Polyethylene glycol (PEG) hydrogels show promise as scaffolds for growth factor delivery to enhance cartilage repair. However, methods to control growth factor release in vivo are needed. We have recently shown that in vitro polymer degradation and in vitro growth factor release kinetics can be altered using PEG crosslinked with different concentrations of genipin. However, the degradation and behavior of PEG-genipin in vivo within the cartilage repair site are unknown. This study was conducted to test the hypotheses that the degradation of PEG-genipin can be altered in vivo within osteochondral defects by changing the concentration of genipin, and that PEG-genipin is biocompatible within the mammalian diarthrodial environment. PEG-genipin cylindrical polymers crosslinked using 8mM, 17.6 mM, or 35.2 mM of genipin were implanted into osteochondral defects made in the trochlea of 24 male Sprague- Dawley rats (48 knees). Rats were sacrificed at 5 weeks and gross, cross-sectional, and histologic assessments were performed. Altering the genipin concentration changed the in vivo degradation properties of the hydrogel ( p < 0.01). Consistent with in vitro findings, polymer degradation was inversely related to the concentration of genipin. Near-complete degradation was seen at 8 mM, intermediate degradation at 17.6 mM, and minimal degradation at 35.2 mM. The results of this study show the degradation of PEGgenipin can be altered in vivo within osteochondral defects by changing the concentration of genipin and that PEG-genipin is biocompatible within osteochondral defects. This new in vivo data support potential use of PEG-genipin polymer as an innovative delivery system to control in vivo release of growth factors for improving articular cartilage repair.
Small intestinal submucosa particles are a favorable scaffold for preadipocytes, allowing ex vivo proliferation on particles small enough to be injected. Delivery of FGF-2 from poly(lactic-co-glycolic acid) microspheres resulted in cell survival and enhanced vascularization.
Injectable scaffolds are promising substrates for regenerative medicine applications. In this study, multi-arm amino-terminated poly(ethylene glycol) (PEG) hydrogels were cross-linked with genipin, a compound naturally derived from the gardenia fruit. In this study, 4-arm and 8-arm amino-terminated PEG hydrogels crosslinked with varying concentrations of genipin were characterized. Both surface and cross-sectional structures of PEG-based hydrogels were observed by scanning electron microscopy. In vitro gelation time, water uptake, swelling and weight loss of PEG hydrogels in phosphate buffered saline at 37°C were studied. Results showed that the 8-arm PEG demonstrated a much slower gelation time compared to the 4-arm PEG, which may be due to the differing structures of the multi-arm PEG hydrogels, which in turn affects the ability of genipin to react with the amine groups. Human adipose-derived stem cells (ASCs) were seeded onto the 4-arm and 8-arm PEG hydrogels in vitro to assess the biological performance and applicability of the gels as cell carriers. The 4-arm PEG hydrogel resulted in enhanced cell adhesion as compared to the 8-arm PEG hydrogel. Overall, these characteristics provide a potential opportunity for multi-arm PEG hydrogels as injectable scaffolds in a variety of tissue engineering applications.
Insulin and dexamethasone were encapsulated in poly(lactic-co-glycolic acid) (PLGA) microspheres to induce adipogenesis for potential applications in soft tissue reconstruction. Release kinetics and bioactivity of the drugs were examined. Surface morphology and diameter of the PLGA microspheres was evaluated using scanning electron microscopy. The release of insulin was determined using ELISA whereas the release of dexamethasone was evaluated spectrophotometrically. The activity of the drugs was assessed by releasing the drugs in the presence of human adipose-derived stem cells. The ability of the cells cultured with microspheres to differentiate into adipocytes was evaluated using Oil Red O stains. Cells treated with the dexamethasone and insulin microspheres demonstrated a significant increase in lipid inclusions compared with control groups. Insulin and dexamethasone microspheres can reproduce the adipogenic effect exerted by differentiation medium, and may represent a clinically relevant method of stimulating adipogenesis in tissue engineering therapies.
We have encapsulated the chemotherapeutic agent doxorubicin into biodegradable polymer microspheres, and incorporated these microspheres into gelatin scaffolds, resulting in a controlled delivery system. Doxorubicin was encapsulated in poly(D,L-lactide-co-glycolide) (PLGA) using a double emulsion/solvent extraction method. Characterization of the microspheres including diameter, surface morphology, and in vitro drug release was determined. The release of doxorubicin up to 30 days in phosphate buffered solution was assessed by measuring the absorbance of the releasate solution. Gelatin scaffolds were crosslinked using glutaraldehyde and microspheres were added to gelatin during gelation. The murine mammary mouse tumor cell line, 4T1, was treated with various doses of doxorubicin. A propidium iodide assay was utilized to visualize dead cells. Using a Transwell basket assay, PLGA microspheres and gelatin constructs were suspended above 4T1 cells for 48 h. Viable cells were determined using the CyQUANT cell proliferation assay. Results indicate that the release was controlled by the incorporation of PLGA microspheres into gelatin constructs. A significant difference was seen in the cumulative release over days 5-16 (p < 0.05). The bioactivity of doxorubicin released from the microspheres and scaffolds was maintained as proven by significant reduction in viable cells after treatment with PLGA microspheres as well as with the gelatin constructs (p < 0.001). The drug-polymer conjugate can be used as a controlled drug delivery system in a biocompatible scaffold that could potentially promote preservation of soft tissue contour.
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