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Parkinson disease (PD) is characterized by a loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). The selective loss of this cell population makes PD a good candidate for cell-based therapies. Clinical trials using cells derived from human fetal ventral midbrain (hVM) have shown that dopamine release could be restored to normal levels and in some PD patients produced substantial long-term clinical improvement. The ethical issues related to the use of human fetal material as well as the low number of hVM tissue available have limited the spread of this approach and have lead researchers to find new sources of cells such as human embryonic stem cells (hESCs) and pluripotent stem cells (IPCs). This study was designed to provide a detailed molecular and functional of the fetal VM tissue in order to give a deeper understanding of its composition and characteristics. The characterization is based on the comparison of standard culture conditions (2D) versus organoids of hVM (3D). We used unbiased single cell RNA sequencing to transcriptionally compare fetal VM tissue in 2D and 3D conditions together with a functional analysis performed using whole cell patch clamp technique. Our analysis determined the molecular and functional properties of the tissue that originates dopaminergic neurons and also confirmed important differences between the culture conditions as well as it . The data can be used for quality assessment of new cell sources and provide new molecular targets for improved subtype specificity during stem cell differentiation.

Parkinson disease (PD) is characterized by a loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). The selective loss of this cell population makes PD a good candidate for cell-based therapies. Clinical trials using cells derived from human fetal ventral midbrain (hVM) have shown that dopamine release was restored to normal levels and in some PD patients produced substantial long-term clinical improvement. Although new sources of cells such as human embryonic stem cells (hESCs) and pluripotent stem cells (IPCs) are now been the focus of studies and new clinical trials, the hVM tissue represent the gold standard for cell based therapies and serve as important comparator for new cell sources. Therefore there is a need of an increased understanding and characterization of this tissue with new technologies such as organoid culture conditions and single cell sequencing. This study is designed to give a detailed characterization of human fetal VM tissue in standard culture conditions (2D) versus organoids (3D). We used immunocytochemistry to study the presence of ventral midbrain markers in 2D versus 3D condition together with single cell RNA sequencing to transcriptionally compare fetal VM tissue in standard or organoid cultures. Our analysis confirmed that the 3D condition preserve the neuronal composition of the tissue more than standard culture where instead, over a long period of time, the majority of the cells are non-neuronal. This suggest that 3D culture represent a better condition to maintain and study dopaminergic neurons in culture.

Parkinson disease (PD) is characterized by a loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). The selective loss of this cell population makes PD a good candidate for cell-based therapies. Clinical trials using cells derived from human fetal ventral midbrain (hVM) have shown that dopamine release could be restored to normal levels and in some PD patients produced substantial long-term clinical improvement. The ethical issues related to the use of human fetal material as well as the low number of hVM tissue available have limited the spread of this approach and have lead researchers to find new sources of cells such as human embryonic stem cells (hESCs) and pluripotent stem cells (IPCs). This study was designed to provide a detailed molecular and functional of the fetal VM tissue in order to give a deeper understanding of its composition and characteristics. The characterization is based on the comparison of standard culture conditions (2D) versus organoids of hVM (3D). We used unbiased single cell RNA sequencing to transcriptionally compare fetal VM tissue in 2D and 3D conditions together with a functional analysis performed using whole cell patch clamp technique. Our analysis determined the molecular and functional properties of the tissue that originates dopaminergic neurons and also confirmed important differences between the culture conditions as well as it . The data can be used for quality assessment of new cell sources and provide new molecular targets for improved subtype specificity during stem cell differentiation.

Parkinson disease (PD) is characterized by a loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). The selective loss of this cell population makes PD a good candidate for cell-based therapies. Clinical trials using cells derived from human fetal ventral midbrain (hVM) have shown that dopamine release was restored to normal levels and in some PD patients produced substantial long-term clinical improvement. Although new sources of cells such as human embryonic stem cells (hESCs) and pluripotent stem cells (IPCs) are now been the focus of studies and new clinical trials, the hVM tissue represent the gold standard for cell based therapies and serve as important comparator for new cell sources. Therefore there is a need of an increased understanding and characterization of this tissue with new technologies such as organoid culture conditions and single cell sequencing. This study is designed to give a detailed characterization of human fetal VM tissue in standard culture conditions (2D) versus organoids (3D). We used immunocytochemistry to study the presence of ventral midbrain markers in 2D versus 3D condition together with single cell RNA sequencing to transcriptionally compare fetal VM tissue in standard or organoid cultures. Our analysis confirmed that the 3D condition preserve the neuronal composition of the tissue more than standard culture where instead, over a long period of time, the majority of the cells are non-neuronal. This suggest that 3D culture represent a better condition to maintain and study dopaminergic neurons in culture. &nbsp

Parkinson disease (PD) is characterized by a loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). The selective loss of this cell population makes PD a good candidate for cell-based therapies. Clinical trials using cells derived from human fetal ventral midbrain (hVM) have shown that dopamine release was restored to normal levels and in some PD patients produced substantial long-term clinical improvement. Although new sources of cells such as human embryonic stem cells (hESCs) and pluripotent stem cells (IPCs) are now been the focus of studies and new clinical trials, the hVM tissue represent the gold standard for cell based therapies and serve as important comparator for new cell sources. Therefore there is a need of an increased understanding and characterization of this tissue with new technologies such as organoid culture conditions and single cell sequencing. This study is designed to give a detailed characterization of human fetal VM tissue in standard culture conditions (2D) versus organoids (3D). We used immunocytochemistry to study the presence of ventral midbrain markers in 2D versus 3D condition together with single cell RNA sequencing to transcriptionally compare fetal VM tissue in standard or organoid cultures. Our analysis confirmed that the 3D condition preserve the neuronal composition of the tissue more than standard culture where instead, over a long period of time, the majority of the cells are non-neuronal. This suggest that 3D culture represent a better condition to maintain and study dopaminergic neurons in culture.

Parkinson disease (PD) is characterized by a loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). The selective loss of this cell population makes PD a good candidate for cell-based therapies. Clinical trials using cells derived from human fetal ventral midbrain (hVM) have shown that dopamine release could be restored to normal levels and in some PD patients produced substantial long-term clinical improvement. The ethical issues related to the use of human fetal material as well as the low number of hVM tissue available have limited the spread of this approach and have lead researchers to find new sources of cells such as human embryonic stem cells (hESCs) and pluripotent stem cells (IPCs). This study was designed to provide a detailed molecular and functional of the fetal VM tissue in order to give a deeper understanding of its composition and characteristics. The characterization is based on the comparison of standard culture conditions (2D) versus organoids of hVM (3D). We used unbiased single cell RNA sequencing to transcriptionally compare fetal VM tissue in 2D and 3D conditions together with a functional analysis performed using whole cell patch clamp technique. Our analysis determined the molecular and functional properties of the tissue that originates dopaminergic neurons and also confirmed important differences between the culture conditions as well as it . The data can be used for quality assessment of new cell sources and provide new molecular targets for improved subtype specificity during stem cell differentiation.