Session Type: Abstract Submissions (ACR)
Scleroderma associated interstitial lung disease (SSc-ILD) is an irreversible and progressive complication and a leading cause of death among SSc patients. Constitutive overexpression of connective tissue growth factor (CTGF) has been observed in both in vivo and in vitro studies of SSc patients. Proteomic analysis of CTGF-activated lung fibroblasts demonstrated that CTGF induces IQ motif containing GTPase activating protein (IQGAP1). IQGAP1 is a multifunctional scaffold protein that integrates diverse signal transduction pathways and regulates fibroblast migration. Our recent findings demonstrate the profibrotic role of IQGAP1 in the bleomycin -induced murine model of SSc-ILD. Here we report our latest data focusing on the molecular mechanism and pathophysiologic action of IQGAP1 in this mouse model of ILD.
Lung injury was induced in female C57BL/6 mice by a single intratracheal instillation of bleomycin (0.05U/mouse). IQGAP1-siRNA and CTGF-siRNA were delivered by intranasal instillation every other day. Mice were sacrificed 3 weeks after bleomycin instillation and lungs were harvested. Lungs were perfused with neutral buffered formaldehyde, embedded in paraffin, stained with hematoxylin and eosin (H&E), and scored for fibrosis. Lung tissue was harvested, lyophilized and run on western blot. IQGAP1 knockout mice were challenged with bleomycin and histology was performed. Assessment of collagen was accomplished by Masson’s trichrome staining and by Sircol Collagen Assay. The role of IQGAP1 in F-actin filament formation was examined by immunofluorescence staining and by actin polymerization assay. The rate of actin polymerization was measured in terms of fluorescence intensity by Fluorometric Imaging Plate Reader.
A profound antifibrotic effect was observed in the bleomycin lung fibrosis model when IQGAP1-siRNA treatment was combined with CTGF-siRNA treatment. Partial reduction of fibrosis was detected with treatment by either of these two siRNA’s alone. Western blot results showed that IQGAP1-siRNA decreased the expression of IQGAP1 by 70% and had no effect on CTGF expression. However, CTGF-siRNA reduced the expression of CTGF by 80% and IQGAP1 by 40%. A similar trend of reduction in fibrosis was observed in IQGAP1 knockout mice. Decreased collagen expression was detected by Masson’s trichrome stain and by Sircol Collagen Assay. Immunofluorescence staining of IQGAP1 and F-actin on human SSc lung fibroblasts demonstrated that IQGAP1 co-localizes with globular actin but not with filamentous stress fiber actin, indicating a crucial role of IQGAP1 in actin rearrangement. The actin polymerization assay demonstrated that the rate of actin polymerization is IQGAP1 dependent.
IQGAP1 forms a signal transduction complex with CTGF in lung fibroblasts, regulates the expression of α-SMA, and promotes pulmonary fibrosis. Inhibition of IQGAP1 has a marked antifibrotic effect in a bleomycin model of pulmonary fibrosis and should be considered as a potential new therapeutic target for the treatment of SSc-ILD.
S. Prince Nelson,
T. A. Morinelli,
R. M. Silver,
G. S. Bogatkevich,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/iqgap1-enhances-contractility-of-scleroderma-lung-fibroblasts-and-promotes-bleomycin-induced-pulmonary-fibrosis/