Identification of dopamine neuron progenitors in the embryonic midbrain and stem cell cultures - Studies on the role of neuronal subtype and differentiation state for cell replacemen in a rodent model of Parkinson's disease

Sammanfattning: Parkinson’s Disease (PD) is a neurodegenerative disorder where the dopamine producing neurons in the ventral mesencephalon (VM) progressively die and result in symptoms such as resting tremors, muscle rigidity, slowness and difficulties in initiating movements.Currently there is no cure for PD and the available drug treatments only offer symptomatic relief and are often associated with severe side effects. Thus, there is an obvious need for
alternative therapies.
A promising alternative approach is cell replacement therapy, which aims to replacethe lost dopamine-producing neurons by transplanting cells with equivalent properties. Clinical transplantations using cells obtained from foetal VM tissue have provided the proof-of-principle that cell replacement therapy can provide a long-lasting recovery. However, the use of foetal tissue involves moral and severe practical issues making it impossible
to standardise the required quantity and quality of cells. In the context of developing an alternative cell source that can offer a consistent and expandable supply of mesDA neuron progenitors, this thesis has investigated the features of foetal VM cells that allow them to survive, innervate and function upon transplantation.
The foetal VM tissue obtained for transplantation contains mesDA neuron progenitors at different differentiation states. By identifying and isolating the mesDA neuron progenitors from different developmental time points and differentiation states, the work in this thesis has shown that the optimal differentiation state for transplantation changes from a proliferative to postmitotic progenitor during development. It is crucial that the mesDA progenitors are harvested within this window of opportunity for the cells to survive, integrate and function upon transplantation. Following transplantation, the foetal VM was shown to give rise to the two main mesDA neuron subtypes of the VM, namely A9 and A10. Furthermore, the data in this thesis demonstrates that a functional graft is explicitly dependent on the generation of A9 neurons, which were shown to be necessary for innervation and connection with the host tissue.
The knowledge obtained from the work with foetal VM was applied to embryonic
stem (ES) cells as the alternative cell source. ES cells posses the feature of being renewable and have the potential to generate mesDA neurons. However, differentiating ES cell cultures also contain other cell types as well as immature cells, which frequently give rise to tumour-like structures within the grafts. The approach of identifying and isolating the ES cell derived mesDA progenitors prior to transplantation resulted in grafts enriched with
mesDA neurons without the concomitant tumour-like growths. The goal of obtaining a safe and alternative cell source for PD cell replacement therapy
has been brought closer with the work presented in this thesis. The use of stem cells is unquestionably promising and may one day be a reality in the clinical treatment of PD patients.