Survival and functional recovery of transplanted human dopaminergic neurons into hemiparkinsonian rats depend on the cannula size of the implantation instrument.

Promising therapeutic strategies for neurodegenerative diseases such as Parkinson's disease include replacement of lost striatal dopaminergic neurons by grafting of embryonic mesencephalic cells. However, the poor survival of the transplanted tissue still limits transplantation of these cells into the human brain in a larger number of patients. We addressed the question, if the diameter of the transplantation cannulas has an effect on the number of surviving transplanted human embryonic mesencephalic cells into the striatum of 6-OHDA lesioned rats. We report a significantly higher number of surviving human cells using an ultrathin micropipette compared to cannulas with wider diameters. Importantly, higher numbers of surviving cells also correlated with a behavioral recovery of the hemiparkinsonian rats.

Establishment of experimental glioma models at the intrinsic brainstem region of the rats.

OBJECTIVES: As the treatment of human intrinsic brainstem gliomas remains challenging, experimental glioma models are needed. METHODS: We developed a rat model of intrinsic brain stem glioma that uses a stereotactic frame to fix the head for the delivery of C6 glioma cells to target sites via a permanently implanted cannula. We inoculated the rat midbrain, pons or cerebral cortex with 5 x 10(4) cells suspended in 1 microl culture medium over the course of 2 minutes. RESULTS: Three days post-implantation, tumor formation was visible in the periaqueductal gray matter in the midbrain and the tegmentum of the pons. On the tenth day, the tumor diameter exceeded over 2 mm; there was no tumor cell seeding into the cerebrospinal fluid space. The tumor manifested the histological features typical of glioblastoma; Ki-67 labeling index was 32%. DISCUSSION: Because in our model the cannula is permanently implanted, additional inocula can be delivered. Here we detail our rat brainstem glioma model and discuss its usefulness for the investigation of these tumor in humans.

Evaluation of surgical techniques for neuronal cell transplantation used in patients with stroke.

Transplantation of cultured neuronal cells was performed in two human clinical trials after safety and efficacy was demonstrated in animal models of stroke. The studies tested the utility of human neuronal cellular transplantation into and around the small stroke volume. We developed a stereotactic surgical technique for cell delivery and evaluated that method in 26 patients with basal ganglia region motor stroke. Human neuronal cells (hNT cells; LBS neurons) were delivered frozen then thawed and formulated on the morning of surgery. Patients in a first trial received 2 or 6 million cells in three or nine implants, and in a second trial, 5 or 10 million in 25 implants. A novel cell delivery cannula was designed, manufactured, tested, and used in surgery. Immediate postoperative CT scans and later serial MR scans were used to evaluate the surgical site. Tests on the cell implantation cannula showed that the cells were not damaged and remained viable after injection. All patients underwent uncomplicated surgeries. Cells could be implanted within a 2-h period, maintaining viability of the preparation. Serial evaluations (maximum 5 years) showed no cell-related adverse serologic or imaging-defined effects. One patient had burr hole drainage of an asymptomatic chronic subdural hematoma. Human neuronal cells can be produced in culture and implanted stereotactically into the brains of patients with stroke. Surgical cell delivery did not lead to new neurological deficits, and imaging studies showed no adverse effects. The cannula used allowed precise injection of the clinical cell dose within a time period that maintained cell viability.

Stereotactic transplantation of a dopamine-producing capsule into the striatum for treatment of Parkinson disease: a preclinical primate study.

OBJECT: The PC12 cells are well known for their ability to secrete dopamine and levodopa. In multiple animal mode encapsulated PC12 cells have been shown to ameliorate parkinsonian symptoms when transplanted into the striatum; technique is expected to be effective clinically as well. The present study was performed using nonhuman primates to ensure that the transplantation of encapsulated PC12 cells is likely to be both safe and effective in human clinical trials. METHODS: Unencapsulated or encapsulated PC12 cells were implanted into the brains of Japanese monkeys (Macaca fuscata). Histological and immunocytochemical analyses were performed 1, 2, 4, and 8 weeks posttransplantation on the unencapsulated cells and 2, 4, and 8 weeks after transplantation on the encapsulated cells. The survival of the PC12 cells inside the capsule was determined by measuring the amounts of dopamine and levodopa released from the capsules a removal from the striatum. Magnetic resonance imaging was performed in both unencapsulated and encapsulated PC12 cell-grafted groups. Due to the immunological reaction of the host brain no unencapsulated PC12 cells remained in the grafted area 8 weeks after transplantation. On the contrary, encapsulated PC12 cells retrieved from the host brain continued to release dopamine and levodopa even 8 weeks after implantation. The host's reaction to the PC12-loaded capsule was much weaker than that to the unencapsulated PC12 cells. CONCLUSIONS: These results suggest that the transplantation of encapsulated PC12 cells could be a safe and effective treatment modality for Parkinson disease in human patients.

Labeling stem cells in vitro for identification of their differentiated phenotypes after grafting into the CNS.

Labeling stem cells for CNS grafting is an empirical process. Specific protocols cannot be given that will work for all cell types and all applications. We have provided the range of conditions under which various labels have been successfully used in CNS grafting studies, and delineated the parameters that have to be empirically established. Given a clear understanding of the limitations of the respective labels, and the expected outcome of the grafting experiment, these labeling guidelines should enable any investigator to develop a successful labeling approach. Our own personal bias is to use labels that cannot be transferred to host cells. We prefer BrdU, or more often, retrovirally delivered EGFP or lacZ. However, each investigator will have to decide what is optimal for their own cell population and experimental design.