TY - JOUR
T1 - Fluorescence in situ hybridization-based approaches for detection of 12p overrepresentation, in particular i(12p), in cell lines of human testicular germ cell tumors of adults
AU - Mostert, M M
AU - van de Pol, M
AU - van Echten, J
AU - Olde Weghuis, D
AU - Geurts van Kessel, A
AU - Oosterhuis, J W
AU - Looijenga, L H
N1 - Funding Information:
This work was supported by Grant No. NKB-DDHK 94-836 from the Dutch Cancer Society (Koningin Wilhelmina Fonds). We thank Barbara Trask and Ger van de Engh (Department of Molecular Biotechnology, University of Washington, Seattle, WA, U.S.A.) for their contribution concerning the flow-sorted “paint” probe and the purification of YAC#5 by pulse-field gel electcophoresis and DOP-PCR. We also thank H. Vuik and A. Kievit (Department of Medical Photography, Dr. Daniel den Hoed Cancer Center) for their contributions in preparation of the figures. Further, we thank H.-J. Schrnoll, I. Damjanov and A. von Keitz for the gift of the cell lines and R. Gemmil for providing YAC#5. Purchase of the CCD camera and screen for the analysis of the in vitro cultures and of the biohazard flowhood was supported by the Nijbakker-Morra Foundation. We thank the Nijbakker-Morra and Maurits en Anna de Kock Foundation for financial support for purchase of hardware for storage of frozen material.
PY - 1996/4
Y1 - 1996/4
N2 - Overrepresentation of the short arm of chromosome 12 is frequently detected in human testicular germ cell tumors of adolescents and adults (TGCT). This overrepresentation mostly results from the formation of an isochromosome: i(12p). Whether the overrepresentation consistently involves the complete 12p arm including the centromere is still unclear. We studied five TGCT-derived cell lines (NT2, 2102Ep, H12.1, NCCIT, and S2), combining conventional chromosome banding, fluorescence in situ hybridization (FISH), and comparative genomic hybridization (CGH) to investigate the suitability of each of these techniques to detect aberrations involving chromosome 12. Karyotyping showed one or more i(12p)s in NT2, 2102Ep, H12.1, and S2. However, FISH with a centromere-specific probe (p alpha 12H8), a 12p "paint" and a 12p11.2--p12.1 region-specific probe yeast artificial chromosome (YAC)#5 and CGH could not confirm the presence of an i(12p) in S2. Additional randomly distributed 12p sequences were detected by FISH in H12.1, NCCIT, and S2. In most of these cases, (a part of) the centromere was included. All overrepresented 12p regions, except for those in S2, showed hybridization with YAC#5. CGH showed increased copy numbers of the complete 12p arm in the cell lines with one or more i(12p)s but no overrepresentation was noted in the cell lines without i(12p). In metaphase spreads, the centromeric block of the i(12p)s differed in size as compared with those of normal chromosomes 12. This was rarely noted in interphase nuclei. A decrease in size of the centromeric block in 2102Ep and H12.1 caused a weak FISH signal, which was difficult to detect, especially in interphase nuclei. The ratio between p alpha 12H8- and YAC#5-derived signals reflected the presence or absence of one or more i(12p)s. Our results indicate that double FISH with a centromere- and a 12p-specific probe can be used to detect 12p overrepresentation [including i(12p)] in TGCT both in metaphase spreads and interphase nuclei. CGH confirmed the relative overrepresentation of 12p sequences as detected by FISH and showed that in these cell lines the complete 12p was involved.
AB - Overrepresentation of the short arm of chromosome 12 is frequently detected in human testicular germ cell tumors of adolescents and adults (TGCT). This overrepresentation mostly results from the formation of an isochromosome: i(12p). Whether the overrepresentation consistently involves the complete 12p arm including the centromere is still unclear. We studied five TGCT-derived cell lines (NT2, 2102Ep, H12.1, NCCIT, and S2), combining conventional chromosome banding, fluorescence in situ hybridization (FISH), and comparative genomic hybridization (CGH) to investigate the suitability of each of these techniques to detect aberrations involving chromosome 12. Karyotyping showed one or more i(12p)s in NT2, 2102Ep, H12.1, and S2. However, FISH with a centromere-specific probe (p alpha 12H8), a 12p "paint" and a 12p11.2--p12.1 region-specific probe yeast artificial chromosome (YAC)#5 and CGH could not confirm the presence of an i(12p) in S2. Additional randomly distributed 12p sequences were detected by FISH in H12.1, NCCIT, and S2. In most of these cases, (a part of) the centromere was included. All overrepresented 12p regions, except for those in S2, showed hybridization with YAC#5. CGH showed increased copy numbers of the complete 12p arm in the cell lines with one or more i(12p)s but no overrepresentation was noted in the cell lines without i(12p). In metaphase spreads, the centromeric block of the i(12p)s differed in size as compared with those of normal chromosomes 12. This was rarely noted in interphase nuclei. A decrease in size of the centromeric block in 2102Ep and H12.1 caused a weak FISH signal, which was difficult to detect, especially in interphase nuclei. The ratio between p alpha 12H8- and YAC#5-derived signals reflected the presence or absence of one or more i(12p)s. Our results indicate that double FISH with a centromere- and a 12p-specific probe can be used to detect 12p overrepresentation [including i(12p)] in TGCT both in metaphase spreads and interphase nuclei. CGH confirmed the relative overrepresentation of 12p sequences as detected by FISH and showed that in these cell lines the complete 12p was involved.
KW - Adult
KW - Chromosome Aberrations
KW - Chromosomes, Human, Pair 12
KW - Humans
KW - In Situ Hybridization, Fluorescence/methods
KW - Karyotyping
KW - Male
KW - Neoplasms, Germ Cell and Embryonal/genetics
KW - Testicular Neoplasms/genetics
KW - Tumor Cells, Cultured
UR - https://www.scopus.com/pages/publications/0029919007
U2 - 10.1016/0165-4608(95)00233-2
DO - 10.1016/0165-4608(95)00233-2
M3 - Article
C2 - 8625271
SN - 0165-4608
VL - 87
SP - 95
EP - 102
JO - Cancer genetics and cytogenetics
JF - Cancer genetics and cytogenetics
IS - 2
ER -