Effects of interferons on proliferation and collagen synthesis of rat palatal wound fibroblasts

Abstract

The purpose was to select drugs that specifically reduce collagen synthesis by palatal granulation fibroblasts without affecting their proliferation. Granulation fibroblasts were obtained from 8-day-old palatal mucoperiosteal wounds and normal fibroblasts from palatal tissue of unwounded rats. Cultured cells were treated with interferon-α2b, interferon-β and interferon-γ (0, 100, 1000, and 10000 U/ml). Cell proliferation was measured by [³H]thymidine incorporation. Collagen synthesis and non-collagenous protein synthesis were determined from the incorporation of [³H]proline. None of the interferons significantly inhibited the proliferation of either type of fibroblasts. Interferon-α2b had no effect on the variables studied at the dosages used. Interferon-β reduced collagen synthesis of granulation fibroblasts without affecting their non-collagenous protein synthesis or protein synthesis by normal fibroblasts. Interferon-γ reduced collagen synthesis of both types of fibroblast and the non-collagenous protein synthesis of granulation fibroblasts. These data show that interferon-β specifically reduces collagen synthesis by oral granulation fibroblasts without affecting normal palatal fibroblasts.

Introduction

Scar formation after cleft palate surgery is one of the major factors causing aberrant maxillary growth and dento-alveolar development (Ross, 1987). Consequently, the inhibition of scar formation seems a feasible way of improving the clinical outcome.

Tissue injury triggers a complex cascade of cellular and biochemical events that finally results in a scar. After injury, there is inflammation, fibroblast proliferation, matrix synthesis and remodelling (Witte and Barbul, 1997). Every phase contributes to the amount of scarring by influencing the rate, quality and total amount of matrix deposition. Oral palatal wounds also pass through these phases of wound healing (Kahnberg and Thilander, 1982). The number of collagen fibres in palatal mucoperiosteal wounds is increased up to 4 weeks after wounding (Searls et al., 1979). This seems to be due not only to an increased number of cells but also to an increase in collagen production per cell (Moriyama et al., 1991).

A comparison of adult and foetal wound healing has shown that foetal wounds heal without scarring (Longaker and Adzick, 1991). This has been attributed to a lower inflammatory response (Whitby and Ferguson, 1991a) and a different cytokine profile (Whitby and Ferguson, 1991b), which suggests that manipulation of wound healing by altering the cytokine profile could lead to scarless healing. Only a few cytokines have been identified that have the potential to reduce scarring; interferons represent the first well-defined group of polypeptides that exert this property (Granstein et al., 1990a).

The ability of interferons to modulate wound healing has been studied in vivo and on a variety of cell types in vitro. IFN-α, IFN-β and IFN-γ reduce collagen synthesis by dermal fibroblasts (Duncan et al., 1995), hypertrophic scar fibroblasts (Tredget et al., 1993) and fibroblasts derived from sclerodermal lesions (Duncan and Berman, 1987). In vivo studies in mice have demonstrated that IFN-γ reduced the increased collagen synthesis associated with the fibrotic response to an implanted foreign body (Granstein et al., 1990a). This effect was also found in bleomycin-induced pulmonary fibrosis and the healing response to cutaneous burn wounds (Granstein et al., 1990a). The therapeutic relevance of these in vitro and in vivo observations has been confirmed in clinical trials. Intralesional injection of IFN-α2b (Berman and Duncan, 1989) or IFN-γ (Granstein et al., 1990b) reduced the sizes of keloid and hypertrophic scars (Larrabee et al., 1990).

The regulation of protein and collagen synthesis in normal fibroblasts by interferon in vitro depends on the anatomical site (Smith and Higgins, 1993, Martens et al., 1992). Moreover, dermal fibroblasts show a modulated response to IFN-γ according to their stage of differentiation during wound healing (Moulin et al., 1998). For wound healing factors such as migration (Irwin et al., 1994) and matrix remodelling (Stephens et al., 1996), it has been shown that there are phenotypic differences between dermal and oral fibroblasts. Therefore it is quite possible that there are differences in the regulation of proliferation and protein synthesis by interferons between dermal and oral fibroblasts, and between fibroblasts from palatal granulation tissue and normal palatal tissue.

Our aim now was to determine the effects of interferons on proliferation, collagen synthesis, and non-collagenous protein synthesis of mucoperiosteal granulation and normal fibroblasts.