Engineering retinal progenitor cell and scrollable poly(glycerol-sebacate) composites for expansion and subretinal transplantation.
ABSTRACT: Retinal degenerations cause permanent visual loss and affect millions world-wide. Presently, a novel treatment highlights the potential of using biodegradable polymer scaffolds to induce differentiation and deliver retinal progenitor cells for cell replacement therapy. In this study, we engineered and analyzed a micro-fabricated polymer, poly(glycerol sebacate) (PGS) scaffold, whose useful properties include biocompatibility, elasticity, porosity, and a microtopology conducive to mouse retinal progenitor cell (mRPC) differentiation. In vitro proliferation assays revealed that PGS held up to 86,610 (+/-9993) mRPCs per square millimeter, which were retained through simulated transplantations. mRPCs adherent to PGS differentiated toward mature phenotypes as evidenced by changes in mRNA, protein levels, and enhanced sensitivity to glutamate. Transplanted composites demonstrated long-term mRPC survival and migrated cells exhibited mature marker expression in host retina. These results suggest that combining mRPCs with PGS scaffolds for subretinal transplantation is a practical strategy for advancing retinal tissue engineering as a restorative therapy.
Project description:Poly(glycerol sebacate) (PGS) is a thermally-crosslinked elastomer suitable for tissue regeneration due to its elasticity, degradability, and pro-regenerative inflammatory response. Pores in PGS scaffolds are typically introduced by porogen leaching, which compromises strength. Methods for producing fibrous PGS scaffolds are very limited. Electrospinning is the most widely used method for laboratory scale production of fibrous scaffolds. Electrospinning PGS by itself is challenging, necessitating a carrier polymer which can affect material properties if not removed. We report a simple electrospinning method to produce distinct PGS fibers while maintaining the desired mechanical and cytocompatibility properties of thermally crosslinked PGS. Fibrous PGS demonstrated 5 times higher tensile strength and increased suture retention compared to porous PGS foams. Additionally, similar modulus and elastic recovery were observed. A final advantage of fibrous PGS sheets is the ability to create multi-laminate constructs due to fiber bonding that occurs during thermal crosslinking. Taken together, these highly elastic fibrous PGS scaffolds will enable new approaches in tissue engineering and regenerative medicine.
Project description:Poly(glycerol sebacate) (PGS) is a biocompatible, biodegradable elastomer that has been shown promise as a scaffolding material for tissue engineering; it is still challenging, however, to produce anisotropic scaffolds by using a thermoset polymer, such as PGS. Previously, we have used aligned sacrificial poly(vinyl alcohol) (PVA) fibers to help produce an anisotropic PGS membrane; a composite membrane, formed by embedding aligned PVA fibers in PGS prepolymer, was subjected to curing and subsequent PVA removal, resulting in aligned grooves and cylindrical pores on the surface of and within the membrane, respectively. PVA, however, appeared to react with PGS during its curing, altering the mechanical characteristics of PGS. In this study, aligned sacrificial fibers made of polylactide (PLA) were used instead. Specifically, PLA was blend-electrospun with polyethylene oxide to increase the sacrificial fiber diameter, which in turn increased the size of the grooves and cylindrical pores. The resultant PGS membrane was shown to be in vitro cyto-compatible and mechanically anisotropic. The membrane's Young's modulus was 1-2 MPa, similar to many soft tissues. In particular, the microscale grooves on the membrane surface were found to be capable of directing cell alignment. Finally, based on the same approach, we fabricated a biomimetic, anisotropic, PGS tubular scaffold. The compliance of the tubular scaffold was comparable to native arteries and in the range of 2% to 8% per 100 mmHg, depending on the orientations of the sacrificial fibers. The anisotropic PGS tubular scaffolds can potentially be used in vascular tissue engineering.
Project description:The development of living heart valves that grow with the patient is a promising strategy for heart valve replacements in pediatric patients. Despite active research in the field of tissue engineered heart valves there have been limited efforts to optimize the balance between biodegradation of the scaffolds and de novo extracellular matrix (ECM) synthesis by cells and study their consequences on the mechanical properties of the cell-seeded construct. This study investigates the effect of in vitro degradation and ECM secretion on the mechanical properties of hybrid polyester scaffolds. The scaffolds were synthesized from blends of fast degrading polyglycerol sebacate (PGS) and slowly degrading polycaprolactone (PCL). PGS-PCL scaffolds were electrospun using a 2:1 ratio of PGS to PCL. Accelerated hydrolytic degradation in 0.1 mM sodium hydroxide revealed 2-fold faster degradation of PGS-PCL scaffolds compared with PCL scaffolds. Thermal analysis and scanning electron microscopy demonstrated marginal change in PCL scaffold properties, while PGS-PCL scaffolds showed preferential mass loss of PGS and thinning of the individual fibers during degradation. Consequently, the mechanical properties of PGS-PCL scaffolds decreased gradually with no significant change for PCL scaffolds during accelerated degradation. Valvular interstitial cells (VICs) seeded on PGS-PCL scaffolds showed higher ECM protein secretion compared with PCL. Thus the mechanical properties of the cell-seeded PCL scaffolds did not change significantly compared with acellular scaffolds, probably due to slower degradation and ECM deposition by VICs. In contrast, the PGS-PCL scaffolds exhibited a gradual decrease in the mechanical properties of the acellular scaffolds due to degradation, which was compensated for by new matrix secreted by VICs seeded on the scaffolds. Our study demonstrated that the faster degrading PGS component of PGS-PCL accelerated the degradation rate of the scaffolds. VICs, on the other hand, were able to remodel the synthetic scaffold, depositing new matrix proteins and maintaining the mechanical properties of the scaffolds.
Project description:Poly(glycerol sebacate) (PGS) is an elastomeric polymer which is attracting increasing interest for biomedical applications, including cartilage regeneration. However, its limited mechanical properties and possible negative effects of its degradation byproducts restrict PGS for in vivo application. In this study, a novel PGS-bioactive glass fiber (F18)-reinforced composite was developed and characterized. PGS-based reinforced scaffolds were fabricated via salt leaching and characterized regarding their mechanical properties, degradation, and bioactivity in contact with simulated body fluid. Results indicated that the incorporation of silicate-based bioactive glass fibers could double the composite tensile strength, tailor the polymer degradability, and improve the scaffold bioactivity.
Project description:Immobilized Candida antarctica lipase B (CALB)-catalyzed polycondensation of glycerol and sebacic acid at mild reaction conditions resulted in branched poly(glycerol sebacate) (PGS). To understand how PGS chains grow and branch, the kinetics of the CALB-catalyzed polycondensation were studied. The influence of the reaction temperature, solvent, CALB amount, and sebacic acid/glycerol feed ratio on the poly(glycerol sebacate) (PGS) molecular weight, degree of branching, and glyceridic repetitive unit distribution was also investigated. PGS architecture changes from linear to branched with the progression of the reaction, and the branching results from the simultaneous CALB-catalyzed esterification and acyl migration. For reactions performed in acetone at the temperature range from 30 to 50 °C, the apparent rate constant increases from 0.7 to 1.5 h-1, and the apparent energy of activation of 32 kJ mol-1 was estimated. The higher mass average molecular weight (16 kDa) and degree of branching (41%) were achieved using the equimolar sebacic acid/glycerol feed ratio in acetone at 40 °C with a CALB amount of 13.6 wt % and in the presence of the molecular sieves.
Project description:Poly(glycerol sebacate) (PGS) is an elastomer used widely in tissue engineering studies due to good biocompatibility. Hereby we report a tyramine functionalized PGS called PGS-TA. Tyramine adds a stronger physical bonding capability to PGS-TA. Tensile tests showed that the softness and toughness of the material were similar to PGS. However, PGS-TA demonstrated 16-folds increase of elastic deformations compared to PGS processed under identical conditions. The in vitro studies demonstrated that the viability, and metabolic activity of baboon smooth muscle cells were the same as those on tissue culture polystyrene. Porous subcutaneous implants of PGS-TA substantially degraded in vivo over two weeks, showing good biodegradability and biocompatibility. We expect PGS-TA to be useful for applications in tissues and organs that are subjected to large reversible mechanical deformations.
Project description:A photocurable thiol-ene network polymer, poly(limonene thioether) (PLT32o), is synthesized, characterized, fabricated into tissue engineering scaffolds, and demonstrated in vitro and in vivo. Micromolded PLT32o grids exhibit compliant, elastomeric mechanical behavior similar to grids made of poly(glycerol sebacate) (PGS), an established biomaterial. Multilayered PL32o scaffolds with regular, geometrically defined pore architectures support heart cell seeding and culture in a manner similar to multilayered PGS scaffolds. Subcutaneous implantation of multilayered PLT32o scaffolds with cultured heart cells provides long-term 3D structural support and retains the exogenous cells, whereas PGS scaffolds lose both their structural integrity and the exogenous cells over 31 d in vivo. PLT32o membrane implants retain their dry mass, whereas PGS implants lose 70 percent of their dry mass by day 31. Macrophages are initially recruited to PLT32o and PGS membrane implants but are no longer present by day 31. Facile synthesis and processing in combination with the capability to support heart cells in vitro and in vivo suggest that PLT32o can offer advantages for tissue engineering applications where prolonged in vivo maintenance of 3D structural integrity and elastomeric mechanical behavior are required.
Project description:Poly(glycerol sebacate) (PGS), produced from renewable monomers such as sebacic acid and glycerol, has been explored extensively for various biomedical applications. However, relatively less attention has been paid to explore PGS as sustainable materials in applications such as elastomers and rigid plastics, primarily because of serious deficiencies in physical properties of PGS. Here, we present two new approaches for enhancing the properties of PGS; (i) synthesizing block copolymers of PGS with poly(tetramethylene oxide)glycol (PTMO) and (ii) preparing a blend of PGS-b-PTMO with a poly(ester-ether) thermoplastic elastomer. The consequence of molar ratio (hard and soft segments) and M n of soft segment on tensile properties of the material was investigated. The PGS-b-PTMO with 25:75 mole ratios of hard and soft segments and having a medium M n soft segment (5350 g mol-1) exhibits an appreciable increase in percentage of elongation that is from 32% for PGS to 737%. Blends of PGS-b-PTMO and a thermoplastic polyester elastomer, Hytrel 3078, form a semi-interpenetrated polymer network, which exhibits increased tensile strength to 2.11 MPa and percentage of elongation to 2574. An elongation of such magnitude is unprecedented in the literature for predominantly aliphatic polyesters and demonstrates that the simple polyester can be tailored for superior performance.
Project description:The objective of this study was to evaluate the long-term performance of cell-free vascular grafts made from a fast-degrading elastic polymer. We fabricated small arterial grafts from microporous tubes of poly(glycerol sebacate) (PGS) reinforced with polycaprolactone (PCL) nanofibers on the outer surface. Grafts were interpositioned in rat abdominal aortas and characterized at 1 year post-implant. Grafts remodeled into "neoarteries" (regenerated arteries) with similar gross appearance to native rat aortas. Neoarteries mimic arterial tissue architecture with a confluent endothelium and media and adventita-like layers. Patent vessels (80%) showed no significant stenosis, dilation, or calcification. Neoarteries contain nerves and have the same amount of mature elastin as native arteries. Despite some differences in matrix organization, regenerated arteries had similar dynamic mechanical compliance to native arteries in vivo. Neoarteries responded to vasomotor agents, albeit with different magnitude than native aortas. These data suggest that an elastic vascular graft that resorbs quickly has potential to improve the performance of vascular grafts used in small arteries. This design may also promote constructive remodeling in other soft tissues.
Project description:In this work, a bio-based copolyester with good mechanical properties was synthesized and characterized in terms of structure, main properties and biodegradability Determining the chemical structure of such materials is important to understand their behavior and properties. Performing an extraction of insoluble cross-linked polymer using different solvents allowed us to analyze how the polymer behaves when subjected to different chemical environments, and to obtain soluble samples suitable for more in-depth analysis. Chemical structure of poly (xylitol sebacate-co-butylene sebacate) was determined by a 1H NMR and FTIR analysis of both prepolymer gel sample and samples obtained by extraction of cross-linked polymer using different solvents. Block structure of the copolymer was confirmed by both NMR and DSC. Gel fraction, swelling value, water contact angle, and mechanical properties were also analyzed. Biodegradability of this material was confirmed by performing enzymatic and hydrolytic degradation. Synthesizing sugar-alcohol based copolyester using three monomers leads to obtaining a material with interesting chemical structure and desirable mechanical properties comparable to conventional elastomers.