Abstract
Transforming Growth Factor β1 Induces αvβ3 Integrin Expression in Human Lung Fibroblasts via a β3 Integrin-, c-Src-, and p38 MAPK-dependent Pathway
Highlights
(ECM)2 in which they proliferate and differentiate into myofibroblasts
transforming growth factor 1 (TGF)1 Increases 3 Protein Expression and Enhances Cell Surface Expression of ␣v3 Integrin on Human Lung Fibroblasts— Recently we showed that TGF1 increased 3 steady-state mRNA expression [4]
In parallel we examined the effect of TGF1 on another ␣v partner, the 5 integrin
Summary
Plasmids and Pharmacological Inhibitors—The pcDNA3Smad plasmid was provided by Dr Steven Mutsaers (University of Western Australia), and the pcDNA3-p38-KM (kinase mutant) plasmid was provided by Dr Kun Liang Guan (University of Michigan). Adherent cells were collected after trypsin/EDTA immersion, and surface expression of integrins was allowed to recover for 30 min in PBS with 10% FBS at room temperature This step served to block nonspecific antibody binding. Preliminary experiments determined that a concentration of 50 –100 particles of Adv-GFP-FRNK and Adv-GFP per cell strongly induced the expression of these proteins (supplemental Fig. 2SB) and infected virtually every fibroblast (ϳ90% of GFP positive cells by flow cytometry) after 48 h of exposure (data not shown). After infection with AdvGFP-FRNK and Adv-GFP, cells were cultured in the presence of serum for 24 h, serum-starved for additional 24 h, and stimulated with TGF1 for the indicated time periods. P38 MAPK phosphorylation was determined using mouse mAb against human phosphorylated p38 MAPK (Thr180/Tyr182) and rabbit polyclonal antibodies against human p38 MAPK (both from Cell Signaling Technology, Danvers, MA).
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