Fabrication of 3D scaffolds based on fully biobased unsaturated polyester resins by microstereo-lithography

Additive Manufacturing (AM) technologies are an effective route to fabricate tailor made scaffolds for tissue engineering (TE) and regenerative medicine with microstereo-lithography (µSLA) being one of the most promising techniques to produce high quality 3D structures. Here, we report the crosslinking studies of fully biobased unsaturated polyesters (UPs) with 2-hydroxyethyl methacrylate (HEMA) as the unsaturated monomer (UM), using thermal and µSLA crosslinking processes. The resulting resins were fully characterized in terms of their thermal and mechanical properties. Determination of gel content, water contact angle (WCA), topography and morphology analysis by atomic force microscopy (AFM) and scanning electron microscopy (SEM) were also performed. The results show that the developed unsaturated polyester resins (UPRs) have promising properties for µSLA. In vitro cytotoxicity assays performed with 3T3-L1 cell lines showed that the untreated scaffolds exhibited a maximum cellular viability around 60 %, which was attributed to the acidic nature of the UPRs. The treatment of the UPRs and scaffolds with ethanol (EtOH) improved the cellular viability to 100%. The data presented in this manuscript contribute to improve the performance of biobased unsaturated polyesters in additive manufacturing.

Biomimetic porous scaffolds containing decellularized small intestinal submucosa and Sr2+/Fe3+ co-doped hydroxyapatite accelerate angiogenesis/osteogenesis for bone regeneration

The design of bone scaffolds is predominately aimed to well reproduce the natural bony environment by imitating the architecture/composition of host bone. Such biomimetic biomaterials are gaining increasing attention and acknowledged quite promising for bone tissue engineering. Herein, novel biomimetic bone scaffolds containing decellularized small intestinal submucosa matrix (SIS-ECM) and Sr2+/Fe3+ co-doped hydroxyapatite (SrFeHA) are fabricated for the first time by the sophisticated self-assembled mineralization procedure, followed by cross-linking and lyophilization post-treatments. The results indicate the constructed SIS/SrFeHA scaffolds are characterized by highly porous structures, rough microsurface and improved mechanical strength, as well as efficient releasing of bioactive Sr2+/Fe3+ and ECM components. These favorable physico-chemical properties endow SIS/SrFeHA scaffolds with an architectural/componential biomimetic bony environment which appears to be highly beneficial for inducing angiogenesis/osteogenesis both in vitro and in vivo. In particular, the cellular functionality and bioactivity of endotheliocytes/osteoblasts are significantly enhanced by SIS/SrFeHA scaffolds, and the cranial defects model further verifies the potent ability of SIS/SrFeHA to accelerate in vivo vascularization and bone regeneration following implantation. In this view these results highlight the considerable angiogenesis/osteogenesis potential of biomimetic porous SIS/SrFeHA scaffolds for inducing bone regeneration and thus may afford a new promising alternative for bone tissue engineering.

Effects of La2O3 contents on microstructure and properties of laser-cladded 5wt%CaB6/HA bioceramic coating

In the present work, 5wt%CaB6/HA bioceramic coatings with different La2O3contents (0-0.6wt%) have been fabricated by a laser cladding technique on Ti-6Al-4V. The effects of La2O3 contents on microstructure and properties of the laser-cladded 5wt%CaB6/HA coatings have been carefully investigated. The results show that the microstructure is obviously refined, and the structure is relatively uniform after doping 0.2-0.4 wt% La2O3. As the La2O3 content increases, the corrosion resistance are found to increase firstly and then gradually decrease. The XRD analysis confirms that the amount of HA and TCP in the coating reaches maximum after doping 0.2wt% La2O3. The La2O3-doped coatings show a significantly higher bone-like apatite precipitation after immersion in SBF compared with La2O3-free coating. In vitro experiment also shows that 5wt%CaB6/HA bioceramic coatings with 0.2-0.4wt% La2O3 are more suitable for the attachment and proliferation of MG63 cells, exhibiting superior bioactivity and biocompatibility.

Preparation and antibacterial properties of an AgBr@SiO2/GelMA composite hydrogel

Pure gelatin hydrogels lack antibacterial function and have poor mechanical properties, which restrict their application in wound dressings. In this study, nanosized silver bromide-doped mesoporous silica (AgBr@SiO2) microspheres with hollow structures were prepared by a modified Stober method. The novel microspheres can not only release silver ions to treat bacteria but also release drugs to treat skin wound. Furthermore, AgBr@SiO2 microspheres were modified with propyl methacrylate, incorporated into methacrylated gelatin (GelMA), and crosslinked by UV light to prepare AgBr@SiO2/GelMA dressings consisting of composite hydrogels. The results showed that the AgBr@SiO2 microspheres could enhance the mechanical properties of the hydrogels. With the increase in the AgBr@SiO2 concentration from 0.5 to 1 mg/mL, the dressings demonstrated effective antimicrobial activity against both Staphylococcus aureus and Escherichia coli. Furthermore, full-thickness skin wounds in vivo wound healing studies with Sprague–Dawley rats were evaluated. When treated with AgBr@SiO2/GelMA containing 1 mg/mL AgBr@SiO2, only 15% of the wound area left on day 10. Histology results also showed the epidermal and dermal layers were better organized. These results suggest that AgBr@SiO2/GelMA-based dressing materials could be promising candidates for wound dressings.

Smart delivery of lethal poly-peptide composite for effective cancer therapy

In the past decade, multifunctional peptides have attracted increasing attention in the biomedical field. Peptides possess many impressive advantages, such as high penetration ability, low cost, and etc. However, the short half-life and instability of peptides limit their application. In this study, a poly-peptide drug loading system (called HKMA composite) was designed based on the different functionalities of four peptides. The peptide compositions of HKMA composite from N-terminal to C-terminal were HCBP1, KLA, MMP-2-cleavable peptide and ABD. The targeting and lethality of HKMA to NSCLC cell line H460 sphere cells and the half-life of the system were measured in vivo. The results showed that the HKMA composite had a long half-life and specific killing effect on H460 sphere cells in vitro and in vivo. Our result proposed smart peptide drug loading system and provided a potential methodology for effective cancer treatment.

Macroporous methacrylated hyaluronic acid hydrogel with different pore sizes for in vitro and in vivo evaluation of vascularization

Vascularization of thick hydrogel scaffolds is still a big challenge, because the submicron- or nano-sized pores seriously restrict endothelial cells adhesion, proliferation and migration. Therefore, porous hydrogels have been fabricated as a kind of promising hydrous scaffolds for enhancing vascularization during tissue repairing. In order to investigate the effects of pore size on vascularization, macroporous methacrylated hyaluronic acid (HAMA) hydrogels with different pore sizes were fabricated by a gelatin microspheres (GMS) template method. After leaching out GMS templates, uniform and highly interconnected macropores were formed in hydrogels, which provided an ideal physical microenvironment to induce human umbilical vein endothelial cells (HUVECs) migration and tissue vascularization. In vitro results revealed that macroporous hydrogels facilitated cells proliferation and migration compared with non-macroporous hydrogels. Hydrogels with middle pore size of 200-250 μm (HAMA250 hydrogels) supported the best cell proliferation and furthest 3D migration of HUVECs. The influences of pore sizes on vascularization were then evaluated with subcutaneous embedding. In vivo results illustrated that HAMA250 hydrogels exhibited optimum vascularization behavior. Highest number of newly formed blood vessels and expression of CD31 could be found in HAMA250 hydrogels rather than in other hydrogels. In summary, our results concluded that the best pore size for endothelial cells migration and tissue vascularization was 200-250 μm. This research provides a new insight into the engineering vascularized tissues and may find utility in designing regenerative biomaterial scaffolds

Effect of passage number of genetically modified TGF-β3 expressing primary chondrocytes on the chondrogenesis of ATDC5 cells in a 3D coculture system

Due to the lack of blood vessels, nerves and lymphatics, articular cartilage is difficult to repair once damaged. Tissue engineering is considered to be a potential strategy for cartilage regeneration. Successful tissue engineering strategies depend on the effective combination of biomaterials, seed cells and biological factors. In our previous study, a genetically modified coculture system with chondrocytes and ATDC5 cells in an alginate hydrogel has exhibited a superior ability to enhance chondrogenesis. In this study, we further evaluated the influence of chondrocytes at various passages on chondrogenesis in the coculture system. The results demonstrated that transfection efficiency was hardly influenced by the passage of chondrocytes. The coculture system with passage 5 (P5) chondrocytes had a better effect on chondrogenesis of ATDC 5 cells, while chondrocytes in this coculture system presented higher levels of dedifferentiation than other groups with P1 or P3 chondrocytes. Therefore, P5 chondrocytes were shown to be more suitable for the coculture system, as they accumulated in sufficient cell numbers with more passages and had a higher level of dedifferentiation, which was prone to form a favorable niche for chondrogenesis of ATDC5 cells. This study may provide fresh insights for future cartilage tissue engineering strategies with a combination of a coculture system and advanced biomaterials.

Investigation of insulin resistance through a multiorgan microfluidic organ-on-chip

The development of hepatic insulin resistance (IR) is a critical factor in developing type 2 diabetes (T2D), where insulin fails to inhibit hepatic glucose production but retains its capacity to promote hepatic lipogenesis. Improving insulin sensitivity can be effective in preventing and treating T2D. However, selective control of glucose and lipid synthesis has been difficult. It is known that excess white adipose tissue is detrimental to insulin sensitivity, whereas brown adipose tissue transplantation can restore it in diabetic mice. However, challenges remain in our understanding of liver-adipose communication because the confounding effects of hypothalamic regulation of metabolic function cannot be ruled out in previous studies. There is a lack of in vitro models that use primary cells to study cellular-crosstalk under insulin resistant conditions. Building upon our previous work on the microfluidic primary liver and adipose organ-on-chips, we report for the first time the development of integrated insulin resistant liver-adipose (white and brown) organ-on-chip. The design of the microfluidic device was carried out using computational fluid dynamics; the experimental studies were conducted by carrying out detailed biochemical analysis RNA-seq analysis on both cell types. Further, we tested the hypothesis that brown adipocytes regulated both hepatic insulin sensitivity and lipogenesis. Our results show effective co-modulation of hepatic glucose and lipid synthesis through a platform for identifying potential therapeutics for IR and diabetes.

Cobalt-doped Ti surface promotes immunomodulation

Here, cobalt-doped plasma electrolytic oxidation (PEO) coatings with different cobalt contents were prepared on Ti implants. The cobalt ions in the PEO coating exhibited a slow and sustainable release and thus showed excellent biocompatibility and enhanced cell adhesion. In vitro ELISA and RT-PCR assays demonstrated that the cobalt-loaded Ti showed immunomodulatory functions to macrophages and upregulated the expression of anti-inflammatory (M1 type) genes and downregulated expression levels of pro-inflammatory (M2 type) genes compared with that of pure Ti sample. High cobalt content induced increased macrophage polarization into the M2 type. Furthermore, the findings from the in vivo air pouch model suggested that cobalt-loaded Ti could mitigate inflammatory reactions. The present work provides a novel strategy to exploit the immunomodulatory functions of implant materials.

Optimized, visible light-induced crosslinkable hybrid gelatin/hyaluronic acid scaffold promotes complete spinal cord injury repair

Severe microenvironmental changes after spinal cord injury (SCI) present serious challenges in neural regeneration and tissue repair. Gelatin (GL)- and hyaluronic acid (HA)-based hydrogels are attractive scaffolds because they are major components of the extracellular matrix and can provide a favorable adjustable microenvironment for neurogenesis and motor function recovery. In this study, three-dimensional hybrid GL/HA hydrogel scaffolds were prepared and optimized. The hybrid hydrogels could undergo in-situ gelation and fit the defects perfectly via visible light- induced crosslinking in the complete SCI rats. We found that the transplantation of the hybrid hydrogel scaffold significantly reduced the inflammatory responses and suppressed glial scar formation in an HA concentration-dependent manner. Moreover, the hybrid hydrogel with GL/HA ratios less than 8/2 effectively promoted endogenous neural stem cell migration and neurogenesis, as well as improved neuron maturation and axonal regeneration. The results showed locomotor function improved 60 days after transplantation, thus suggesting that GL/HA hydrogels can be considered as a promising scaffold for complete SCI repair.