title: Non-invasive, neuron-specific gene therapy by focused ultrasound-induced blood-brain barrier opening in Parkinson's disease mouse model.
abstract: Focused ultrasound (FUS)-induced with microbubbles (MBs) is a promising technique for noninvasive opening of the blood-brain barrier (BBB) to allow targeted delivery of therapeutic substances into the brain and thus the noninvasive delivery of gene vectors for CNS treatment. We have previously demonstrated that a separated gene-carrying liposome and MBs administration plus FUS exposure can deliver genes into the brain, with the successful expression of the reporter gene and glial cell line-derived neurotrophic factor (GDNF) gene. In this study, we further modify the delivery system by conjugating gene-carrying liposomes with MBs to improve the GDNF gene-delivery efficiency, and to verify the possibility of using this system to perform treatment in the 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced animal disease model. FUS-BBB opening was verified by contrast-enhanced MRI, and GFP gene expression was verified via in vivo imaging system (IVIS). Western blots as well as enzyme-linked immunosorbent assay (ELISA) were conducted to measure protein expression, and immunohistochemistry (IHC) was conducted to test the Tyrosine hydroxylase (TH)-neuron distribution. Dopamine (DA) and its metabolites as well as dopamine active transporter (DAT) were quantitatively analyzed to show dopaminergic neuronal dopamine secretion/activity/metabolism. Motor performance was evaluated by rotarod test weekly. Results demonstrated that the LpDNA-MBs (gene-liposome-MBs) complexes successfully serve as gene carrier and BBB-opening catalyst, and outperformed the separated LpDNA/MBs administration both in terms of gene delivery and expression. TH-positive IHC and measurement of DA and its metabolites DOPAC and HVA confirmed improved neuronal function, and the proposed system also provided the best neuroprotective effect to retard the progression of motor-related behavioral abnormalities. Immunoblotting and histological staining further confirmed the expression of reporter genes in neuronal cells. This study suggests that FUS exposures with the administration of LpDNA-MBs complexes synergistically can serve as an effective gene therapy strategy for MPTP-animal treatment, and may have potential for further application to perform gene therapy for neurodegenerative disease.
authors.affiliation: Medical Imaging Research Center, Institute for Radiological Research, Chang Gung University/Chang Gung Memorial Hospital, Taoyuan 333, Taiwan; Department of Nephrology, Division of Clinical Toxicology, Chang Gung Memorial Hospital, Lin-Kou Medical Center, Taoyuan 333, Taiwan. Electronic address: winwood5782@gmail.com.
authors.forename: Chung-Yin
authors.initials: CY
authors.lastname: Lin
authors.affiliation: Department of Electrical Engineering, Chang Gung University, Taoyuan 333, Taiwan.
authors.forename: Han-Yi
authors.initials: HY
authors.lastname: Hsieh
authors.affiliation: Department of Neurology, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan.
authors.forename: Chiung-Mei
authors.initials: CM
authors.lastname: Chen
authors.affiliation: Institute of Oral Medicine, National Cheng Kung University, Tainan 701, Taiwan.
authors.forename: Shang-Rung
authors.initials: SR
authors.lastname: Wu
authors.affiliation: Department of Electrical Engineering, Chang Gung University, Taoyuan 333, Taiwan.
authors.forename: Chih-Hung
authors.initials: CH
authors.lastname: Tsai
authors.affiliation: Department of Neurosurgery, Chang Gung Memorial Hospital, Linkou Medical Center and College of Medicine, Chang Gung University, Taoyuan 333, Taiwan.
authors.forename: Chiung-Yin
authors.initials: CY
authors.lastname: Huang
authors.affiliation: Department of Chemical and Material Sciences, Chang Gung University, Taoyuan 333, Taiwan.
authors.forename: Mu-Yi
authors.initials: MY
authors.lastname: Hua
authors.affiliation: Department of Neurosurgery, Chang Gung Memorial Hospital, Linkou Medical Center and College of Medicine, Chang Gung University, Taoyuan 333, Taiwan.
authors.forename: Kuo-Chen
authors.initials: KC
authors.lastname: Wei
authors.affiliation: Department of Biomedical Engineering and Environmental Sciences, National Tsing Hua University, Hsinchu 300, Taiwan.
authors.forename: Chih-Kuang
authors.initials: CK
authors.lastname: Yeh
authors.affiliation: Medical Imaging Research Center, Institute for Radiological Research, Chang Gung University/Chang Gung Memorial Hospital, Taoyuan 333, Taiwan; Department of Electrical Engineering, Chang Gung University, Taoyuan 333, Taiwan. Electronic address: haoliliu@mail.cgu.edu.tw.
authors.forename: Hao-Li
authors.initials: HL
authors.lastname: Liu
mesh_terms: D015632:1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; D000818:Animals; D001921:Brain / Q000378:metabolism; D004247:DNA / Q000008:administration & dosage*; D004195:Disease Models, Animal; D004298:Dopamine / Q000378:metabolism; D050483:Dopamine Plasma Membrane Transport Proteins / Q000378:metabolism; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D051100:Glial Cell Line-Derived Neurotrophic Factor / Q000235:genetics* / Q000378:metabolism; D008081:Liposomes; D008297:Male; D008807:Mice, Inbred BALB C; D045423:Microbubbles; D009474:Neurons / Q000378:metabolism; D009498:Neurotoxins; D010302:Parkinson Disease, Secondary / Q000139:chemically induced / Q000235:genetics / Q000378:metabolism / Q000628:therapy*; D010957:Plasmids; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism; D000069453:Ultrasonic Waves
chemical_list: D050483:Dopamine Plasma Membrane Transport Proteins; C495040:Gdnf protein, mouse; D051100:Glial Cell Line-Derived Neurotrophic Factor; D008081:Liposomes; D009498:Neurotoxins; D004247:DNA; D015632:1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; D014446:Tyrosine 3-Monooxygenase; D004298:Dopamine
keywords: Blood-brain barrier opening; Focused ultrasound; GDNF; Gene therapy; Microbubbles