In vitro and in vivo biocompatibility evaluation of k-carrageenan hydrogels aimed at applications in regenerative medicine

Abstract

Introduction The development of biomaterials for biomedical applications always requires extensive biological testing to demonstrate the safety of both the material and its degradation components. K-carrageenan is a naturally occurring polysaccharide which forms a hydrogel with potassium ions and the temperature-induced gelation enables its application as an in vitro cell-carrier or as an in vivo injectable system. The aim of this study was to evaluate these novel systems as biomaterials by in vitro biological screening and by in vivo implantation to assess for the inflammatory response.

Methods

In vitro evaluation: The cytotoxicity of the hydrogels was evaluated under standard tests using the L929 cell. Furthermore, the viability and proliferation of encapsulated human adipose stem cells (hASCs) was determined by fluorescence staining and DNA quantification. In vivo evaluation: discs of k-carrageenan were subcutaneous implanted in a wistar rats for up to 15 days. The materials (agarose-control material and k-carrageenan) were positioned into each pocket. Control animals with empty defect and empty defect injected with lipopolysaccharide were used. After each time period, the biomaterial, surrounding tissue and nearby lymph nodes were explanted and used for histological analysis and molecular biology evaluation.

Results

The cytotoxicity test and biological evaluation of k-carrageenan revealed that this polymer is not cytotoxic and enables long term viability and proliferation of cells in vitro. At implant retrieval there were no macroscopic signs of a considerable inflammatory reaction in any of the animals and no cellular exudates was formed around the implants.

Conclusions

The results indicated that k-carrageenan is a biocompatible material and does not cause a severe host response. These results together with those obtained regarding the properties of k-carragenan investigated under other studies indicate that theses hydrogels may be successfully applied in tissue engineering approaches.

Acknowledgements: The authors gratefully acknowledge Portuguese Foundation for Science and Technology (FCT) for the PhD grants of Popa EG (SFRH/BD/64070/2009), Carvalho P.P. (SFRH/BD/44128/2008), Santo VE (SFRH / BD / 39486 / 2007) and Frias AM (SFRH /BPD/45206/2008).