Timed Release of Cerebrolysin Using Drug-Loaded Titanate Nanospheres Reduces Brain Pathology and Improves Behavioral Functions in Parkinson’ s Disease Asya Ozkizilcik 1 & Aruna Sharma 2,3,6 & Dafin F. Muresanu 4,5 & José V. Lafuente 6,7,8 & Z. Ryan Tian 9 & Ranjana Patnaik 10 & Herbert Mössler 5,11 & Hari S. Sharma 2,3,6 # Springer Science+Business Media, LLC 2017 Abstract Previous studies from our laboratory show that intraperitoneal injections of 1-metyl-4-phenyl-1,2,3,6- tetrahydropyridin (MPTP, 20 mg/kg) daily within 2-h intervals for 5 days in mice induce Parkinson’ s disease (PD)-like symptoms on the 8th day. A significant de- crease in dopamine (DA) and its metabolites 3,4- dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) along with a marked decrease in the num- ber of tyrosine hydroxylase (TH)-positive cells in the substantia nigra pars compacta (SNpc) and striatum (STr) confirms the validity of this model for studying PD. Since cerebrolysin (CBL) is a well-balanced com- position of several neurotrophic factors and active pep- tide fragments, in the present investigation we examined the timed release of CBL using titanate nanospheres (TiNS) in treating PD in our mouse model. Our obser- vations show that TiNS-CBL (in a dose of 3 ml/kg, i.v.) given after 2 days of MPTP administration for 5 days resulted in a marked increase in TH-positive cells in the SNpc and STr as compared to normal CBL. Also, TiNS-CBL resulted in significantly higher levels of DA, DOPAC, and HVA in SNpc and STr on the 8th day as compared to normal CBL therapy. TiNS-CBL also thwarted increased α-synuclein levels in the brain and in the cerebrospinal fluid (CSF) as well as neuronal nitric oxide synthase (nNOS) in the in PD brain as compared to untreated group. Behavioral function was also significantly improved in MPTP-treated animals that received TiNS-CBL. These observations are the first to demonstrate that timed release of TiNS-CBL has far more superior neuroprotective effects in PD than normal CBL. Keywords 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridin (MPTP) . Parkinson’ s disease (PD) . Titanate nanospheres (TiNS) . Cerebrolysin . Neuroprotection . Alpha-synuclein . Neuronal nitoic oxide synthase (nNOS) . Cerebrospinal fluid (CSF) * Hari S. Sharma Sharma@surgsci.uu.se 1 Department of Biomedical Engineering, University of Arkansas, Fayetteville, AR, USA 2 Department of Surgical Sciences, Anesthesiology & Intensive Care Medicine, Uppsala University Hospital, Uppsala University, SE-75185 Uppsala, Sweden 3 International Experimental CNS Injury & Repair (IECNSIR), University Hospital, Uppsala University, Frödingsgatan 12, Bldg. 28, SE-75421 Uppsala, Sweden 4 Department of Clinical Neurosciences, University of Medicine & Pharmacy, Cluj-Napoca, Romania 5 “RoNeuro” Institute for Neurological Research and Diagnostic, 37 Mircea Eliade Street, 400364 Cluj-Napoca, Romania 6 LaNCE, Department of Neuroscience, University of the Basque Country (UPV/EHU), Leioa, Biscay, Spain 7 Nanoneurosurgery Group, BioCruces Health Research Institute, 48903 Barakaldo, Biscay, Spain 8 Faculty of Health Science, Universidad Autónoma de Chile, Santiago de Chile, Chile 9 Chemistry & Biochemistry, University of Arkansas, Fayetteville, AR, USA 10 School of Biomedical Engineering, Department of Biomaterials, Indian Institute of Technology, Banaras Hindu University, Varanasi, India 11 EVER Neuro Pharma, Oberburgau, Austria Mol Neurobiol DOI 10.1007/s12035-017-0747-4