| Stem cell transplantation has been widely used in tissue repair and its research.However,due to the difference between pathological microenvironment and pHysiological environment,how to improve the microenvironment of transplantation area to make it more conducive to the growth and differentiation of stem cells has been widely followed in the development of stem cell products and basic research.Irrational diets or hemorrhagic diseases can bring about tissue iron overload,which can lead to oxidative damage of tissues.Transplantation of stem cells in iron overload areas can also affect the growth and differentiation of stem cells.Based on the iron overload environment after intracerebral hemorrhage,a core-shell designed keratin nerve scaffold is constructed as a carrier for stem cell transplantation.The shell is composed of low molecular weight MH-loaded keratose hydrogel(KG).The purpose of the shell is to chelate iron ions overloaded in brain tissue to improve brain environment.Besides,the low molecular weight keratin also has the characteristics of fast degradation.The core uses high molecular weight keratin as a carrier,and encapsulates bone marrow mesenchymal stem cells and PLGA nanoparticles loaded with growth factors,Furthermore,the degradation of high molecular weight keratin is slow,which can support the growth and differentiation of stem cells in the iron overload region.Core-shell designed keratin nerve scaffold can provide a new strategy for stem cell therapy after intracerebral hemorrhage,and also offer experimental basis for stem cell therapy in the iron overload area.The main research contents and corresponding conclusions are as follows:(1)TGA was used as a reducing agent to extract keratin from human hair.Ultrafiltration plate membrane equipment was used to classify the molecular weight of keratin.SDS-PAGE showed that high molecular weight keratins(> 50 kDa)and low molecular weight keratins(30-50 kDa)were obtained.The core scaffold needed a long time to support the growth and differentiation of stem cells.Through the rheological property experiment,the core scaffold was composed of high molecular weight keratin,with a concentration of 35%;and the shell scaffold was composed of low molecular weight keratin,and its concentration was 45% determined by the iron ion adsorption experiment.The above two concentrations of keratin scaffold were observed by SEM,and the results showed that they both had high porosity.Degradation experiments showed that the two scaffolds had different degradation rates in vitro.Core-shell designed nerve scaffolds were prepared by stratified injection in vitro and observed.The results showed that the bilayer structure was complete and uniform.At the same time,PLGA nanoparticles loaded with growth factor EGF/b-FGF were prepared by W1/O/W2.Drug release experiments in vitro showed that PLGA nanoparticles had the effect of slowly releasing the growth factor.(2)After the construction of core-shell keratin nerve scaffolds,their preliminary biological properties were evaluated at the cellular level.Bone mesenchymal stem cells were extracted by the whole bone marrow method.The stem cell toxicity of the core-shell keratin nerve scaffolds was studied.The protective effect of MH-loaded KG on the growth and differentiation of BMSCs under iron overload environment was investigated.Iron overload model was established with the heme concentration of 25 ?M,and the MH-loaded KG was added to cell culture medium during the growth process.The results showed that the survival rate of stem cells increased with the increase of the concentration of MH-loaded KG.When the concentration of MH-loaded KG reached 400 mg/mL,the survival rate of stem cells increased to 80.65±2.36%(P<0.01).During the induction and differentiation of stem cells,an iron overload model was also established,and the MH-loaded KG was used to protect it.Immunofluorescence staining showed that the MH-loaded KG had a strong antagonistic effect on iron overload toxicity.(3)The primary biological properties of core-shell keratin nerve scaffolds were further evaluated at the animal model level.An iron overload model was established by injecting FeCl2 into the brain of SD rats.At the same time,core-shell keratin nerve scaffolds and core keratin nerve scaffolds were transplanted.Perls' staining showed that core-shell nerve scaffolds could effectively remove overloaded iron ions from brain tissues.Brain water content showed that the implantation of core-shell scaffolds could alleviate brain edema.Nissl staining showed that core-shell scaffolds could reduce brain atropHy.Forelimb-placing and corner turn experiment showed that the core-shell scaffolds transplantation group had a good effect on the recovery of nerve function.Brdu-NeuN immunofluorescence showed that the growth and differentiation of stem cells transplanted under the core-shell scaffolds were better than those of the core scaffolds.Therrfore,the transplantation of core-shell keratin nerve scaffolds had a good effect in repairing brain tissues. |
PDF Full Text Request