Controlled Release Of Growth Factors Research Articles

Bioactive polymer composite scaffolds fabricated from 3D printed negative molds enable bone formation and vascularization

Scaffolds for bone defect treatment should ideally support vascularization and promote bone formation, to facilitate the translation into biomedical device applications. This study presents a novel approach utilizing 3D-printed water-dissolvable polyvinyl alcohol (PVA) sacrificial molds to engineer polymerized High Internal Phase Emulsion (polyHIPE) scaffolds with microchannels and distinct multiscale porosity. Two sacrificial mold variants (250 µm and 500 µm) were generated using fused deposition modeling, filled with HIPE, and subsequently dissolved to create polyHIPE scaffolds containing microchannels. In vitro assessments demonstrated significant enhancement in cell infiltration, proliferation, and osteogenic differentiation, underscoring the favorable impact of microchannels on cell behavior. High loading efficiency and controlled release of the osteogenic factor BMP-2 were achieved, with microchannels facilitating release of the growth factor. Evaluation in a mouse critical-size calvarial defect model revealed enhanced vascularization and bone formation in microchanneled scaffolds containing BMP-2. This study not only introduces an accessible method for creating multiscale porosity in polyHIPE scaffolds but also emphasizes its capability to enhance cellular infiltration, controlled growth factor release, and in vivo performance. The findings suggest promising applications in bone tissue engineering and regenerative medicine, and are expected to facilitate the translation of this type of biomaterial scaffold. Statement of significanceThis study holds significance in the realm of biomaterial scaffold design for bone tissue engineering and regeneration. We demonstrate a novel method to introduce controlled multiscale porosity and microchannels into polyHIPE scaffolds, by utilizing 3D-printed water-dissolvable PVA molds. The strategy offers new possibilities for improving cellular infiltration, achieving controlled release of growth factors, and enhancing vascularization and bone formation outcomes. This microchannel approach not only marks a substantial stride in scaffold design but also demonstrates its tangible impact on enhancing osteogenic cell differentiation and fostering robust bone formation in vivo. The findings emphasize the potential of this methodology for bone regeneration applications, showcasing an interesting advancement in the quest for effective and innovative biomaterial scaffolds to regenerate bone defects.

Near-Infrared Responsive Properties of Bone Repair Scaffolds Facilitated by Specific Osteoinductive Photothermal Converters for Highly Efficient Bone Repair.

The effective repair of bone defects has long been a major challenge in clinical practice. Currently, research efforts mostly focus on achieving sufficiently good bone repair, with little attention paid to achieving both good and fast repair. However, achieving highly efficient (H-efficient) bone repair, which is both good and fast, can shorten the treatment cycle and facilitate rapid patient recovery. Therefore, the development of a H-efficient bone repair material is of significant importance. This study incorporated the previously developed osteoinductive photothermal agent (PTA) BPICT into printing paste to prepare a near-infrared (NIR)-responsive BPICT scaffold. Subsequently, the effects of photothermal therapy (PTT) on bone repair and drug release were assessed in vitro. To further validate the H-efficient bone repair properties of the BPICT scaffold, the scaffold was implanted into bone defects and its ability to promote bone repair in vivo was evaluated through radiology and histopathological analysis. The results indicated that compared to scaffolds containing only Icaritin (ICT), the BPICT scaffold can achieve PTT to promote bone repair through NIR irradiation, while also enabling the controlled release of ICT from the scaffold to enhance bone repair. Within the same observation period, the BPICT scaffold achieves more efficient bone repair than the ICT scaffold, significantly shortening the bone repair cycle while ensuring the effectiveness of bone repair. Therefore, the NIR-responsive scaffold based on PTT-mediated controlled release of bone growth factors represents a feasible solution for promoting H-efficient bone repair in the area of bone defects.

467 Enhancing Cell Infiltration and Controlled Growth Factor Release for a Customized 3D-Printed Bone Graft Composite

OBJECTIVES/GOALS: Annually, 1.5 million global patients receive maxillofacial reconstruction. The gold standard, involving bone particulate, lacks reproducibility. To improve this, we have developed a custom 3D-printable, porous cover-core design. This study optimizes the hydrogel core properties and growth factor (GF) release for enhanced bone regeneration. METHODS/STUDY POPULATION: Different ratios of Methacrylated Gelatin (GelMa), Methacrylated Alginate (AlgMa) and tricalcium phosphate (α²-TCP) were combined to optimize cell viability, GF sequestration and mechanical stability. Material characterization was performed using a rheometer to determine the viscoelastic properties of the blends. Release from disks loaded with FGF-containing PLGA microparticles was quantified with an ELISA kit. Furthermore, scanning electron microscopy (SEM) was conducted to quantify hydrogel porosity. In vitro studies were performed using NIH 3T3 murine fibroblasts in Corning Transwells while immunofluorescent, metabolic and osteogenic studies were performed in 96 well plates to investigate cell infiltration, cell adhesion, viability and differentiation, respectively. RESULTS/ANTICIPATED RESULTS: By adjusting the AlgGelMa ratio, we manipulated matrix properties. GelMa possesses excellent durability and cell adhesion due to intrinsic RGD-binding motifs. AlgMa enhanced swelling by 30%, growth factor sequestration by 50% in 24hrs, and matrix storage modulus without increasing the loss modulus which could cause cell migration away from the hydrogel. Varying the AlgGelMa ratio lowered pH, promoted cell infiltration, and reduced fibronectin accumulation. The addition of β-TCP is anticipated to improve cell differentiation towards an osteogenic lineage due to improved elastic modulus, calcium and phosphate ion concentration improving mineral deposition. DISCUSSION/SIGNIFICANCE: These findings suggest through the use of this composite, early cell infiltration can be increased and promoted due to FGF release, leading to increased osteointegration. Our porous cover-core design ensures efficient clot integration and early cell infiltration, enhancing osteointegration through FGF release.

Physicochemical, in vitro and in vivo evaluation of VEGF loaded PCL-mPEG and PDGF loaded PCL-Chitosan dual layered vascular grafts.

The present study has attempted to evaluate the endothelialization and smooth muscle regeneration efficiency of a novel dual-layer small-diameter vascular graft. Two types of layers (PCL-mPEG-VEGF and PCL-Chitosan-PDGF) were fabricated to find out the best layer giving endothelialization support for the lumen and unique contractile function for outer layer of blood vessels. Platelet-derived growth factor (PDGF) and chitosan were immobilized onto PCL surface by aminolysis-based surface modification technique. Besides, Poly (ethylene glycol) methyl ether (mPEG) and vascular endothelial growth factor (VEGF) were directly blended with PCL. Morphological analysis of membranes ensured consistency of average fibers diameter with native extracellular matrix. A favorable interaction of PCL-mPEG-VEGF with cow pulmonary endothelial cells (CPAEs) and PCL-Chitosan-PDGF with rat bone marrow mesenchymal stem cells (RBMSCs) was obtained during in vitro study. Controlled growth factor release patterns were found from both layers. Further, PCL-mPEG-VEGF exhibited endothelial markers expression properties from RBMSCs. Up-regulation of SMCs markers expression was significantly ensured by the PCL-Chitosan-PDGF membrane. Thus, PCL-mPEG-VEGF and PCL-Chitosan-PDGF were preferred as inner and outer layers respectively of a finally prepared tubular hybrid tissue engineered small diameter vascular graft. Finally, the dual-layer vascular graft was implanted onto a rat abdominal aorta model for 2 months. The extracted samples exhibited the presence of endothelial marker (ICAM 1) in the inner layer and smooth muscle cell marker (αSMA) in the outer layer as well as substantial amount of collagen deposition was observed in the both layers.

Chitosan nanoparticles encapsulated into PLA/gelatin fibers for bFGF delivery

Abstract Electrospinning is a trendy method because of the ease of use and the high surface-to-volume ratio. The mechanical and biological properties of polylactic acid (PLA) make it one of the most enticing polymers. Gelatin and PLA together are thought to enhance cellular behavior and hydrophilicity of scaffolds. Furthermore, chitosan nanoparticles (CNPs) can be incorporated into PLA fibers to achieve controlled growth factor release. This study utilized PLA–gelatin nanofibrous scaffolds in which CNPs were encapsulated within PLA fibers to achieve a controlled release of basic fibroblast growth factor (bFGF). To produce CNPs, a simple ionic gelation reaction was used. The optimal diameter of CNPs was determined by investigating chitosan to tricalciumphosphatesodium (TPP) ratio and TPP concentration. Using a spectrophotometer, we measured the release rate of bFGF from CNPS and scaffolds. Images from a scanning electron microscope (SEM) were used to assess the effect of various concentrations of PLA and gelatin on fiber diameter. The results showed that PLA–gelatin scaffolds could stimulate the release of growth factors and promote cell proliferation. Using a two-jet electrospinning device to produce PLA–gelatin fibers in combination with CNPs incorporated within PLA fibers to release the bFGF growth factor is the novelty of this study.

Long-Term Controlled Growth Factor Release Using Layer-by-Layer Assembly for the Development of In Vivo Tissue-Engineered Blood Vessels.

The development of a well-designed tissue-engineered blood vessel (TEBV) still remains a challenge. In recent years, approaches in which the host response to implanted biomaterials is used to generate vascular constructs within the patient’s body have gained increasing interest. The delivery of growth factors to these in situ-engineered vascular grafts might enhance myofibroblast recruitment and the secretion of essential extracellular matrix proteins, thereby optimizing their functional properties. Layer-by-layer (LbL) coating has emerged as an innovative technology for the controlled delivery of growth factors in tissue engineering applications. In this study, we combined the use of surface-etched polymeric rods with LbL coatings to control the delivery of TGF-β1, PDGF-BB, and IGF-1 and steer the foreign body response toward the formation of a functional vascular graft. Results showed that the regenerated tissue is composed of elastin, glycosaminoglycans, and circumferentially oriented collagen fibers, without calcification or systemic spill of the released growth factors. Functional controlled delivery was observed, whereas myofibroblast-rich tissue capsules were formed with enhanced collagen and elastin syntheses using TGF-β1 and TGF-β1/PDGF-BB releasing rods, when compared to control rods that were solely surface-engineered by chloroform etching. By combining our optimized LbL method and surface-engineered rods in an in vivo bioreactor approach, we could regulate the fate and ECM composition of in situ-engineered vascular grafts to create a successful in vivo vascular tissue-engineered replacement.

Spatially and Temporally Controllable BMP-2 and TGF-β3 Double Release From Polycaprolactone Fiber Scaffolds via Chitosan-Based Polyelectrolyte Coatings.

Temporally and spatially controlled growth factor release from a polycaprolactone fiber mat, which also provides a matrix for directional cell colonization and infiltration, could be a promising regenerative approach for degenerated tendon-bone junctions. For this purpose, polycaprolactone fiber mats were coated with tailored chitosan-based nanogels to bind and release the growth factors bone morphogenetic protein 2 (BMP-2) and transforming growth factor-β3 (TGF-β3), respectively. In this work we provide meaningful in vitro data for the understanding of the drug delivery performance and sterilizability of novel implant prototypes in order to lay the foundation for in vivo testing. ELISA-based in vitro release studies were used to investigate the spatial and temporal control of release, as well as the influence of radiation sterilization on protein activity and release behavior. Layer-by-layer coatings based on BMP-2-containing chitosan tripolyphosphate nanogel particles and negatively charged alginate showed a good sustainment of BMP-2 release from chemically modified polycaprolactone fiber mats. Release control improved with increasing layer numbers. The approach of controlling the release via a barrier of cross-linked chitosan azide proved less promising. By using a simple, partial immersion-based dip-coating process, it was possible to apply opposing gradients of the growth factors BMP-2 and TGF-β3. Final radiation sterilization of the growth factor-loaded implant prototypes resulted in a radiation dose-correlated degradation of the growth factors, which could be prevented by lyophilization into protective matrices. For the manufacture of sterile implants, the growth factor loading step must probably be carried out under aseptic conditions. The layer-by-layer coated implant prototypes provided sustained release from opposing gradients of the growth factors BMP-2 and TGF-β3 and thus represent a promising approach for the restoration of tendon-bone defects.

Surface Modification of Polycaprolactone Scaffold With Improved Biocompatibility and Controlled Growth Factor Release for Enhanced Stem Cell Differentiation

Polycaprolactone (PCL) has been widely used as a scaffold material for tissue engineering. Reliable applications of the PCL scaffolds require overcoming their native hydrophobicity and obtaining the sustained release of signaling factors to modulate cell growth and differentiation. Here, we report a surface modification strategy for electrospun PCL nanofibers using an azide-terminated amphiphilic graft polymer. With multiple alkylation and pegylation on the side chains of poly-L-lysine, stable coating of the graft polymer on the PCL nanofibers was achieved in one step. Using the azide-alkyne “click chemistry”, we functionalized the azide-pegylated PCL nanofibers with dibenzocyclooctyne-modified nanocapsules containing growth factor, which rendered the nanofiber scaffold with satisfied cell adhesion and growth property. Moreover, by specific immobilization of pH-responsive nanocapsules containing bone morphogenetic protein 2 (BMP-2), controlled release of active BMP-2 from the PCL nanofibers was achieved within 21 days. When bone mesenchyme stem cells were cultured on this nanofiber scaffold, enhanced ossification was observed in correlation with the time-dependent release of BMP-2. The established surface modification can be extended as a generic approach to hydrophobic nanomaterials for longtime sustainable release of multiplex signaling proteins for tissue engineering.

Biomimetic chitosan scaffolds with long-term controlled release of nerve growth factor repairs 20-mm-long sciatic nerve defects in rats.

Although autogenous nerve transplantation is the gold standard for treating peripheral nerve defects of considerable length, it still has some shortcomings, such as insufficient donors and secondary injury. Composite chitosan scaffolds loaded with controlled release of nerve growth factor can promote neuronal survival and axonal regeneration after short-segment sciatic nerve defects. However, the effects on extended nerve defects remain poorly understood. In this study, we used chitosan scaffolds loaded with nerve growth factor for 8 weeks to repair long-segment (20 mm) sciatic nerve defects in adult rats. The results showed that treatment markedly promoted the recovery of motor and sensory functions. The regenerated sciatic nerve not only reconnected with neurons but neural circuits with the central nervous system were also reconstructed. In addition, the regenerated sciatic nerve reconnected the motor endplate with the target muscle. Therefore, this novel biomimetic scaffold can promote the regeneration of extended sciatic nerve defects and reconstruct functional circuits. This provides a promising method for the clinical treatment of extended peripheral nerve injury. This study was approved by the Animal Ethics Committee of Capital Medical University, China (approval No. AEEI-2017-033) on March 21, 2017.

Hydroxyapatite sonosensitization of ultrasound-triggered, thermally responsive hydrogels: An on-demand delivery system for bone repair applications.

While bones have the innate capability to physiologically regenerate, in certain cases regeneration is suboptimal, too slow, or does not occur. Biomaterials-based growth factor delivery systems have shown potential for the treatment of challenging bone defects, however, achieving controlled growth factor release remains a challenge. The objective of this study was to develop a thermally responsive hydrogel for bone regeneration capable of ultrasound-triggered on-demand delivery of therapeutic agents. Furthermore, it was hypothesized that incorporation of hydroxyapatite (HA) into the hydrogel could increase sonosensitization, augmenting ultrasound sensitivity to enable controlled therapeutic release to the target tissue. Alginate thermally responsive P(Alg-g-NIPAAm) hydrogels were fabricated and varying quantities of HA (1, 3, 5, and 7% wt./vol.) incorporated. All hydrogels were highly injectable (maximum injection force below 6.5N) and rheological characterization demonstrated their ability to gel at body temperature. The study demonstrated the ultrasound-triggered release of sodium fluorescein (NaF), bovine serum albumin (BSA), and bone morphogenetic protein 2 (BMP-2) from the hydrogels. Release rates of BSA and BMP-2 were significantly enhanced in the HA containing hydrogels, confirming for the first time the role of HA as a son sensitizer. Together these results demonstrate the potential of these ultrasound-triggered thermally responsive hydrogels for on-demand delivery of therapeutic agents for bone regeneration.

Supercritical emulsion extraction fabricated PLA/PLGA micro/nano carriers for growth factor delivery: Release profiles and cytotoxicity

Controlled delivery of human growth factors is still a challenge in tissue engineering protocols, and poly-lactic acid and poly-lactic-co-glycolic acid carriers have been recently proposed for this purpose. Here, the microencapsulation of two human growth factors, namely Growth Differentiation Factor −5 (hGDF-5) and Transforming Growth Factor β1 (hTGF-β1) was tested, by processing different emulsions with Supercritical Emulsion Extraction (SEE) technology. Polymer molecular weight, co-polymer ratio and surfactant amount in aqueous phases as well as phases mixing rate were varied to fabricate carriers with suitable size and loadings.Carriers with different mean sizes from 0.4 ± 0.09 μm up to 3 ± 0.9 μm were obtained by SEE technology when processing emulsions with different formulations; carriers were loaded with 3 μg/g and 7 μg/g for hGDF-5 and hTGF-β1 with controlled growth factor release over 25 days. Carriers displayed extremely low cytotoxicity when evaluated in Chinese Hamster Ovary cells (CHO-K1). Further, they also exhibited reduced cytotoxicity with respect to carriers obtained by conventional evaporation techniques, and low reactivity on human peripheral blood mononuclear cells (hPBMCs), suggesting their safety and potential use in tissue engineering protocols.

Biomimicking Antibacterial Opto-Electro Sensing Sutures Made of Regenerated Silk Proteins.

Surgical sutures play an important role across a wide range of medical treatments and a wide variety exist, differing in strength, size, composition, and performance. Recently, increasing interest has been paid to bioactive and electronic sutures made of synthetic polymers, owing to their ability to reduce inflammation as well as medically and/or electronically facilitate wound healing. However, integrating sensing capabilities into bioactive sutures without adversely affecting their mechanical strength, biocompatibility, and/or bioactivity remains challenging. In this work, a set of biomimicking, antibacterial, and sensing sutures based on the regenerated silk fibroin is designed and fabricated. These sensing sutures, inspired by the "core-shell" multilayered structure of natural spider-silk fibers, are hierarchically structured and heterogeneously functionalized to allow for the integration of multiple, clinically favorable functions into one suture device. These functions included: reducing inflammation and bacterial infection in wound sites, measuring tension of both the tissue and suture, and aiding tissue healing via multi-modal controlled drug and growth factor release. Critically, these functions are coupled with real-time optical and electronic monitoring capabilities. This approach provides greater insight into multifunctional sutures with inherent sensing capabilities and offers enormous potential in both therapeutic and diagnostic applications.

Evaluation of dual release of stromal cell-derived factor-1 and basic fibroblast growth factor with nerve conduit for peripheral nerve regeneration: An experimental study in mice.

The development of drug delivery systems has enabled the release of multiple bioactive molecules. The efficacy of nerve conduits coated with dual controlled release of stromal cell-derived factor-1 (SDF-1) and basic fibroblast growth factor (bFGF) for peripheral nerve regeneration was investigated. Sixty-two C57BL6 mice were used for peripheral nerve regeneration with a nerve conduit (inner diameter, 1 mm, and length, 7 mm) and an autograft. The mice were randomized into five groups based on the different repairs of nerve defects. In the group of repair with conduits alone (n = 9), a 5-mm sciatic nerve defect was repaired by the nerve conduit. In the group of repair with conduits coated with bFGF (n = 10), SDF-1 (n = 10), and SDF-1/bFGF (n = 10), it was repaired by the nerve conduit with bFGF gelatin, SDF-1 gelatin, and SDF-1/bFGF gelatin, respectively. In the group of repair with autografts (n = 10), it was repaired by the resected nerve itself. The functional recovery, nerve regeneration, angiogenesis, and TGF-β1 gene expression were assessed. In the conduits coated with SDF-1/bFGF group, the mean sciatic functional index value (-88.68 ± 10.64, p = .034) and the axon number (218.8 ± 111.1, p = .049) were significantly higher than the conduit alone group, followed by the autograft group; in addition, numerous CD34-positive cells and micro vessels were observed. TGF-β1 gene expression relative values in the conduits with SDF-1/bFGF group at 3 days (7.99 ± 5.14, p = .049) significantly increased more than the conduits alone group. Nerve conduits coated with dual controlled release of SDF-1 and bFGF promoted peripheral nerve regeneration.

Sustained release of growth factors from photoactivated platelet rich plasma (PRP)

The goal of this study is to specify the ability of polychromatic light source (PAC), providing effective wavelengths in the range of 600–1200 nm (near-infrared region, NIR), to activate human platelets in platelet-rich plasma (PRP) and to achieve sustained and controlled release of growth factors from photoactivated platelets. PRP was isolated from human blood and treated with PAC in different time intervals during 1, 5 and 10 min from 10 cm distance to the platelets. ATP secretion and then, calcium release from platelets significantly increased after light application. Photostimulation of platelets triggered lamellipodia extension, numerous filopodia formation, and platelet agglomeration as activation indicators. P-selectin expression was significantly increased after the application of PAC. In conclusion, PRP was successfully activated with PAC for 10 min and realized activation-dependent sustained growth factor release during 28 days. We proved that PAC which has a great potential of activation of PRP enables sustained growth factor release from PRP with a periodic use for therapeutic applications of PRP.

Controlled Growth Factor Release in 3D-Printed Hydrogels.

Growth factors (GFs) are critical components in governing cell fate during tissue regeneration. Their controlled delivery is challenging due to rapid turnover rates in vivo. Functionalized hydrogels, such as heparin-based hydrogels, have demonstrated great potential in regulating GF release. While the retention effects of various concentrations and molecular weights of heparin have been investigated, the role of geometry is unknown. In this work, 3D printing is used to fabricate GF-embedded heparin-based hydrogels with arbitrarily complex geometry (i.e., teabag, flower shapes). Simplified cylindrical core-shell structures with varied shell thickness are printed, and the rates of GF release are measured over the course of 28 days. Increasing the shell layers' thickness decreases the rate of GF release. Additionally, a mathematical model is developed, which is found capable of accurately predicting GF release kinetics in hydrogels with shell layers greater than 0.5 mm thick (R2 > 0.96). Finally, the sequential release is demonstrated by printing two GFs in alternating radial layers. By switching the spatial order, the delivery sequence of the GFs can be modulated. This study demonstrates how 3D printing can be utilized to fabricate user-defined structures with unique geometry in order to control the rate of GF release in hydrogels.

Microparticles for controlled growth differentiation factor 6 delivery to direct adipose stem cell-based nucleus pulposus regeneration.

Currently, there is no effective long‐term treatment for intervertebral disc (IVD) degeneration, making it an attractive candidate for regenerative therapies. Hydrogel delivery of adipose stem cells (ASCs) in combination with controlled release of bioactive molecules is a promising approach to halt IVD degeneration and promote regeneration. Growth differentiation factor 6 (GDF6) can induce ASC differentiation into anabolic nucleus pulposus (NP) cells and hence holds promise for IVD regeneration. Here, we optimised design of novel poly(DL‐lactic acid‐co‐glycolic acid) (PLGA)–polyethylene glycol–PLGA microparticles to control GDF6 delivery and investigated effect of released GDF6 on human ASCs differentiation to NP cells. Recombinant human (rh)GDF6 was loaded into microparticles and total protein and rhGDF6 release assessed. The effect of microparticle loading density on distribution and gel formation was investigated through scanning electron microscopy. ASC differentiation to NP cells was examined after 14 days in hydrogel culture by quantitative polymerase chain reaction, histological, and immunohistochemical staining in normoxic and IVD‐like hypoxic conditions. RhGDF6 microparticles were distributed throughout gels without disrupting gelation and controlled rhGDF6 release over 14 days. Released GDF6 significantly induced NP differentiation of ASCs, with expression comparable with or exceeding media supplemented rhGDF6. Microparticle‐delivered rhGDF6 also up‐regulated sulphated glycosaminoglycan and aggrecan secretion in comparison with controls. In hypoxia, microparticle‐delivered rhGDF6 continued to effectively induce NP gene expression and aggrecan production. This study demonstrates the effective encapsulation and controlled delivery of rhGDF6, which maintained its activity and induced ASC differentiation to NP cells and synthesis of an NP‐like matrix suggesting suitability of microparticles for controlled growth factor release in regenerative strategies for treatment of IVD degeneration.

Microcapsule Technology for Controlled Growth Factor Release in Musculoskeletal Tissue Engineering.

Tissue engineering strategies have relied on engineered 3-dimensional (3D) scaffolds to provide architectural templates that can mimic the native cell environment. Among the several technologies proposed for the fabrication of 3D scaffold, that can be attractive for stem cell cultivation and differentiation, moulding or bioplotting of hydrogels allow the stratification of layers loaded with cells and with specific additives to obtain a predefined microstructural organization. Particularly with bioplotting technology, living cells, named bio-ink, and additives, such as biopolymer microdevices/nanodevices for the controlled delivery of growth factors or biosignals, can be organized spatially into a predesigned 3D pattern by automated fabrication with computer-aided digital files. The technologies for biopolymer microcarrier/nanocarrier fabrication can be strategic to provide a controlled spatiotemporal delivery of specific biosignals within a microenvironment that can better or faster address the stem cells loaded within it. In this review, some examples of growth factor-controlled delivery by biopolymer microdevices/nanodevices embedded within 3D hydrogel scaffolds will be described, to achieve a bioengineered 3D interactive microenvironment for stem cell differentiation. Conventional and recently proposed technologies for biopolymer microcapsule fabrication for controlled delivery over several days will also be illustrated and critically discussed.

Supramolecular Platform Stabilizing Growth Factors.

High concentrations of supplemented growth factors can cause oversaturation and adverse effects in in vitro and in vivo studies, though these supraphysiological concentrations are often required due to the low stability of growth factors. Here we demonstrate the stabilization of TGF-β1 and BMP4 using supramolecular polymers. Inspired by heparan sulfate, sulfonated peptides were presented on a supramolecular polymer to allow for noncovalent binding to growth factors in solution. After mixing with excipient molecules, both TGF-β1 and BMP4 were shown to have a prolonged half-life compared to the growth factors free in solution. Moreover, high cellular response was measured by a luciferase assay, indicating that TGF-β1 remained highly active upon binding to the supramolecular assembly. The results demonstrate that significant lower concentrations of growth factors can be used when supramolecular polymers bearing growth factor binding moieties are implemented. This approach can also be exploited in hydrogel systems to control growth factor release.

Layered PEGDA hydrogel for islet of Langerhans encapsulation and improvement of vascularization

Islets of Langerhans need to maintain their round morphology and to be fast revascularized after transplantation to preserve functional insulin secretion in response to glucose stimulation. For this purpose, a non-cell-adhesive environment is preferable for their embedding. Conversely, nutrient and oxygen supply to islets is guaranteed by capillary ingrowth within the construct and this can only be achieved in a matrix that provides adhesion cues for cells. In this study, two different approaches are explored, which are both based on a layered architecture, in order to combine these two opposite requirements. A non-adhesive islet encapsulation layer is based on polyethyleneglycole diacrylate (PEGDA). This first layer is combined with a second hydrogel based on thiolated-gelatin, thiolated-heparin and thiolated-hyaluronic acid providing cues for endothelial cell adhesion and acting as a growth factor releasing matrix. In an alternative approach, a conformal PEGDA coating is covalently applied on the surface of the islets. The coated islets are subsequently embedded in the previously mentioned hydrogel containing thiolated glycosaminoglycans. The suitability of this approach as a matrix for controlled growth factor release has been demonstrated by studying the controlled release of VEGF and bFGF for 14 days. Preliminary tube formation has been quantified on the growth factor loaded hydrogels. This approach should facilitate blood vessel ingrowth towards the embedded islets and maintain islet round morphology and functionality upon implantation.Graphical abstract

A pro-angiogenic degradable Mg-poly(lactic-co-glycolic acid) implant combined with rhbFGF in a rat limb ischemia model.

Magnesium (Mg)-based implant has been already used in patients with critical limb ischemia. Site-specific controlled release of recombinant human basic fibroblast growth factor (rhbFGF), has become a promising approach to improve peripheral vascular disease. We report here on a novel combination implant composed of spiral magnesium and a coating made using poly(lactic-co-glycolic acid) (PLGA) with encapsulated rhbFGF (Mg-PLGA-rhbFGF). The preparation method does not involve any complex processes and results in a high encapsulation efficiency (approximately 100%). The degradation of metal Mg raise the microclimate pH in the PLGA polymer, which could well preserve the bioactivity of rhbFGF incorporated in the implant. Mg-PLGA-based, sustained local delivery of rhbFGF promotes post-ischemic angiogenesis and blood flow recovery in rat limb ischemic model. This work marks the first report for controlled release of rhbFGF in combination with metal Mg, and suggests potential therapeutic usefulness of Mg-PLGA-rhbFGF for tissue ischemia.