Human neural stem cell transplantation in experimental brain trauma

Abstract: Traumatic brain injury (TBI) is the leading cause of death and disability in young adults in the industrialized world. Human neural stem/progenitor cells (hNSCs) can be expanded in culture and have the ability to differentiate into the major cell types of the adult brain. These properties make them candidates for therapeutic transplantation in cases of neurological injuries and diseases that involve cell loss. The project was initiated to expand the knowledge on, and refine the techniques used in, hNSC xenografting to the traumatized brain. The weight-drop cortical contusion model was evaluated with focus on the delayed cell death. Neurons were found to be the cell type predominantly affected by the delayed cell death in this experimental model of TBI. Furthermore, an increase and redistribution of the pro-apoptotic protein Bax was evident at 6 days post injury indicating a wide window for potential therapeutic intervention. The model simulates aspects of the clinical histopathology and the subsequent xenograft investigations were carried out in this experimental model for TBI. Cryo-preserved hNSCs were xenografted to the medial limit of the lesion and the ipsilateral hippocampus immediately after injury in immunosuppressed rats. The human cells survived and increased in number between 2 and 6 weeks post surgery indicating that the xenografted cells proliferated. This was corroborated by the expression of the proliferation antigen Ki-67. A low proportion of the hNSC-derived cells differentiated to astrocytes and neurons. An increase in the proportion of human cells that expressed the neuronal marker NeuN was observed between 2 and 6 weeks in the hippocampus. A marker for degenerating neurons, Fluoro-Jade, was used to investigate if there was any difference in host neuroprotection between viable and non-viable hNSCs xenografted to the lesion. The results indicated that non-viable hNSCs increase neurodegeneration while viable hNSCs exerted some neuroprotection at 6 days post surgery. Immunosuppressive treatment has several serious side effects. To determine if immunosuppression can be shortened two immunosuppression regimes were compared; one group was given immunosuppression throughout the whole 6 weeks of the experiment while the other group was taken of immunosuppressant at 3 weeks post surgery. There were no differences in number of hNSC-derived cells, proliferation, migration or differentiation. In addition, 5/8 animals in a pilot group that was allowed to survive for six months after 3 weeks of immunosuppression still had surviving grafts. The ability to control grafted hNSCs in the environment of the traumatized brain is desirable. Polymer rods with slow-release kinetics was manufactured and implanted in the center of the lesion to investigate if administration of trophic factors could affect the xenografted hNSCs. The animals were immunosuppressed during the first 3 weeks and were allowed to survive 3 and 6 months. The pooled values from the 3 and 6 month groups were used for comparisons. Release of basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) did not increase the survival of xenografted hNSCs significantly. However, grafts receiving rods with rat serum albumin displayed increased survival compared to grafts receiving rods without any factor. Moreover, retinoic acid decreased the proportion of nestin-positive human cells. In conclusion, hNSCs xenografted to a clinically relevant model of TBI survive, differentiate and have effects on host neuroprotection. Decreased immunosuppression can be achieved without short-term effects on the xenografted hNSCs and both long-term survival and differentiation of hNSCs can be modulated by short-term release of bioactive molecules in situ.