Constitutional aneuploidy in the normal human brain

Abstract

The mouse brain contains genetically distinct cells that differ with respect to chromosome number manifested as aneuploidy (Rehen et al., 2001); however, the relevance to humans is not known. Here, using double-label fluorescence in situ hybridization for the autosome chromosome 21 (chromosome 21 point probes combined with chromosome 21 "paint" probes), along with immunocytochemistry and cell sorting, we present evidence for chromosome gain and loss in the human brain. Chromosome 21 aneuploid cells constitute approximately 4% of the estimated one trillion cells in the human brain and include non-neuronal cells and postmitotic neurons identified by the neuronspecific nuclear protein marker. In comparison, human interphase lymphocytes present chromosome 21 aneuploidy rates of 0.6%. Together, these data demonstrate that human brain cells (both neurons and non-neuronal cells) can be aneuploid and that the resulting genetic mosaicism is a normal feature of the human CNS.

Figure 1.

Chromosome 21 gain and loss is observed in cells isolated from the normal human brain. Human lymphocyte labeling demonstrates the specificity of the paint and point probes in metaphase spreads (A) and interphase nuclei (B). Neural progenitor cells from a Down's syndrome fetus have three copies of chromosome 21, as seen in a metaphase chromosome spread (C) and in an interphase nucleus (D), demonstrating the sensitivity of the FISH probes with regard to detecting numerical abnormalities of chromosome 21. E-H, Nuclei isolated from the brains of different patients containing one (E), two (F), three (G), or four (H) copies of chromosome 21. Blue indicates 4′,6-diamidino-2-phenylindole, whole-chromosome paint is shown in green, and the chromosome 21 point probe is shown in red. There is complete overlap between the paint and the point probe, as seen at higher magnification in the insets. Arrowheads indicate the numbers of chromosome 21 per cell. Scale bar, 5 μm.

Figure 2.

NeuN immunolabeling and cell sorting. A, Isolated normal human brain nuclei populations (P1) were gated based on forward scatter and side scatter and gated into NeuN+ (P4) and NeuN- (P5) sort populations based on Alexa Fluor 488 signal. The NeuN+ population is generally composed of larger nuclei than NeuN- nuclei based on forward scatter, although there is a significant size overlap. B, A representative preparation of sorted nuclei demonstrating >99.5% purity of separation between NeuN+ (P4) and NeuN- (P5) populations.