Cellular consequences of small supernumerary marker chromosome derived from chromosome 12: mosaicism in daughter and father

Constitutional genomic imbalances are known to cause malformations, disabilities, neurodevelopmental delay, and dysmorphia and can lead to dysfunctions in the cell cycle. In extremely rare genetic conditions such as small supernumerary marker chromosomes (sSMC), it is important to understand the cellular consequences of this extra marker, as well the factors that contribute to their maintenance or elimination through successive cell cycles and phenotypic impact. The study of chromosomal mosaicism provides a natural model to characterize the effect of aneuploidy on genome stability and compare cells with the same genetic background and environment exposure, but differing in the presence of sSMC. Here, we report the functional characterization of different cell lines from two familial patients with mosaic sSMC derived from chromosome 12. We performed studies of proliferation dynamics, stability, and variability of these cells using fluorescent in situ hybridization (FISH), sister chromatid exchanges (SCE), and conventional staining. We also quantified the telomere-related genomic instability of sSMC cells using 3D telomeric profile analysis by quantitative-FISH. sSMC cells exhibited differences in the cell cycle dynamics compared to normal cells. First, the sSMC cells exhibited lower proliferation index and higher frequency of SCE than normal cells, associated with a higher level of chromosomal instability. Second, sSMC cells exhibited more telomeric-related genomic instability. Lastly, the differences of sSMC cells distribution among tissues could explain different phenotypic repercussions observed in patients. These results will help in our understanding of the sSMC stability, maintenance during cell cycle, and the cell cycle variables involved in the different phenotypic manifestations.

Molecular Cytogenetic Characterization of Supernumerary Marker Chromosomes by Chromosomal Microarray

PURPOSE: Supernumerary marker chromosome (SMC) could be associated with various phenotypic abnormalities based on the chromosomal origin of SMCs. The present study aimed to determine the genomic contents of SMCs using chromosomal microarray and to analyze molecular cytogenetic characterizations and clinical phenotypes in patients with SMCs. MATERIALS AND METHODS: Among patients with SMCs detected in routine chromosomal analysis, SMCs originating from chromosome 15 were excluded from the present study. CGH-based oligonucleotide chromosomal microarray was performed in 4 patients. RESULTS: The chromosomal origins of SMCs were identified in 3 patients. Case 1 had a SMC of 16.1 Mb in 1q21.1-q23.3. Case 2 showed 21 Mb gain in 19p13.11-q13.12. Case 3 had a 4.5 Mb-sized SMC rearranged from 2 regions of 2.5 Mb in 22q11.1-q11.21 and 2.0 Mb in 22q11.22-q11.23. CONCLUSION: Case 1 presented a wide range of phenotypic abnormalities including the phenotype of 1q21.1 duplication syndrome. In case 2, Asperger-like symptoms are apparently related to 19p12-q13.11, hearing problems and strabismus to 19p13.11 and other features to 19q13.12. Compared with cat-eye syndrome type I and 22q11.2 microduplication syndrome, anal atresia in case 3 is likely related to 22q11.1-q11.21 while other features are related to 22q11.22-q11.23. Analyzing SMCs using high-resolution chromosomal microarray can help identify specific gene contents and to offer proper genetic counseling by determining genotype-phenotype correlations.

Identification of marker chromosomes by reverse painting fluorescence in situ hybridization and comparative genomic hybridization / 대한산부인과학회지

OBJECTIVE: Although marker chromosome is defined as an abnormal chromosome in which no part can be identified, derivative chromosomes with structural abnormalities of unknown origin are also called as marker chromosomes conventionally. The clinical significance of a marker chromosome is determined according to the origin of marker chromosome. In this study reverse painting fluorescence in situ hybridization (FISH), and comparative genomic hybridization (CGH) methods were employed to elucidate the origin of marker chromosomes in 5 clinical cases. METHODS: Reverse painting probes were generated from five copies of each marker chromosomes microdissected with micromanipulator, amplified with DOP-PCR, and labeled with fluorochromes. The probes were hybridized to normal metaphases. For CGH, normal control and patients' DNA were directly labeled with spectrum-red-dUTP and spectrum-green-dUTP by CGH nick translation kit, and hybridized to normal reference metaphases. The CGH images were captured with a computer controlled fluorescence microscope equipped with a CCD camera and analyzed by Cytovision workstation. RESULTS: Five marker chromosomes were identified as follows (1) derivative chromosome 15 inducing partial trisomy of 15pter->q21, (2) isochromosome of 18p causing 18p tetrasomy, (3) short arm of chromosome 5 causing 5p trisomy (4) small accessory chromosome originated from centromeric region of chromosome Xq11->q12 (5) der(17) with inverted duplication of the short arm of chromosome 17. In all cases the origin of each marker chromosomes were identified successfully with reverse painting FISH, and these results were concordant with the CGH profiles. CONCLUSION: Our results indicate that combined reverse painting FISH and CGH is a rapid, convinient and powerful tool to identify the origin of marker chromosomes and derivative chromosomes caused by various chromosome abnormalities such as translocation, duplication, deletion.

Establishment and Characterization of Chromosome Aberrations in Hman Colangiocarcinoma Cell Line, PCK1 / 대한해부학회지

The analysis of recurring chromosome aberrations has become an integral part of the diagnostic and prognostic workup of many human cancers, and their molecular analyses have facilitated the identification of genes related to the pathogenesis of cancer. Cholangiocarcinoma (CC), a malignant neoplasm of the biliary epithelium, is usually fatal because of the difficulty in early diagnosis and unavailability of effective therapy. Furthermore, little is known about the genetics and biology of CC. Only few reports concerning cytogenetic studies of CC have been published and few cell lines have been established. We recently established CC cell line, designated as PCK1. The purpose of this study is to establish in detail karyotype of PCK1 cell line. The origins of the unidentified marker chromosomes were analyzed by G-banding, cross species color banding (RxFISH), and human chromosome-specific painting. In PCK1 cell line, gains involved chromosomes and chromosome regions, 4, 5, 9, 12, 16, 21, 1q, 7q11-q22, 8q, 12p, 14q11-q22, 15q21-qter, 17p11-qter, and 18p. Losses involved Y, 7q31-qter, 8p, 14q23-qter, 17p12-pter, and 18q. Established PCK1 cell line will be able to use the basic research of cholangiocarcinoma and the abnormal chromo-somes may be the candidate regions for isolation of the genes related to CC.