[Joint tumors: rare but important differential diagnoses of malignant and benign tumors as well as pseudotumors in rheumatology]. / Gelenktumoren: Seltene, aber wichtige Differenzialdiagnosen von malignen und benignen Tumoren sowie Pseudotumoren in der Rheumatologie.

This review article elucidates the differential diagnostics of malignant and benign joint tumors, pseudotumors of the joints and the peri-implant tissue, which are rare but important entities in rheumatology and orthopedic rheumatology. The tissue of origin includes the synovium, peri-implant tissue, peri-articular fibrous tissue and peri-articular osseous tissue. Pseudotumors can be viewed as independent but heterogeneous entities. These are essentially manifested as tumor-like depositions of crystals, calcareous deposits, vascular malformations, ectasia of the synovia and joint capsule tissue and pseudocysts. Other causes for pseudotumors are focal destructive inflammation (e.g. induced by foreign bodies), high grade synovitis and focal fibrinoid necrosis (i.e. rheumatoid nodules). Methodologically, these diagnostics are based on conventional standard staining methods, immunohistochemical analyses of formalin-fixed and paraffin-embedded materials and on molecular diagnostic procedures. The latter are of great importance in cases of benign and malignant joint tumors. The most important immunohistochemical markers with respect to joint tumors are S100, SM-actin, CD68, CD34, STAT6, clusterin, Muc­4, beta-catenin and MDM2-FISH. The following markers are recommended for the differential diagnostics and typing of periarticular tumor metastases in the pathology of rheumatic diseases: AE1/AE3, CK8, p63, TTF­1, TGB, PSA, androgen receptor, GATA, CD56, chromogranin, CDX­2, SAT-B2, SALL4, estrogen and progesterone receptors, CD45LCA, CD30, CD79a and S100. Necrosis, inflammatory infiltrations and reparative inflammatory changes may complicate the histopathological classification. Therefore, a correlation with clinical, microbiological and radiological data in the sense of interdisciplinary synergistic diagnostics may be required.

[Molecular pathological diagnostics of infections in orthopedic pathology]. / Molekularpathologische Infektionsdiagnostik in der orthopädischen Pathologie.

The diagnosis of infections in patients with arthritis and/or in joint prostheses requires interdisciplinary cooperation and the application of up-to-date methods. The histological investigation of the synovial membrane allows the differentiation of acute, chronic and granulomatous synovialitis. Detection of conserved regions of the microbial genome by PCR, especially 16S rRNA for bacteria and 18S rRNA for fungi, is a broad approach for the classification of pathogens which cannot be cultured. Acute infectious arthritis and periprosthetic infections share the spectrum of pathogens with sepsis, therefore multiplex PCR-based methods for the detection of sepsis can be employed. Molecular diagnostics can detect minimal infections in periprosthetic tissues even after antibiotic therapy. The anamnesis (enteral or urogenital infection), clinical picture (oligoarthritis) and further parameters (e.g. HLA B27 status) are important for the diagnosis of reactive arthritis. In many cases of reactive arthritis, molecular methods allow the detection of bacterial DNA or RNA in synovial fluid or tissue samples. The low sensitivity of histopathological methods may be compensated by application of PCR techniques, especially in the differential diagnosis of granulomatous synovitis including mycobacterial infections. Molecular methods can be used to support the differential diagnosis of septic and reactive arthritis. MicroRNA techniques combined with PCR for detection of pathogens support the differential diagnosis of rheumatoid arthritis with severe inflammatory activity compared to infectious arthritis. Proteomic methods could expand the methodological spectrum for the diagnosis of infections.

[Applications of molecular pathology in the diagnosis of joint infections]. / Molekularpathologische Diagnostik von Gelenkinfektionen.

The diagnosis of infections in patients with arthritis and/or joint prostheses requires interdisciplinary cooperation and the use of up-to-date methods. Massive bacterial infection can be identified by bacterial culture, and minimal infection can be detected by molecular pathological methods. These processes include specific enrichment of bacterial and fungal DNA, amplification, and identification of the DNA by gel electrophoresis, sequencing techniques, and chip technologies.Anamnesis (enteral or urogenital infection), the clinical picture (oligoarthritis), and further parameters (e.g., HLA B27 status) are important for the diagnosis of reactive arthritis. In many cases of reactive arthritis, molecular methods allow detection of bacterial DNA or RNA in synovial fluid or tissue. Molecular pathological methods allow the fast and reliable differential diagnosis of granulomatous synovialitis without prior cultivation of bacteria or fungi. The development of new molecular pathological methods for detecting bacterial and fungal nucleic acids will increase diagnostic accuracy.

Bronchioloalveolar carcinoma: a new cancer in Peutz-Jeghers syndrome.

Besides gastrointestinal hamartomatous polyposis and melanin spots in the skin and mucosa, patients with the Peutz-Jeghers syndrome (PJS) have repeatedly been observed with a variety of tumours, including lung cancer. Available data indicate an increased cancer risk among PJS patients, which suggests that the gene involved in PJS, STK11 on chromosome 19p13.3, may be a tumour suppressor gene. Herein, bronchioloalveolar carcinoma (BAC) of mucinous type is reported in a 22-year old male PJS patient with a novel germline frameshift insertion in exon 2 at codon 118 of the STK11 gene. Molecular studies of his BAC indicated loss of heterozygosity (LOH) in the region of STK11 on chromosome 19p13.3. This observation supports the hypothesis that STK11 is a tumour suppressor gene which is involved in the development of lung adenocarcinoma.

[Nuclear localization of heterochromatic regions varies in hyperplastic and preneoplastic lesions of the breast]. / Unterschiedliche intranukleäre Lokalisation von Heterochromatinregionen in hyperplastischen und neoplastischen Läsionen der Mamma.

Previous molecular cytogenetic studies in breast cancer revealed numerous chromosomal changes and identified alterations involving the chromosomes 1 and 16 as early incidents in mammary carcinogenesis. Since both chromosomes reveal pericentromeric heterochromatic areas, these chromosomal alterations might result from instable heterochromatin caused by DNA hypomethylation. In the present study, we investigated whether hyperplastic and neoplastic lesions of the breast differ regarding the distance between the heterochromatic areas of chromosomes 1 and 16 within the nuclei. We hybridized differently fluorescence-labeled DNA samples specific for the heterochromatic regions of chromosomes 1 and 16 to formalin-fixed tissue sections. Histological classification of the lesions was supported by immunohistochemical staining using cytokeratin-specific antibodies. The methylation state of the heterochromatic regions was tested by staining with an antibody specific for methylated cytidin. Our results revealed an increased frequency of paired intranuclear signals specific for chromosomes 1 and 16 in neoplastic lesions (atypical ductal hyperplasia, ductal carcinoma in situ) compared to ductal hyperplasia and normal glandular epithelium. Staining with the methylation-specific antibody reavealed a weaker staining in neoplastic lesions compared to hyperplastic lesions and normal cells. We conclude that atypic ductal hyperplasia represents the histomorphological equivalent for the beginning of tumor genome evolution that progresses in ductal carcinoma in situ and infiltrating carcinoma.

Protein and mRNA expression of uPAR and PAI-1 in myoepithelial cells of early breast cancer lesions and normal breast tissue.

Myoepithelial cells (MEs), which surround ducts and acini of the breast glands, exhibit an anti-invasive phenotype and form a natural border separating proliferating tumour cells of ductal carcinoma in situ (DCIS) from basement membrane (bm) and underlying stroma. Invasion requires penetration of these host cellular and extracellular matrix barriers. This destruction is caused by proteolytic activity of tumour cells and host bystander cells. There is substantial evidence that high concentrations of the urokinase plasminogen-activating system are conducive to tumour cell spread and metastasis. Prompted by the conspicuous absence of studies examining the role of the ME in breast cancer progression, we studied the expression of the urokinase plasminogen activator receptor (uPAR) and plasminogen activator inhibitor type-1 (PAI-1) in MEs of 60 DCIS samples. Our results show that nearly all MEs of DCIS and normal breast glands exhibit the uPAR antigen, whereas the PAI-1 antigen was mainly expressed in MEs of high-grade DCIS. In one intermediate DCIS numerous ducts showed an incomplete myoepithelial layer expressing uPAR and PAI-1. We conclude that uPAR in MEs may be necessary to attach them to the bm by uPAR/vitronectin (Vn) interaction. The strong expression of PAI-1, which is known to resolve the uPAR/Vn binding, may be involved in the detachment of MEs of DCIS. Although the role of PAI-1 acting as cell detachment factor could not be demonstrated in our study, we speculate that the loss of the anti-invasive ME layer in DCIS may be triggered by PAI-1 and could be an early sign of subsequent tumour cell infiltration.

Variation in performance of the tree fern Cyathea caracasana (Cyatheaceae) across a successional mosaic in an Andean cloud forest.

In Andean forests, Cyathea caracasana grows across a range of successional habitats. This study documents variation in several measures of plant performance (stem growth, leaf production, leaf longevity, and spore production) in C. caracasana growing in open habitat, low-canopy secondary forest, and high-canopy secondary forest, based on 33 mo of observation. In open habitat, C. caracasana displayed significantly higher growth rates, leaf production rates, and leaf turnover than in either of the forested habitats. The highest rates of spore production were also observed in open-habitat individuals, with only one plant in the forest understory producing spores during the study. Despite low growth and no reproduction, I observed no mortality among ferns in the forest understory. These data show that C. caracasana performs best under conditions of full sun but can persist under the closed canopy. This suggests a life history in which periods of rapid growth, spore production, and recruitment in forest gaps alternate with low growth rate and persistence in the understory. A phylogenetic perspective suggests that the habitat flexibility, which might conventionally mark C. caracasana as a habitat generalist, is better interpreted as specialization for the dynamic forest in which it grows.

Identification of frequent chromosomal aberrations in ductal adenocarcinoma of the pancreas by comparative genomic hybridization (CGH).

Despite the continuous progress in molecular methodology, the genetic events involved in the initiation and progression of ductal adenocarcinoma of the pancreas remain largely unknown. In this study, 33 pancreatic ductal adenocarcinomas were screened for genomic alterations by comparative genomic hybridization (CGH). To date, most CGH studies of pancreatic cancer have been based on cell lines. To emphasize genetic imbalances that are involved in the in vivo development and progression of pancreatic carcinoma only fresh-frozen or paraffin-embedded tumour samples were analysed in the present study. Twenty-two tumours (67%) showed genomic alterations involving up to three (12%) or more (55%) chromosomal regions. The number and nature of the genetic imbalances did not, however, correlate with tumour stage or grade. Chromosome 18 was preferentially altered in the tumours analysed. Frequent chromosomal losses were found at 18q, 10q, 8p, and 13q. Commonly gained regions were located on 8q and 3q. Moreover, high copy number amplifications of the chromosomal regions 5p, 8q22-ter, 12p12-cen, 19q12-13.2, and 20q were identified. These data provide evidence for the occurrence of characteristic genomic alterations which are of biological relevance for the genesis of pancreatic cancer. The identified altered chromosomal regions may harbour tumour genes which involved in the multistep process of pancreatic carcinogenesis.

Comparative genomic hybridization (CGH) discloses chromosomal and subchromosomal copy number changes in Merkel cell carcinomas.

We analyzed three Merkel cell carcinomas (MCC), applying comparative genomic hybridization (CGH) with DNA from paraffin-embedded and cultured tumor material as the probes. By this method, numerous changes in chromosome copy numbers were observed in each tumor investigated. Recurrent gains of chromosomes 1, 6, 18q and 20 were detected in two tumors. A third tumor showed complex chromosomal copy number changes, including gain of chromosome 8 and 9. These gains, as well as gain of chromosome 1 in the first two tumors, were confirmed by fluorescence in situ hybridization to paraffin tissue sections. Our results support the view that important genes for MCC development may be located on chromosomes 1, 6, 18q and 20.

Detection of complex genetic alterations in human glioblastoma multiforme using comparative genomic hybridization.

The aim of the present study was to detect complex genetic alterations in human glioblastoma multiforme (GBM) by comparative genomic in situ hybridization (CGH). Of the 24 GBM that were examined, increased fluorescence intensities indicating chromosomal polysomy of chromosome 7 and gene amplification at chromosome 7p were found in 42% of the tumors. In addition, signal enhancement of chromosome 19 was present in 29% and at 12q13-15 in 21% of the tumors. We also detected reduction of fluorescence intensities indicating gross deletions on chromosomes 10 (58%), 9p (46%), and 13 (29%). There was a close correlation of CGH results when compared with Southern analysis of the EGFR gene localized on chromosome 7 and loss of heterozygosity detection of chromosome 9 and 10 by microsatellite PCR. A close correlation was also observed between copy number changes of chromosome 7 and deletions of chromosome 10. Amplification of chromosome 12q and deletions of chromosomes 9p and 13 seemed to be complementary in the tumors investigated in the present study.

[Detection of amplified DNA sequences by comparative genomic in situ hybridization with human glioma tumor DNA as probe]. / Nachweis amplifizierter DNA-Sequenzen mittels Komparativer Genomischer in situ Hybridisierung und Tumor-DNA humaner gliome als Probe.

Comparative genomic hybridization (CGH) provides a new possibility for the investigation of genetic alterations in tumour genomes. In our experiments CGH was carried out using genomic DNA from human glioblastoma multiforme (GBM) as a probe for chromosomal in situ suppression hybridization. Amplified DNA sequences contained in the tumour DNA showed specific signals, revealing the chromosomal positions of these sequences. Using this approach we detected amplifications of different chromosomal segments in individual GBM specimens. In accordance with the results from Southern analysis demonstrating amplification of the EGFR gene in 45% of human GBM, CGH signals in different GBM mapped to the region of this gene on chromosome 7p. Other signals detected by CGH involved chromosome 12q and 8q. Our data demonstrate CGH as a novel comprehensive and rapid approach for the analysis of complex genomic alterations in glial tumours.