Biomedical subjects

M Aubele

Publications and source records attributed to M Aubele.

At least 19 recordsLinked to original sources

PubMed2001

Mapping the chromosome 16 cadherin gene cluster to a minimal deleted region in ductal breast cancer.

The cadherin family of cell adhesion molecules has been implicated in tumor metastasis and progression. Eight family members have been mapped to the long arm of chromosome 16. Using radiation hybrid mapping, we have located six of these genes within a cluster at 16q21-q22.1. In invasive lobular carcinoma of the breast frequent LOH and accompanying mutation affect the CDH1 gene, which is a member of this chromosome 16 gene cluster. CDH1 LOH also occurs in invasive ductal carcinoma, but in the absence of gene mutation. The proximity of other cadherin genes to 16q22.1 suggests that they may be affected by LOH in invasive ductal carcinomas. Using the mapping data, microsatellite markers were selected which span regions of chromosome 16 containing the cadherin genes. In breast cancer tissues, a high rate of allelic loss was found over the gene cluster region, with CDH1 being the most frequently lost marker. In invasive ductal carcinoma a minimal deleted region was identified within part of the chromosome 16 cadherin gene cluster. This provides strong evidence for the existence of a second 16q22 suppressor gene locus within the cadherin cluster.

Base Sequence↗

PubMed2001

Specific steps in aneuploidization correlate with loss of heterozygosity of 9p21, 17p13 and 18q21 in the progression of pre-malignant laryngeal lesions.

Laryngeal squamous-cell carcinoma is often preceded by pre-malignant lesions. In this study, pre-malignant as well as malignant laryngeal lesions were analyzed using p53 immunohistochemistry and in situ hybridization for chromosomes 1, 7, 9, 17 and 18. Microsatellite analysis was performed on laser-microdissected tissue fragments with the aim of studying loss of heterozygosity (LOH) of 9p21, 17p13 and 18q21. Sequential biopsies were analyzed from a few cases to study genetic progression in more detail. The following genetic progression patterns were observed: (i) histologically normal mucosa and hyperplastic lesions without malignant progression were typically disomic for all chromosomes tested and showed no or only basal cell layer positivity for p53 and no allelic loss; (ii) hyperplastic lesions preceding dysplastic/invasive growth frequently showed trisomy for chromosome 7 and LOH of 9p21 and 17p13, and small foci within these lesions sometimes showed tetraploidization and p53 positivity; (iii) dysplastic lesions were characterized by a tetraploid chromosome content, LOH of 9p21 and 17p13 and p53 positivity; (iv) carcinoma in situ lesions and invasive laryngeal carcinomas showed a more unbalanced chromosome pattern and an additional 18q21 LOH. These results show that different steps in aneuploidization correlate with LOH of 9p21, 17p13 and 18q21 in early laryngeal carcinogenesis. These genomic changes could be of potential use in the diagnosis and prognosis of pre-malignant laryngeal lesions.

Aneuploidy↗

PubMed2001

Tissue microdissection techniques in quantitative genome and gene expression analyses.

Current advances in quantitative genome and gene expression analyses allow precise molecular genetic fingerprinting of tumor tissues. A crucial factor for the reliability of the data obtained with these refined techniques is the use of morphologically well-defined cell populations. Microdissection technology has been developed to procure pure cell populations from specific areas of tissue sections under microscopic control. This review covers techniques of tissue microdissection in the context of commonly used methods of quantitative genome and gene expression analysis. The first part of the review will summarize the technical aspects of various methods developed for tissue microdissection. In the latter part, current applications of quantitative genome and gene expression analysis techniques employed in microdissected tissue samples will be described.

Animals↗

PubMed2001

Her-2/neu gene amplification, elevated mRNA expression, and protein overexpression in the metaplasia-dysplasia-adenocarcinoma sequence of Barrett's esophagus.

SUMMARY: The importance of alterations of the Her-2/neu oncogene in the tumorigenesis of Barrett's adenocarcinoma (BCA) is discussed controversially. In the present study, we evaluated for the first time the Her-2/neu status in the metaplasia-dysplasia-adenocarcinoma sequence of BCA simultaneously at the DNA, mRNA, and protein level using resection specimens of 25 patients. The locus-specific Her-2/neu gene status was quantified by performing fluorescence in situ hybridization, and information about the ploidy status of chromosome 17 was obtained. Tissue sections from the same areas were used for quantitative RT-PCR (TaqMan RT-PCR) of laser-microdissected tumor cells and for immunohistochemistry to quantify Her-2/neu mRNA and oncoprotein expression. Her-2/neu gene amplification was observed in 35% of BCA, and all of these samples showed strong overexpression of both mRNA and oncoprotein. A polysomy 17 without Her-2/neu gene amplification was observed in 52% of BCA, showing a normal or moderately elevated mRNA expression and no or weak immunopositivity. From 13 areas of high-grade dysplasia (HGD) we found four to be amplified for the Her-2/neu locus, whereas five showed a polysomy 17. All four samples of HGD areas with Her-2/neu gene amplification displayed mRNA and strong oncoprotein overexpression; however, lower mRNA levels were seen than in the amplified BCA areas. None of the samples with low-grade dysplasia (LGD) showed a locus-specific Her-2/neu amplification, but polysomy 17 was present in four of eight cases. No changes were detected in BCA-associated intestinal metaplasia and squamous epithelium. In summary, only a locus-specific Her-2/neu gene amplification was associated with strong mRNA overexpression and strong membranous Her-2/neu immunostaining in BCA and HGD. A chromosome 17 polysomy, as found in the majority of BCA, led to no or weak mRNA overexpression and no or weak immunopositivity. In the metaplasia-dysplasia-adenocarcinoma sequence, a chromosome 17 polysomy without Her-2/neu gene amplification was already present in LGD. This may be a result of an early polyploidization, preceding the later genetic events, such as Her-2/neu gene amplification in HGD and BCA.

Adenocarcinoma↗

PubMed2001

Chromosomal changes during development and progression of prostate adenocarcinomas.

Chromosomal copy number changes were investigated in 16 prostate carcinomas, 12 prostatic intraepithelial neoplasias (PIN; 4 low-grade and 8 high-grade) adjacent to the invasive tumour areas, and 5 regional lymph node metastases. For this purpose, comparative genomic hybridization (CGH) was performed and a copy number karyotype for each histomorphological entity was created. CGH on microdissected cells from non-neoplastic glands was carried out on 3 different cases to demonstrate the reliability of the overall procedure. None of the non-neoplastic tissue samples revealed chromosome copy number changes. In PIN areas, chromosomal imbalances were detected on chromosomes 7, 8q, Xq (gains), and on 4q, 5q, 8p, 13q and 18q (losses). In the primary tumours, recurrent (at least 25% of cases) gains on chromosomes 12q and 15q, and losses on 2q, 4q, 5q, Xq, 13q and 18q became apparent. Losses on 8p and 6q as well as gains on 8q and of chromosome 7 were also detected at lower frequencies than previously reported. The pooled CGH data from the primary carcinomas revealed a novel region of chromosomal loss on 4q which is also frequently affected in other tumour entities like oesophageal adenocarcinomas and is supposed to harbour a new tumour suppressor gene. Gains on chromosome 9q and of chromosome 16 and loss on chromosome 13q were observed as common aberrations in metastases and primary tumours. These CGH results indicate an accumulation of chromosomal imbalances during the PIN-carcinoma-metastasis sequence and an early origin of tumour-specific aberrations in PIN areas.

Adenocarcinoma↗

PubMed2001

[Oncogene amplification and genetic heterogeneity in the metaplasia-dysplasia-adenocarcinoma sequence of Barrett esophagus].

AIMS: Information about numerical genomic alterations in the tumorigenesis of Barrett's adenocarcinoma (BCA) is still limited. In order to search for gene amplification and ploidy status, a series of locus-specific DNA probes and associated centromere probes was analysed in the metaplasia-dysplasia-adenocarcinoma-sequence. METHODS: Fluorescence in situ hybridisation (FISH) was performed on paraffin sections with locus-specific DNA probes for D7S486, c-myc, cyclin D1, Her-2/neu, 20q13.2 and associated chromosomes 7, 8, 11, 17 and 20. Corresponding areas of intestinal metaplasia (IM, n = 5), low grade dysplasia (LGD, n = 9), high grade dysplasia (HGD, n = 15) and BCA (n = 16) were analysed. RESULTS: Gene amplification of c-myc, Cyclin D1, Her-2/neu and 20q13.2 was observed in 15-35% of BCA. Coincident amplification of genes was also present. Polysomies for all investigated centromere probes were highly prevalent (up to 85%). Gene amplification was also demonstrated in HGD lesions. Polysomies were observed in HGD in high frequency (up to 80%). Extensive genetic heterogeneity was observed in both, BCA and HGD displaying different levels of amplification. None of the samples with LGD showed a locus-specific amplification, but polysomies for all investigated chromosomes were present in 18-48% of LGD. No changes were detected in BCA associated IM and squamous epithelium. CONCLUSIONS: Our data indicate that oncogene amplification of c-myc, cyclin D1, Her-2/neu, and 20q13.2 occurring in BCA and less frequently in HGD is a late event in the tumorigenesis. Polysomies of chromosomes 7, 8, 11, 17 and 20, which were highly prevalent in BCA and HGD occur already at the stage of LGD. This may be a result of an early polyploidization, preceding the later genetic events, such as gene amplification in HGD and BCA. The detection of shared numerical genomic changes and the detected extensive genetic heterogeneity in the metaplasia-dysplasia-carcinoma-sequence in Barrett's esophagus supports the hypothesis of a process of multiclonal expansion underlying this progression.

Adenocarcinoma↗

PubMed2000

Extensive ductal carcinoma In situ with small foci of invasive ductal carcinoma: evidence of genetic resemblance by CGH.

Although ductal carcinoma in situ (DCIS) of the breast is accepted as a potential precursor lesion for invasive ductal cancer (IDC), the critical genetic events associated with the tumor progression remain unknown. Since some extensive DCIS may show a small focus of IDC, these cases seem to be particularly suitable to investigate the primary abnormalities that determine the progression from in situ to early invasive cancer. We combined laser-microdissection with degenerative oligonucleotide-primed PCR (DOP-PCR) and comparative genomic hybridization (CGH) to detect copy number changes in 7 cases of extensive (>4 cm) DCIS with 1 small adjacent invasive focus. In 3 of the cases, single lymph node metastases (LN) were already present and were also investigated. Analysis of DCIS, IDC and LN components in the same patients revealed several consistent chromosomal changes present at all 3 sites: 1q, 7q, 8q, 16, 17, 19, 20q, 21q and 22q, the most frequent losses on 4q, 11q and 13q. DNA gain on 3p and 12q were more frequently found in IDC than in DCIS, suggesting the presence of proto-oncogenes activated during the progression to invasive cancer on these regions. Using paired analysis, resemblence of alterations found in DCIS and IDC could be quantified (odds ratio 7.0, p< or = 0.01). Gains on 6p, 10q, 14q and 15q and losses on 9p were identified in DCIS and IDC but not in LN, which may, therefore, represent early events in the carcinogenic process. Additional losses were found in the LNs on 2q, 3q, 5q, 6q, 12q and 16q. CGH results on chromosome 1 and 20 were confirmed by FISH and on chromosomal region 9p by microsatellite analyses. Our findings strongly underline the precursor status of high-grade DCIS, in which most of the chromosomal changes identified in IDC are already present. However, although the early stages of breast cancer, i.e., DCIS and the small foci of IDC were mainly characterized by DNA gains, the progression to metastatic tumor (LN) must have involved additional DNA losses on several regions.

Breast Neoplasms↗

PubMed2000

Increasing chromosome 1 copy number parallels histological progression in breast carcinogenesis.

Chromosome 1 copy number in the benign breast lesions hyperplasia and atypical duct hyperplasia (ADH) was investigated using fluorescence in situ hybridization on paraffin sections. Progression of chromosome 1 changes occurring in parallel with histological progression from normal through hyperplasia and ADH to ductal carcinoma in situ (DCIS) and invasive carcinoma was also assessed, both overall and within individual patients. The mean signal number for normal cells was 1.14, while that for hyperplasia was 1.56 and ADH was 1.5, while values for DCIS of 1.95 and invasive duct carcinoma of 1.79, were higher (P < 0.001). Six of the seven cases also showed a significant trend towards an increasing proportion of cells with greater than 2 signals per nucleus occurring with histological progression (P < 0.001). These results support the concept that benign proliferative breast disease is a biological precursor of in-situ and invasive ductal carcinoma, the early histological changes possibly indicating a field effect with further genetic changes required for the development of a malignant phenotype.

Breast Neoplasms↗

PubMed2000

Heterogeneous chromosomal aberrations in intraductal breast lesions adjacent to invasive carcinoma.

There is evidence that breast cancer is a heterogeneous disease phenotypically as well as molecular biologically. So far, heterogeneity on the molecular biological level has not been investigated in potential precursor lesions, such as ductal hyperplasia (DH) and ductal carcinoma in situ (DCIS). In this study we applied comparative genomic hybridization (CGH) to formalin-fixed, paraffin-embedded breast tissue with DH and DCIS, adjacent to invasive ductal carcinoma (IDC), to screen these potential precursor lesions for whole genomic chromosomal imbalances. Laser-microdissection was used to select pure cell populations from the sections. Isolated DNA was amplified by degenerate oligonucleotide primed PCR (DOP-PCR) and further processed for CGH analysis. Investigating multiple samples (n = 25) from four patients we found an average of 5.6 +/- 0.9 (mean +/- SEM) chromosomal imbalances already present in DH. In the twelve DCIS lesions an average of 10.8 (+/- 0.9) aberrations was identified with 14.8 (+/- 0.8) aberrations in the four adjacent IDC lesions. The increasing number of chromosomal changes in parallel with the histopathological sequence corroborate the hypothesis, that the carcinomas may have developed through a sequential progression from normal to proliferative epithelium and eventually into carcinoma. However, heterogeneous results were identified in the multiple samples per entity from the same patient, demonstrated mainly in the DCIS samples in the chromosomal regions 6p, 9p, 11q, 16p and 17q, in the DH samples by 3p, 16p and 17q. This heterogeneous findings were most pronounced within the DH and was less in the DCIS and IDC samples. The only aberration consistently found in all samples-even in all DH sample-was amplification of the 20q13 region. Our results demonstrate, that the applied combination of laser-microdissection, DOP-PCR and CGH, may serve to analyse breast carcinogenesis pathways in suitable histological material. However, so far, it is unclear how to handle heterogeneous results and these make identification of relevant changes more difficult. Setting a threshold and evaluating only those chromosomal changes which are present in a majority of samples may be one possibility. This involves however, the risk that infrequent but possibly significant aberrations may be missed. Figures on http://www.esacp.org/acp/2000/20-1/aubele. htm.

Breast Neoplasms↗

PubMed2000

Evaluation of c-erbB-2 overexpression and Her-2/neu gene copy number heterogeneity in Barrett's adenocarcinoma.

Amplification of the Her-2/neu gene is accompanied by overexpression of its cell surface receptor product, c-erbB-2 protein. To investigate the degree of intratumoural heterogeneity we applied immunohistochemistry in primary Barrett's adenocarcinoma (BCA) (n = 6) and dysplasia adjacent to the carcinoma (n = 4). In addition, fluorescence in situ hybridisation (FISH) was performed in primary BCA (n = 5) and dysplastic areas (n = 4). For an objective evaluation digital image analysis and laser scanning microscopy were used. Five of six BCA showed a marked intratumoral heterogeneous staining pattern ranging from areas in which the tumour cells were negative or faintly positive to tumour areas with a strong staining of the entire membrane. Among the two dysplastic areas also a heterogeneous staining pattern was observed. FISH analysis revealed marked heterogeneity of intratumoral gene copy number changes in all BCA showing populations with different fractions of cells with polysomy, low level amplification and high level amplification. One dysplasia showed a minor population with Her-2/neu signal clusters. In conclusion, we observed marked intratumoural heterogeneity of c-erbB-2 protein overexpression and Her-2/neu gene copy number in the majority of the primary BCA analyzed. Digital image analysis and laser scanning microscopy were helpful in quantifying the variations in protein expression and DNA copy number in individual tumour cells. The observed heterogeneity could hamper the exact diagnostic determination of the c-erbB-2 status in small biopsies and possibly influence the effectiveness of a potential c-erbB-2 targeting therapy. Figures on http://www.esacp.org/acp/2000/20-1/walch.htm+ ++.

Adenocarcinoma↗

PubMed1999

Intratumoral heterogeneity in breast carcinoma revealed by laser-microdissection and comparative genomic hybridization.

To evaluate the potential cytogenetic heterogeneity in breast carcinoma, several small cell groups (each consisting of 20 to 50 cells) were investigated within paraffin sections. By laser-microdissection, three to seven cell groups were taken per case. The DNA was amplified by degenerate oligonucleotide primed PCR (DOP-PCR), and the samples were analyzed by CGH for chromosomal gains and losses. Two ductal invasive breast carcinomas, one of them with two lymphnode metastases, were investigated. To compare the results from the small samples, CGH was also performed on DNA isolated from the tumorous regions of three to five serial sections (10(7) to 10(6) cells). The aberrations observed in the microdissected tumor samples were multiple and involved up to 14 different chromosomal or subchromosomal regions. The most frequent changes were gains on chromosomes 12q (14/20) and 20q (16/20), and loss on 13q (12/20). Some aberrations have rarely been detected (e.g., loss on 2p, gain on 8q). Comparing chromosomal imbalances in primary tumors and lymph node metastases, more consistent changes were found between the primary tumor and its corresponding metastases than between both primary tumors. The laser-microdissected samples in general showed more chromosomal aberrations than DNA isolated from several tumor sections. Our CGH results were confirmed by fluorescence in situ hybridization (FISH) for the chromosomal regions of centromere 1 and 20, and 20q13. In addition, microsatellite analyses on 31 samples confirmed our CGH findings for selected chromosome regions 2p and 11q. It can be concluded that there is a distinct intratumoral heterogeneity in primary breast tumors as well as in the corresponding lymph node metastases. The combination of microdissection and CGH enabled us to detect cytogenetic aberrations from important clones which are missed when analyzing DNA extracted from large cell numbers.

Breast Neoplasms↗

PubMed1999

20q13.2 amplification in intraductal hyperplasia adjacent to in situ and invasive ductal carcinoma of the breast.

The 20q13 region harboring recently described putative oncogenes is frequently amplified in invasive ductal carcinoma (IDC). The aim of this study was to examine the 20q13 copy number in intraduct hyperplasia (IH), atypical duct hyperplasia (ADH), and ductal carcinoma in situ (DCIS) adjacent to IDC. In 5 patients, comparative genomic hybridization (CGH) after laser microdissection revealed 20q13 amplification in four of five cases of IH, in all of three cases of IH with atypia, all five of DCIS, and all five of IDC. Fluorescence in situ hybridization (FISH) confirmed the amplification at 20q13.2 in IH in the two specimens analyzed. The amplification rate, however, was higher in DCIS and IDC. In phenotypically normal ductal epithelium normal values were found for 20q13 copy number by FISH (n=2) and CGH (n=5). Although the number of cases presented here is small, our results suggest that mutations in the 20q13.2 region in IH may be associated with accelerated proliferation and hyperplasia of the ductal epithelium. Progression to DCIS and ICD is accompanied by a further increase in the 20q13.2 copy number.

Breast Neoplasms↗

PubMed1999

Heterogeneity in breast cancer and the problem of relevance of findings.

Many attempts are made to identify critical genetic events responsible for the development and progression of breast cancer. There is increasing evidence that breast cancer is a heterogeneous disease, both, phenotypically as well as with respect to its molecular biologically. It is, therefore, extremely difficult to establish a diagnostically and prognostically relevant tumourigenesis model. Emerging new techniques such as microarrays, will provide us with a wealth of additional data over the next years. The precise sampling of tumour material in clearly defined histopathological lesions will be a prerequisite for the assignment of specific genetic alterations to defined stages of breast disease.

Breast Neoplasms↗

PubMed1998

Distinct cytogenetic alterations in squamous intraepithelial lesions of the cervix revealed by laser-assisted microdissection and comparative genomic hybridization.

BACKGROUND: It has been established that comparative genomic hybridization (CGH) on Papanicolaou-stained cervical smears can be used to identify chromosomal imbalances. METHODS: In this study, the authors identified normal and dysplastic squamous epithelial cells cytologically, eliminated surrounding bacteria or leukocytes by a ultraviolet laser microbeam under microscopic control, and scraped out the cell groups of interest by a microdissection system. In 3 cases of squamous intraepithelial lesions (SIL), a total of 9 samples of dysplastic (n = 6) and nontumorous cells (n = 3) were investigated, each of them consisting of 3-40 cells. The DNA was amplified by degenerate oligonucleotide primed PCR (DOP-PCR) and used for CGH. RESULTS: Analyses of all nontumorous cell groups resulted in fluorescence ratio profiles that showed no deviation from the normal range, confirming that no methodologic artefacts have been produced. The CGH profiles from dysplastic cells, however, showed various chromosomal imbalances affecting six to nine different chromosomes. The most frequent gains in DNA were observed on chromosomes 1p, 2q, 4, and 5, whereas losses were found on chromosomes 6q and 13q. CONCLUSIONS: The results of this study demonstrate the feasibility and reliability of CGH on microdissected cell samples of routinely processed cervical smears. To the authors' knowledge, this is the first study reporting the use of CGH on cervical routine smears. This approach offers the opportunity to investigate sequence copy number changes in small, morphologically well-defined groups of dysplastic cells. It may, therefore, serve as a cytogenetic screening test for identifying chromosomal aberrations in precancerous lesions that are associated with a high risk for progression to invasive cancer.

Chromosome Aberrations↗

PubMed1998

Genetic heterogeneity in a prostatic carcinoma and associated prostatic intraepithelial neoplasia as demonstrated by combined use of laser-microdissection, degenerate oligonucleotide primed PCR and comparative genomic hybridization.

We combined laser-assisted microdissection from H&E-stained paraffin sections, degenerated oligonucleotide-primed polymerase chain reaction (DOP-PCR), and comparative genomic hybridization (CGH) to analyse chromosomal imbalances in small tumour areas consisting of 50-100 cells. This approach was used to investigate intratumour genetic heterogeneity in a case of metastatic prostatic adenocarcinoma and chromosomal changes in areas of prostatic intraepithelial neoplasia (PIN) adjacent to the invasive tumour. In four microdissected invasive tumour areas with different histological patterns (acinar, cribriform, papillary and solid) marked intratumour heterogeneity was found by CGH. Recurrent chromosomal imbalances detected in at least two microdissected tumour areas were gains on 1p32-->p36, 2p22, 3q21, 7, 8q21-->q24, 11q12-->q13, 16p12-->p13, 17, 19 and loss on 16q23. Additional chromosomal changes were found in only one of the microdissected areas (gains on 16q21-->q23, 20q22 and losses on 8p21-->p23, 12p11-->q12, 12q21-->q26, 13q21-->q34, 16q12, and 18q22). In PIN, gains on chromosomes 8q21-->q24 and 17 were found in both samples investigated (low and high grade PIN), while gains on chromosomes 7, 11q, 12q, 16p, and 20q and losses on 2p, 8p21-->p23, 12q were found only in one PIN area. Controls to ensure reliable CGH results consisted in CGH analyses of (i) approximately 80 microdissected normal epithelial cells, which showed no aberrations after DOP-PCR and (ii) larger cell numbers (approximately 10(5) or 10(7) cells) of the primary tumour investigated without DOP-PCR and partially displaying the chromosomal imbalances (gain on 16p12-->p13, losses on 2p25, 8p21-->p23, 12p11-->p12, 12q21-->q26, 18q22) found in the small microdissected areas. Microsatellite and FISH analyses further confirmed our CGH results from microdissected cells. The combined approach of laser-assisted microdissection, DOP-PCR and CGH is suitable to identify early genetic changes in PIN and chromosomal imbalances associated with the particular histological patterns of invasive prostatic adenocarcinoma.

Adenocarcinoma↗

PubMed1997

Comparative FISH analysis of numerical chromosome 7 abnormalities in 5-micron and 15-micron paraffin-embedded tissue sections from prostatic carcinoma.

Interphase fluorescence in situ hybridization (FISH) was performed on 15-micron-thick paraffin sections from prostatic carcinomas using a chromosome 7-specific alpha-satellite DNA probe. A confocal laser scanning microscope (CLSM) was used for optical sectioning of the thick sections and reconstruction of 3D images. The number of FISH signals was determined by a gallery of optical sections evaluating only complete nuclei. To investiate the influence of section thickness and truncation and nuclei on scoring results, we compared the FISH data from 15-micron sections with signal counts obtained from 5-micron sections. The latter were evaluated by conventional fluorescence microscopy in the same tumor regions previously defined and marked on the slides. After statistical analysis of spot frequencies in tumor and non-tumorous cells (chi 2 test), we transferred the signal frequencies into a cytogenetic classification (-7, +7, polysomy 7). Based on this classification, most cases showed more than one chromosome 7 aberration type. Trisomy 7 (+7) became apparent in 15-micron thick sections in all 19 tumors, polysomy 7 (> 3 spots) in 18/19 cases, and monosomy 7 (-7) in 13/19 cases. In 5-micron sections, however, trisomy 7 and polysomy 7 were found in only 7/19 and 13/19 cases, respectively, and monosomy 7 in 7/19 cases. When comparing the classification results of tumor cells of the same tumor regions originating either from 5-micron or 15-micron sections, the following discrepancies were noted: in 15-micron sections exclusively, in 12/19 tumors, trisomy 7 was found; in 6/19 cases, polysomy 7; in 8/19 cases, monosomy 7. The high proportion of cases with tumor nuclei expressing only one hybridization signal of chromosome 7 in 15-micron sections could be confirmed as monosomy 7 in five selected cases by double-hybridization using centromere-specific probes for chromosomes 7 and 12. These results demonstrate that numerical chromosome 7 aberrations are more frequently observed in thick (15-micron) paraffin-embedded tissue sections by evaluating only complete nuclei. The use of routine sections (5-micron) for interphase cytogenetic analyses is compromised by a remarkable underestimation of the real chromosome copy numbers.

Adenocarcinoma↗

PubMed1997

Groping for quantitative digital 3-D image analysis: an approach to quantitative fluorescence in situ hybridization in thick tissue sections of prostate carcinoma.

In molecular pathology numerical chromosome aberrations have been found to be decisive for the prognosis of malignancy in tumours. The existence of such aberrations can be detected by interphase fluorescence in situ hybridization (FISH). The gain or loss of certain base sequences in the desoxyribonucleic acid (DNA) can be estimated by counting the number of FISH signals per cell nucleus. The quantitative evaluation of such events is a necessary condition for a prospective use in diagnostic pathology. To avoid occlusions of signals, the cell nucleus has to be analyzed in three dimensions. Confocal laser scanning microscopy is the means to obtain series of optical thin sections from fluorescence stained or marked material to fulfill the conditions mentioned above. A graphical user interface (GUI) to a software package for display, inspection, count and (semi-)automatic analysis of 3-D images for pathologists is outlined including the underlying methods of 3-D image interaction and segmentation developed. The preparative methods are briefly described. Main emphasis is given to the methodical questions of computer-aided analysis of large 3-D image data sets for pathologists. Several automated analysis steps can be performed for segmentation and succeeding quantification. However tumour material is in contrast to isolated or cultured cells even for visual inspection, a difficult material. For the present a fully automated digital image analysis of 3-D data is not in sight. A semi-automatic segmentation method is thus presented here.

Humans↗

PubMed1997

Canine neuroendocrine tumors of the pancreas: a study using image analysis techniques for the discrimination of metastatic versus nonmetastatic tumors.

Canine pancreatic neuroendocrine tumors were studied using different image analysis techniques (nuclear image histometry, analysis of argyrophilic proteins of nucleolar organizer regions, determination of the mouse anti-Ki 67 antigen proliferation index, and DNA densitometry) to correlate their biological behavior with objective phenotypic markers. The methods were compared to determine the best method for distinguishing between metastatic and nonmetastatic tumors. Discrimination between the two types of tumor was possible using nuclear image histometry in combination with morphometric analysis of argyrophilic proteins of nucleolar organizer regions. In contrast, the mouse anti-Ki 67 antigen proliferation index, DNA measurement, and immunohistochemical parameters revealed no significant difference between the two types of tumors.

Animals↗