Effects of Passage Number and Differentiation Protocol on the Generation of Dopaminergic Neurons from Rat Bone Marrow-Derived Mesenchymal Stem Cells.

Gabrielle Shall,Priya Nethala,Ute Hochgeschwender,Ryan Welchko,Julien Rossignol,Gary Dunbar,Michael Sandstrom,Megan Menosky,Sarah Decker,Xavier Leveque,Ming Lu

doi:10.3390/ijms19030720

Abstract

Multiple studies have demonstrated the ability of mesenchymal stem cells (MSCs) to differentiate into dopamine-producing cells, in vitro and in vivo, indicating their potential to be used in the treatment of Parkinson’s disease (PD). However, there are discrepancies among studies regarding the optimal time (i.e., passage number) and method for dopaminergic induction, in vitro. In the current study, we compared the ability of early (P4) and later (P40) passaged bone marrow-derived MSCs to differentiate into dopaminergic neurons using two growth-factor-based approaches. A direct dopaminergic induction (DDI) was used to directly convert MSCs into dopaminergic neurons, and an indirect dopaminergic induction (IDI) was used to direct MSCs toward a neuronal lineage prior to terminal dopaminergic differentiation. Results indicate that both early and later passaged MSCs exhibited positive expression of neuronal and dopaminergic markers following either the DDI or IDI protocols. Additionally, both early and later passaged MSCs released dopamine and exhibited spontaneous neuronal activity following either the DDI or IDI. Still, P4 MSCs exhibited significantly higher spiking and bursting frequencies as compared to P40 MSCs. Findings from this study provide evidence that early passaged MSCs, which have undergone the DDI, are more efficient at generating dopaminergic-like cells in vitro, as compared to later passaged MSCs or MSCs that have undergone the IDI.

Highlights

Parkinson’s disease (PD) is the second most common neurodegenerative disorder after Alzheimer’s disease, with the incidence of PD rapidly increasing in individuals over the age of 60 [1,2].The pathological hallmark of PD is the loss of dopaminergic neurons from the substantia nigra, pars compacta that innervate the striatum through the nigrostriatal pathway [3]
P4 and P40 undifferentiated mesenchymal stem cells (MSCs) were negative for the hematopoietic stem cell marker, CD34, and the major histocompatibility complex class II (MHC II) antigen (Table 1), which is consistent with MSC characterization
This study investigated the influence of serial passaging on the ability of MSCs to differentiate into dopaminergic neurons

Summary

Introduction

The pathological hallmark of PD is the loss of dopaminergic neurons from the substantia nigra, pars compacta that innervate the striatum through the nigrostriatal pathway [3]. This degeneration is often accompanied by an intraneuronal accumulation of α-synuclein-containing Lewy bodies in the remaining, intact nigral neurons [2,3,4,5]. Transplantations of iPSCs introduce the risk of tumor formation in vivo [28,29,30,31,32,33] These issues warrant the use of alternative sources for cell replacement therapies

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Results

Conclusion