Microsphere Scaffolds Research Articles

Microfluidic-engineered Chinese herbal nanocomposite hydrogel microspheres for diabetic wound tissue regeneration

Microfluidic-engineered hydrogel microspheres have emerged as a promising avenue for advancements in tissue engineering and regenerative medicine, particularly through the precise manipulation of fluids to achieve personalized composite biomaterials. In this study, we employed microfluidic technology to fabricate hydrogel microspheres (HMs) using Chinese herbal Bletilla striata polysaccharide (BSP) as the primary material. Concurrently, the natural active ingredient 20(S)-protopanaxadiol (PPD) was encapsulated within the HMs in the form of liposomes (PPD-Lipo), resulting in the formation of nanocomposite hydrogel microspheres (PPD-Lipo@HMs) intended for diabetic wound tissue repair. PPD-Lipo@HMs are characterized by the expansive specific surface area, adjustable mechanical properties, and exceptional biocompatibility. PPD-Lipo@HMs can stimulate the production of vascular endothelial factors, which in turn enhances the migration of endothelial cells, the creation of tubes, angiogenesis, and tissue repair. Moreover, the PPD-Lipo@HMs accumulation produces a microsphere scaffold that effectively covers damaged tissues, promoting the attachment, spread, and multiplication of fibroblast and endothelial cells. The polysaccharide material BSP within PPD-Lipo@HMs can modulate the immune microenvironment of the damaged tissue, reducing inflammation, encouraging re-epithelialization and granulation tissue formation, accelerating angiogenesis and collagen deposition, ultimately leading to tissue repair. The findings highlight the superior therapeutic efficacy of the microfluidic-engineered PPD-Lipo@HMs in addressing the complex challenges of diabetic wound tissue repair, thereby affirming the significant potential of microfluidic engineering technology in tissue repair applications.

Mussel-inspired multi-bioactive microsphere scaffolds for bone defect photothermal therapy

Hydrogel microspheres hold great promise as scaffolds for bone repair. Their hydrated matrix, biocompatibility, and functional properties make them an attractive choice in regenerative medicine. However, the irregularity of defect requires shape adaptability of the microspheres. Additionally, there is still room for improvement regarding the component of the microspheres to achieve sufficient bioactivity. Here, we prepare multi-bioactive microspheres composed of methacrylated silk fibroin (SFMA) via microfluidic electrospray. Magnesium ascorbyl phosphate (MAP) is encapsulated within the microspheres, whose sustained release facilitates angiogenesis and osteogenic differentiation. The microspheres are further coated with a polydopamine (PDA) layer, allowing them to assemble in situ into a scaffold that conforms to the non-uniform contours of bone defects. The photothermal conversion capability of PDA also provides mild photothermal stimulation to further promote bone regeneration. Based on the synergistic effects, our in vivo experiments demonstrated that the microsphere scaffold effectively promotes bone defect healing. Thus, this multi-bioactive scaffold offers a versatile strategy for bone repair with promising clinical potential.

The induction of bone formation by 3D-printed PLGA microsphere scaffolds in a calvarial orthotopic mouse model: a pilot study.

Polymeric biodegradable microspheres are readily utilized to support targeted drug delivery for various diseases clinically. 3D printed tissue engineering scaffolds from polymer filaments with embedded microspheres or nanoparticles, as well as bulk microsphere scaffolds, have been investigated for regenerative medicine and tissue engineering. However, 3D printed scaffolds consisting only of a homogenous microsphere size with an optimized architecture that includes a unique micro- and macroporosity, have been challenging to produce and hence, have not been assessed in the literature yet. Utilizing our recently established 3D-MultiCompositional Microsphere-Adaptive Printing (3D-McMap) method, the present study evaluated the effectiveness of 3D-printed poly (lactic-co-glycolic acid) (PLGA) microsphere scaffolds, consisting of microsphere sizes 50, 100, or 200μm, on the induction of bone formation when implanted in the calvarial murine regeneration model. Our results showed that PLGA microsphere scaffolds possess unique properties that support bone regeneration by supporting osteoconduction and stimulating, in our opinion, true spontaneous osteoinduction. The study demonstrated that PLGA microsphere-based scaffolds support bone growth in the absence of additional growth factors and promote osteogenesis primarily via their unique geometric configuration. The larger the microspheres were, the greater de novo bone formation was. This proves that bone tissue engineering scaffolds 3D printed from microspheres, enabled by the 3D-McMap method, are superior over bulk material printed scaffolds, as they possess the unique capability of spontaneous induction of new bone formation. With the addition of encapsulated modulatory bone-forming biomolecules they can substantially improve the spatiotemporal control of tissue morphogenesis, potentially leading to new innovative clinical tissue repair therapies that regenerate bone in large defects correctly and fully.

The 3D-McMap Guidelines: Three-Dimensional Multicomposite Microsphere Adaptive Printing.

Microspheres, synthesized from diverse natural or synthetic polymers, are readily utilized in biomedical tissue engineering to improve the healing of various tissues. Their ability to encapsulate growth factors, therapeutics, and natural biomolecules, which can aid tissue regeneration, makes microspheres invaluable for future clinical therapies. While microsphere-supplemented scaffolds have been investigated, a pure microsphere scaffold with an optimized architecture has been challenging to create via 3D printing methods due to issues that prevent consistent deposition of microsphere-based materials and their ability to maintain the shape of the 3D-printed structure. Utilizing the extrusion printing process, we established a methodology that not only allows the creation of large microsphere scaffolds but also multicomposite matrices into which cells, growth factors, and therapeutics encapsulated in microspheres can be directly deposited during the printing process. Our 3D-McMap method provides some critical guidelines for issues with scaffold shape fidelity during and after printing. Carefully timed breaks, minuscule drying steps, and adjustments to extrusion parameters generated an evenly layered large microsphere scaffold that retained its internal architecture. Such scaffolds are superior to other microsphere-containing scaffolds, as they can release biomolecules in a highly controlled spatiotemporal manner. This capability permits us to study cell responses to the delivered signals to develop scaffolds that precisely modulate new tissue formation.

Characterization and in-vitro assessment of silicon-based apatite microspheres for bone tissue engineering applications.

In the field of bone tissue engineering, silicon (Si) has been found as an essential element for bone growth. However, the use of silicon in bioceramics microspheres remains limited. In this work, different weight percentages (0.8, 1.6, and 2.4 wt %) of silicon was incorporated into hydroxyapatite and fabricated into microspheres. 2.4 wt % of Si incorporated into HAp microspheres (2.4 SiHAp) were found to enhance functional properties of the microspheres which resulted in improved cell viability of human mesenchymal stem cells (hMSCs), demonstrating rapid cell proliferation rates resulting in high cell density accumulated on the surface of the microspheres which in turn permitted better hMSCs differentiation into osteoblasts when validated by bone marker assays (Type I collagen, alkaline phosphatase, osteocalcin, and osteopontin) compared to apatite microspheres of lower wt % of Si incorporated and non-substituted HAp (2.4 SiHAp >1.6 SiHAp >0.8 SiHAp > HAp). SEM images displayed the densest cell population on 2.4 SiHAp surfaces with the greatest degree of cell stretching and bridging between neighboring microspheres. Incorporation of silicon into apatite microspheres was found to accelerate the rate and number of apatite nucleation sites formed when subjected to physiological conditions improving the interface between the microsphere scaffolds and bone forming cells, facilitating better adhesion and proliferation.

Injectable and Self-Adaptive Gel Scaffold Based on Heparin Microspheres for Adipogenesis of Human Adipose-Derived Stem Cells.

An injectable and self-adaptive heparin microsphere-based cell scaffold was developed to achieve adipose regeneration. Simultaneously, the cell scaffold exhibited a dynamic architecture, self-regulated glucose levels, sustained insulin delivery, and steady viscoelastic properties for adipogenesis. The dynamic cell scaffold is cross-linked by the boronate-diol interaction among heparin-based microspheres, which have boronate and maltose groups. Because of the boronate-maltose ester bonds, the gelatinous complex would be partially dismantled and readily display glucose-sensitive performance by free glucose via competitive displacement. The dynamic cross-linking heparin microsphere scaffold can deliver the lipogenic drug insulin to enhance lipid filling, which has an impact on fat tissue enhancement. A 4-week in vitro cell culture demonstrated that the dynamic heparin microsphere-based cell scaffold, through loading with insulin, showed significantly higher efficiency in promoting ASC differentiation compared with traditional 3D culture methods. In vivo histological results further demonstrated that there was a significant increase in adipose in the proposed cell scaffold, which proved to be statistically significant compared with traditional biomaterials. Notable stain expression of the FABP4 and PPAR-γ genes was also observed in the dynamic cell scaffold containing insulin, which was more similar to natural fat.

Optimizing fabrication parameters via Taguchi method for production of high yield hydroxyapatite microsphere scaffolds using Drop-on-Demand inkjet method.

Drop on demand (DOD) inkjet method is a cost-efficient way of producing hydroxyapatite (HAp) microsphere scaffolds with narrow size distribution. However, DOD fabrication parameters may influence the yield and characteristics of the microsphere scaffolds. Testing different permutations and combinations of fabrication parameters is costly and time consuming. Taguchi method could be used as a predictive tool for optimizing the key fabrication parameters to produce HAp microspheres with desired yield and properties, minimizing the number of experimental combinations to be tested. The aim of this study is to investigate the influence of the fabrication parameters on the characteristics of the microspheres formed and determine optimum parameter conditions for producing high yield HAp microsphere scaffolds with the desired properties intended to serve as potential bone substitutes. We aimed to achieve microspheres with high production yield, microsphere size of <230 μm, micropore sizes <1 μm, rough surface morphology and high sphericity. Experiments were conducted using Taguchi method with a L9 orthogonal array at three levels per parameter to determine optimum parameter values for (1) operating pressure, (2) shutter speed duration, (3) nozzle height and (4) CaCl2 concentration. Based on signal-to-noise (S/N) ratio analysis, the identified optimum parameter conditions for operating pressure, shutter speed duration, nozzle height and CaCl2 concentration to be 0.9-1.3 bar, 100 ms, 8 cm and 0.4 M respectively. The microspheres obtained had an average size of 213 μm, 0.45 μm micropore size, high sphericity index of 0.95 and high production yield of 98%. Confirmation tests and ANOVA results affirms the validity of Taguchi method in optimizing HAp microspheres with high yield, desired size, micropore size and shape. HAp microsphere scaffolds produced by optimum conditions were subjected to a 7-day in-vitro study. Cells remained viable and continued to proliferate (increased 1.2-fold) over 7 days with microspheres maintaining high cell density with cells bridging between microspheres. Alkaline phosphatase (ALP) assay increased 1.5-fold from day 1, suggesting good osteogenic potency of HAp microspheres as potential bone substitutes.

Elastomeric Porous Poly(glycerol sebacate) Methacrylate (PGSm) Microspheres as 3D Scaffolds for Chondrocyte Culture and Cartilage Tissue Engineering.

Cartilage defects can be difficult to treat; therefore, tissue engineering of cartilage is emerging as a promising potential therapy. One interesting area of research explores the delivery of cells to the cartilage defect via scaffold-based cell delivery vehicles and microsurgery. This study explores the use of novel poly(glycerol sebacate) methacrylate (PGSm)-polymerised high internal phase emulsion (polyHIPE) microspheres as scaffolds with embedded cells for cartilage tissue engineering. Porous microsphere scaffolds (100 µm-1 mm diameter) were produced from emulsions consisting of water and a methacrylate-based photocurable resin of poly(glycerol sebacate). These resins were used in conjunction with a T-junction fluidic device and an ultraviolet (UV) curing lamp to produce porous microspheres with a tuneable size. This technique produced biodegradable PGSm microspheres with similar mechanical properties to cartilage. We further explore these microspheres as scaffolds for three-dimensional culture of chondrocytes. The microspheres proved to be very efficient scaffolds for primary chondrocyte culture and were covered by a dense extracellular matrix (ECM) network during the culture period, creating a tissue disk. The presence of glycosaminoglycans (GAGs) and collagen-II was confirmed, highlighting the utility of the PGSm microspheres as a delivery vehicle for chondrocytes. A number of imaging techniques were utilised to analyse the tissue disk and develop methodologies to characterise the resultant tissue. This study highlights the utility of porous PGSm microspheres for cartilage tissue engineering.

Regenerative engineering of long bones using the small molecule forskolin

Bone grafting procedures have become increasingly common in the United States, with approximately 500,000 cases occurring each year at a societal cost exceeding $2.4 billion. Recombinant human bone morphogenetic proteins (rhBMPs) are therapeutic agents that have been widely used by orthopedic surgeons to stimulate bone tissue formation alone and when paired with biomaterials. However, significant limitations such as immunogenicity, high production cost, and ectopic bone growth from these therapies remain. Therefore, efforts have been made to discover and repurpose osteoinductive small-molecule therapeutics to promote bone regeneration. Previously, we have demonstrated that a single-dose treatment with the small-molecule forskolin for just 24 h induces osteogenic differentiation of rabbit bone marrow-derived stem cells invitro, while mitigating adverse side effects attributed with prolonged small-molecule treatment schemes. In this study, we engineered a composite fibrin-PLGA [poly(lactide-co-glycolide)]-sintered microsphere scaffold for the localized, short-term delivery of the osteoinductive small molecule, forskolin. Invitro characterization studies showed that forskolin released out of the fibrin gel within the first 24 h and retained its bioactivity toward osteogenic differentiation of bone marrow-derived stem cells. The forskolin-loaded fibrin-PLGA scaffold was also able to guide bone formation in a 3-mo rabbit radial critical-sized defect model comparable to recombinant human bone morphogenetic protein-2 (rhBMP-2) treatment, as demonstrated through histological and mechanical evaluation, with minimal systemic off-target side effects. Together, these results demonstrate the successful application of an innovative small-molecule treatment approach within long bone critical-sized defects.

Magnetic Microsphere Scaffold-Based Soft Microbots for Targeted Mesenchymal Stem Cell Delivery.

A soft microbot assembled from individual magnetic microsphere scaffold (MMS) beads carrying mesenchymal stem cells (MSC) is navigated under magnetic actuation, where an oscillating field induces mechanical flexion to propel the microbot toward the target site. A seven-bead microbot attained a top translational speed of 205.6µms-1 (0.068 body lengths-1 ) under 10mT and 2Hz field oscillation. The shallow flexion angle (10-24.5°) allows precision movements required to navigate narrow spaces. Upon arrival at the target site, the MMS beads unload their MSC cargo following exposure to a phosphate-buffered saline (PBS) solution, mimicking the extracellular fluid's sodium concentration. The released stem cells have excellent viability and vitality, promoting rapid healing (i.e., 83.2% vs 49%) in a scratch-wound assay. When paired with minimally invasive surgical methods, such as laparoscopy and endoscopic surgery, the microbot can provide precise stem cell delivery to hard-to-reach injury sites in the body to promote healing. Moreover, the microbot is designed to be highly versatile, with individual MMS beads customizable for cargoes of live cells, biomolecules, bionanomaterials, and pharmaceutical compounds for various therapeutic requirements.

Enhanced Osteogenic Properties of Bone Repair Scaffolds through Synergistic Effects of Mechanical and Biochemical Stimulation

Herein, the osteogenic properties of bone repair scaffolds are improved by combining mechanical and biochemical stimulations. In the bone microenvironment, fluid shear stress (FSS) is the primary mechanical stimulation. The FSS of the cells is closely related to the scaffold structure. Scaffolds with the same wall thickness (400 μm) and different pore sizes (700, 800, and 900 μm) are established. The FSS of the cells on the scaffold surface is analyzed using the two‐way fluid–structure interaction (FSI) method to obtain the ideal structure. The 400–900 polylactic acid (PLA) scaffold with optimal mechanical stimulation is fabricated using selective laser sintering (SLS). BMP‐2/PLGA microspheres are loaded onto the 400–900 scaffold to further enhance the osteogenic properties through the synergistic effect of biochemical stimulations. The results indicate that the FSS of cells decreases with an increase in the pore size, and the pore size of 900 μm yields the best osteogenic differentiation. At 8 weeks after implantation, the 400–900 microspheres scaffold have the highest ratio of bone volume to total volume (BV/TV) and a uniform distribution of new bone. The results show that BMP‐2/PLGA microspheres can accelerate the formation of new bone.

3D Gelatin Microsphere Scaffolds Promote Functional Recovery after Spinal Cord Hemisection in Rats.

Spinal cord injury (SCI) damages signal connections and conductions, with the result that neuronal circuits are disrupted leading to neural dysfunctions. Such injuries represent a serious and relatively common central nervous system condition and current treatments have limited success in the reconstruction of nerve connections in injured areas, especially where sizeable gaps are present. Biomaterial scaffolds have become an effective alternative to nerve transplantation in filling these gaps and provide the foundation for simulating the 3D structure of solid organs. However, there remain some limitations with the application of 3D bioprinting for preparation of biomaterial scaffolds. Here, the approach in constructing and testing mini-tissue building blocks and self-assembly, solid 3D gelatin microsphere (GM) scaffolds with multiple voids as based on the convenient preparation of gelatin microspheres by microfluidic devices is described. These 3D GM scaffolds demonstrate suitable biocompatibility, biodegradation, porosity, low preparation costs, and relative ease of production. Moreover, 3D GM scaffolds can effectively bridge injury gaps, establish nerve connections and signal transductions, mitigate inflammatory microenvironments, and reduce glial scar formation. Accordingly, these 3D GM scaffolds can serve as a novel and effective bridging method to promote nerve regeneration and reconstruction and thus recovery of nerve function after SCI.

Construction and osteogenic effects of 3D-printed porous titanium alloy loaded with VEGF/BMP-2 shell-core microspheres in a sustained-release system.

The repair and reconstruction of bone defects remain a challenge in orthopedics. The present study offers a solution to this problem by developing a vascular endothelial growth factor (VEGF)/bone morphogenetic protein 2 (BMP-2) shell-core microspheres loaded on 3D-printed porous titanium alloy via gelatin coating to prepare a titanium-alloy microsphere scaffold release system. The composite scaffold was characterized via scanning electron microscope (SEM) and energy disperse spectroscopy (EDS), and the effect of the composite scaffold on the adhesion, proliferation, and differentiation of osteoblasts were determined in vitro. Furthermore, a rabbit femoral defect model was established to verify the effect of the composite scaffold on osteogenesis and bone formation in vivo. The results demonstrated that the composite scaffold could release VEGF and BMP-2 sequentially. Meanwhile, the composite scaffold significantly promoted osteoblast adhesion, proliferation, and differentiation (p < 0.05) compared to pure titanium alloy scaffolds in vitro. Furthermore, the composite scaffold can exhibit significant osteogenic differentiation (p < 0.05) than gelatin-coated titanium alloy scaffolds. The in vivo X-rays demonstrated that the implanted scaffolds were in a good position, without inflammation and infection. Micro-CT and quantitative results of new bone growth illustrated that the amount of new bone in the composite scaffold is significantly higher than that of the gelatin-coated and pure titanium alloy scaffolds (p < 0.05). Similarly, the fluorescence labeling and V-G staining of hard tissue sections indicated that the bone integration capacity of the composite scaffold was significantly higher than the other two groups (p < 0.05). This research suggests that VEGF/BMP-2 shell-core microspheres loaded on 3D-printed titanium alloy porous scaffold through gelatin hydrogel coating achieved the sequential release of VEGF and BMP-2. Most importantly, the in vitro and in vivo study findings have proven that the system could effectively promote osteogenic differentiation and osseointegration.

Microspheres and their Potential in Endodontic Regeneration Application.

Microspheres have been widely utilised as versatile carriers in biomedical applications. In recent years, as a new type of injectable scaffold, microspheres have attracted increasing attention in the field of regenerative medicine owing to their various advantages including their small size, large specific surface area and mimicry of the 3D native environment. These characteristics enable them to adopt the narrow and irregular anatomy of the tooth and become an ideal scaffold for endodontic regeneration. Microspheres play an important role in carrying biologics (cells, biomolecules and drugs), which effectively regulate the fate of stem cells and control the release of growth factors and drugs. Cell-laden microspheres, which can be divided into microcarriers and microcapsules, have great application prospects in dental pulp regeneration. This paper summarises the properties and characteristics of microsphere scaffolds used in tissue engineering, placing emphasis on their advantages and applications in endodontic regeneration.

Investigating the degradation behaviour and characteristic changes of phase pure hydroxyapatite (HAp) microsphere scaffolds under static and dynamic conditions

ABSTRACT Hydroxyapatite (HAp) has been widely researched on due to its chemical similarity to native bone. HAp microspheres can encourage good cellular attachment due to their high surface area to volume ratio, facilitating further proliferation and differentiation of cells at later stages. To ensure the reliability and effectiveness of HAp microspheres in physiological conditions, we investigated their behaviour and key characteristic changes in both static and dynamic conditions. We found both environments exhibiting similar results but noticeably the microspheres in the dynamic environment resulted in faster rate of degradation. The degradation mechanism follows that of a surface degradation process. Microspheres underwent reduction in size and weight loss upon release of Ca2+ ions into Dulbecco’s Phosphate Buffered Saline medium (DPBS). Apatite reprecipitation was visible in the form of granular deposits in both environments from day 1 and continued throughout 21 days which altered the size of the microspheres but not its shape.

Integrated polycaprolactone microsphere-based scaffolds with biomimetic hierarchy and tunable vascularization for osteochondral repair

Osteochondral lesion potentially causes a variety of joint degenerative diseases if it cannot be treated effectively and timely. Microfracture as the conservative surgical choice achieves limited results for the larger defect whereas cartilage patches trigger integrated instability and cartilage fibrosis. To tackle aforementioned issues, here we explore to fabricate an integrated osteochondral scaffold for synergetic regeneration of cartilage and subchondral bone in one system. On the macro level, we fabricated three integrated scaffolds with distinct channel patterns of Non-channel, Consecutive-channel and Inconsecutive-channel via Selective Laser Sintering (SLS). On the micro level, both cartilage zone and subchondral bone zone of integrated scaffold were made of small polycaprolactone (PCL) microspheres and large PCL microspheres, respectively. Our findings showed that Inconsecutive-channel scaffolds possessed integrated hierarchical structure, adaptable compression strength, gradient interconnected porosity. Cartilage zone presented a dense phase for the inhibition of vessel invasion while subchondral bone zone generated a porous phase for the ingrowth of bone and vessel. Both cartilage regeneration and subchondral bone remodeling in the group of Inconsecutive-channel scaffolds have been demonstrated by histological evaluation and immunofluorescence staining in vivo. Consequently, our current work not only achieves an effective and regenerative microsphere scaffold for osteochondral reconstruction, but also provides a feasible methodology to recover injured joint through integrated design with diverse hierarchy. Statement of significanceRecovery of osteochondral lesion highly depends on hierarchical architecture and tunable vascularization in distinct zones. We therefore design a special integrated osteochondral scaffold with inconsecutive channel structure and vascularized modulation. The channel pattern impacts on mechanical strength and the infiltration of bone marrow, and eventually triggers synergetic repair of osteochondral defect. The cartilage zone of integrated scaffolds consisted of small PCL microspheres forms a dense phase for physical restriction of vascularized infiltration whereas the subchondral bone zone made of large PCL microspheres generates porous trabecula-like structure for promoting vascularization. Consequently, the current work indicates both mechanical adaptation and regional vascularized modulation play a pivotal role on osteochondral repair.

Honeycomb-Like Hydrogel Microspheres for 3D Bulk Construction of Tumor Models.

A two-dimensional (2D) cell culture-based model is widely applied to study tumorigenic mechanisms and drug screening. However, it cannot authentically simulate the three-dimensional (3D) microenvironment of solid tumors and provide reliable and predictable data in response to in vivo, thus leading to the research illusions and failure of drug screening. In this study, honeycomb-like gelatin methacryloyl (GelMA) hydrogel microspheres are developed by synchronous photocrosslinking microfluidic technique to construct a 3D model of osteosarcoma. The in vitro study shows that osteosarcoma cells (K7M2) cultured in 3D GelMA microspheres have stronger tumorous stemness, proliferation and migration abilities, more osteoclastogenetic ability, and resistance to chemotherapeutic drugs (DOX) than that of cells in 2D cultures. More importantly, the 3D-cultured K7M2 cells show more tumorigenicity in immunologically sound mice, characterized by shorter tumorigenesis time, larger tumor volume, severe bone destruction, and higher mortality. In conclusion, honeycomb-like porous microsphere scaffolds are constructed with uniform structure by microfluidic technology to massively produce tumor cells with original phenotypes. Those microspheres could recapitulate the physiology microenvironment of tumors, maintain cell-cell and cell-extracellular matrix interactions, and thus provide an effective and convenient strategy for tumor pathogenesis and drug screening research.

Silk fibroin/chitosan/TGF-β1-loaded microsphere scaffolds for cartilage reparation

Transforming growth factor-β1 (TGF-β1) plays an important role in chondrocyte growth and the synthesis of extracellular matrix (ECM). Due to the rapid metabolism, controlled release systems for TGF-β1 have attracted increasing interest recently. In this study, a silk fibroin (SF)/chitosan (CS) scaffold incorporated with TGF-β1-loaded microspheres (MSs) was created for cartilage reparation. The optimal proportion of the SF/CS composite scaffold was determined by evaluating their micromorphology and the proliferation rate of fibroblasts on the surface. Then, SF/CS/TGF-β1-loaded MS scaffolds were prepared by the adsorption method. TGF-β1 release capacity, degradation patterns, cytocompatibility and in vivo implantation were evaluted. The SF/CS/TGF-β1-loaded MS scaffold showed good TGF-β1 release over more than 16 days, which could sequentially stimulate chondrocyte synthetic activity. In vitro cell proliferation experiments showed the SF/CS/TGF-β1-loaded MS scaffold could promote chondrocytes adhesion, growth, proliferation and maintained the cellular morphology. An in vivo study demonstrated that a low inflammatory response was observed in rats and that the materials exhibited good biocompatibility. the results indicated that our SF/CS/TGF-β1-loaded MS scaffold constitute a promising therapeutic option for cartilage reparation.

Integrated Osteochondral Microsphere Scaffolds with Biomimetic Architecture and Regional Vascularization for Joint Repair

Osteochondral lesion potentially causes a variety of joint degenerative diseases if it cannot be treated effectively and timely. Microfracture as the first line surgical choice achieves limited results for the larger defect whereas cartilage patches trigger integrated instability and cartilage fibrosis. To tackle aforementioned issues, here we explore to construct an integrated osteochondral scaffold for synergetic regeneration of cartilage and subchondral bone in one system. On the macro level, we fabricated three integrated scaffolds with distinct channel structure of Non-channel, Consecutive-channel and Inconsecutive-channel via Selective Laser Sintering (SLS). On the micro level, cartilage zone and subchondral bone zone of integrated scaffold consisted of small polycaprolactone (PCL) microspheres with the diameter of 50 μm and large PCL microspheres with the diameter of 150 μm, respectively. As a result, cartilage zone exhibited a dense phase for the inhibition of vessel invasion while subchondral zone generated a porous phase for the ingrowth of bone and vessel. Our findings showed that Inconsecutive-channel scaffolds possessed biomimetic hierarchical structure, adequate mechanical properties, gradient interconnected porosity in comparison with Non-channel and Consecutive-channel groups. Moreover, Inconsecutive-channel scaffolds obviously induced cartilage regeneration and subchondral bone remodeling. Consequently, this work not only achieves an effective and regeneratable microsphere scaffold for osteochondral reconstruction, but also provides a feasible methodology to recover injured joint through integrated design pattern. Funding Information: This work was financially supported by the National Key Research and Development Program of China (2017YFC1103900), the National Natural Science Foundation of China (32171323, 91939111), Key Research and Development Program of Hubei Province (2020BCB056), International Science and Technology Cooperation Projects of Shenzhen (GJHZ20200731095211035), Sanming Project of Medicine in Shenzhen (SZSM201812055), and the Fundamental Research Funds for the Central Universities (2019kfyRCPY103). Declaration of Interests: The authors declare no conflict of interest. Ethics Approval Statement: All procedures for animal experiments were approved by the Institutional Animal Care and Use Committee of Huazhong University of Science and Technology (SYXK20170094).

Novel microsphere-packing synthesis, microstructure, formation mechanism and in vitro biocompatibility of porous gelatin/hydroxyapatite microsphere scaffolds

In this study, three types of porous gelatin/HAp composite-based scaffolds were prepared and their microstructure, mechanical property, and in vitro biocompatibility were evaluated. FD-gelatin/HAp scaffolds were prepared via a conventional freeze-drying (FD) method, while DCPD-bound gelatin/HAp (DCPD-gelatin/HAp) microsphere scaffolds and HAp-bound gelatin/HAp (HAp-gelatin/HAp) microsphere scaffolds were prepared via the combined method of emulsion technique and novel microsphere-packing method. It was found that the microstructure and mechanical property of porous gelatin/HAp scaffolds were highly dependent on the preparation method and type of scaffolds. FD-gelatin/HAp scaffolds had few pores with a comparatively flat surface and had a compressive strength was 24 ± 1 MPa. DCPD-gelatin/HAp microsphere scaffolds had a porous structure with the average pore size of 121 ± 12 μm and had the compressive strength of 50 ± 3 MPa. HAp-gelatin/HAp microsphere scaffolds also had a porous structure with the average pore size of 190 ± 26 μm and had the compressive strength of 17 ± 4 MPa. In vitro biocompatibility was performed by culturing the osteoblast-like MC3T3-E1 cells with all types of scaffolds. The results showed that all types of scaffolds were biocompatible to support cell attachment and proliferation. However, a significant difference in cell behavior was clearly observed. Most cells were grown on the surface of FD-gelatin/HAp scaffolds due to the lack of the porous structure, while those were grown on the surface and inside the pore of both DCPD-gelatin/HAp microsphere scaffolds and HAp-gelatin/HAp microsphere scaffolds due to the presence of the porous structure. Thus, the porous gelatin/HAp microsphere-based scaffolds have a potential as novel bone tissue regenerative materials.