プルキンエ様細胞への間葉系および神経幹細胞の分化：形態学的研究

Purkinje cells are a critical class of inhibitory neurons in the central nervous system, and their dysfunction has been implicated in a range of neurological conditions. Purkinje cell isolation and expansion have enabled advances in regenerative neuroscience for cell-based therapies to restore the inhibitory balance in diseased neural circuits. The aim of this study was to develop a simple in vitro method for generating Purkinje-like cells from two neural stem cell sources: neural stem cells derived from bone marrow mesenchymal stem cells (bmMSCs) and neural stem cells isolated from the olfactory bulb, with potential relevance for future clinical applications.

We isolated MSCs from mouse bone marrow and induced their differentiation into NSCs. In addition, we isolated NSCs directly from the olfactory bulb in mice. Purkinje cells were isolated via magnetic beads. The isolated Purkinje cells were used to induce differentiation of NSCs into Purkinje-like cells via cell lysate and coculture methods. The characteristics and phenotypes of the resultant cells were determined according to branch number and length, axon length, and immunocytochemistry.

MSCs, NSCs, and Purkinje-like cells were successfully isolated and expanded in culture, as demonstrated through immunocytochemical analyses indicating positivity for related phenotypic markers, such as CD105 and CD95 for MSCs; nestin for NSCs; and calbindin for Purkinje cells. The produced Purkinje cells also had greater axon lengths, and branch numbers and lengths, than the other cell types. Moreover, both the cell lysate and coculture methods successfully expanded Purkinje-like cells in vitro.

Our methods involving coculturing isolated purified Purkinje cells and/or using their cell lysates successfully differentiated NSCs into phenotypically Purkinje-like cells.