Microenvironment Regulation And Biomarker Detection Of Neural Cell Of Parkinson’s Disease

Parkinson’s disease(PD)is mainly due to dopaminergic neurons losing in the substantia nigra,Pathology features of PD include intracellular oxidative stress,dysfunction of mitochondrial and abnormal neurotransmission release,etc.These influence the state and function of neurons.More than 4 million people all over the world are suffering from PD.Therefore,diagnosis and treatment of PD has become a high profile issue.Here,we focus on the pathological state of cells.Firstly,we designed a new fluorescent probe to detect the nitroreductase in hypoxia cells.Secondly,we designed and synthesized a library of GSH responsive lipids to deliver protein into cells and control protein release.Thirdly,we used CRISPR/Cas9 technology to edit genes in neuro cells and recorded the neurotransmitters release by electrochemical method.The main studies are summarized as following:1)Detecting and imaging nitroreductase in neurons with a new fluorescent probe.Nitroreductase is one of the significant biomarkers in hypoxia cells.Smallmolecule fluorescent probes are commonly utilized for nitroreductase imaging and detection in living systems due to high sensitivity,temporal-spatial resolution.Herein,a ratiometric fluorescent probe for nitroreductase was synthesized,using sulfonyl-benzoxadiazole(SBD)as a fluorophore and p-nitrobenzyl carbamate group as a recognition domain for nitroreducatse.This probe’s reaction with nitroreducase causes red shift of emission wavelength.Experiment results indicate that this probe possesses great sensitivity,selectivity and biocompatibility for bioimaging nitroreducatse in living cells.2)Targeting delivery of protein using GSH responsive cationic lipid nanoparticles in neural cell.One of the toughest challenges in development of protein therapeutics is the efficient and targeting delivery of protein into mammalian cells.There reported more and more studies about bio-reducible lipid nanoparticles as a vector of protein.Here,we designed and synthesized a library of GSH responsive cationic lipids.Saporin,a model protein,was delivered into cells by these lipids.In addition,we demonstrate GSH responsiveness of the lipids and its feasibility for therapeutics protein intracellular delivery.3)Investigation of neural cell gene editing based on the CRISPR/Cas9 technology.The synaptic vesicle plays a significant role in the process of neuronal communication.A deep insight into the character and release of neurotransmission is of great importance in diagnosing PD.Therefore,we utilize CRISPR/Cas9 technology targeting exogenous genes to edit neural cell and neural system in vivo.Moreover,signal of the single cell neurotransmitter was measured and recorded by patch clamp.We are able to efficiently induce gene mutant in PD model cells.The systematic work established here can act as a good-performance tool in mammalian systems for research applications and therapeutics.