Objective/Rationale:
Recently, genetic manipulation of human cells from various organs, e.g. skin, enabled direct generation of human nerve cells (so-called induced neurons – iNs) that potentially can be used for cell replacement therapies in neurodegenerative diseases, such as Parkinson’s disease (PD). However, it remains unclear whether transplanted human iNs are capable of functionally integrating into the existing brain circuitry and thereby exerting therapeutic effect in the PD patients. Using recently developed optogenetic technique will enable us addressing this important question in previously unprecedented and very specific way.
Project Description:
The studies will be performed both in cell and tissue cultures and pre-clinical models of PD. We will generate human iNs from skin cells and transduce these cells with light-activated bacterial proteins, so-called opsins, enabling selective activation of trasnpalanted iNs by light illumination. In separate experiments, the recipient brain cells will be transduced with these opsins, which will allow for their specific activation by light. Such optogenetic approach will make it possible to study in great detail the interaction of transplanted iNs with the host brain cells, and explore whether transplanted iNs are able to functionally integrate into the host circuitry.
Relevance to Diagnosis/Treatment of Parkinson’s Disease:
Generation of iN cells from patients own skin eliminates ethical problems related to human embryonic stem cells, and minimizes the risk for tumor formation associated with other human-derived stem-like cells. We will generate human iNs that produce and release dopamine (DA), a neurotransmitter progressively lost in PD due to degeneration of patients own DA-producing cells in midbrain. The study will lay the foundation for developing this approach towards clinical applications, and provide with better understanding how these iNs will function after transplantation in patients’ brain.
Anticipated Outcome:
The proposed project will address one of the major unresolved issues in regenerative medicine by promoting patient-specific iN-based therapies and improving our understanding of the basic cellular, synaptic and molecular mechanisms of their regenerative capacity. It is expected that, using optogenetic approaches, which are novel and most powerful strategies, the project will establish the role of synaptic integration of grafted iNs for achieving therapeutic effects, and provide means for enhancing these effects by stimulating grafted iN DA cells.
Final Outcome
We have shown that human iN cells when differentiated in vitro express neuronal properties and can generate action potentials, a hallmark of neurons. The factors used for such differentiation are defined and tested, and characterized to yield TH-positive dopaminergic neurons that are needed for replacement cell therapy approaches in Parkinson’s disease. These cells are also able to express opsins used for their optogenetic stimulation, which may be useful for fine-tuning their functional impact. Currently these human iN cells are transplanted into the pre-clinical model’s host brain and in few months their properties will be analyzed in detail.