Research reportNestin-containing cells express glial fibrillary acidic protein in the proliferative regions of central nervous system of postnatal developing and adult mice
Introduction
It is well documented that the expression of nestin protein in the prenatal and postnatal developing central nervous system (CNS) of mammals reflects the differentiation or proliferative state of neural precursors [6], [18], [19], [28], [36], [38], [49], [51]. Embryonic intermediate filament (IF) nestin belongs to class III IFs, which also include vimentin and glial fibrillary acidic protein (GFAP), and, structurally, it is closely related to class IV IFs, such as neurofilaments and α-internexin [9], [15], [19], [27], [28], [37]. At the embryonic neurulation stage, multipotential neural precursors or neuroepithelial precursor cells temporarily express nestin protein. Substitution of nestin protein by vimentin and GFAP or neurofilaments takes place sequentially during the maturation or differentiation of neural precursor cells. Nestin is down-regulated either at the onset of GFAP or neurofilament expression or during subsequent differentiation of multipotential neural precursor into astrocytes or neurons [6], [17], [18], [24], [27], [28], [31], [36], [37], [51]. The transient abundant expression of embryonic intermediate filament nestin is extensively recognized as a marker for multipotential neural precursor cells in developing CNS of mammals [6], [20], [22], [25], [28], [32], [41], [43].
Previous evidence has shown that nestin protein is abundantly expressed in proliferative regions of embryonic or developing CNS, and is also predominately localized in the radial glial cells that are recognized as neural precursors [6], [13], [27], [29], [32], [34], [38], [50], [52]. Furthermore, re-expression of nestin was reported to occur in traumatic brain injury, ischemic or excitotoxic cerebral lesions, as well as pathologically deafferented target regions [2], [5], [8], [21], [30], [39], [44], [45]. Significant up-regulation of nestin was also demonstrated in reactive astrocytes after systemic administration of the neurotoxicant 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) [4]. Re-expression of nestin in astroglial cells of adult is thus recognized as a sensitive marker for activation of astroglial cells in the CNS. It implies that, to certain extent, these cells expressing nestin reflect their reiterating or sustaining active state of embryonic precursor cells, and may implicate in the neurogenesis, remodeling and repairing processes of developing and adult CNS [12], [25], [35], [42], [43], [45]. Though in situ hybridization was carried out for the localization of nestin mRNA developing brain [6], the detail distribution pattern of nestin protein in adult and postnatal CNS remains to be delineated. Further studies are also needed to characterize the precursor property of nestin-expressing cells and their differentiation fate. In the present study, immunocytochemical approach was utilized to demonstrate the expression patterns of nestin protein, and its co-expression with GFAP or neuronal nuclear specific protein (NeuN) within the brain and spinal cord of adult and postnatal mice.
Section snippets
Materials and methods
Forty-two BALB/c mice were used in the present study. Six groups of animal were used: postnatal day 2 (P2, n=8), P7 (n=8), P14 (n=8), P21 (=6), P30 (n=6), and adult (P60, n=6). All animal procedures conformed to the guidelines of the National Institute of Health for the care and use of laboratory animals (NIH Publication No. 80-23), and all efforts were made to minimize animal suffering. The animals deeply anesthetized with sodium pentobarbital (40 mg/kg, i.p.) were transcardially perfused with
Characterization of nestin-, GFAP-, or NeuN-immunoreactivity
Nestin-, GFAP-, or NeuN-immunoreactivity was detected in distinct regions of the brain and spinal cord. Nestin- or GFAP-immunoreactivity was predominately localized in somata and processes of glial cells (Fig. 1), while NeuN-immunoreactivity was exclusively localized in nuclei of neurons. The distribution of nestin-immunoreactivity was found to overlap partially with that of GFAP- or NeuN-immunoreactivity in distinct CNS regions, e.g. the cerebral cortex, hippocampus, hypothalamus, cerebellar
Discussion
Our present data demonstrated the distribution of nestin-containing cells in distinct proliferative CNS regions of postnatal developing mice. Double immunofluorescence further indicated that a large percentage (77%) of nestin-containing cells in adult express glial cell marker GFAP. These results revealed the distribution pattern of nestin-containing neural precursors of the postnatal CNS and their possible differentiation to neurons and astrocytes, suggesting that these nestin-expressing cells
Acknowledgements
The authors thank Dr. N.H. Jing, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, for his gift of rabbit antiserum to nestin protein. This work was supported by grants from the National Natural Science Foundation of China (30040012), The Foundation of Fourth Military Medical University (CX01A013), and Hong Kong Research Grants Council.
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