Synthetic polymeric materials to be used for biomedical applications should be biocompatible and should not be carcinogenic or immunogenic when implanted into the host.16
For tissue engineering applications, biodegradability of synthetic materials is also crucial, and the degradation rate need to be required to match the rate of tissue formation. Bioactivity is another requirement for synthetic materials to interact with surrounding cells towards wound healing or tissue regeneration.17
In the previous report, we have developed new exciting class of biodegradable hydrogels based on PEG and sebacic acid (e.g., PEGSDA) and have shown to be biocompatible and biodegradable in vitro and were equally bioactive with or without modification with cell adhesion peptides, such as RGDS. As mentioned previously, these hydrogels have two main building blocks, PEG and sebacic acid and PEG is supposed to render resulting macromers biocompatible due to its well known biocompatibility and sebacic acid makes the hydrogels absorb less water, thus increase mechanical properties due to its hydrophobic nature.9,18
We also evaluated the cytotoxicity of initial PEGSDA macromers before crosslinking and final degradation products by culturing rat-derived bone marrow stromal cells (MSCs) in the presence of these molecules and found no change in metabolic activity of the cells compared to those cultured on control tissue culture polystyrene.9
Although in vitro cellular responses to PEGSDA-based hydrogels appear to be promising, the evaluation of in vivo performance of the materials is a pre-requisite to eliminate the biocompatibility concern of the materials.
In current study, we further evaluated in vivo biodegradability and biocompatibility of PEGSDA hydrogels using a cage implantation system to assess inflammatory response of the materials and then we also evaluated tissue response using direct subcutaneous implantation of the materials. One of the advantages of the cage implantation system is the ability to evaluate the dynamic nature of inflammatory cell function at the implant site throughout the time course without sacrificing animals at each time period.19
Therefore, it is possible to determine the duration and extent of the inflammatory responses, which indicate in vivo biocompatibility of the implanted materials.4
Another advantage of the system is that it provides additional in vivo biodegradation characteristics of the materials.
The stainless steel cages containing test articles were implanted into rats for up to 12 weeks to determine inflammatory response, as well as in vivo biodegradation characteristics. We expected that in vivo degradation rates of the PEGSDA hydrogel were faster than in vitro, as demonstrated by others with other materials due to enzymatic and/or cellular interaction at the interface of the material and tissue.13
As compared with our previously reported in vitro degradation profiles of PEGSDA hydrogels,9
there was no significant difference between in vitro and in vivo degradation rate of this specific hydrogel. We can speculate that non-specific protein adsorption such as albumin or immunoglobulin G (IgG), both of which are known to hinder the cellular interaction with the materials may prevent to accelerate in vivo degradation due to the hydrophobicity of the materials.20
Regarding the effect of RGD peptide on the in vivo degradation rate, there was no significant difference in degradation profiles was detected between PEGSDA and PEGSDA+RGD hydrogels, although the trend showed that the weight loss of PEGSDA+RGD was greater than PEGSDA throughout the test periods. In general, RGD peptide is known to trigger significant interactions with surrounding cells which can trigger rapid degradation of a material.21
However, the peptide modification in this study appeared to show minimal interaction between surrounding cells and the materials likely due to the small amount of the peptide in the polymer network.22
Further, it is also possible to assume that since these materials are hydrophobic, non-specific protein adsorption may essentially ‘hide’ any cellular interaction with the materials.
The inflammatory cell analysis in the exudates from the cages revealed that there was no significant difference between each test group and the control (empty cage). Total leukocyte concentration data showed a typical acute and chronic inflammatory response over time regarding the number of leukocytes in the exudates (), in which PMNs were predominant during the first 7 days (acute inflammatory response), and macrophages and lymphocytes are dominant thereafter, which represents typical chronic inflammatory response.23
Further evaluation may be needed to determine the extracellular matrix protein concentration, such as alkaline phosphatase in the exudates, which may be helpful in understanding the interaction between the constituents of the exudates and the surface of the implants.19
In addition to the cage implant system, which provides a simple and effective means to examine in vivo biocompatibility of biomaterials by monitoring inflammatory cell responses of the exudates withdrawn from the animal, we further evaluated wound healing response associated with the implants by using histology and immunohistochemistry. Tissue response to the implanted materials showed a substantial amount of inflammatory cells in the tissues, regardless of the type of implants at two months after the surgeries (), likely due to an apparent active in vivo biodegradation process.22
Further, histology and immunohistochemistry revealed that PLGA implant showed normal wound healing response as evidenced by the significant presence of mononuclear cells (stained with CD45) such as lymphocytes which is considered chronic inflammation as well as macrophages (stained with CD68),23
possibly foreign body giant cells (FBGCs) at the surface of the implant (). However, PEGSDA-based implants showed significantly less inflammatory cells, compared to PLGA implant. Fibrous capsule formation around PEGSDA and PEGSDA+RGD hydrogel implants was similar or significantly less as compared to the PLGA implant. In general, thick fibrous capsule formation should be avoided to facilitate mass transfer between the implants and surrounding tissues, which maintains the implant function.14
RGD peptide incorporation into the hydrogel appears to have minimal effect on the in vivo tissue responses, probably due to the relatively small amount of peptide incorporated into the hydrogel network (0.4% w/w), which is consistent to the previous study reported by others.1
There was neither massive inflammatory response nor thick fibrous capsule formation which should be avoided to facilitate mass transfer between the implants and surrounding tissues.14
Overall, the implant materials were found to be biocompatible in small animal model. These exciting novel biodegradable and biocompatible hydrogels may be useful in various tissue engineering applications. However, in vivo response studies in large animal models would be necessary before this exciting new class of biodegradable hydrogels can be realized in clinical settings.