin also co-localized with elastin in vessel walls and additionally with amorphous elastin deposits in fibrotic lung. Analysis of primary lung fibroblast isolates in vitro confirmed the down-regulation of clusterin expression in fibrotic compared with control lung fibroblasts and further demonstrated that TGF-ß1 is capable of down-regulating fibroblast clusterin expression. shRNA-mediated down-regulation of clusterin did not affect TGF-ß1-induced fibroblast-myofibroblast differentiation but inhibited fibroblast proliferative responses and sensitized to apoptosis. Down-regulation of clusterin in fibrotic lung fibroblasts at least partly due to increased TGF-ß1 may therefore represent an appropriate but insufficient response to limit fibroproliferation. Reduced expression of clusterin in the lung may also limit its extracellular chaperoning activity contributing to dysregulated deposition of extracellular matrix proteins.

Cladribine Exposure Results in a Sustained Modulation of the Cytokine Response in Human Peripheral Blood Mononuclear Cells.

BACKGROUND AND OBJECTIVES: Cladribine is a cytotoxic drug which ameliorates the clinical course of relapsing-remitting multiple sclerosis. In addition to cytotoxicity, the mode of action may include immunomodulatory mechanisms. This in vitro study was designed to investigate cladribine's effects on cell function after the removal of cladribine to distinguish cytotoxic versus immunomodulatory effects. METHODS: Cells were incubated in the absence or presence of cladribine (1 × 10(-8) M to 1 × 10(-5) M) for 72 h. Cladribine was removed from the cell culture and surviving peripheral blood mononuclear cells were cultured up to 58 days to determine the immunomodulatory effects of cladribine on cell function (e.g., proliferation and cytokine release). RESULTS: In the long-term, brief cladribine exposure did not impair the proliferation of surviving peripheral blood mononuclear cells. However, it induced an anti-inflammatory shift in the cytokine milieu with significantly enhanced release of IL-4 (Days 9 and 44, p<0.01; Day 58, p<0.05) and IL-5 (Day 9, p<0.01), resulting in an increased IL-4/INF-gamma ratio (Days 9 and 44, p<0.01; Day 58, p<0.05). Additionally, a trend towards an increased IL-10 production was observed. No changes were found in the production of IFN-gamma, TNF-alpha, IL-6, IL-8, IL-17A, IL-23 or NGF-beta. CONCLUSIONS: In vitro cladribine exposure induces a sustained anti-inflammatory shift in the cytokine profile of surviving peripheral blood mononuclear cells. This immunomodulatory action might contribute to cladribine's beneficial effects in the treatment of multiple sclerosis.