Chromosomal instability and cytoskeletal defects in oral cancer cells - PubMed (original) (raw)

Chromosomal instability and cytoskeletal defects in oral cancer cells

W S Saunders et al. Proc Natl Acad Sci U S A. 2000.

Abstract

Oral squamous cell carcinomas are characterized by complex, often near-triploid karyotypes with structural and numerical variations superimposed on the initial clonal chromosomal alterations. We used immunohistochemistry combined with classical cytogenetic analysis and spectral karyotyping to investigate the chromosomal segregation defects in cultured oral squamous cell carcinoma cells. During division, these cells frequently exhibit lagging chromosomes at both metaphase and anaphase, suggesting defects in the mitotic apparatus or kinetochore. Dicentric anaphase chromatin bridges and structurally altered chromosomes with consistent long arms and variable short arms, as well as the presence of gene amplification, suggested the occurrence of breakage-fusion-bridge cycles. Some anaphase bridges were observed to persist into telophase, resulting in chromosomal exclusion from the reforming nucleus and micronucleus formation. Multipolar spindles were found to various degrees in the oral squamous cell carcinoma lines. In the multipolar spindles, the poles demonstrated different levels of chromosomal capture and alignment, indicating functional differences between the poles. Some spindle poles showed premature splitting of centrosomal material, a precursor to full separation of the microtubule organizing centers. These results indicate that some of the chromosomal instability observed within these cancer cells might be the result of cytoskeletal defects and breakage-fusion-bridge cycles.

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Figures

Figure 1

Figure 1

(A) Display image of metaphase chromosomes from a Colcemid-arrested UPCI:SCC172 cell after SKY analysis. Arrowhead identifies the marker chromosome “c” shown in B. (B) Normal chromosomes 3, 4, 8, and 11 from this culture are shown. Marker chromosomes, also from UPCI:SCC172, are indicated as a, b, and c. (C) A representative dicentric chromosome is shown from a Colcemid-arrested UPCI:SCC131 cell C-banded to highlight the centromeres.

Figure 2

Figure 2

UPCI:SCC131 cells were immunolabeled with Abs to tubulin (yellow), and kinetochores (red in A, D, and E) or NuMA (purple in C) and counterstained with the DNA dye, DAPI (blue). A normal metaphase from this culture is shown in A, and multipolar metaphase spindles in B–E. (C, Inset) The anti-tubulin image alone of the pole marked with an arrowhead. (D and E) Arrows mark minor spindle poles. (Bars, 5 μm.) (F) A tissue section from a formalin-fixed, paraffin-embedded, laryngeal carcinoma was stained with hematoxylin and eosin. An atypical tripolar mitosis (arrow) and a normal bipolar mitotic figure (arrowhead) are shown.

Figure 3

Figure 3

UPCI lines SCC131, SCC172, and SCC003 were treated with anti-tubulin immunolabeling and DAPI staining and examined for five segregational abnormalities. The percentages of metaphase cells with multipolar spindles, interphase cells with micronuclei, metaphase, and anaphase cells with lagging chromosomes, and anaphase cells with chromatin bridges are given, as well as the percentage of metaphase cells with normal spindles. The mean and SEM from three coverslips are shown. A total for each cell line of 25–81 anaphase cells was counted for anaphase bridges and lagging chromosomes, 86–164 metaphase cells for multipolar spindles and lagging chromosomes, 1264–1593 interphase cells for micronuclei, and 110–370 metaphase cells for normal spindles.

Figure 4

Figure 4

Anaphase bridges containing centromeres and chromosome 11. Immunolabeling with Abs to tubulin (yellow), centromeres (red), and with DAPI (blue) and FISH with a chromosome 11 paint probe (green). (B) Arrows point to centromeres trapped in the forming midbody as these late telophase cells divide. (D) Arrow points to the trapped lagging chromosome excluded from the reforming nucleus of the cell on the right. (E) Some micronuclei are immunonegative for anti-centromere Abs. Arrow points to negative micronucleus, and arrowhead points to positive. Examples are from UPCI:SCC131.

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