Abstract
We previously reported that CD44-positive cells were candidates for astrocyte precursor cells in the developing cerebellum, because cells expressing high levels of CD44 selected by fluorescence-activated cell sorting (FACS) gave rise only to astrocytes in vitro. However, whether CD44 is a specific cell marker for cerebellar astrocyte precursor cells in vivo is unknown. In this study, we used immunohistochemistry, in situ hybridization, and FACS to analyze the spatial and temporal expression of CD44 and characterize the CD44-positive cells in the mouse cerebellum during development. CD44 expression was observed not only in astrocyte precursor cells but also in neural stem cells and oligodendrocyte precursor cells (OPCs) at early postnatal stages. CD44 expression in OPCs was shut off during oligodendrocyte differentiation. Interestingly, during development, CD44 expression was limited specifically to Bergmann glia and fibrous astrocytes among three types of astrocytes in cerebellum, and expression in astrocytes was shut off during postnatal development. CD44 expression was also detected in developing Purkinje and granule neurons but was limited to granule neurons in the adult cerebellum. Thus, at early developmental stages of the cerebellum, CD44 was widely expressed in several types of precursor cells, and over the course of development, the expression of CD44 became restricted to granule neurons in the adult.
Highlights
The cerebellum is composed of distinct layers: the external germinal layer (EGL), the molecular layer (ML), the Purkinje cell layer (PCL), the granule layer (GL), and the white matter (WM) [1]
CD44high cells isolated from glial-enriched cellular fraction of postnatal day 3 (P3) mouse cerebellum by fluorescence-activated cell sorting (FACS) were positive for astrocyte-lineage markers (BLBP, GLAST) and the neural stem cell marker but were negative for the mature astrocyte marker (GFAP), the immature oligodendrocyte marker (O4) or the neuronal marker (Tuj1)
We focused on the expression profile of CD44 during cerebellar development in order to determine whether CD44 expression is restricted to astrocyte-lineage cells
Summary
The cerebellum is composed of distinct layers: the external germinal layer (EGL), the molecular layer (ML), the Purkinje cell layer (PCL), the granule layer (GL), and the white matter (WM) [1]. There are three types of astrocytes in the murine cerebellar cortex: Bergmann glia in the Purkinje cell layer, fibrous astrocyte in the white matter, and protoplasmic astrocyte in the granule layer [8]. By utilizing this specific characteristic, we can identify those astrocytes judging from their morphologies and locations, we focused on developing cerebellum as a good model to examine glial development. How these different types of cerebellar astrocytes are generated remains poorly understood. We previously have shown that cells with high CD44 expression (CD44high cells), purified from the large-cell fraction (enriched in glia) of mouse postnatal day 3 (P3) cerebellum, were astrocyte-restricted precursor cells in vitro [9]
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