Distribution of chromosome 18 and X centric heterochromatin in the interphase nucleus of cultured human cells

In situ hybridization of human chromosome 18 and X-specific alphoid
DNA-probes was performed in combination with three dimensional (3D)
and two dimensional (2D) image analysis to study the interphase
distribution of the centric heterochromatin (18c and Xc) of these
chromosomes in cultured human cells. 3D analyses of 18c targets
using confocal laser scanning microscopy indicated a nonrandom
disposition in 73 amniotic fluid cell nuclei. The shape of these
nuclei resembled rather flat cylinders or ellipsoids targets were
preferentially arranged in a domain around the nuclear center, but
close to or associated with the nuclear envelope. Within this
domain, however, positionings of the two targets occurred
independently from each other, i.e., the two targets were observed
with similar frequencies at the same (upper or lower) side of the
nuclear envelope as those on opposite sides. This result strongly
argues against any permanent homologous association of 18c. A 2D
analytical approach was used for the rapid evaluation of 18c
positions in over 4000 interphase nuclei from normal male and
female individuals, as well as individuals with trisomy 18 and
Bloom's syndrome. In addition to epithelially derived amniotic
fluid cells, investigated cell types included in vitro cultivated
fibroblastoid cells established from fetal lung tissue and
skin-derived fibroblasts. In agreement with the above 3D
observations 18c targets were found significantly closer (P <
0.01) to the center of the 2D nuclear image (CNI) and to each other
in all these cultures compared to a random distribution derived
from corresponding ellipsoid or cylinder model nuclei. For
comparison, a chromosome X-specific alphoid DNA probe was used to
investigate the 2D distribution of chromosome X centric
heterochromatin in the same cell types. Two dimensional Xc-Xc and
Xc-CNI distances fit a random distribution in diploid normal and
Bloom's syndrome nuclei, as well as in nuclei with trisomy X. The
different distributions of 18c and Xc targets were confirmed by the
simultaneous staining of these targets in different colors within
individual nuclei using a double in situ hybridization approach.