Proteasome Inhibition Reduces Proliferation, Collagen Expression, and Inflammatory Cytokine Production in Nasal Mucosa and Polyp Fibroblasts

Abstract

Proteasome inhibitors, used in cancer treatment for their proapoptotic effects, have anti-inflammatory and antifibrotic effects on animal models of various inflammatory and fibrotic diseases. Their effects in cells from patients affected by either inflammatory or fibrotic diseases have been poorly investigated. Nasal polyposis is a chronic inflammatory disease of the sinus mucosa characterized by tissue inflammation and remodeling. We tested the hypothesis that proteasome inhibition of nasal polyp fibroblasts might reduce their proliferation and inflammatory and fibrotic response. Accordingly, we investigated the effect of the proteasome inhibitor Z-Leu-Leu-Leu-B(OH) 2 (MG262) on cell viability and proliferation and on the production of collagen and inflammatory cytokines in nasal polyp and nasal mucosa fibroblasts obtained from surgery specimens. MG262 reduced the viability of nasal mucosa and polyp fibroblasts concentration-and time-dependently, with marked effects after 48 h of treatment. The proteasome inhibitor bortezomib provoked a similar effect. MG262-induced cell death involved loss of mitochondrial membrane potential, caspase-3 and poly(ADP-ribose) polymerase activation, induction of c-Jun phosphorylation, and mitogen-activated protein kinase phosphatase-1 expression. Low concentrations of MG262 provoked growth arrest, inhibited DNA replication and retinoblastoma phosphorylation, and increased expression of the cell cycle inhibitors p21 and p27. MG262 concentrationdependently inhibited basal and transforming growth factor-␤-induced collagen mRNA expression and interleukin (IL)-1␤-induced production of IL-6, IL-8, monocyte chemoattractant protein-1, regulated on activation normal T cell expressed and secreted, and granulocyte/macrophage colony-stimulating factor in both fibroblast types. MG262 inhibited IL-1␤/tumor necrosis factor-␣-induced activation of nuclear factor-B. We conclude that noncytotoxic treatment with MG262 reduces the proliferative, fibrotic, and inflammatory response of nasal fibroblasts, whereas high MG262 concentrations induce apoptosis.

Effect of Lipopolysaccharide on Glucocorticoid Receptor Function in Control Nasal Mucosa Fibroblasts and in Fibroblasts from Patients with Chronic Rhinosinusitis with Nasal Polyps and Asthma

“…Thirty to 50% confluent nasal fibroblasts were transfected with 20 nM of either MKP-1 or GILZ Silencer Select pre-designed siRNA or Silencer Select negative control siRNA using lipofectamine RNAiMAX, as previously reported [ 29 ]. Twenty-four hours after transfection, cells were pre-incubated in the absence or presence of LPS (10 μg/ml) for 24 hours.…”

“…Total RNA was isolated and reverse transcribed to cDNA [ 29 ]. GRα , GRβ , MKP-1 , GILZ , and RNA polymerase II (RPII) mRNA expression was analyzed by real-time PCR using TaqMan Fast Universal PCR Master Mix and either customized (for GRα ) or predesigned (for all other genes) TaqMan Gene Expression Assays (Life Technologies) [ 25 ].…”

Roles of the Skp2/p27 axis in the progression of chronic nephropathy

“…We have shown that p21 accumulation in UUO kidneys is moderately enhanced by Skp2 deficiency [20]. In addition, the accumulation of p21 and p27 as a result of proteasome inhibition is associated with inhibition of interstitial fibroblast proliferation [92]. Therefore, p21 and p27 are negative regulators of interstitial cell proliferation while upregulated Skp2 in the UUO kidney enhances their degradation to promote interstitial fibroblast proliferation and myofibroblast formation as critical stages in the EMT.…”

“…In these cell lines, treatment with a proteasome inhibitor suppressed fibrosis together with reduced MCP-1 production and TGF-β- and TNF-α-induced collagen mRNA expression. Moreover, the inflammatory response in fibroblasts is inhibited by suppression of IL-1β-/TNF-α-induced NF-κB activation and IL-1β-induced IL-6/8 production [92]. These results suggest that the accumulated p21 and p27 in fibroblasts can inhibit tissue inflammation and progressive fibrosis.…”

“…It has been reported that renal fibrosis is ameliorated by proteasome inhibitors in rat obstructive nephropathy [120]. Therefore, Neubert et al [121] suggested that proteasome inhibitors are effective for treatment of nephropathy, and Pujols et al [92] reported that a proteasome inhibitor could reduce proliferation, collagen production, and inflammatory responses in nasal fibroblasts. However, proteasome inhibitors reportedly show severe side effects because they accumulate many proteins by inhibition of proteasome-mediated degradation [122].…”