Effects of Unilateral Intrastriatal Dopaminergic Grafts in MPTP-Treated Hemiparkinsonian Monkeys

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Nicolass Bohnen, M.D., Ph.D.

Department of Neurology
University of Pittsburgh School of Medicine

Abstract

There are approximately 2 million patients with Parkinson’s disease (PD) in the United States. Although many can be treated for a period of time with pharmacotherapy (e.g., L-DOPA), these treatments are usually not effective in all patients or for all symptoms and even when effective they usually fail after several years. Thus, a more complete and permanent restoration of function is an extremely important objective. Cell implantation is a very promising approach to brain repair, which has the potential to lead to the cure of this disabling neurodegenerative disorder.

Recent advances in stem cell technology have generated new optimism that PD can be cured in the relative near future. Previous cell transplantation studies in PD have revealed variable results. There is a clear need to improve pre-transplant cell preparation and also expand our understanding of the factors associated with post-transplantation cell viability.

One of the most promising strategies is the use of neurotrophic factor in vitro treatments of cellular grafts, pre-implantation. Previous studies have shown that telencephalic tissues from aborted fetal tissue are a reliable and abundant source of many types of cells. In vitro, the fetal brain cultures differentiate into all major types of human brain cells. Studies in nonhuman primates are needed to answer basic questions such as whether immunization is needed, the nature of pretreatment with neurotrophic factors, the amount of cells implanted, the best brain target for controlling clinical symptoms, and other factors affecting survival of cells.

The MPTP-treated nonhuman primate is the most accurate model of PD in terms of neuochemical anatomy and response to symptomatic and restorative treatment. Unilateral carotid arterial infusion of MPTP has proven to be a good model of subhuman primate hemiparkinsonism (affecting only one side of the body).

Dr. Bohnen will exploit his extensive experience with human fetal brain cells in vitro and in vivo to study the benefit of using fetal tissues in restoring the dopaminergic pathways in a primate model of PD. His approach will focus on in vitro treatments leading to pre-grafting enrichment in dopaminergic neurons by using neurotrophic factors. In addition to the traditional autopsy studies, Dr. Bohnen and his associates will assess in vivo the function of the human cell grafts by using PET imaging.

Progress Report (as of 8/2002)

Specific aims. To perform in vivo PET assessment of striatal dopamine transporter (DAT) binding using [C-11]CFT in MPTP-treated hemi-parkinsonian monkeys before and two months after growth factor pretreated stem cell grafting

Hypothesis. (1) MPTP-treated hemi-parkinsonian monkeys will have ipsilateral striatal reduction of DAT binding; (2) Ipsilateral striatal stem cell grafting in MPTP-treated hemi-parkinsonian monkeys will lead to improved motor symptoms and increased DAT binding when compared to pre-transplant DAT symptoms and binding.

Methods. Unilateral infusion of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in a dose of 0.4 mg/kg will be given into the left carotid artery. The monkeys will be evaluated within 2-3 days and then weekly up to 1 months for the development of hemiparkinsonism. Prior to grafting, animals will be imaged by PET using the [C-11]-CFT ligand to assess the loss of dopamine re-uptake following MPTP injection. For transplantation, we will use one week in vitro old brain cell aggregates that will be implanted unilaterally, on the lesion side in the putamen of two MPTP monkeys (Macaca mulatta) at one month post injection of the chemical. One animal will be implanted with BDNF pretreated cell agreggates while the other one will be grafted with untreated cell aggregates. PET imaging will be repeated 2 months after grafting. The left and right hemispheres will be sampled separately. ROI analysis on the parametric images will be performed on the following striatal (sub-) regions: whole striatum, caudate nucleus and putamen. Pre-transplant images will be analyzed by comparing the ipsilateral (MPTP-treated) striatum to the contralateral striatum. Pre- and post-transplant images will be analyzed using non-parametric repeated measures testing per ipsilateral brain region. In addition, post-transplant striatal ipsilateral (transplanted) and contralateral (control) striatal BP values will be also compared. Pre- and post-transplant DAT binding will be compared to clinical motor behavior before and after transplantation. Dr. Achim will perform post-transplantation autopsy studies of graft cytoarchitecture, DA neuronal survival and differentiation.

Preliminary findings.
We have imaged two monkeys using the [C-11]-CFT DAT tracer pre-MPTP lesion and at 6 weeks post-MPTP lesion. Repeat PET studies demonstrates prominent striatal tracer activity before lesion and an approximately 40-44% decrease in tracer binding after a single unilateral intracarotid MPTP injection. Comparing 2 week and 6 week post-lesion PET images, we see no regeneration of the DA system which would be revealed by an increase in {C-11}CFT DAT binding. The 2 week and 6 week images are identical (data not shown). Our collaborator, Dr. Douglas Kondziolka, implanted a human cell graft in the anterior putamen of the first animal, without immunosuppression, and there were no post surgery complications or clinical changes. A repeat PET study did not show evidence for viability of the graft. We are now in the process of exploring technical factors to improve the surgical technique.

Significance. Restoration of function is critical for people who now have PD. Cell implantation is a very promising approach to brain repair, which has the potential to lead to the cure of this disabling neurodegenerative disorder.

Pilot data for new NIH grant applications. Yes. We have used the preliminary MPTP-lesioning PET imaging findings as part of two NIH grant submissions as of 6/1/02. We hope that these preliminary data obtained by this seed grant will help us to secure funding to extend our studies.

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