Long-term health of dopaminergic neuron transplants in Parkinson's disease patients.

To determine the long-term health and function of transplanted dopamine neurons in Parkinson's disease (PD) patients, the expression of dopamine transporters (DATs) and mitochondrial morphology were examined in human fetal midbrain cellular transplants. DAT was robustly expressed in transplanted dopamine neuron terminals in the reinnervated host putamen and caudate for at least 14 years after transplantation. The transplanted dopamine neurons showed a healthy and nonatrophied morphology at all time points. Labeling of the mitochondrial outer membrane protein Tom20 and α-synuclein showed a typical cellular pathology in the patients' own substantia nigra, which was not observed in transplanted dopamine neurons. These results show that the vast majority of transplanted neurons remain healthy for the long term in PD patients, consistent with clinical findings that fetal dopamine neuron transplants maintain function for up to 15-18 years in patients. These findings are critically important for the rational development of stem-cell-based dopamine neuronal replacement therapies for PD.

Transplantation of GABAergic cells derived from bioreactor-expanded human neural precursor cells restores motor and cognitive behavioral deficits in a rodent model of Huntington's disease.

Huntington's disease (HD) is a neurodegenerative disorder that is characterized by progressive dementia, choreiform involuntary movements, and emotional deterioration. Neuropathological features include the progressive degeneration of striatal γ-aminobutyric acid (GABA) neurons. New therapeutic approaches, such as the transplantation of human neural precursor cells (hNPCs) to replace damaged or degenerated cells, are currently being investigated. The aim of this study was to investigate the potential for utilizing telencephalic hNPCs expanded in suspension bioreactors for cell restorative therapy in a rodent model of HD. hNPCs were expanded in a hydrodynamically controlled and homogeneous environment under serum-free conditions. In vitro analysis revealed that the bioreactor-expanded telencephalic (BET)-hNPCs could be differentiated into a highly enriched population of GABAergic neurons. Behavioral assessments of unilateral striatal quinolinic acid-lesioned rodents revealed a significant improvement in motor and memory deficits following transplantation with GABAergic cells differentiated from BET-hNPCs. Immunohistochemical analysis revealed that transplanted BET-hNPCs retained a GABAergic neuronal phenotype without aberrant transdifferentiation or tumor formation, indicating that BET-hNPCs are a safe source of cells for transplantation. This preclinical study has important implications as the transplantation of GABAergic cells derived from predifferentiated BET-hNPCs may be a safe and feasible cell replacement strategy to promote behavioral recovery in HD.

Duration of lipid peroxidation after acute spinal cord injury in rats and the effect of methylprednisolone.

OBJECT: Oxidative stress leading to lipid peroxidation is a major cause of secondary injury following spinal cord injury (SCI). The objectives of this study were to determine the duration of lipid peroxidation following acute SCI and the efficacy of short-and long-term administration of methylprednisolone on decreasing lipid peroxidation. METHODS: A total of 226 female Wistar rats underwent clip-compression induced SCI. In the first part of the study, spinal cords of untreated rats were assayed colorimetrically for malondialdehyde (MDA) to determine lipid peroxidation levels at various time points between 0 and 10 days. In the second part of the study, animals were treated with methylprednisolone for either 24 hours or 7 days. Control animals received equal volumes of normal saline. Treated and control rats were killed at various time points between 0 and 7 days. RESULTS: The MDA levels initially peaked 4 hours postinjury. By 12 hours, the MDA levels returned to baseline. A second increase was observed from 24 hours to 5 days. Both peak values differed statistically from the trough values (p < 0.008). The methylprednisolone reduced MDA levels (p < 0.04) within 12 hours of injury. No effect was seen at 24 hours or later. CONCLUSIONS: The results of this study indicate that oxidative stress persists for 5 days following SCI in rats, and although methylprednisolone reduces MDA levels within the first 12 hours, it has no effect on the second lipid peroxidation peak.

Functional improvement with low-dose dopaminergic grafts in hemiparkinsonian rats.

OBJECTIVE: The beneficial functional effects of neural transplantation in Parkinson's disease are often directly attributed to the number of surviving dopaminergic cells within a graft. However, recent clinical trials of fetal neural transplantation suggest that a high number of dopaminergic cells may induce serious side effects. In this study, we explored the ability of low-dose dopaminergic grafts to produce functional benefits in the 6-hydroxydopamine rodent model of Parkinson's disease over a long period of observation. METHODS: Twelve rats received either 50,000 or 400,000 fetal ventral mesencephalic cells implanted into the striatum. Rotational behavior was assessed after the lesion and at 3, 6, 9, and 12 weeks after transplantation. Twelve weeks after transplantation, animals were perfused, and microtome sections were stained for tyrosine hydroxylase, glial fibrillary acidic protein, heat-shock protein 27, and vimentin. RESULTS: The low-dose group had a three-fold increase in tyrosine hydroxylase-positive cell survival rate compared with the high-dose group rate. The low-dose group also had a mean cell diameter significantly higher than the high-dose group. There was no significant difference between groups in fiber density; however, a higher percentage of longer fibers was encountered in the low-dose group. The low-dose group had a lower degree of trauma in the striatum, as assessed by optical density scores from glial fibrillary acidic protein, heat-shock protein 27, and vimentin staining. There was significant improvement in rotational behavior in the high-dose group at 3 weeks after transplantation, whereas the rotational behavior normalized in the low-dose group at 6 weeks after grafting. There was no significant difference in rotational behavior scores between groups at 6 weeks after grafting. CONCLUSION: This study demonstrates that over time, a low-dose dopaminergic graft has the capability of eliciting the same functional effect as a high-dose graft. Furthermore, low-dose grafts may increase graft survival, fiber outgrowth, and dopamine production and decrease trauma to the brain.

Liposomal formulations of tacrolimus and rapamycin increase graft survival and fiber outgrowth of dopaminergic grafts.

The immunosuppressive drugs tacrolimus (TAC) and rapamycin (RAPA) have both been found to have neuroprotective effects on dopaminergic neurons. The purpose of the present study was to investigate whether liposomal formulations of these drugs administered directly into the brain improve cell survival and fiber outgrowth. Rats with unilateral 6-hydroxydopamine lesions were transplanted with 800,000 fetal rat ventral mesencephalic cells and randomly divided to one of four groups. Group 1 received a transplant containing cells only; group 2 received a cell suspension containing 0.68 microM liposomal RAPA (LRAPA); group 3 received a cell suspension containing 2.0 microM liposomal TAC (LTAC); and group 4 received a cell suspension containing a liposomal formulation of both 0.68 microM RAPA and 2.0 microM TAC (LRAPATAC). Rats were sacrificed after 6 weeks, and cell survival and fiber outgrowth were assessed using tyrosine hydroxylase (TH) immunohistochemistry. The animals receiving a cell suspension containing either LTAC or LRAPATAC were found to have significantly more surviving TH-immunoreactive (TH-ir) cells than the control group receiving cells only. The group receiving LTAC had significantly longer fibers, the group receiving LRAPA had significantly more fibers close to the graft, and the group receiving LRAPATAC had significantly more fibers at all distances. This study shows the feasibility of using liposomal formulations of neuroimmunophilins directly in the brain at the time of implantation to improve graft survival and fiber outgrowth. Furthermore, we have shown that the combination of LTAC and LRAPA has a synergistic effect. These compounds may play an important role in optimizing graft survival and host reinnervation in cell-mediated brain repair strategies for the treatment of neurological conditions.

Intraspinal transplantation of hNT neurons in the lesioned adult rat spinal cord.

BACKGROUND: The role of neural transplantation as a restorative strategy for spinal cord injury continues to be intensely investigated. Ideally, the tissue source for transplantation must be readily available, free of disease and able to survive and mature following implantation into the adverse environment created by the injury. We have studied the use of a commercially available cell line of cultured human neurons (hNT neurons) as a tissue source for neural transplantation in spinal cord injury. METHODS: Following a left lateral thoracic hemisection, 54 immunosuppressed, female Wistar rats were randomly allocated into different treatment groups; hemisection only or hemisection and hNT cell transplantation (via a bridge, double or triple graft). Grafting occurred three days after spinal cord injury. After thirteen weeks the animals were sacrificed and tissue sections were stained with human neuron specific enolase and human specific neural cell adhesion molecule. RESULTS: Immunohistochemical evidence of graft survival was displayed in 66.7% of the surviving, grafted animals. Fibre outgrowth, greatest in the bridge and triple grafts, was observed in both rostral and caudal directions essentially bridging the lesion. Double grafts were smaller, displaying less fibre outgrowth, which did not cross the lesion. Long fibre outgrowth was evident up to 2 cm from the graft as assessed by tracing and immunohistochemical studies. CONCLUSIONS: Bridge and triple grafts displayed greater growth and enabled the hNT graft to essentially bridge the lesion. This suggests that hNT neurons have the potential to structurally reconnect the proximal and distal spinal cord across the region of injury.