Increase in cell migration and angiogenesis in a composite silk scaffold for tissue-engineered ligaments.

The purpose of this study was to evaluate the biocompatibility of silk and collagen-hyaluronan (HA) in vitro by assessing anterior cruciate ligament (ACL) cell and T-lymphocyte cultures on scaffolds. The use of composite scaffolds as artificial ligaments in ACL reconstruction and their effects on angiogenesis were evaluated in vivo. The silk scaffold was knitted by hand and dry coated with collagen-HA, whereas the composite silk scaffold was made by covering a silk scaffold with a lyophilized collagen-HA substrate. The initial attachment and proliferation of human ACL cells on the composite silk scaffold was superior to the attachment and proliferation observed on the silk scaffold. The immune response was higher in both scaffolds after 72 h (p < 0.05) compared with the control culture condition without scaffolding, as assessed by T-lymphocyte cultures in vitro. There was no significant difference in the immune response in vitro between the silk and composite silk scaffolds. Silk and composite silk scaffolds were implanted as artificial ligaments in ACLs removed from the knees of dogs, and they were harvested 6 weeks after implantation. On gross examination, the onset of an inflammatory tissue reaction, such as synovitis, was seen in both the silk scaffold and the composite silk scaffold groups. An histological evaluation of the artificial ligament implants revealed the presence of monocytes in the silk composite scaffold and the absence of giant cells in all cases. MT staining in the composite silk scaffold-grafted group showed granulation tissue consisting of fibroblasts, lymphocytes, monocytes, and collagen fibers. In addition, CD31 staining revealed the formation of new blood vessels. On the other hand, no reparative tissues, such as blood vessels, collagen, and cells, were observed in the silk scaffold-grafted group. These results suggest that the lyophilized collagen-HA substrate is biocompatible in vitro and enhances new blood vessel and cell migration in vivo.