The 3D-printing Fabrication And Property Study Of Biomimetic Anti-inflammatory Hydrogel

Skin provides a major barrier to protect human body.Skin damage caused by burns,wounds and severe ulcers will lead to the loss of tissue fluid in patients and infection if it is not properly treated,which could be life-threatening in some serious cases.Skin wound healing includes four stages,i.e.hemostasis,inflammatory response,proliferation and remodeling.Inflammatory response and proliferation are the key stages that can be intervened to promote wound healing.Spermidine has been shown to have a variety of beneficial biological functions,such as anti-rejection,anti-oxidation,inhibition of inflammation,inhibition of cell necrosis,prolongation of individual life,etc.,but there are no relevant reports on the use of spermidine to inhibit the inflammatory response in the process of skin wound repair.3D-printing is a technology based on the layered manufacturing of rapid prototyping,which can accurately deposit biological materials and cells into specific locations and hence construct tissue-like substitutes with three-dimensional bionic structure and function.In this study,a bionic anti-inflammatory hydrogel has been fabricated by 3D-bioprinting using CTS/GEL as the matrix biomaterial and SPD crosslinker as the crosslinking agent,and the physicochemical properties,biological activities and anti-inflammatory activity of hydrogel have been characterized.With the consideration of the physical properties and forming mechanism of the biomaterials used,as well as the characteristics and advantages of various existing 3Dprinting equipments,a micro-valve 3D-printing platform has independently designed and built this research.Subsequently,a bionic anti-inflammatory CTS/GEL hydrogel(SPD-CTS/GEL hydrogel)has been prepared with natural biopolymer chitosan(CTS)and gelatin(GEL)as the matrix biomaterials and a crosslinking agent(SPD crosslinker)based on a bioactive polyamine,spermidine(SPD),through a method combining 3Dprinting with spraying.The microstructure,physicochemical properties and biological activities of the SPD-CTS/GEL hydrogel have been characterized.First,the key processing parameters,such as the setting for 3D-printing,the formula of CTS/GEL bio-ink and the concentration of SPD crosslinker,have been optimized.Then a series of hydrogels have been fabricated with different ratios of CTS/GEL and concentrations of SPD crosslinker by 3D-printing.The micropore size,swelling ratio,mechanical properties and in vitro degradability of hydrogels have been studied to understand the effects of CTS/GEL ratio and SPD crosslinker concentration on the microstructure and various physicochemical properties of the 3D-printed hydrogels.It is found that the 3D-printed SPD-CTS/GEL hydrogels have interconnected micropores,controllable mechanical properties,tunable swelling ratio and adjustable degradability.The biological activities of the 3D-printed SPD-CTS/GEL hydrogels have been assessed in vitro by Live/Dead staining,CCK-8 cell proliferation assay and scratch test on NIH/3T3 mouse fibroblasts.The 3D-printed SPD-CTS/GEL hydrogels have demonstrated excellent cytocompatibility,enhanced cell proliferation and improved cell migration,indicating that the obtained hydrogel has the ability to promote wound repair.The anti-inflammatory function of the SPD-CTS/GEL hydrogel has been evaluated in an inflammation model based on lipopolysaccharide(LPS)-stimulated Raw264.7 cells(mouse macrophage cell line).The SPD-CTS/GEL hydrogel has shown a significant reduction of secreted proinflammatory cytokines,such as TNF-α and IL-6,and an evident decrease of nitric oxide(NO)level in cell culture compared to the control group.In summary,a bionic anti-inflammatory hydrogel with both microscopic porous structure and tunable macroscopic mechanical strength has been fabricated by 3Dbioprinting using CTS/GEL as the matrix biomaterial and SPD crosslinker as the crosslinking agent.The obtained SPD-CTS/GEL hydrogel has controllable degradability,excellent cytocompatibility and desirable anti-inflammatory activity.It can not only promote the fibroblast proliferation and migration,but also inhibit the expression of relevant inflammatory factors by macrophages.It has the potential to reduce the inflammatory response in skin grafting and wound healing,holding a good promise for clinical application of skin wound repair.