FGFs and their receptors, in vitro and in vivo studies: New FGF receptor in the brain, FGF‐1 in muscle, and the use of functional analogues of low‐affinity heparin‐binding growth factor receptors in tissue repair

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Abstract
Several heparin‐binding growth factors (HBGFs) are thought to play a key role in the natural processes of tissue homeostasis, regeneration or repair. The HBGFs are active upon release from neighbouring inflammatory or circulating cells, as well as upon release from heparan sulfate proteoglycosaminoglycans that are associated with the extracellular matrix (ECM). To better understand the physiological role of these HBGFs, we have focused our effort on studying a subset of HBGFs, namely FGF‐1 and FGF‐2 and their receptors. We present the purification and characterisation of a new form of heparinbinding FGF receptor from adult bovine brain (Perderiset et al., 1992). This receptor has now been purified to homogeneity. Ligand blot and cross‐linking experiments performed with labeled FGF‐1 or FGF‐2 revealed 80‐kd and 130‐kd bands. Preliminary sequence information indicates that receptor is different from the receptors, FGFR‐1 to ‐4, but it may be related the cysteine‐rich‐FGF receptor (CFR).We have previously shown that FGF‐1, but not FGF‐2, is specifically expressed in myoblastic satellite cells during the proliferating phase preceding myoblast alignment and fusion. We have now transfected primary cultures of rat myoblastic satellite cells with FGF‐1 cDNA and expressed this growth factor constitutively. The transfected cells were no longer able to form myotubes. Transfection with antisense FGF‐1 induced myotube formation suggesting that endogenous expression of FGF‐1 is associated with myoblastic cell differentiation.Numerous studies have concluded that the ECM represents a natural reservoir for various HBGFs. The HBGFs are stored through their interaction with matrix‐associated heparan sulfates and can become available for stimulating cell migration, multiplication, and differentiation during tissue repair. In order to better understand how the control of the bioavailability of HBGFs plays a role in wound healing, we have studied the healing effect of various chemically substituted dextrans (CMDBS) selected for their affinity for HBGFs, alone and in association with HBGFs. The CMDBS were obtained by substitution of methyl carboxylic (CM), benzylamide (B), and benzylamine sulfonate (S) groups. We expected that the CMDBS could act in the following ways: (1) to potentiate the biological activity of FGF‐1 and FGF‐2; (2) to protect FGF‐1 and FGF‐2 against thermal or pH inactivation; or (3) to protect FGF‐1 and FGF‐2 against proteolytic degradation (Tardieu et al., 1992). Selected CMDBSs were tested alone in cutaneous and flat bone wound‐healing models. Rats were skin punched and skin regeneration was studied by morphometric and histological analysis. The wounds (6‐mm diameter) were filled with collagen plaster alone or soaked with CMDBS. The CMDBS in collagen plaster was able to induce a remarkable effect both on the kinetics and on the quality of the restored skin.We have also looked at the effect of CMDBS in the healing of calvarian bone defects. Adult rats were trephined (5‐mm diameter) and the healing of their defects was determined after 35 days. Only those treated with CMDBS show appreciable new bone formation. The filling of defects was almost complete, and the response was dose‐dependent. Optimal doses were at 50‐100 m̈g CMDBS, while at 400 m̈g CMDBS, less bone was formed. Interestingly, when the defect included bone suture, the suture was restored after CMDBS treatment. We have measured by biochemical, immunological, and histological methods the content and distribution of several HBGFs in the granulation tissue formed during healing, in the presence or absence of CMDBS. Our results suggest that the endogenous HBGFs naturally released during the regeneration process could be trapped, protected, and released by CMDBS. In conclusion, biopolymers can be designed to mimic some of the mechanisms regulating the availability of growth factors and so could be used as wound‐healing agents.