Abstract
Specific sites and sequences in collagen to which cells can attach, either directly or through protein intermediaries, were identified using Toolkits of 63-amino acid triple-helical peptides and specific shorter GXX′GEX″ motifs, which have different intrinsic affinity for integrins that mediate cell adhesion and migration. We have previously reported that collagen type I (COL-I) was able to prime in vitro the respiratory burst and induce a specific set of immune- and extracellular matrix-related molecules in phagocytes of the teleost fish gilthead seabream (Sparus aurata L.). It was also suggested that COL-I would provide an intermediate signal during the early inflammatory response in gilthead seabream. Since fibroblasts are highly involved in the initiation of wound repair and regeneration processes, in the present study SAF-1 cells (gilthead seabream fibroblasts) were used to identify the binding motifs in collagen by end-point and real-time cell adhesion assays using the collagen peptides and Toolkits. We identified the collagen motifs involved in the early magnesium-dependent adhesion of these cells. Furthermore, we found that peptides containing the GFOGER and GLOGEN motifs (where O is hydroxyproline) present high affinity for SAF-1 adhesion, expressed as both cell number and surface covering, while in cell suspensions, these motifs were also able to induce the expression of the genes encoding the proinflammatory molecules interleukin-1β and cyclooxygenase-2. These data suggest that specific collagen motifs are involved in the regulation of the inflammatory and healing responses of teleost fish.
Highlights
Collagens are major components of the extracellular matrix (ECM) of all metazoans (Kramer, 1994)
We showed in the teleost fish gilthead seabream (Sparus aurata L.) that native Collagen type I (COL-I) can act as a damage associated molecular pattern (DAMP) by increasing the respiratory burst in leukocytes and the mRNA levels of gene coding for IL-1 and other pro-inflammatory molecules in professional phagocytes (Castillo-Briceno et al, 2009)
The end-point adhesion assay results showed that SAF-1 cells had higher affinity for GLOGER, GFOGER- and GLKGENGLOGENcontaining peptides in COL-II (Fig. 1, Supplementary Table 2) and COL-III (Fig. 2, Supplementary Table 3) Toolkits
Summary
Collagens are major components of the extracellular matrix (ECM) of all metazoans (Kramer, 1994). Fibrillar collagen types I and II are the most abundant proteins of ECM (Wess, 2005), and like the other fibril-forming collagens (types III, V, XI, XXIV and XXVII), they are able to form continuous triple-helical domains (Heino et al, 2009) These collagens can form stable large fibrils and complex fibrous superstructures that are responsible for the tensile strength of the tissues (Perumal et al, 2008; Heino et al, 2009; Herr and Farndale, 2009). We showed in the teleost fish gilthead seabream (Sparus aurata L.) that native COL-I can act as a damage associated molecular pattern (DAMP) by increasing the respiratory burst in leukocytes and the mRNA levels of gene coding for IL-1 and other pro-inflammatory molecules in professional phagocytes (Castillo-Briceno et al, 2009)
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