TGFβ functionalized starPEG-heparin hydrogels modulate human dermal fibroblast growth and differentiation

Abstract

Hydrogels are promising biomaterials that can adapt easily to complex tissue entities. Furthermore, chemical modifications enable these hydrogels to become an instructive biomaterial to a variety of cell types. Human dermal fibroblasts play a pivotal role during wound healing, especially for the synthesis of novel dermal tissue replacing the primary fibrin clot. Thus, the control of growth and differentiation of dermal fibroblasts is important to modulate wound healing. In here, we utilized a versatile starPEG-heparin hydrogel platform that can be independently adjusted with respect to mechanical and biochemical properties for cultivating human dermal fibroblasts. Cell-based remodeling of the artificial matrix was ensured by using matrix metalloprotease (MMP) cleavable crosslinker peptides. Attachment and proliferation of fibroblasts on starPEG-heparin hydrogels of differing stiffness, density of pro-adhesive RGD peptides and MMP cleavable peptide linkers were tested. Binding and release of human TGFβ1 as well as biological effect of the pre-adsorbed growth factor on fibroblast gene expression and myofibroblast differentiation were investigated. Hydrogels containing RGD peptides supported fibroblast attachment, spreading, proliferation matrix deposition and remodeling compared to hydrogels without any modifications. Reversibly conjugated TGFβ1 was demonstrated to be constantly released from starPEG-heparin hydrogels for several days and capable of inducing myofibroblast differentiation of fibroblasts as determined by induction of collagen type I, ED-A-Fibronectin expression and incorporation of alpha smooth muscle actin and palladin into F-actin stress fibers. Taken together, customized starPEG-heparin hydrogels could be of value to promote dermal wound healing by stimulating growth and differentiation of human dermal fibroblasts.

Statement of significance: The increasing number of people of advanced age within the population results in an increasing demand for the treatment of non-healing wounds. Hydrogels are promising biomaterials for the temporary closure of large tissue defects: They can adapt to complex tissue geometry and can be engineered for specific tissue needs. We used a starPEG-heparin hydrogel platform that can be independently adjusted to mechanical and biochemical characteristics. We investigated how these hydrogels can support attachment, proliferation and differentiation of dermal fibroblasts. After introducing adhesive peptides these hydrogels support cell attachment and proliferation. Moreover, TGFβ - an essential growth and differentiation factor for fibroblasts - can be immobilized reversibly and functionally on these hydrogels. Thus, starPEG-heparin hydrogels could be developed to bioactive temporary wound dressings.

Keywords: Alpha-smooth muscle actin (αSMA); Extracellular matrix (ECM); Heparin; Hydrogels; Metalloprotease (MMP); Myofibroblast; Wound healing; starPEG.