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The CXC chemokine cCAF stimulates precocious deposition of ECM molecules by wound fibroblasts, accelerating development of granulation tissue.

Feugate JE, Wong L, Li QJ, Martins-Green M - BMC Cell Biol. (2002)

Bottom Line: In contrast, stimulation of fibronectin and collagen I both require the entire molecule and do not involve changes in gene expression.Fibronectin accumulation appears to be linked to tenascin production, and collagen I to decreased MMP-1 levels.In addition, cCAF is chemotactic for fibroblasts and accelerates their migration.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell Biology and Neuroscience, University of California, Riverside, California, USA. feugate@citrus.ucr.edu

ABSTRACT

Background: During wound repair, fibroblasts orchestrate replacement of the provisional matrix formed during clotting with tenascin, cellular fibronectin and collagen III. These, in turn, are critical for migration of endothelial cells, keratinocytes and additional fibroblasts into the wound site. Fibroblasts are also important in the deposition of collagen I during scar formation. The CXC chemokine chicken Chemotactic and Angiogenic Factor (cCAF), is highly expressed by fibroblasts after wounding and during development of the granulation tissue, especially in areas where extracellular matrix (ECM) is abundant. We hypothesized that cCAF stimulates fibroblasts to produce these matrix molecules.

Results: Here we show that this chemokine can stimulate precocious deposition of tenascin, fibronectin and collagen I, but not collagen III. Studies in culture and in vivo show that tenascin stimulation can also be achieved by the N-terminal 15 aas of the protein and occurs at the level of gene expression. In contrast, stimulation of fibronectin and collagen I both require the entire molecule and do not involve changes in gene expression. Fibronectin accumulation appears to be linked to tenascin production, and collagen I to decreased MMP-1 levels. In addition, cCAF is chemotactic for fibroblasts and accelerates their migration.

Conclusions: These previously unknown functions for chemokines suggest that cCAF, the chicken orthologue of human IL-8, enhances healing by rapidly chemoattracting fibroblasts into the wound site and stimulating them to produce ECM molecules, leading to precocious development of granulation tissue. This acceleration of the repair process may have important application to healing of impaired wounds.

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Effect of cCAF on FN levels in culture. Embryonic connective tissue fibroblasts immunolabeled for FN. (A) Untreated 3 day-old cultures show FN fibers over the cells. (B) Cultures treated with cCAF for 3 days show more fibers, and the staining is more intense than those in the controls. (C,D) Cells treated with the same amounts of N-peptide or C-peptide have levels of FN similar to control. (E) Immunoblot analysis of cells treated with cCAF show higher levels of FN than untreated or peptide-treated cells. All lanes contain equal amounts of total protein, as measured by the DC protein assay (BioRad). (F) Northern blot analysis for FN shows no difference in mRNA levels between control and cells treated with cCAF for times up to 48 hours.
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Figure 3: Effect of cCAF on FN levels in culture. Embryonic connective tissue fibroblasts immunolabeled for FN. (A) Untreated 3 day-old cultures show FN fibers over the cells. (B) Cultures treated with cCAF for 3 days show more fibers, and the staining is more intense than those in the controls. (C,D) Cells treated with the same amounts of N-peptide or C-peptide have levels of FN similar to control. (E) Immunoblot analysis of cells treated with cCAF show higher levels of FN than untreated or peptide-treated cells. All lanes contain equal amounts of total protein, as measured by the DC protein assay (BioRad). (F) Northern blot analysis for FN shows no difference in mRNA levels between control and cells treated with cCAF for times up to 48 hours.

Mentions: Cellular FN is abundantly present in granulation tissue, is produced primarily by the wound fibroblasts and myofibroblasts. Fibroblast cultures treated as for TN, produced much more FN, as shown by immunolabeling (Figs. 3A &3B) and immunoblot analysis (Fig. 3E). Unlike TN, this effect can only be accomplished by the whole cCAF molecule; immunolabeling and immunoblot analysis of cells treated with the N-peptide (Fig. 3C &3E) or C-peptide (Fig. 3D &3E) revealed similar FN levels to control. Unlike TN, cCAF did not affect FN mRNA levels (Fig. 3F). In vivo, immunostaining for FN, 3 days after making excision wounds, showed that levels of this protein are high in the healing tissues and that treatment with cCAF increased FN accumulation in the granulation tissue (Fig. 4A &4B). Immunoblot analysis for FN extracted from excision wounds showed that by 3 days after injury the cCAF-treated wounds contained more FN than the control wounds but that the levels by 7 days are similar to those of the control (Fig. 4C).


The CXC chemokine cCAF stimulates precocious deposition of ECM molecules by wound fibroblasts, accelerating development of granulation tissue.

Feugate JE, Wong L, Li QJ, Martins-Green M - BMC Cell Biol. (2002)

Effect of cCAF on FN levels in culture. Embryonic connective tissue fibroblasts immunolabeled for FN. (A) Untreated 3 day-old cultures show FN fibers over the cells. (B) Cultures treated with cCAF for 3 days show more fibers, and the staining is more intense than those in the controls. (C,D) Cells treated with the same amounts of N-peptide or C-peptide have levels of FN similar to control. (E) Immunoblot analysis of cells treated with cCAF show higher levels of FN than untreated or peptide-treated cells. All lanes contain equal amounts of total protein, as measured by the DC protein assay (BioRad). (F) Northern blot analysis for FN shows no difference in mRNA levels between control and cells treated with cCAF for times up to 48 hours.
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Figure 3: Effect of cCAF on FN levels in culture. Embryonic connective tissue fibroblasts immunolabeled for FN. (A) Untreated 3 day-old cultures show FN fibers over the cells. (B) Cultures treated with cCAF for 3 days show more fibers, and the staining is more intense than those in the controls. (C,D) Cells treated with the same amounts of N-peptide or C-peptide have levels of FN similar to control. (E) Immunoblot analysis of cells treated with cCAF show higher levels of FN than untreated or peptide-treated cells. All lanes contain equal amounts of total protein, as measured by the DC protein assay (BioRad). (F) Northern blot analysis for FN shows no difference in mRNA levels between control and cells treated with cCAF for times up to 48 hours.
Mentions: Cellular FN is abundantly present in granulation tissue, is produced primarily by the wound fibroblasts and myofibroblasts. Fibroblast cultures treated as for TN, produced much more FN, as shown by immunolabeling (Figs. 3A &3B) and immunoblot analysis (Fig. 3E). Unlike TN, this effect can only be accomplished by the whole cCAF molecule; immunolabeling and immunoblot analysis of cells treated with the N-peptide (Fig. 3C &3E) or C-peptide (Fig. 3D &3E) revealed similar FN levels to control. Unlike TN, cCAF did not affect FN mRNA levels (Fig. 3F). In vivo, immunostaining for FN, 3 days after making excision wounds, showed that levels of this protein are high in the healing tissues and that treatment with cCAF increased FN accumulation in the granulation tissue (Fig. 4A &4B). Immunoblot analysis for FN extracted from excision wounds showed that by 3 days after injury the cCAF-treated wounds contained more FN than the control wounds but that the levels by 7 days are similar to those of the control (Fig. 4C).

Bottom Line: In contrast, stimulation of fibronectin and collagen I both require the entire molecule and do not involve changes in gene expression.Fibronectin accumulation appears to be linked to tenascin production, and collagen I to decreased MMP-1 levels.In addition, cCAF is chemotactic for fibroblasts and accelerates their migration.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell Biology and Neuroscience, University of California, Riverside, California, USA. feugate@citrus.ucr.edu

ABSTRACT

Background: During wound repair, fibroblasts orchestrate replacement of the provisional matrix formed during clotting with tenascin, cellular fibronectin and collagen III. These, in turn, are critical for migration of endothelial cells, keratinocytes and additional fibroblasts into the wound site. Fibroblasts are also important in the deposition of collagen I during scar formation. The CXC chemokine chicken Chemotactic and Angiogenic Factor (cCAF), is highly expressed by fibroblasts after wounding and during development of the granulation tissue, especially in areas where extracellular matrix (ECM) is abundant. We hypothesized that cCAF stimulates fibroblasts to produce these matrix molecules.

Results: Here we show that this chemokine can stimulate precocious deposition of tenascin, fibronectin and collagen I, but not collagen III. Studies in culture and in vivo show that tenascin stimulation can also be achieved by the N-terminal 15 aas of the protein and occurs at the level of gene expression. In contrast, stimulation of fibronectin and collagen I both require the entire molecule and do not involve changes in gene expression. Fibronectin accumulation appears to be linked to tenascin production, and collagen I to decreased MMP-1 levels. In addition, cCAF is chemotactic for fibroblasts and accelerates their migration.

Conclusions: These previously unknown functions for chemokines suggest that cCAF, the chicken orthologue of human IL-8, enhances healing by rapidly chemoattracting fibroblasts into the wound site and stimulating them to produce ECM molecules, leading to precocious development of granulation tissue. This acceleration of the repair process may have important application to healing of impaired wounds.

Show MeSH
Related in: MedlinePlus