Injectable Hydrogel Dressing With Thermosensitivity And Bacterial Resistance For Diabetic Foot Ulcers Treatment

Diabetic foot ulcer(DFU)is one of the common diabetic complications.Compared with ordinary wounds,once DFU is formed,it is extremely difficult to cure,will results in infection,gangrene,amputation,and even death,brings great pain to patients.In the face of this complex chronic wound,the general wound dressing is difficult to fully adapt to the location,depth and shape of the wound.In addition changing the wound dressing is extremely hard,which will cause secondary injury to the wound and impede the wound healing.For this reason,a thermosensitive antibacterial injectable hydrogel dressing(insulin@PLGA,Ag)/ CPN was constructed for the treatment of diabetic foot ulcers.It has the following advantages:1.It can adapt irregularly shaped wound through it’s injectability;2.It can be transformed into Gel quickly at body temperature,and when the wound dressing need to be changed,it can be easily removed because it’s thermosensitivity.3.The loaded insulin@PLGA and AgNPs endow the hydrogel with insulin sustained release and antibacterial properties,which is beneficial to wound healing.The detailed studies are as follows:(1)Synthesis and characterization of injectable thermosensitive wound dressing CPN.Poly(N-isopropylacrylamide)-carboxylic acid(PNIPAM-COOH)was synthesized by spontaneous polymerization of N-isopropylacrylamide(NIPAM)in the presence of initiator AIBN and chain transfer agent MAA.Increasing the molar ratio of MAA to NIPAM can reduce the number-average molecular weight of PNIPAM-COOH.When the molar ratio of NIPAM,AIBN and MAA is 100/1/10,the molecular weight of PNIPAM-COOH1 is the largest,which is 1.9×104.The relationship between temperature and viscosity of PNIPAM-COOH with different molecular weight was measured by rotary rheometer.The results show that the higher the molecular weight of PNIPAM-COOH is,the lower the LCST is.The LCST of PNIPAM-COOH1,with the highest molecular weight is 31.60 ℃.PNIPAM-COOH1 was grafted onto chitosan(CS)by the reaction of carboxyl and amino groups to form CPN hydrogel.The test results of rotary rheometer showed that the dosage of CS did not affect the LCST,of CPN hydrogel,but the CPN12.93 hydrogel with high CS content had higher viscosity and was more suitable to be used as wound dressing.By comparing the CPN12.93 hydrogels with different mass fractions,it was found that the gel time of CPN12.93 hydrogels with 15% mass fraction was 18 s,suitable for being wound dressing.SEM test shows that CPN12.93 has interconnected pore structure.Temperature sensitivity test showed that CPN12.93 hydrogel could be converted repeatedly from solution(25 ℃)to gel(37 ℃),and its structure and properties were stable.(2)Preparation and study of insulin@PLGA MicrospheresInsulin-loaded insulin@PLGA microspheres were prepared by oil / water / oil emulsification method.The pore size of PLGA microspheres without insulin loading was 63.99 nm measured by BET,which meant it can be loaded with insulin.SEM results showed that the average diameter of insulin@PLGA microspheres was 39 μ m.The insulin loading of insulin@PLGA microspheres measured by BCA and TG was7.8%,and the entrapment efficiency was 87.4%.The in vitro release curve showed that the insulin release of insulin@PLGA microspheres in the first three days was82.1%,which could easily meet the needs of insulin concentration.(3)Preparation and properties of(insulin@PLGA,Ag)/ CPN hydrogel.AgNPs,was synthesized by non-in-situ synthesis and compounded with insulin@PLGA in CPN12.93 hydrogel to prepare(insulin@PLGA,Ag)/ CPN hydrogel.EDS test showed that the content of Ag element in hydrogel was 1.994%.The rotational rheometer test shows that the LCST of(insulin@PLGA,Ag)/ CPN is31.60 ℃ the same as CPN12.93,and its viscosity and viscoelasticity are better than CPN12.93,and it can also change many times between(25 ℃)and gel(37 ℃).It has good injectability and temperature sensitivity,and its structure and properties are stable.The antibacterial activity of(insulin@PLGA,Ag)/ CPN was tested by Escherichia coli and Staphylococcus aureus.It was found that(insulin@PLGA,Ag)/CPN had stronger antibacterial activity than CPN12.93,and the antibacterial ability against Staphylococcus aureus was stronger than that of Escherichia coli.The culture results of mouse L929 fibroblasts showed that the cell survival rate of(insulin@PLGA,Ag)/ CPN hydrogel reached more than 90% after being cultured for 24 h and72 h,so it has good biocompatibility.