Angiogenic potential of gellan gum-based hydrogels for application in nucleus pulposus regeneration: In vivo study

Hydrogels for nucleus pulposus (NP) regeneration should be able to comprise a non-angiogenic or even anti-angiogenic feature. Gellan gum-based hydrogels have been reported to possess adequate properties for being used as NP substitutes in acellular and cellular strategies, due to its ability to support cells encapsulation, adequate mechanical properties and non-cytotoxicity. In this study, the angiogenic response of gellan gum-based hydrogels was investigated by performing the chorioallantoic membrane (CAM) assay. The convergence of macroscopic blood vessels toward the gellan gum (GG), ionic- (iGG-MA) and photo-crosslinked methacrylated gellan gum (phGG-MA) hydrogel discs was quantified. Gelatin sponge (GSp) and filter paper (FP) alone and with VEGF, were used as controls of angiogenesis. The images obtained were digitally processed and analyzed by three independent observers. The macroscopic blood vessel quantification demonstrated that the gellan gum-based hydrogels are not angiogenic as compared to FP controls. No statistical differences between the gellan gum-based hydrogels tested in respect to its angiogenic ability were observed. Haematoxylin & eosin staining and SNA-lectin immunohistochemistry assay indicated that the iGG-MA and phGG-MA hydrogels do not allow the ingrowth of chick endothelial cells, following 4 days of implantation. On the contrary, GG, GSp and FP controls allowed cells infiltration. The histological data also indicated that the gellan gum-based hydrogels do not elicit any acute inflammatory response. The results showed that the GG, iGG-MA and phGG-MA hydrogels present different permeability to cells but functioned as a physical barrier for vascular invasion. These hydrogels present promising and tunable properties for being used as NP substitutes in the treatment of degenerative intervertebral disc.