| Traumatic brain injury(TBI)leads to cognitive and emotional dysfunction of patients,which seriously harms their physical and mental health.According to statistics,mild and moderate TBI accounts for up to 90% of all TBI patients,and the number of patients suffering from TBI is expected to increase in the future.Due to the complexity of the pathogenesis and course of TBI,although some therapeutic drugs,including antiinflammatory drugs,free radical scavengers,acetylcholinesterase inhibitors,have been used to alleviate a series of injuries and concurrent symptoms caused by TBI,so far there are no specific and effective drug for the treatment of TBI.Previous studies have shown that nerve growth factor(NGF)can protect injured neurons,guide and accelerate axonal regeneration,reduce nerve inflammation,inhibit astrocyte activation and scar formation in the central nervous system(CNS).However,some factors restrict the clinical application of NGF,such as the existence of blood brain barrier(BBB),which limits the penetration of NGF protein into brain tissue,and the change of the spatial structure of NGF protein can easily lead to the decrease of its biological activity.At present,many researches are committed to delivering polypeptide and protein drugs into the brain through BBB,among which cell penetrating peptide(CPP)shows unique advantages.As a drug delivery carrier,CPP has many advantages such as simple synthesis,high tissue permeability and compatibility with other drug delivery systems.CPP generally combines drugs by direct coupling or recombinant fusion expression and delivers them into cells and tissues.However,some studies have shown that drugs can also be delivered into cells by non-covalent combination of CPP and drugs,this combination method has the advantages of simple and easy operation,and does not change the biological activity of the combined drugs.Therefore,this study aims to select CPP that can effectively promote NGF protein to penetrate the cell membrane and observe the therapeutic effect of CPP combined with NGF on moderate TBI mice.First of all,we screened CPP through literature research and synthesized peptides by solid-phase synthesis peptide synthesis.Secondly,to analyze the effect of different CPP combined with NGF on membrane penetration and select the best mixing ratio by immunocytofluorescence technique on cells in vitro,according to the different CPPs included,NGF group,NGF/Pep-1 group and NGF/Penetratin group were established respectively.Subsequently,to analyze the behavioral changes of mice in each group after administration on the established moderate TBI mouse model,two behavioral tests are used for the study of coordinative motor ability and the spatial memory function: rotarod performance test and morris water maze test,respectively.Immunohistochemical staining was used to observe the positive content of nerve cells and reactive astrocytes around the focus of brain injury in mice.The results of immunocytofluorescence assay show that the content of NGF protein in cells of NGF/Penetratin group was significantly higher than that of other groups,and the fluorescence intensity is the highest at a molar ratio of 20:1between Penetratin and NGF.The results of animal experiments show that the coordination motor function,spatial memory ability and the expression of Neu N protein in brain tissue of mice in NGF/Penetratin group were significantly higher than those in TBI group and NGF group,the expression level of GFAP protein in brain tissue was significantly lower than that in TBI group and NGF group.The results of this experiment pointed out that Penetratin can promote NGF to the damaged brain tissue through the blood-brain barrier(BBB),and has a good therapeutic effect on moderate TBI mice. |
PDF Full Text Request