| Diabetic foot ulcers(DFU)are one of the most common complications of diabetes,difficult to heal,prone to infection,and severe deterioration leading to amputation or death,which is a common clinical problem.Reactive oxygen species(ROS)and bacterial infectioninduced inflammatory responses are the two most prominent detrimental conditions that impede angiogenesis and wound healing.Based on the drug treatment of 4-octyl itaconate(4OI),different hydrogel preparation techniques were used to design different hydrogel delivery systems by analyzing the characteristics of the material and the characteristics of the disease,so as to achieve multi-dimensional promotion of wound repair such as antibacterial,anti-inflammatory,antioxidant and pro-vascular reconstruction.The design of the entire project is as follows.The main research content is divided into two parts.(1)Injectable and self-healing hydrogel delivering 4OI with antibacterial,antioxidant,and anti-inflammatory activity to promote diabetic wound repairObjective: In order to meet the disease and wound shape characteristics of diabetic wounds,this study constructed the injectable hydrogel through the dynamic self-healing of Ag-S bond,which meet the delivery functions of 4OI drugs and played an antioxidant,antiinflammatory,antibacterial and angiogenesis effects.Methods:1.The dynamic self-healing injectable hydrogel of Ag-S bond was constructed by the mixed reaction of 4-PEG-SH and Ag NO3.The hydrogel was characterized by scanning electron microscope and rheometer,and its drug release curve was explored.2.The antibacterial properties of hydrogels were detected by agar plate experiment,SYTO 9/PI and bacterial scanning electron microscope.3.The biocompatibility of the hydrogels was detected by live-dead staining and CCK8,and the angiogenesis ability of the hydrogels under oxidative stress was detected by tubeforming experiments and WB.4.ROS probe,apoptosis detection,mitochondrial membrane potential,WB and mitochondrial detection kits were used to explore the antioxidant capacity and mitochondrial function changes of hydrogels under oxidative stress.5.Diabetic rat wound model,HE staining,Masson staining,immunofluorescence,RT-PCR and laser speckle imaging were used to explore the pro-vascular formation function of hydrogels in vivo and evaluate the wound repair effect of hydrogels.Result:1.The hydrogel synthesized in this experiment has a porous structure observed by SEM.The injectability and self-healing properties of the hydrogel were verified from multiple angles by rheometer and syringe testing.The drug release relationship between Ag+ and 4OI was determined by drug release curve.2.The agar plate experiment showed that the antibacterial performance of the hydrogel was greater than 70%.SEM observation found that the PEG hydrogel could make the bacteria collapse and die,and the live-dead staining also obtained the same result.3.The good biocompatibility of PEG hydrogels was verified by CCK8 and Live-Dead staining.Tube formation experiments,mitochondrial-related assays and WB showed that 4OI@PEG could promote the recovery of mitochondrial morphology and function in cells under oxidative stress through activating the antioxidant system Keap1-Nrf2.4.A rat diabetic wound model was successfully constructed.Laser speckle imaging showed that 4OI@PEG could promote the recovery of vascular blood flow in diabetic wounds.The 14-day observation showed that the 4OI@PEG group had fastest wound healing rate and the largest area.Conclusion: An injectable and self-healing hydrogel was successfully constructed,which reversed the oxidative stress of diabetic wounds,inhibited local inflammation,promoted vascular reconstruction,epithelial regeneration,and wound healing in diabetic rats.(2)Black phosphorus based and 4OI loaded hydrogel with NIR responsive antibacterial,antioxidant,and anti-inflammatory activity for infected diabetic wound treatmentObjective: Eliminating bacterial infections and improving the oxidative stress state of endothelial cells are strategies to promote diabetic wound repair.Therefore,it is necessary to develop a multifunctional hydrogel with antioxidant,anti-inflammatory and antibacterial properties.Black phosphorus(BP)has a unique structure and a large body surface with photothermal and photodynamic abilities.Through the surface modification of BP,a hydrogel delivery platform of 4OI was constructed to promote the repair of diabetic wounds.Methods:1.Modifying with PEG-SH,the surface charge of BP was changed to load 4OI,and then an injectable light-responsive BP-Gel MA hydrogel was constructed.2.The BP,BP-PEG and 4OI-BP@Gel were characterized by TEM,AFM,FITR,Zeta potential meter,Raman,etc.,and the release relationship of 4OI was determined by the drug release curve.3.The photothermal properties and photostability properties of 4OI-BP@Gel under NIR irradiation were observed by thermal imager,and the photodynamic properties of 4OIBP@Gel were detected by ESR and DPBF.4.The antibacterial properties of hydrogels were detected by agar plate experiment,SYTO 9/PI and scanning electron microscope.5.ROS probe,apoptosis detection,mitochondrial membrane potential,WB and mitochondrial detection kits were used to explore the antioxidant capacity and mitochondrial function changes of hydrogels under oxidative stress.6.A diabetic infection wounds rat model,HE staining,Masson staining,Giemsa staining,immunofluorescence and laser speckle imaging were performed to explore the angiogenesis and antibacterial abilities of hydrogels in vivo and to evaluate the wound repair effect of hydrogels.Results:1.The successful combination of PEG-SH and BP NSs was obtained by TEM,AFM,energy spectrum detection,and FITR detection.Zeta potential found that the negative potential of BP-PEG tends to be neutral,and the curve of BP FITR is affected by the amplitude of PEG.SEM shows that 4OI-BP@Gel has a distinct pore structure.The release curve confirmed that the BP-containing hydrogel had a sustained release effect on 4OI.2.Biocompatibility testing of the hydrogel showed that it has no cytotoxicity,and 4OIBP@Gel can promote angiogenesis by tube formation assay and WB detection.ROS probes and mitochondrial membrane potential and morphology assays showed that it has the ability to resist oxidation damage and promote the recovery of mitochondrial function.3.Infrared monitoring showed that 4OI-BP@Gel could raise the temperature to more than 50℃ in vitro and in vivo under the irradiation of NIR(808nm,1W cm-2,5 min),while maintaining stability.ESR and DPBF show that it has obvious photodynamic effect.4.Antibacterial experiments show that 4OI-BP@Gel has a significant antibacterial effect under the stimulation of NIR in vivo and vitro,and the antibacterial efficacy is greater than 90%.The results of an infectious diabetic wound model in rats showed that the 4OIBP@Gel could promote angiogenesis,increase the blood supply for wound repair,reduce the level of inflammation in the wound and promote wound repair rapidly.Conclusion: We successfully developed a biocompatible and biodegradable BPincorporated Gel MA hydrogel for diabetic wound healing.Surface modification by PEG–NH2 rendered the BP–PEG NSs less easily degraded and endowed the hydrogel with photostable properties.Owing to its photothermal and photodynamic properties,the hydrogel had sufficient antibacterial ability to kill bacteria.At the same time,4OI-BP@Gel can release 4OI slowly with anti-oxidation,anti-inflammatoryand angiogenesis ability and promote the rapid repair of diabetic infected wounds.4OI–BP@Gel has the above advantages and has become a promising dressing for wound healing in diabetic infections.Therefore,the gradual treatment of diabetic wound infections provides a reference for the design of diabetic wound dressings. |
