[Principles of intervertebral disc assessment in private accident insurance]. / Grundlagen der Bandscheibenbegutachtung in der privaten Unfallversicherung (I).

Due to the spread of intervertebral disc degeneration, insurance companies and experts are regularly confronted with related assessments of insured persons under their private accident insurance. These claims pose a particular challenge for experts, since, in addition to the clinical assessment of the facts, extensive knowledge of general accident insurance conditions, case law and current study findings is required. Each case can only be properly assessed through simultaneous consideration of both the medical and legal facts. These guidelines serve as the basis for experts and claims.managers with respect to the appropriate individual factual assessment of intervertebral disc degeneration in private accident insurance.

Deregulated expression of p16INK4a and p53 pathway members in benign and malignant myoepithelial tumours of the salivary glands.

AIMS: Myoepithelial salivary gland tumours are uncommon and follow an unpredictable biological course. The aim was to examine their molecular background to acquire a better understanding of their clinical behaviour. METHODS AND RESULTS: Expression of protein (E2F1, p16(INK4a), p53, cyclin D1, Ki67 and Polycomb group proteins BMI-1, MEL-18 and EZH2) was investigated in 49 benign and 30 primary malignant myoepithelial tumours and five histologically benign recurrences by immunohistochemistry and the findings correlated with histopathological characteristics. Benign tumours showed a higher percentage of cells with expression of p16(INK4a) pathway members [p16(INK4a) and E2F1 (both P < 0.001), and cyclin D1, P = 0.002] compared with normal salivary gland. Furthermore, malignant tumours expressed p53 (P = 0.003) and EZH2 (P = 0.09) in a higher percentage. Recurrences displayed more p53 + tumour cells (P = 0.02) than benign primaries. Amongst the benign tumours, the clear cell type had the highest proliferation fraction (P = 0.05) and a higher percentage of EZH2 was detected in the plasmacytoid cell type (P = 0.002). CONCLUSIONS: This study is the first to demonstrate that deregulation of the p16(INK4a) senescence pathway is involved in the development of myoepithelial tumours. We propose that additional inactivation of p53 in malignant primaries and benign recurrences contributes to myoepithelial neoplastic transformation and aggressive tumour growth.

[Significance of molecular-cytogenetic findings in mucoepidermoid carcinoma as an example of salivary gland tumors]. / Bedeutung molekular-zytogenetischer Befunde bei Speicheldrüsentumoren am Beispiel des Mukoepidermoidkarzinoms.

Chromosome translocations in tumors frequently give rise to fusion genes encoding proteins with oncogenic activities. Mucoepidermoid carcinomas (MEC) are characterized by a t(11;19)(q21-22;p13) translocation found in approximately 60% of the tumors. This t(11;19) translocation results in a fusion gene consisting of exon 1 of the MECT 1 gene and exons 2-5 of the MAML 2 gene. As a result of the t(11;19) a fusion protein is generated which, independent of NOTCH-ligands, activates the transcription of the NOTCH target gene HES 1. The altered function of MAML 2 causes a disruption of NOTCH signalling which suggests a novel mechanism of tumorigenesis. Pending the elucidation of the t(11;19) at the molecular level of an apparently identical chromosomal translocation in Warthin's tumor, the identification of the translocation in MEC by FISH- and/or RT-PCR-analyses may become important in diagnosis and might have prognostic relevance. Warthin's tumors are benign salivary gland neoplasms with a distinctive histomorphology and histogenesis completely different from MEC.

[Immunohistochemical characterization of salivary gland tumors with tissue micro-arrays]. / Immunphänotypische Charakterisierung von Speicheldrüsentumoren unter Verwendung von Gewebe-Micro-Arrays.

Systematic analysis of gene expression in salivary gland tumors is necessary to identify genes associated with specific tumor types. From the salivary gland register of the University Hospital Hamburg-Eppendorf sufficient samples of various tumors were available to generate Tissue Micro-Arrays (TMA). In light of the considerable heterogeneity of salivary gland tumors, this study was aimed at evaluating the suitability of TMA in salivary gland diagnostics and research. Epithelial antigens are not sufficient for a tumor-type-specific characterization. Myoepithelial markers are suitable for distinguishing biphasic tumor types from purely epithelial tumors. The detection of amylase in acinic cell carcinomas, and the detection of steroid hormone receptors in these and other malignant salivary gland tumors particularly in combination with the expression of transcription factors, oncogenes and proliferation associated antigens result in characteristic expression profiles. These may prove to be valuable for further investigations, especially on the molecular level.

Translocations t(11;18)(q21;q21) and t(14;18)(q32;q21) are the main chromosomal abnormalities involving MLT/MALT1 in MALT lymphomas.

The recently discovered MLT/MALT1 gene is fused with the API2 gene in the t(11;18)(q21;q21), which characterizes about one-third of MALT lymphomas. In order to screen for variant translocations and amplifications of MLT/MALT1, we have developed a novel, undirected two-color interphase fluorescence in situ hybridization (FISH) assay with two PAC clones flanking MLT/MALT1. This assay was applied to 108 marginal zone B-cell lymphomas (MZBCLs), including 72 extranodal MALT lymphomas, 17 nodal, and 19 splenic MZBCL. In 19 MALT lymphomas (26%), but in none of the nodal or splenic MZBCL, separated hybridization signals of the MLT/MALT1 flanking probes, were found. Further FISH analyses showed that 12 of these 19 cases displayed the classical t(11;18) and the remaining seven cases revealed the novel t(14;18)(q32;q21), involving the MLT/MALT1 and IGH genes. The frequency at which these translocations occurred varied significantly with the primary location of disease. The t(11;18) was mainly detected in gastrointestinal MALT lymphomas, whereas the t(14;18) occurred in MALT lymphomas of the parotid gland and the conjunctiva. Amplification of MLT/MALT1 was not observed in any of the lymphomas analyzed. We conclude that the translocations t(11;18)(q21;q21) and t(14;18)(q21;q32) represent the main structural aberrations involving MLT/MALT1 in MALT lymphomas, whereas true amplifications of MLT/MALT1 occur rarely in MZBCL.

Expression and distribution of basement membrane proteins in rat larynx and trachea following irradiation.

OBJECTIVE: Basement membranes-(BM) influence polarization, differentiation, migration and proliferation of cell and play an important role in maintaining structural and functional tissue integrity. While BM alterations have been reported in various lesions (e.g. inflammation, tumors) of laryngeal-tracheal tissues, reports on radiogenic BM alterations are rare. External irradiation (IRR) of advanced head and neck tumors often includes "normal tissues" (tissues without cancer) of the larynx. In these normal tissues both single-cell damage (necrosis, apoptosis, functional cell death) and interstitial damage (edema, fibrosis, vascular alterations, cellular infiltrations) resulting in tissue remodeling can occur, depending on various IRR parameters. In this study, we set out to add to our knowledge on the phenotypic characterization of the radiogenic BM expression pattern in laryngo-tracheal tissues. MATERIALS AND METHODS: In 63 laryngo-tracheal specimens from Wistar rats, we investigated the laminin (LA) and collagen IV (CIV) expression profile and distribution pattern depending on the IRR dose (fractionated IRR, 2 Gy/day, up to a total dose of 20, 40, or 60 Gy), the time since IRR (6 months vs 12 months) and animal age (1 year vs 1.5 years) using immunohistochemical methods, semiquantitative assessment, and multivariate analysis. RESULTS: In specimens irradiated with more than 20 Gy, both BM constituents predominantly showed dose-dependent increases and sometimes fluctuations in staining at slight to moderate levels. The expression differed in frequency and level among the various tissue structures. In some structures there was decreased expression. In the vocalis muscle, laryngeal and esophageal nerve endings, recurrent laryngeal nerve and laryngeal and tracheal muscles, LA was detected at levels significantly stronger than in controls. BM surrounding gland structures, nerve endings of the piriform sinus and esophageal muscles showed a marked tendency towards increased LA expression. However, the BM underlying the mucosal layer of the supra- and subglottic region revealed decreasing LA immunoreaction up to 40 Gy IRR, but a distinct increase in expression at 60 Gy. In the esophageal and tracheal muscles, tracheal perichondrium, recurrent laryngeal nerve and capillaries, CIV was detected at significantly stronger levels than in the controls. The vocal ligament exhibited positive CIV immunoreactions adjacent to interstitial and infiltrate cells and CIV-positive BM condensations, resulting in increased staining of these structures. CIV reactions of laryngeal and hypopharyngeal nerve endings tended towards increased expression. In contrast, BM staining surrounding vocal muscle cells revealed significantly decreased expression. In addition, there was a tendency towards decreased expression for supraglottic, subglottic and hypopharyngeal muscle cells. Age and time since irradiation had no significant effect on staining. CONCLUSION: The BM constituents laminin and collagen IV showed prominent dose-dependent increases and sometimes fluctuations in expression. This expression pattern persisted up to one year after completion of the irradiation. Thus, these findings must be related to late radiation effects. The altered BM expression may play a role, at least in part, in structural (e.g. laryngeal edema) and functional (voice disorders) changes associated with irradiation of the head and neck area.

The expression pattern of collagen I in irradiated mandibular salivary glands of rats.

UNLABELLED: Irradiation damage to salivary glands leads to loss of function and fibrosis. Immunohistochemical analysis of extracellular matrix proteins might give a more precise insight into the irradiation damage of glands. Collagen I (C-I) is a major component of the extracellular space. The aim of this study was to analyse the effects of irradiation on the distribution pattern of C-I in the salivary glands of rats. MATERIALS AND METHODS: Sixty female Wistar rats were fractionated irradiated up to 60 Gy (left side of the neck; 2 Gy/d, 5d/week; total dosage either 20, 40 or 60 Gy). The glands were explanted after 6 or 12 months following supravital anesthesia, the shielded right gland serving as internal control. C-I was detected immunohistochemically. RESULTS: In non-irradiated animals the immunoreaction was mainly homogeneous and slight around the ductal epithelia, in the area of the capsule and septae and the peri- and epineurium of nerves. A statistically significant difference was identified in the irradiated rats vs control animals and comparing in-the-radiation field (left side) vs outside-the-radiation field (right side) situated glands. Multivariate analysis revealed a statistically significant increase in staining of irradiated rats concerning the excretory ducts, the area of the capsule and septae, the nerves and striated ducts and adventitia of vessels [p = 0.0001]. The increase of immunoreaction in irradiated glands started above 20 Gy total dosage and was at its maximum after 60 Gy. However, the expression profile was inhomogenous following 20 Gy exposure and did not differ statistically from glands of control animals. Neither the age of the animals nor the latency period following exposure to the radiation source yielded a statistically significant effect on the immunoreaction. CONCLUSION: Studies on irradiation damage to the salivary glands require a more detailed description of the proteins accumulating in the extracellular space, thereby forming the so-called "fibrosis". These accumulations of proteins, e.g. C-I, may both support apoptosis and support a hypoxic environment giving rise to transformed cells.

[Involvement of the maxillary sinus in Langerhans cell histiocytosis]. / Kieferhöhlenbeteiligungen im Rahmen der Langers-Zell-Histiozytose.

AIM: A clinical presentation of Langerhans cell histiocytosis (LCH) in the maxillary sinus of two patients is given. LCH in the maxillary sinus is a rare occurrence. Our aim was to compare the different treatment alternatives available and to suggest a classification as well as a therapeutic regime. PATIENTS AND METHOD: Records and clinical data of two patients treated between 1994 and 2001 were retrospectively evaluated. Both patients suffered from LCH in the maxillary sinus and the maxilla regions. Only surgical treatment was used. After resection, a large defect of the maxillary sinus, which did not allowing coverage, was seen in both cases. After reconstructive operations, closure was finally achieved. Both patients underwent follow-ups, whereby one suffered from a relapse after 15 months. RESULTS: Although one of the patients under investigation showed a recurrence of LCH, we are of the opinion that surgical treatment is very effective in the elimination of this condition. A proposal for a classification of LCH in the oral-maxillo-facial-region is made. CONCLUSIONS: The evaluation of our clinical study suggests that LCH is a disease that should be treated surgically. Only in very severe cases should the surgical treatment be complimented by either radiotherapy or chemotherapy. In disseminated cases, especially chemotherapy seems to improve the outcome. Surgery offers the possibility of eliminating systemic side effects.

[Experimental and histomorphologic studies of healing processes of anastomoses of the carotid artery in the Wistar rat animal model with immunohistochemical imaging of collagen IV]. / Experimentelle und histomorphologische Untersuchungen zu Heilungsvorgängen an Anastomosen der A. carotis am Tiermodell der Wistar-Ratte mit immunhistochemischer Darstellung von Kollagen IV.

INTRODUCTION: The main risk in free flap tissue transfer is posed by the microvascular anastomosis. The dangers confronting the anastomosis are thrombosis, aneurysm, and vascular insufficiency. Collagen IV is associated with the basement membrane complex. It plays an essential role in vascular tissue organization and supports mechanical properties of vessels. We used a long-term animal experiment to gain a new understanding of histomorphological apposition of anastomosis and distribution of collagen IV. METHODS: Seventy Wistar rats, seven groups of ten animals each, were operated. A 4-mm long segment of the common carotid artery was isolated and reinserted. After a varying length of time (between directly after the operation and 6 months later), the common carotid arteries including bifurcation were isolated after cardiovascular perfusion. Carotid arteries were embedded, and cross-sections were stained with hematoxylin and eosin, Verhoeff's tissue elastin stain, and immunohistochemical anti-collagen IV antibody. By using the above-described technique in each section four anastomoses were examined. The histomorphology and intensity of the anti-collagen IV stainings were evaluated. RESULTS: Compression, shift, and dehiscence as forms of vessel apposition were often seen. We observed an overexpression of collagen IV in the media in cases of compression and shift. The grade of expression of collagen IV in the anastomosis depends on the extent of the injury. CONCLUSIONS: A well-performed microvascular anastomosis is clinically important not only for the acute phase after the operation, but also for the long-term course. The application of antibodies to identify the collagen IV distribution is valuable for studying vascular healing. Further applications could be used in follow-up studies of vascular prostheses.

Angiogenesis, vascular endothelial growth factor and platelet-derived growth factor-BB expression, iron deposition, and oxidation-specific epitopes in stented human coronary arteries.

Pathogenesis of in-stent restenosis remains poorly understood because information from human histopathologic studies is scarce. We used an improved saw-grinding and cutting method on methacrylate-embedded samples containing metal stents, which allows in situ hybridization and immunohistochemical analysis of in-stent restenosis. Twenty-one samples were collected 3 hours to 3 years after stenting from 6 patients aged 36 to 81 years. Except in very early samples collected within hours after the stent deployment, neovascularization was present in all segments studied. At advanced stages, extensive neovascularization was located mainly at the luminal side of the stent struts and was only rarely accompanied by inflammatory cells. The neovessels colocalized with vascular endothelial growth factor (VEGF)-A mRNA and protein expression as well as with iron deposits and oxidation-specific epitopes, which imply the presence of chronic oxidative stress. VEGF-A expression was detected in the same areas containing macrophages, endothelial cells, and, to a lesser extent, smooth muscle cells, which also showed platelet-derived growth factor-BB expression. We conclude that in-stent restenosis features neovascularization, VEGF-A and platelet-derived growth factor-BB expression, and iron deposition, which is most probably derived from microhemorrhages. These mechanisms may play an important role in the development of neointimal thickening and could provide useful targets for the prevention and treatment of in-stent restenosis.

[About the prognostic value of Her-2 gene-amplification and cell-proliferation in salivary duct carcinoma of the major salivary glands - a pilot-study]. / Pilotstudie zur prognostischen Wertigkeit der Her-2 Genamplifikation und Proliferationsaktivität beim Speichelgangkarzinom der grossen Kopfspeicheldrüsen1.

INTRODUCTION: The salivary duct carcinoma (sdc) represents a rare variant of the group of adeno-carcinomas of the salivary glands. Histopathologically, it is marked by solid and cribriform cell nests with central necrosis, displaying distinct similarity with the ductal carcinoma of the breast, where prognosis can be correlated with Her-2 gene-amplification. Based on this histopathological similarity, the prognostic value of Her-2 gene amplification in SDC was examined in the presented pilot-study. PATIENTS AND METHODS: Four own patients with different clinical courses were examined in regard to their histopathological features, Her-2 gene-amplification and proliferation (Ki67). RESULTS: Three of the four patients died tumor related 2.4, 5.5 and 8.2 years after initial diagnosis. The remaining patient died tumor-free 6 year after diagnosis (myocardial infarct). The two patients with an early recurrent disease and distant metastasis showed a high Her-2 expression and proliferation (Ki67), compared to the other two patients. CONCLUSION: In the presented pilot-study a distinct correlation between Her2-gene-amplification, proliferation (Ki67) and clinical course could be observed. Additional analysis to evaluate this aspect seems rectified, especially under recognition of therapy decisions.

Oxidized implants and their influence on the bone response.

Surface oxide properties are regarded to be of great importance in establishing successful osseointegration of titanium implants. Despite a large number of theoretical questions on the precise role of oxide properties of titanium implants, current knowledge obtained from in vivo studies is lacking. The present study is designed to address two aspects. The first is to verify whether oxide properties of titanium implants indeed influence the in vivo bone tissue responses. The second, is to investigate what oxide properties underline such bone tissue responses. For these purposes, screw-shaped/turned implants have been prepared by electrochemical oxidation methods, resulting in a wide range of oxide properties in terms of: (i) oxide thickness ranging from 200 to 1000 nm, (ii) the surface morphology of barrier and porous oxide film structures, (iii) micro pore configuration - pore sizes<8 microm by length, about 1.27 microm2 to 2.1 microm2 by area and porosity of about 12.7-24.4%, (iv) the crystal structures of amorphous, anatase and mixtures of anatase and rutile type, (v) the chemical compositions of TiO2 and finally, (vi) surface roughness of 0.96-1.03 microm (Sa). These implant oxide properties were divided into test implant samples of Group II, III, IV and V. Control samples (Group I) were turned commercially pure titanium implants. Quantitative bone tissue responses were evaluated biomechanically by resonance frequency analysis (RFA) and removal torque (RT) test. Quantitative histomorphometric analyses and qualitative enzyme histochemical detection of alkaline (ALP) and acidic phosphatase (ACP) activities were investigated on cut and ground sections after six weeks of implant insertion in rabbit tibia. In essence, from the biomechanical and quantitative histomorphometric measurements we concluded that oxide properties of titanium implants, i.e. the oxide thickness, the microporous structure, and the crystallinity significantly influence the bone tissue response. At this stage, however, it is not clear whether oxide properties influence the bone tissue response separately or synergistically.

A new approach to demonstrate cellular activity in bone formation adjacent to implants.

Bone tissue repeatedly formed in titanium 6-aluminum 4-vanadium rabbit bone harvest implants was collected in vivo at various times between 12 days and 5 weeks. Qualitative and quantitative examinations on undecalcified thin sections were performed in the light microscope. The amount of bone tissue was calculated on routinely stained sections. Alkaline (ALP) and acid phosphatase (ACP) enzyme activities were investigated. We also performed immunohistological detection of bone matrix proteins. Increasing bone density as well as an increasing mineralization of the tissue was observed in the biopsies with increasing time. The ALP and ACP activities were similar at short times (12 days and 2 weeks). With increasing time the ALP activity was stronger than that of ACP. The results from the immunohistochemical detection of osteonectin, osteopontin, bone sialoprotein, and collagen I and II demonstrated changes in the tissue differentiation with time. The tissue formation in the canal became more mature with time of ingrowth, as observed with the various techniques used in this study. Owing to these methodical developments, undecalcified ground sections may be used for detailed analysis of various phases of tissue formation in close proximity to implants.

Integration of periorbital titanium implants in irradiated bone: case report and histologic evaluation.

Extraoral implants are used increasingly frequently in the wake of ablative tumor surgery and adjuvant radiation or chemotherapy for craniofacial rehabilitation with facial prostheses and epitheses. However, high rates of nonintegration and implant loss have been reported for extraoral implants, especially for those in the periorbital region following irradiation. This case report and corresponding histologic evaluation describe the osseointegration pattern in irradiated periorbital bone, based on the example of 3 retrieved, clinically integrated, stable titanium screw implants.

Bone-tissue formation and integration of titanium implants: an evaluation with newly developed enzyme and immunohistochemical techniques.

BACKGROUND: Examination of the tissue surrounding retrieved implants involve routine investigations on cut and ground sections. Undecalcified sections with implants in situ are histologically stained followed by qualitative and quantitative observations of the tissue response to the implants by light microscopy. PURPOSE: A novel technique that allows for the accurate definition and quantification of enzymes involved in bone formation (alkaline phosphatase) and resorption (acid phosphatase) in the tissue is presented. MATERIALS AND METHODS: Commercially pure titanium and titanium alloy (Ti6Al4V) implants were retrieved after 6 and 12 weeks of healing in rabbit bone. In addition, 4-week specimens from commercially pure titanium bone harvest chambers placed in rabbit bone were used. Undecalcified cut and ground sections were produced and evaluated with enzyme and immunohistochemical staining techniques. RESULTS: The titanium implants retrieved after 6 weeks of insertion in rabbit bone revealed a higher activity of both alkaline phosphatase and acid phosphatase activity compared to the implants followed for 12 months. The former samples revealed ongoing bone-tissue remodeling in the interface, whereas the latter ones showed steady-state bone conditions. Applying the new technique allowed for investigation of various bone proteins present in the tissue that had formed inside titanium canals of harvest chambers at various times of follow-up. CONCLUSIONS: The combination of routine histologic stainings with enzyme and immunohistochemical technique of cut and ground specimens is a valuable tool in the investigations of retrieved implants from humans and animals. This novel technique now may be used to describe the state of bone regeneration in the interface zone associated with implant research.

The spectrum of giant cells in tumours of the salivary glands: an analysis of 11 cases.

In view of the different terminology for salivary gland tumours with giant cells, eleven cases were analysed by histopathology and immunocytochemistry. Four cases (three pleomorphic adenomas, one carcinosarcoma in a pleomorphic adenoma) were classified as having a foreign-body giant cell reaction, and five cases (two mucoepidermoid carcinomas, one acinic cell carcinoma, two carcinomas in pleomorphic adenomas) as having a sarcomatoid osteoclast-like giant cell reaction. In two further cases a giant cell tumour and a giant cell granuloma were associated with carcinomas in pleomorphic adenomas. All giant cells showed characteristic expression of CD68 as a typical marker for histiocytes and macrophages with their origin in mononuclear haematopoetic stem cells. There was no evidence for an epithelial origin of the giant cells because all those examined had a negative reaction to cytokeratin. Foreign-body cells were characterized by cytoplasmic vacuoles and irregularly dispersed nuclei. They showed a focally circumscribed reaction mostly outside the connective tissue pseudocapsule of the tumours. The sarcomatoid osteoclast-like giant cell reactions in carcinomas were distinctly intermingled with the carcinomatous patterns. In contrast, the associated osteoclast-like giant cell tumour was distinctly separate from the salivary gland tumour tissue and was composed of numerous larger osteoclast-like giant cells with a greater number of nuclei (more than 20); these giant cells were uniformly distributed throughout the tumour tissue. The giant cell granuloma was also separate from the carcinoma and was composed of nests of smaller, more irregularly distributed giant cells.