Chitosan and chitosan sulfate have opposing effects on collagen-fibroblast interactions

Soon after injury, hyaluronan is prominent in granulation tissue. As hyaluronan wanes, sulfated glycosaminoglycans predominate. The temporal relationship between the transition from unsulfated to sulfated glycosaminoglycans and the phenotypic changes in fibroblasts in the wound bed suggest that these two events are interrelated. This possibility was investigated using chitosan and its sulfated product as model compounds. The ability of cultured human foreskin fibroblasts to bind and to contract lattices of collagen, collagen-chitosan, and collagen-chitosan sulfate was determined. Fibroblast adherence to substrates after 24 hours was determined by the MTT assay at A570. Adherence to the collagen-chitosan substrate was markedly reduced (mean A570 +/- SD; 0.16 +/- 0.05, n = 6) (p < 0.01) compared to collagen alone (0.92 +/- 0.04) or to collagen-chitosan sulfate (0.84 +/- 0.05). Kinetics of contraction of lattices by enmeshed fibroblasts was determined by planimetric measurements, 0-48 hours after loosening the lattices. Contraction of the collagen-chitosan lattices (n = 5) was less at all time points than for the other two lattices. After 48 hours, the collagen- chitosan lattices contracted significantly (p < 0.01) less (30.0% +/- 4.4) compared to collagen alone (66.9% +/- 4.7) and collagen-chitosan sulfate (71.6% +/- 7.7). Scanning electron microscopy of the acellular lattices showed fibers of the collagen-chitosan mixture to be the thickest and with altered organization. These results show that chitosan sulfation markedly enhances fibroblast adhesion and promotes contraction of a collagen lattice compared to the unsulfated material. By analogy to the in vivo sequence of hyaluronan followed by sulfated glycosaminoglycans in wounds, the results suggest that glycosaminoglycan sulfation may be a contributing signal for phenotypic transformation during wound healing.