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Biomaterials

Biomaterials - Poster Session E

Poster N3 - Animal study: Biocompatibility and Biodistribution of Novel Hydrogels

Saturday, October 26, 2024
10:00 AM - 11:00 AM EST
Location: Exhibit Hall E, F & G

Introduction:  Voice disorders due to pathological conditions related to vocal folds can impair quality of life because healthy vocal folds are essential for voice production and phonation. Our research laboratory has been innovating numerous biomaterials for vocal fold repair and regeneration.
Safety and efficacy are the most significant features of innovative biomaterials. The safety of a biomaterial is generally addressed in terms of its biocompatibility, which is mainly determined using histological analysis studying inflammatory/foreign body response. For such studies, the biomaterial is implanted in an animal model, and tissue is collected after the animal has been euthanized at the end of the study. Because histological analysis is an end-point measurement providing one data point per animal, other analytical techniques, such as in vivo fluorescence imaging (IVFI), are being used to get multiple data points from the same animals over a period of time. This practice reduces the number of animals used for the study and the cost of the study. Additionally, it allows for the analysis of the biomaterial at different stages of the inflammatory process within the same animals. Additionally, IVFI can also be used to assess the biodistribution and biodegradation of the materials over a period of time.
In the present study, three hydrogels (Chitosan-based porous viscoelastic hydrogel; PEGDA-GELMA, Polyethylene Glycol Deacetylate - Gelatin methacrylate hydrogel; Silk Fibroin-DAHA, dopamine-grafted Aldehyde functionalized Hyaluronic acid hydrogel) innovated for vocal fold repair and regeneration underwent rat trials to analyze their biocompatibility and biodistribution using IVFI.

Materials and

Methods:  An animal protocol (AUP: MCGL-8275) using 5-20 weeks old Sprague Dawley rats to study biocompatibility and biodistribution of the experimental hydrogels (Chitosan based porous viscoelastic hydrogel, G; PEGDA-GELMA hydrogel, S1; Silk Fibroin-DAHA Hyaluronic acid hydrogel, S2) was approved by Comparative Medicine and Animal Resource Centre (CMARC), McGill University. Subcutaneous injections of hydrogels (G, S1, S2) and controls (commercially available Restylane by Galderma, saline, and blank) were performed on the rat’s dorsal skin (Figure 1). Each animal received six injections on their dorsal surfaces under isoflurane anesthesia. The biocompatibility and biodegradation of the gels were assessed using the In Vivo Imaging System, IVIS100 (PerkinElmer), and histological analysis. A total of 27 animals underwent trials: 5 for pilot trials and 22 underwent actual study. Of 22 rats, 11 animals each underwent a biocompatibility and a biodistribution study.
For biodistribution studies, hydrogels labeled with Cy5.5 fluorescent tags were injected. The biodegradation was monitored using IVIS (Figure 2) to visualize the fluorescent markers at day 3, 7, 14, and 21 time points.
For biocompatibility studies, the hydrogels without fluorescent tags were injected, and the foreign body reactions were monitored by injecting fluorescent antibody tags (ProSense 680 for cathepsin activity representing foreign body reaction) 24 hours before in vivo imaging. The IVIS imaging (Figure 3) was performed at day 3, 7, and 21 time points.
At the end of 21 days, all the animals were euthanized, and the tissue was collected for histological analysis.

Results, Conclusions, and Discussions: 

Results: The biodegradation analysis showed statistically significant differences between the area covered by S2 & S1, and S2 & G on day 3. Days 7, 14, and 21 showed no statistically significant differences. Additionally, S2 showed significantly lower area coverage on days 7, 14, and 21 than on day 3, while other experimental groups (S1 and G) did not show differences. Generally, the area covered by all the groups reduced from day 3 to 21, except for the control, which remained constant.
The biocompatibility analysis showed no statistically significant differences; the cathepsin activity demonstrated by the experimental gels was equivalent to that of commercially available Restylane by Galderma. Generally, the cathepsin activity increased from day 3 to 21.
Histological analysis is ongoing and will be instrumental in comparing biocompatibility analyzed by IVIS. Additionally, we will be able to use histology to explore the Cy5.5 toxicity in the animals analyzed for the biodegradation study.

Discussion: All the components of the experimented hydrogels are known to be compatible; hence, the biocompatibility analysis did not reveal any significant differences among the groups. Significant differences were seen in the biodegradation studies. Although all the animals received an equal quantity of hydrogels (200µl gels) for all the experimental and control groups, S2 showed the highest area distribution on day 3 than S1 and G, but S2 also showed a significant area reduction through 21 days. These findings can be co-related to the degradability profile of the hydrogels/hydrogel components. Chitosan and PEGDA take longer to degrade than silk fibroin, hyaluronic acid, and GELMA. Therefore, S2, a combination of silk fibroin and hyaluronic acid, quickly got biodegraded than other gels: Chitosan-based porous viscoelastic hydrogel, G and PEGDA-GELMA hydrogel, S1.

Conclusion: All the experimental gels showed equivalent biocompatibility to a commercially available hydrogel. S2 hydrogel showed the quickest area reduction, signifying the fastest biodegradation among the experimental and control groups.

Acknowledgements (Optional):  NIH 1R01DC01852