Biocompatibility and Immunotoxicology of the Preclinical Implantation of a Collagen-based Artificial Dermal Regeneration Matrix

Objective Graft rejection, with the possibility of a violent immune response, may be severe and life threatening. Our aim was to thoroughly investigate the biocompatibility and immunotoxicology of collagen-based dermal matrix(DM) before assessment in clinical trials. Methods DM was subcutaneously implanted in BALB/c mice in two doses to induce a potential immune response. The spleen and lymph nodes were assessed for shape, cell number, cell phenotype via flow cytometry, cell activation via CCK8 kit, Annexin V kit, and Ki67 immunostaining. Serum samples were used to measure antibody concentration by enzyme-linked immunosorbent assay. Local inflammation was analyzed by histology and immunohistochemistry staining. Data analysis was performed by one-way ANOVA and non-parametric tests. Results Our data illustrate that the spleen and lymph node sizes were similar between the negative control mice and mice implanted with DM. However, in the high-dose DM(DM-H) group, the total cell populations in the spleen and lymph nodes, T cells and B cells in the spleen had slight increases in prophase, and the low-dose DM(DM-L) group did not display gross abnormities. Moreover, DM-H initiated moderate cell activation and proliferation in the early phase post-immunization, whereas DM-L did not. Neither DM-H nor DM-L implantation noticeably increased IgM and IgG serum concentrations. Examination of the local cellular response revealed only benign cell infiltration and TNF-α expression in slides of DM in the early phase. Conclusion Overall, DM-H may have induced a benign temporary acute immune response post-implantation, whereas DM-L had quite low immunogenicity. Thus, this DM can be regarded as a safe product.

In vivo evaluation of additively manufacturedmulti-layered scaffold for the repair of large osteochondral defects

The repair of osteochondral defects is one of the major clinical challenges in orthopaedics.Well-established osteochondral tissue engineering methods have shown promising results for the early treatment of small defects.However,less success has been achieved for the regeneration of large defects,which is mainly due to the mechanical environment of the joint and the heterogeneous nature of the tissue.In this study,we developed a multi-layered osteochondral scaffold to match the heterogeneous nature of osteochondral tissue by harnessing additive manufacturing technologies and combining the established art laser sintering and material extrusion techniques.The developed scaffold is based on a titanium and polylactic acid matrix-reinforced collagen“sandwich”composite system.The microstructure and mechanical properties of the scaffold were examined,and its safety and efficacy in the repair of large osteochondral defects were tested in an ovine condyle model.The 12-week in vivo evaluation period revealed extensive and significantly higher bone in-growth in the multi-layered scaffold compared with the collagen–HAp scaffold,and the achieved stable mechanical fixation provided strong support to the healing of the overlying cartilage,as demonstrated by hyaline-like cartilage formation.The histological examination showed that the regenerated cartilage in the multi-layer scaffold group was superior to that formed in the control group.Chondrogenic genes such as aggrecan and collagen-II were upregulated in the scaffold and were higher than those in the control group.The findings showed the safety and efficacy of the cell-free“translation-ready”osteochondral scaffold,which has the potential to be used in a one-step surgical procedure for the treatment of large osteochondral defects.

Carbon nanotubes-reinforced polylactic acid/hydroxyapatite porous scaffolds for bone tissue engineering

In the field of bone defect repair,critical requirements for favorable cytocompatibility and optimal mechanical properties have propelled research efforts towards the development of composite materials.In this study,carbon nanotubes/polylactic acid/hydroxyapatite(CNTs/PLA/HA)scaffolds with different contents(0.5,1,1.5 and 2 wt.%)of CNTs were prepared by the thermally induced phase separation(TIPS)method.The results revealed that the composite scaffolds had uniform pores with high porosities over 68%and high through performances.The addition of CNTs significantly enhanced the mechanical properties of resulted PLA/HA,in which the 1.5 wt.%CNTs/PLA/HA composite scaffold demonstrated the optimum mechanical behaviors with the bending elastic modulus of(868.5±12.34)MPa,the tensile elastic modulus of(209.51±12.73)MPa,and the tensile strength of(3.26±0.61)MPa.Furthermore,L929 cells on the 1.5 wt.%CNTs/PLA/HA scaffold displayed good spreading performance and favorable cytocompatibility.Therefore,it is expected that the 1.5 wt.%CNTs/PLA/HA scaffold has potential applications in bone tissue engineering.

The effect of carbon nanotubes on osteogenic functions of adipose-derived mesenchymal stem cells in vitro and bone formation in vivo compared with that of nano-hydroxyapatite and the possible mechanism

It has been well recognized that the development and use of artificial materials with high osteogenic ability is one of the most promising means to replace bone grafting that has exhibited various negative effects.The biomimetic features and unique physiochemical properties of nanomaterials play important roles in stimulating cellular functions and guiding tissue regeneration.But efficacy degree of some nanomaterials to promote specific tissue formation is still not clear.We hereby comparatively studied the osteogenic ability of our treated multiwalled carbon nanotubes(MCNTs)and the main inorganic mineral component of natural bone,nano-hydroxyapatite(nHA)in the same system,and tried to tell the related mechanism.In vitro culture of human adiposederived mesenchymal stem cells(HASCs)on the MCNTs and nHA demonstrated that although there was no significant difference in the cell adhesion amount between on the MCNTs and nHA,the cell attachment strength and proliferation on the MCNTs were better.Most importantly,the MCNTs could induce osteogenic differentiation of the HASCs better than the nHA,the possible mechanism of which was found to be that the MCNTs could activate Notch involved signaling pathways by concentrating more proteins,including specific bone-inducing ones.Moreover,the MCNTs could induce ectopic bone formation in vivo while the nHA could not,which might be because MCNTs could stimulate inducible cells in tissues to form inductive bone better than nHA by concentrating more proteins including specific bone-inducing ones secreted from M2 macrophages.Therefore,MCNTs might be more effective materials for accelerating bone formation even than nHA.

The physiological polyphosphate as a healing biomaterial for chronic wounds:Crucial roles of its antibacterial and unique metabolic energy supplying properties

Insufficient metabolic energy,in the form of adenosine triphosphate(ATP),and bacterial infections are among the main causes for the development of chronic wounds.Previously we showed that the physi-ological inorganic polymer polyphosphate(polyP)massively accelerates wound healing both in animals(diabetic mice)and,when incorporated into mats,in patients with chronic wounds.Here,we focused on a hydrogel-based gel formulation,supplemented with both soluble sodium polyP(Na-polyP)and amor-phous calcium polyP nanoparticles(Ca-polyP-NP).Exposure of human epidermal keratinocytes to the gel caused a significant increase in extracellular ATP level,an effect that was even enhanced when Na-polyP was combined with Ca-polyP-NP.Furthermore,it is shown that the added polyP in the gel is converted into a coacervate,leading to encapsulation and killing of bacteria.The data on human chronic wounds showed that the administration of hydrogel leads to the complete closure of these wounds.Histological analysis of biopsies showed an increased granulation of the wounds and an enhanced microvessel forma-tion.The results indicate that the polyP hydrogel,due to its properties to entrap bacteria and generate metabolic energy,is a very promising formulation for a new therapy for chronic wounds.

Osteochondral scaffolds for early treatment of cartilage defects in osteoarthritic joints:from bench to clinic

Osteoarthritis is a degenerative joint disease,typified by the loss in the quality of cartilage and bone at the interface of a synovial joint,resulting in pain,stiffness and reduced mobility.The current surgical treatment for advanced stages of the disease is joint replacement,where the non-surgical therapeutic options or less invasive surgical treatments are no longer effective.These are major surgical procedures which have a substantial impact on patients’quality of life and lifetime risk of requiring revision surgery.Treatments using regenerative methods such as tissue engineering methods have been established and are promising for the early treatment of cartilage degeneration in osteoarthritis joints.In this approach,3-dimensional scaffolds(with or without cells)are employed to provide support for tissue growth.However,none of the currently available tissue engineering and regenerative medicine products promotes satisfactory durable regeneration of large cartilage defects.Herein,we discuss the current regenerative treatment options for cartilage and osteochondral(cartilage and underlying subchondral bone)defects in the articulating joints.We further identify the main hurdles in osteochondral scaffold development for achieving satisfactory and durable regeneration of osteochondral tissues.The evolution of the osteochondral scaffolds–from monophasic to multiphasic constructs–is overviewed and the osteochondral scaffolds that have progressed to clinical trials are examined with respect to their clinical performances and their potential impact on the clinical practices.Development of an osteochondral scaffold which bridges the gap between small defect treatment and joint replacement is still a grand challenge.Such scaffold could be used for early treatment of cartilage and osteochondral defects at early stage of osteoarthritis and could either negate or delay the need for joint replacements.