IGF-I increases IGFBP-5 and collagen α1(I) mRNAs by the MAPK pathway in rat intestinal smooth muscle cells

2004 ◽  
Vol 286 (5) ◽  
pp. G777-G783 ◽  
Author(s):  
Xiping Xin ◽  
Yong Tai Hou ◽  
Lina Li ◽  
Phyllissa Schmiedlin-Ren ◽  
Gregory M. Christman ◽  
...  
Keyword(s):  
Gene Regulation ◽  
Smooth Muscle ◽  
Crohn’S Disease ◽  
Smooth Muscle Cells ◽  
Mapk Pathway ◽  
Muscle Cells ◽  
Intestinal Smooth Muscle ◽  
Concurrent Treatment ◽  
Igf I ◽  
Docking Proteins

IGF-I is a potent fibrogenic growth factor that stimulates proliferation of intestinal smooth muscle cells and increases synthesis of collagen and IGF-I-binding proteins by the cells. These processes contribute to intestinal fibrosis that develops in patients with Crohn's disease and in Lewis-strain rats with experimental Crohn's disease. The aim of this study was to determine which early docking proteins are associated with IGF-I receptor signal transduction and which transduction pathway is involved in IGF-I-mediated gene regulation in intestinal smooth muscle cells. Primary cultures of smooth muscle cells isolated from the muscularis externa of the distal colon of Lewis rats were treated with IGF-I (100 ng/ml). Immunoprecipitation studies demonstrated that IGF-I stimulation resulted in tyrosine phosphorylation of IRS-1, IRS-2, and Shc. Coimmunoprecipitation demonstrated a close association between the IGF-I receptor and these three early docking proteins. Concurrent treatment with the MAPK inhibitor PD98059 (10 μM) resulted in an inhibition of the IGF-I-mediated increase in IGFBP-5 and collagen α1(I) mRNAs, while concurrent treatment with the phosphatidylinositol 3-kinase (PI3-K) inhibitor wortmannin (100 nM) had no effect. In additional experiments, cells were transiently transfected with adenoviral vectors dominantly expressing inactive mutant Akt or constitutively expressing wild-type Akt. In both cases, the IGF-I-mediated increase in collagen I protein did not differ from that observed in control cultures that had been transfected with an adenoviral vector carrying the LacZ reporter gene. These results suggest that the MAPK pathway is key to IGF-I-mediated gene regulation in intestinal smooth muscle cells, whereas data do not suggest a role for the Akt-dependent pathway in our system.

IGFBP-3 activates TGF-β receptors and directly inhibits growth in human intestinal smooth muscle cells

2004 ◽  
Vol 287 (4) ◽  
pp. G795-G802 ◽  
Author(s):  
John F. Kuemmerle ◽  
Karnam S. Murthy ◽  
Jennifer G. Bowers
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Nuclear Translocation ◽  
Thymidine Incorporation ◽  
Muscle Cells ◽  
Serine Phosphorylation ◽  
Intestinal Smooth Muscle ◽  
Igf I ◽  
Tgf Β1 ◽  
Igfbp 3

We have shown that human intestinal smooth muscle cells produce IGF-I and IGF binding protein-3 (IGFBP-3). Endogenous IGF-I acts in autocrine fashion to stimulate growth of these cells. IGFBP-3 inhibits the binding of IGF-I to its receptor and thereby inhibits IGF-I-stimulated growth. In several carcinoma cell lines and some normal cells, IGFBP-3 regulates growth independently of IGF-I. Two mechanisms for this effect have been identified: IGFBP-3 can directly activate transforming growth factor-β (TGF-β) receptors or it can undergo direct nuclear translocation. The aim of the present study was to determine whether IGFBP-3 acts independently of IGF-I and to characterize the mechanisms mediating this effect in human intestinal smooth muscle cells. The direct effects of IGFBP-3 were determined in the presence of an IGF-I receptor antagonist to eliminate its IGF-I-dependent effects. Affinity labeling of TGF-β receptors (TGF-βRI, TGF-βRII, and TGF-βRV) with 125I-labeled TGF-β1 showed that IGFBP-3 displaced binding to TGF-βRII and TGF-βRV in a concentration-dependent fashion. IGFBP-3 stimulated TGF-βRII-dependent serine phosphorylation (activation) of both TGF-βRI and of its primary substrate, Smad2(Ser465/467). IGFBP-3 also caused IGF-I-independent inhibition of basal [3H]thymidine incorporation. The effects of IGFBP-3 on Smad2 phosphorylation and on smooth muscle cell proliferation were independent of TGF-β1 and were abolished by transfection of Smad2 siRNA. Immunoneutralization of IGFBP-3 increased basal [3H]thymidine incorporation, implying that endogenous IGFBP-3 inhibits proliferation. We conclude that endogenous IGFBP-3 directly inhibits proliferation of human intestinal smooth muscle cells by activation of TGF-βRI and Smad2, an effect that is independent of its effect on IGF-I-stimulated growth.

Endogenous IGF-I regulates IGF binding protein production in human intestinal smooth muscle cells

2000 ◽  
Vol 278 (5) ◽  
pp. G710-G717 ◽  
Author(s):  
John F. Kuemmerle
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Map Kinase ◽  
Binding Protein ◽  
Muscle Cells ◽  
Intestinal Smooth Muscle ◽  
Igf I ◽  
Igf Binding ◽  
Igf Binding Protein ◽  
Igfbp 3

Human intestinal smooth muscle in culture produces insulin-like growth factor (IGF)-I and IGF binding protein (IGFBP)-3, IGFBP-4, and IGFBP-5, which modulate the effects of IGF-I. This study examined the regulation of IGFBP production by endogenous IGF-I. R3-IGF-I, an agonist unaffected by IGFBPs, elicited concentration-dependent increase in growth, measured by [3H]thymidine incorporation, and production of IGFBP-3, IGFBP-4, and IGFBP-5, measured by Western blot. Antagonists of the IGF-I receptor, IGF-I Analog or monoclonal antibody 1H7, elicited concentration-dependent inhibition of growth and decrease in IGFBP-3, IGFBP-4, and IGFBP-5 production, implying that endogenous IGF-I stimulated growth and IGFBP production. R3-IGF-I-induced increase in IGFBP-3, IGFBP-4, and IGFBP-5 production was partially inhibited by a mitogen-activated protein (MAP) kinase or a phosphatidylinositol-3-kinase (PI 3-kinase) inhibitor and abolished by the combination. We conclude that endogenous IGF-I stimulates growth and IGFBP-3, IGFBP-4, and IGFBP-5 production in human intestinal smooth muscle cells. Regulation of IGFBP production by IGF-I is mediated by activation of distinct MAP kinase and PI 3-kinase pathways, the same pathways through which IGF-I stimulates growth.

scholarly journals Dual Role of Src Homology Domain 2-Containing Inositol Phosphatase 2 in the Regulation of Platelet-Derived Growth Factor and Insulin-Like Growth Factor I Signaling in Rat Vascular Smooth Muscle Cells

Endocrinology ◽  
2003 ◽  
Vol 144 (9) ◽  
pp. 4204-4214 ◽  
Author(s):  
Toshiyasu Sasaoka ◽  
Kosei Kikuchi ◽  
Tsutomu Wada ◽  
Akira Sato ◽  
Hiroyuki Hori ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Smooth Muscle Cells ◽  
Phosphatase Activity ◽  
Mapk Pathway ◽  
Muscle Cells ◽  
Sh2 Domain ◽  
Pi3 Kinase ◽  
Inositol Phosphatase ◽  
Igf I

Abstract Src homology domain 2 (SH2)-containing inositol phosphatase 2 (SHIP2) possesses 5-phosphatase activity and an SH2 domain. The role of SHIP2 in platelet-derived growth factor (PDGF) and IGF-I signaling was studied by expressing wild-type (WT-) and a catalytically defective (ΔIP-) SHIP2 into rat aortic smooth muscle cells by adenovirus-mediated gene transfer. PDGF- and IGF-I-induced tyrosine phosphorylation of their respective receptors and phosphatidylinositol 3-kinase (PI3-kinase) activity were not affected by the expression of either WT- or ΔIP-SHIP2. SHIP2 possessed 5′-phosphatase activity to hydrolyze the PI3-kinase product phosphatidylinositol 3,4,5-trisphosphate in vivo. Akt and glycogen synthase kinase 3β are known to be downstream molecules of PI3-kinase, leading to the antiapoptotic effect. Overexpression of WT-SHIP2 inhibited PDGF- and IGF-I-induced phosphorylation of these molecules and the protective effect of poly(ADP-ribose) polymerase degradation, whereas these phosphorylations and the protective effect were enhanced by the expression of ΔIP-SHIP2, which functions in a dominant negative fashion. Regarding the Ras-MAPK pathway, PDGF- and IGF-I-induced tyrosine phosphorylation of Shc was not affected by the expression of either WT- or ΔIP-SHIP2, whereas both expressed SHIP2 associated with Shc. Importantly, PDGF and IGF-I stimulation of Shc/Grb2 binding, MAPK activation, and 5-bromo-2′-deoxyuridine incorporation were all decreased in both WT- and ΔIP-SHIP2 expression. These results indicate that SHIP2 plays a negative regulatory role in PDGF and IGF-I signaling in vascular smooth muscle cells. As the bifunctional role, our results suggest that SHIP2 regulates PDGF- and IGF-I-mediated signaling downstream of PI3-kinase, leading to the antiapoptotic effect via 5-phosphatase activity, and that SHIP2 regulates the growth factor-induced Ras-MAPK pathway mainly via the SH2 domain.

Regulation of IGFBP-4 levels in human intestinal muscle by an IGF-I-activated, confluence-dependent protease

2000 ◽  
Vol 279 (5) ◽  
pp. G975-G982 ◽  
Author(s):  
John F. Kuemmerle ◽  
Baiqin Teng
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Protease Activity ◽  
Muscle Cells ◽  
Intestinal Smooth Muscle ◽  
Mrna Levels ◽  
Proliferating Cells ◽  
Intact Cells ◽  
Igf I

Human intestinal smooth muscle cells in culture produce insulin-like growth factor-I (IGF-I), IGF binding protein-3 (IGFBP-3), IGFBP-4, and IGFBP-5, which can modulate the effects of IGF-I on growth. This study examined the role of IGFBP-4 on IGF-I-induced growth and the mechanisms regulating IGFBP-4 levels. IGFBP-4 inhibited IGF-I-induced [3H]thymidine incorporation. IGFBP-4 mRNA levels were not altered by IGF-I. IGF-I caused a concentration-dependent activation of an endogenous IGFBP-4 protease, resulting in time-dependent degradation of intact IGFBP-4 into inactive fragments. Protease activity was measured in a cell-free assay using smooth muscle cell conditioned medium containing the IGFBP-4 protease. The protease was inhibited by EDTA and benzamidine. Protease activity was highest in proliferating cells and lowest in postconfluent cells. The role of endogenous IGF-I in regulating IGFBP-4 degradation was confirmed by the ability of an IGF-I antagonist to inhibit IGF-I-activated IGFBP-4 proteolysis in intact cells. We conclude that in human intestinal smooth muscle cells levels of secreted IGFBP-4 are determined by the confluence-dependent production of a cation-dependent serine protease that is activated by endogenous IGF-I.

Redox Imbalance in Crohn's Disease Intestinal Smooth Muscle Cells Causes NF-κB-Mediated Spontaneous Interleukin-8 Secretion

2001 ◽  
Vol 21 (6) ◽  
pp. 349-359 ◽  
Author(s):  
Ramesh Natarajan ◽  
Shobha Ghosh ◽  
Bernard J. Fisher ◽  
Robert F. Diegelmann ◽  
Amy Willey ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Crohn’S Disease ◽  
Crohn's Disease ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Interleukin 8 ◽  
Intestinal Smooth Muscle ◽  
Redox Imbalance

IGF-I induces collagen and IGFBP-5 mRNA in rat intestinal smooth muscle

1997 ◽  
Vol 273 (4) ◽  
pp. G875-G882 ◽  
Author(s):  
E. M. Zimmermann ◽  
L. Li ◽  
Y. T. Hou ◽  
M. Cannon ◽  
G. M. Christman ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Type I Collagen ◽  
Muscle Cells ◽  
Northern Analysis ◽  
Rat Colon ◽  
Intestinal Smooth Muscle ◽  
Type I ◽  
Igf I ◽  
Cultured Smooth Muscle Cells

Insulin-like growth factor (IGF) binding protein 5 (IGFBP-5) mRNA was studied in intestines of rats with peptidoglycan-polysaccharide enterocolitis by Northern analysis and in situ hybridization. IGFBP-5 mRNA was increased 2.4 ± 0.5-fold in inflamed rat colon compared with controls and was highly expressed in smooth muscle. Cultured rat intestinal smooth muscle cells were used to study the regulation of IGFBP-5 and type I collagen synthesis. IGF-I (100 ng/ml) increased IGFBP-5 mRNA (1.9 ± 0.1-fold) and collagen type α1(I) mRNA (1.6 ± 0.2-fold) in cultured smooth muscle cells. IGF-I induced a dose- and time-dependent increase in IGFBP-5 in conditioned medium by Western ligand blot and by immunoblot. IGF-I did not affect the IGFBP-5 mRNA decay rate after transcriptional blockade. Cycloheximide abolished IGFBP-5 mRNA. In conclusion, IGFBP-5 mRNA is expressed by intestinal smooth muscle and is increased during chronic inflammation. IGF-I increases IGFBP-5 and collagen mRNAs in intestinal smooth muscle cells.

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