Effect of granulocyte macrophage-colony stimulating factor on extracellular matrix deposition by dermal fibroblasts from patients with scleroderma.
OBJECTIVE
. To investigate the in vitro effect of granulocyte macrophage-colony stimulating factor (GM-CSF) on the deposition of extracellular matrix (ECM) in fibroblasts obtained from the skin of patients with systemic sclerosis (Ssc), compared to healthy controls.


METHODS
Dermal fibroblasts obtained from 14 patients with SSc (7 with the diffuse form and 7 with CREST syndrome) and from 7 controls were studied. Both SSc and normal skin fibroblast cultures were stimulated for 4 and 8 days with 100 ng/ml GM-CFS. GM-CSF receptor (GM-CSFR) expression was determined by Western blot of cell lysates. Immunofluorescence was used to determine GM-CSFR expression and to investigate the deposition of ECM (type I collagen, fibronectin, and tenascin). Quantitative analysis of ECM was performed by ELISA. Expression of type I collagen and metalloproteinase 1 (MMP-1) mRNA was determined by real-time quantitative PCR.


RESULTS
Deposition of ECM by normal fibroblasts appeared not to be influenced by stimulation with GM-CSF; in contrast, after stimulation with GM-CSF SSc fibroblasts showed increased deposition of fibronectin and tenascin, while type I collagen production was decreased; these results were found with both immunofluorescence and ELISA. Quantitative PCR revealed that GM-CSF inhibited the expression of mRNA type I collagen in SSc fibroblasts but not in normal fibroblasts, whereas levels of the main collagenolytic enzyme, MMP-1, were not affected.


CONCLUSIONS
These results suggest that in SSc fibroblasts GM-CSF exerts a blocking effect on the deposition of type I collagen, through an inhibitory action on mRNA, while the production of other components of ECM such as fibronectin and tenascin is increased by stimulation with this cytokine.
Postiglione
Loredana
L
Dipartimento di Biologia e Patologia Cellulare e Molecolare, Università degli Studi Federico II, Naples, Italy. lorposti@unima.it
Ladogana
Paolo
P
Montagnani
Stefania
S
di Spigna
Gaetano
G
Castaldo
Clotilde
C
Turano
Mimmo
M
Bruno
Eugenia Maria
EM
Di Meglio
Franca
F
Riccio
Antonio
A
Rossi
Guido
G
D000328:Adult; D000368:Aged; D017675:CREST Syndrome / Q000378:metabolism* / Q000473:pathology; D002478:Cells, Cultured; D024042:Collagen Type I / Q000378:metabolism; D020405:Dermis / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D004305:Dose-Response Relationship, Drug; D005109:Extracellular Matrix / Q000187:drug effects* / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D005353:Fibronectins / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology*; D006801:Humans; D008297:Male; D020781:Matrix Metalloproteinase 1 / Q000235:genetics / Q000378:metabolism; D008875:Middle Aged; D012333:RNA, Messenger; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor / Q000187:drug effects / Q000378:metabolism; D045743:Scleroderma, Diffuse / Q000378:metabolism* / Q000473:pathology; D019063:Tenascin / Q000378:metabolism
D024042:Collagen Type I; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D012333:RNA, Messenger; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor; D019063:Tenascin; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D020781:Matrix Metalloproteinase 1