bluuebunny/crossref_metadata_2025 huggingface dataset:
Ductular structures in acute hepatitis with panacinar necrosis
Authors: Delladetsima Johanna K. , Kyriakou Vasiliki , Vafiadis Irene , Karakitsos Petros , Smyrnoff Tatiana , Tassopoulos Nikolaos C.
Published: June 2005

The development of ductular structures in acute hepatitis with panacinar necrosis was studied in 15 cases of fulminant hepatitis with variable clinical duration, using immunohistochemical markers. The immunophenotype of ductular structures was assessed by the expression of two bile duct epithelium determinants, wide spectrum cytokeratin and epithelial membrane antigen (EMA), and by their glycoconjugate expression using the specific binding lectins Dolichos biflorus agglutinin (DBA) and soybean agglutinin (SBA). Ductular structures showed a predilective, but not a strictly selective location in acinar zone 1 and at the periphery of newly formed parenchymal nodules. All were positive for keratin, while EMA and the lectins were identified less frequently. Cytokeratin expression was additionally observed in hepatic cells with no other phenotypic alteration: this occurred along isolated hepatic cords, within parenchymal remnants, in the spared parenchyma in acinar zone 1 and occasionally at the periphery of parenchymal nodules. The presence of cytokeratin expression in liver cell plates in association with intermediate morphological stages of tubular remodelling speaks in favour of biliary metaplasia of hepatocytes. This process may represent a phenotypic‐functional accommodation of hepatocytes to an altered microenvironment, due to loss of parenchymal integrity. During the phenotypic shift, altered cytokeratin expression appears as one of the earliest biliary features, while EMA and the expression of glycoconjugates represent maturation markers.
An improved method for the species‐specific assessment of mycobacteria in routinely formalin‐fixed and paraffin‐embedded tissues
Authors: Richter Elvira , Schlüter Carsten , Duchrow Michael , Hahn Margrit , Rüsch‐Gerdes Sabine , Galle Jürgen , Flad Hans‐D. , Gerdes Johannes
Published: June 2005

A polymerase chain reaction (PCR) assay for the rapid and species‐specific diagnosis of mycobacterial infections in paraffin‐embedded clinical specimens was developed using oligonucleotide primers to amplify a fragment of the DNA coding for the ribosomal 16S RNA of mycobacteria. The oligonucleotide primers amplified DNA from all 14 species of mycobacteria tested. By means of a reamplification protocol, as few as one to two mycobacteria could be detected in the presence of human DNA. The method of DNA isolation and amplification was applied on sections of routinely formalin‐fixed and paraffin‐embedded tissues. PCR for the β‐actin gene served as a control for successful DNA isolation. Mycobacterial DNA could be detected in cases of mycobacterial infections. The mycobacterial species was determined by additional sequencing of the PCR fragment. This PCR method may be a powerful tool for the diagnosis of mycobacterial infections from histopathological material and for the assessment of those mycobacteria that cannot readily be cultured, such as Mycobacterium leprae.
Accumulation of p53 protein in normal, dysplastic, and neoplastic Barrett's oesophagus
Authors: Krishnadath Kausilia K. , Tilanus Hugo W. , van Blankenstein Mark , Bosman Fre T. , Mulder Andries H.
Published: June 2005

Accumulation of p53 protein was determined by immunohistochemisty in archival material of biopsy specimens from 102 patients with Barrett's oesophagus with different grades of dysplasia, in 24 oesophageal adenocarcinomas associated with Barrett's oesophagus, and in 23 cases of metaplatic epithelium adjacent to these carcinomas. Immunostaining for the p53 protein was found in 23/102 (23 per cent) cases of the Barrett's oesophagus biopsies and in 12/23 (52 per cent) cases of Barrett's oesophagus adjacent to adenocarcinoma. Significant correlations were found between the grade of dysplasia and p53 immunoreactivity in both Barrett's biopsies without adenocarcinoma (P<0.001) and Barrett's oesophagus adjacent to adenocarcinoma (P<0.05). In the adenocarcinomas, intense nuclear immunohistochemical staining for p53 was diffusely or focally present in 20/24 (83 per cent) of the specimens. In Barrett's oesophagus, p53 is a progression marker with high expression in high‐grade dysplasia (89 per cent) and adenocarcinoma (83 per cent).
p53 expression in normal, dysplastic, and neoplastic laryngeal epithelium. Absence of a correlation with prognostic factors
Authors: Nadal Alfons , Campo Elías , Pinto José , Mallofré Carme , Palacín Antonio , Arias Carlos , Traserra José , Cardesa Antonio
Published: June 2005

p53 expression has been examined in 89 squamous cell carcinomas of the larynx (34 glottic, 28 supraglottic, 18 transglottic, 8 pyriform sinus, and 1 subglottic) obtained from 88 patients surgically treated in our centre. In addition, 59 laryngeal samples including normal respiratory epithelium and non‐invasive squamous cell lesions were also tested. Frozen sections were immunostained with PAb 1801 and the results were correlated with pathological features, DNA ploidy and S‐phase of the tumours, disease‐free interval, and survival of the patients. p53 immunoreactivity was observed in 57 (64 per cent) carcinomas. None of the eight samples of normal respiratory epithelium was positive. p53‐positive cells were seen in 8 of 23 (35 per cent) squamous cell metaplasias, 6 of 19 (32 per cent) low‐grade dysplasias and 5 of 10 (50 per cent) high‐grade dysplasias. No correlation was found between p53 expression in carcinomas and their clinical and pathological characteristics, DNA ploidy, or proliferative activity. Neither disease‐free nor overall survival showed differences between p53‐positive and p53‐negative cases. These findings indicate that p53 may play a role in an early stage of malignant transformation of a subset of squamous cell carcinomas of the larynx, but seems not to be associated with further progression of the tumours.
PCR‐mismatch analysis of p53 gene mutation in Hodgkin's disease
Authors: Xerri Luc , Parc Patricia , Bouabdallah Reda , Camerlo Jacques , Hassoun Jacques
Published: June 2005

Expression of the p53 protein can be detected by immunohistochemistry in Reed‐Sternberg (RS) cells, the presumed neoplastic component of Hodgkin's disease (HD) lesions. At present, there is no clear molecular evidence that p53 positive immunostaining in HD correlates with the presence of mutations or other structural alterations of the p53 gene. To address this question, 49 cases of HD have been investigated for p53 expression by immuno‐histochemistry, using the DO1 monoclonal antibody on paraffin sections. Thirty‐seven out of 49 cases (75 per cent) exhibited positive immunostainining restricted to RS cells and variants. Among these 37 positive cases, ten cases were selected on the basis of a rich content of RS cells showing virtually 100 per cent DO1 positivity. A PCR‐mismatch strategy was chosen for the detection of p53 mutations. The threshold level of sensitivity was assessed on positive cell‐line controls. A proportion of 10–15 per cent p53 mutated cells mixed in a normal population could be identified. Total genomic DNA was extracted from the ten selected HD cases and PCR amplification of exons 5–8 of the p53 gene was performed. Heteroduplex mismatch analysis revealed no structural alterations of the p53 gene in any case. In view of these findings, it appears unlikely that the sensitivity of the method by itself can fully explain the negative results, although this possibility cannot be completely ruled out. Thus, it is conceivable that p53 positive immunostaining in HD may not necessarily imply genomic alterations in the classic ‘hot spot’ regions and may be related to another mechanism of p53 protein stabilization.
Loss of heterozygosity in ductal carcinoma in situ of the breast
Authors: Stratton Michael R. , Collins Nadine , Lakhani Sunil R. , Sloane John P.
Published: June 2005

Loss of heterozygosity (LOH) at loci reported to show allele loss in invasive breast cancers was examined in ductal in situ carcinomas of the breast using polymorphic short tandem repeats and the polymerase chain reaction (PCR). LOH was detected at all loci examined in at least 11 per cent of the samples examined. The proportion of cases of in situ carcinoma showing LOH at these loci was similar to that previously reported in invasive cancers. Cases of pure in situ cancer without an invasive component exhibited an overall lower frequency of allele loss. LOH at more than one locus was observed in some intraductal cancers. In a small number of cases, LOH was present in the invasive but not in the intraductal component of the tumour, suggesting that mutation at the locus concerned was associated with development of invasive behaviour.
Expression of c‐myc protein is related to cell proliferation and expression of growth factor receptors in transitional cell bladder cancer
Authors: Lipponen Pertti K.
Published: June 2005

Archival biopsy specimens from transitional cell bladder tumours (n=185) were analysed immunohistochemically for expression of c‐myc protein. The results were compared with compared with histopathological and clinical parameters and survival. Forty‐three per cent of the tumours were negative for c‐myc protein and weak, moderate, or strong cytoplasmic expression was found in 34, 14, and 9 per cent of cases, respectively. Nuclear positivity for c‐myc protein was detected in 35 per cent of tumours and nuclear opositivity was related to overexpression of c‐erb B‐2 (P=0.01) and a high proportion of nuclei were also positive for p53 oncoprotein (p<0.05). Cytoplasmic expression of c‐myc protein was related to histological grade (P=0.005), papillary status (P=0.007), the S‐phase fraction (P=0.008), the mitotic index (P=0.021), overexpression of epidermal growth factor receptor (P=0.045), and c‐erb B‐2 (P=0.17). Expression of c‐myc protein was not significantly related to the progression of tumours and it had no prognostic value in survival analysis. Independent predictors were the T‐category (P<0.001), papillary status. (P=0.001), and S‐phase fraction (P=0.061). The results show that while c‐myc gene product participates in growth regulation of human bladder cancer cells, it has no independent prognostic significance.
MDM2 and CDK4 gene amplification in Ewing's sarcoma
Authors: Ladanyi Marc , Lewis Ronald , Jhanwar Suresh C. , Gerald William , Huvos Andrew G. , Healey John H.
Published: June 2005

Amplification of the MDM2 gene, which maps to chromosome band 12q13 and encodes a p53‐binding protein, may result in functional inactivation of p53 and has been observed in various bone and soft tissue sarcomas. Published studies have included few cases of Ewing's sarcoma (ES) or peripheral neuroectodermal tumour (PNET), a tumour group in which alterations of the p53 pathway have so far not been extensively studied. We examined two ES cell lines, RD‐ES and SK‐ES‐1, and 30 specimens from 27 patients (24 ES, 6 PNET; 19 primary, 4 local recurrence, 7 metastasis) for MDM2 gene amplification by Southern blot analysis. All 30 clinical specimens had been confirmed to contain sufficient ES/PNET DNA by the demonstration of a rearrangement of the t(11;22)‐associated EWS gene using an EWS cDNA probe on the same blots. MDM2 gene amplification was detected in 3 of 30 specimens (10 per cent), including two ES and one PNET, but in neither of the cell lines. The three cases with amplification were morphologically typical primary tumours. Two of the three cases also showed co‐amplification of the CDK4 gene, which endoces a cyclin‐dependent kinase and also maps to band 12q13. Clinically, all three cases had metastatic disease at diagnosis, compared with only 1 of 15 MDM2‐negative cases where the primary tumour was studied. The difference was statistically significant (P=0.005), suggesting an association of MDM2 amplification with advanced stage. Further accural and multivariate analysis of ES/PNET cases with MDM2 gene amplification will be necessary to confirm the clinical significance of these findings. The results suggest that the prevalence of MDM2 gene amplification in ES/PNET is comparable to other sarcomas, and implicate dysfunction of the p53 pathway in a subset of ES/PNET. The biological significance of CDK4 co‐amplification remains to be determined.
Epstein‐Barr virus in B‐cell non‐Hodgkin's lymphomas: Unexpected infection patterns and different infection incidence in low‐ and high‐grade types
Authors: Hummel Michael , Anagnostopoulos Ioannis , Korbjuhn Petra , Stein Harald
Published: June 2005

Two hundred and eight cases of B‐cell non‐Hodgkin's lymphoma (B‐NHL) occurring in Europeans without any signs of HIV infection were investigated for their association with an Epstein‐Barr virus (EBV) infection. The polymerase chain reaction (PCR) was applied for EBV‐DNA detection, in situ hybridization (ISH) for the cellular localization of EBV‐encoded small nuclear RNAs (EBER) and immediate‐early RNAs (BHLF), and immunohistology (IH) for the detection of EBV‐encoded latent membrane protein (LMP) and EBV nuclear antigen 2 (EBNA2) expression. PCR and EBER‐ISH produced congruent results in those cases with amplifiable DNA. EBV was present overall in 26 per cent (54/208) of the B‐NHL cases. Through EBER‐ISH, the virus could be localized merely in rare non‐neoplastic bystander lymphocytes in 27 and additionally in tumour cells of 27 cases. Unexpectedly, the proportion of EBV‐infected tumour cells present in the different cases varied between 1 and 100 per cent. All but three of the cases with infected tumour cells were of high‐grade malignancy. Correlation with the morphological and immunological tumour phenotype revealed that all cases with more than 80 per cent EBER‐positive tumour cells were either B‐anaplastic large cell lymphomas (B‐ALCL), sporadic Burkitt's lymphomas, or B‐NHLs with partial or full plasmacellular differentiation. LMP was consistently absent from Burkitt's lymphomas and constantly expressed in B‐ALCLs with EBER‐positive tumour cells, while in all other instances it varied greatly and was rarer than EBER expression. The above findings suggest that in cases where EBV is present in 80–100 per cent of tumour cells, EBV infection takes place before malignant transformation and thus may have contributed to the malignant phenotype, whereas in the other cases with only a smaller number of infected tumour and single bystander cells, EBV infection may have occurred following malignant transformation. In these cases, infection would appear to be of little or no significance in tumourigenesis.
Identification of tumour‐specific translocations in archival material
Authors: Williams G. H. , Williams E. D.
Published: June 2005

A reverse transcriptase nested polymerase reaction (RT‐nPCR) method has been developed to detect the presence of a tumour‐specific translocation in archival formalin‐fixed, paraffin‐embedded tissue. This technique can be applied to the large numbers of tumours in pathology departments' archives, allowing correlation of translocations with morphological diagnosis, response to therapy, and survival.
Immunohistochemical analysis of adhesion molecules in the micro‐environment of portal tracts in relation to aberrant expression of PDC‐E2 and HLA‐DR on the bile ducts in primary biliary cirrhosis
Authors: Yasoshima Mitsue , Nakanuma Yasuni , Tsuneyama Kouichi , van de Water Judy , Eric Gershwin M.
Published: June 2005

We have examined immunohistochemically the expression of adhesion molecules in the micro‐environment of portal tracts and their relationship to the expression of the pyruvate dehydrogenase E2 complex (PDC‐E2) and HLA‐DR in liver biopsy specimens.

Ten cases of primary biliary cirrhosis (PBC) and 19 controls were examined, including four cases of extrahepatic biliary obstruction, six of chronic viral hepatitis, and nine normal livers. In PBC, the damaged small bile ducts demonstrated an increased expression of PDC‐E2 and an aberrant expression of HLA‐DR; about half of these damaged bile ducts also expressed intercellular adhesion molecules (ICAM)‐1 and a few expressed vascular adhesion molecule (VCAM)‐1. In addition, lymphocyte function‐associated antigen (LFA)‐1 and very late antigen (VLA)‐4 were expressed on infiltrating lymphocytes around these bile ducts. In contrst, in control livers, these alterations in antigen expression on the bile ducts were either not observed or were only focal and weak, when present. These findings suggest that ICAM‐1/LFA‐1 and also VCAM‐1/VLA‐4 linkages between the damaged bile ducts and lymphocytes may facilitate antigen‐specific reactions such as the presentation of antigens, possibly PDC‐E2, to the periductal lymphocytes in PBC. ICAM‐1, VCAM‐1, and E‐selectin were strongly expressed on the endothelial cells of some vessels in the portal tracts in PBC, suggesting the facilitation of the recruitment of lymphocytes around the of some vessels in the portal tracts in PBC, suggesting the facilitation of the recruitment of lymphocytes around the bile ducts of PBC. VCAM‐1, a member of the immunoglobulin superfamily, has not hitherto been reported on bile ducts.
In vitro effects of interleukins on human mesangial cells: Implications for glomerulonephritis
Authors: Chen W.‐P. , Chen A. , Lin C.‐Y.
Published: June 2005

Intrinsic glomerular cells, especially mesangial cells, are considered to be actively involved in the pathogenesis of glomerulonephritis (GN), but the precise mechanism(s) remains elusive. We have previously demonstrated that nephritogenic IgA immune complex can stimulate human mesangial cells (HMCs) to increase their production of interleukin‐1 (IL‐1) and interleukin‐6 (IL‐6). In order to evaluate the roles of cytokines such as IL‐1 and/or IL‐6 and mesangial cells as mediators of renal injury in GN, we have now examined the changes of HMCs and their secreted products in vitro, after stimulation with various concentrations of IL‐1 and IL‐6. Cytokine‐activated HMCs showed the following changes: (1) increased cell size, with intracytoplasmic vaculoes, dilated endoplasmic reticulum, increased free ribosomes and polysomes, and mitochondrial swelling; (2) increased cell proliferation, reflected in thymidine incorporation and an increased proportion of S and G2/M phase cells by cell cycle analysis; (3) enhancement of IL‐6 mRNA expression in HMCs with stimulation of IL‐6 alone or IL‐1 plus IL‐6; and (4) release of large amounts of platelet activating factor (PAF), thromboxane B2 (TxB2), and superoxide anion. Taken together, these results strongly suggest that mesangial cell proliferation and increased production of immune/chemical mediators and superoxide anion can be directly induced by IL‐1 plus IL‐6. These changes may lead to ongoing renal injury.
Interphase cytogenetic analysis of distinct X‐chromosomal translocation breakpoints in synovial sarcoma
Authors: Janz Martin , de Leeuw Bertie , Weghuis Danielle Olde , Werner Martin , Nolte Martina , van Kessel Ad Geurts , Nordheim Alfred , Hipskind Robert A.
Published: June 2005

Synovial sarcomas show a specific translocation involving chromosomes X and 18, t(X;18)(p11.2;q11.2). Two distinct X‐chromosomal breakpoints occur in different synovial sarcoma tumour samples. These breakpoints are located within two related genomic regions containing ornithine aminotransferase‐like sequences, termed OATAL1 and OATL2. Preliminary observations indicated the potential correlation of OATL1‐associated breakpoints with biphasic tumours and OATL2‐associated breakpoints with monophasic fibrous tumours. The present study uses interphase cytogenetics to investigate the nature of chromosomal aberrations in frozen synovial sarcoma tissue samples. Two‐colour fluorescence in situ hybridization (FISH) was performed using probes specific for the centromeres of chromosome X or 18, along with yeast artificial chromosome probes corresponding to the distinct breakpoint regions on Xp. One monophasic epithelial and two monophasic fibrous synovial sarcomas showed an OATL2‐associated breakpoint, while a biphasic tumour revealed a hybridization pattern indicating a breakpoint within the OATL1 region. These results confirm our previous suggestion of a relationship between alternative breakpoints in Xp11.2 and different histological phenotypes observed in synovial sarcomas. They also demonstrate the utility of the two‐colour hybridization approach for the identification of chromosomal changes in interphase nuclei isolated from frozen tissues.
Experimental ulceration leads to sequential expression of spasmolytic polypeptide, intestinal trefoil factor, epidermal growth factor and transforming growth factor alpha mRNAs in rat stomach
Authors: Alison Malcolm R. , Chinery Rebecca , Poulsom Richard , Ashwood Paul , Longcroft Janet M. , Wright Nicholas A.
Published: June 2005

A model of gastric ulceration in the rat has been used to determine the expression of four messenger RNAs (mRNAs) encoding peptides considered to play active parts in the healing response. The trefoil peptides, rat spasmolytic polypeptide (rSP) and rat intestinal trefoil factor (rITF), along with epidermal growth factor (EFG) and transforming growth factor alpha (TGFα) were the molecules studied. Ulceration was caused under anaesthesia by brief application of a liquid nitrogen‐filled cryoprobe to the gastric serosal surface and RNA expression was monitored over the next 10 days. Each mRNA was quantified by ribonuclease protection assay, and mRNAs encoding rSP and rITF were localized within tissue sections by hybridization in situ with 35S antisense riboprobes. Ulceration induced the very rapid expression of first rSP and then rITF mRNA, whereas the mRNAs encoding EGF and TGFα increased at later times, with maxima recorded at 3 and 6 days, respectively. Hybridization in situ detected extensive rSP mRNA expression in the regenerative epithelia. The pronounced, but temporally different patterns of mRNA induction after ulceration suggest that the trefoil peptides may fulfil different and more ummediate roles than the more ‘traditional’ healing proteins EGF and TGAα.
Tenascin immunoreactivity in cryptogenic fibrosing alveolitis
Authors: Wallace W. A. H. , Howie S. E. M. , Lamb D. , Salter D. M.
Published: June 2005

Tenascin is a hexameric extracellular matrix (ECM) glycoprotein which has been demonstrated to have a temporal relationship with active scar formation in adult tissues. We hypothesized that this ECM protein might therefore serve to identify areas of active scarring in lung biopsies from patients with cryptogenic fibrosing alveolitis (CFA). The distribution of tenascin was examined in open lung biopsies from ten patients with CFA, six patients with sarcoidosis, and six pulmonary resection specimens from patients with no evidence of interstitial lung disease, using an immunohistochemical technique. Immunoreactive tenascin was not identified in histologically normal control lung parenchyma and was only focally found around large aggregates of granulomas in sarcoidosis. In the CFA, tenascin production was demonstrated in minimally damaged alveolar walls and areas of active disease but not in end‐stage scarred lung. There was considerable local heterogeneity of staining within cases, which did not appear to relate to the density of the local inflammatory infiltrate. Large plaques of tenascin were noted to be particularly associated with hyperplastic type II alveolar epithelial lining cells, which are recognized to produce fibrogenic cytokines. The examination of tenascin expression in open lung biopsies from patients with CFA may be useful in assessing fibrogenic activity and may thus provide additional prognostic information.
Standardized staining and analysis of argyrophilic nucleolar organizer region associated proteins (AgNORs) in radically resected colorectal adenocarcinoma—correlation with tumour stage and long‐term survival
Authors: Öfner Dietmar , Riedmann Bernhard , Maier Hans , Hittmair Anton , Rumer Astrid , Tötsch Martin , Spechtenhauser Bernhard , Böcker Werner , Schmid Kurt W.
Published: June 2005

Quantification of silver‐stained nucleolar organizer region associated proteins (AgNORs) was introduced in histopathology as a marker of cellular and nucleolar activity. However, due to the poor staining quality obtained on routinely processed archival material, the method yielded controversial and sometimes non‐reproducible results. The recent introduction of wet autoclave pretreatment has reliably improved AgNOR staining quality on routinely formalin‐fixed and paraffin‐embedded tissues. In the present study, 92 routinely processed colorectal carcinomas were investigated, applying this novel staining technique. Subsequent standardized morphometric analysis revealed, irrespective of common tumour staging or grading classifications, a statistically highly significant correlation between AgNOR parameters and clinical course. The usefulness of standardized AgNOR parameters for the independent prediction of patient survival was proven by uni‐ and multivariate analysis.
The pathogenesis of fat embolism
Authors: Hulman Geoffrey
Published: June 2005

Fat embolism is a common autopsy finding in patients with or without a history of trauma. There are two basic mechanisms causing fat to embolize. Depot‐derived fat embolism arises by disruption of depot fat, usually as a result of trauma, allowing direct entry into the bloodstream. Plasma‐derived fat embolism is caused by agglutination of endogenous or infused exogenous fat such as Intralipid, with consequent embolism. Chylomicrons and Intralipid liposomes are known to undergo calcium‐dependent agglutination by C‐reactive protein (CRP), and this may play a role in vivo in this type of fat embolism.
Demonstration of DNA damage/repair in individual cells using in situ end labelling: Association of p53 with sites of DNA damage
Authors: Coates Philip J. , Save Vicki , Ansari Bijan , Hall Peter A.
Published: June 2005

We describe the development and application of in situ end labelling (ISEL) to identity sites of damaged DNA in the nuclei of individual cells. In cell culture, exposure to a variety of genotoxic agents induced a dose and time‐dependent increase in nuclear labelling. In addition, examination of histological sections of human skin exposed to solar‐simulated UV light showed ISEL in both keratinocytes and superficial dermal cells, with the same spatial and temporal distribution as that of a marker of DNA repair, PCNA (proliferating cell nuclear antigen). Using co‐localization techniques and confocal microscopy, we found increased levels of p53 in many ISEL‐positive cells in vitro, with a similar distribution of labelling in the nucleus. This observation provides further evidence for a direct role of p53 in the recognition of damaged DNA. Thus, ISEL should prove a convenient method for demonstrating genotoxic insult in individual cells and in histological material, and may have value in toxicological screening. This high‐resolution microscopy technique can also be used to compare the spatial distribution of various proteins implicated in the response to DNA damage with the sites of the lesion.
Role of apoptosis in modulation of the growth of human colorectal tubular and villous adenomas
Authors: Arai Tomio , Kino Isamu
Published: June 2005

To estimate the effect of cell proliferation and deletion on the growth of the human colorectal adenoma, 27 tubular adenomas and eight villous adenomas were examined. Tubular adenomas were categorized into three grades of cellular atypia: mild, moderate, and severe. Villous adenomas were give a single grade. Morphological characteristics of apoptosis (nuclear condensation and budding) were used to quantify an apoptotic index, AI (percentage of apoptotic cells in viable adenomatous cells). The apoptotic cells were found mainly among the basal site of the adenomatous tubules. The AIs of tubular adenomas with mild, moderate, and severe grades of atypia were 3·5, 5·7 and 8·8 per cent, respectively. The AI of villous adenomas was 1·8 per cent, which was significantly lower than that of tubular adenomas. The mitotic indices, MIs (percentage of mitotic cells in viable adenomatous cells) of tubular adenomas were 0·41 per cent (mild), 0·58 per cent (moderate), and 0·83 per cent (severe), and for villous adenomas the MI was 0·38 per cent. There was a close positive relationship between the AI and MI of tubular adenomas, which paralleled the grade of atypia. These results indicate that both cell proliferation and death were more frequent in adenomas with severe atypia than in adenomas with mild atypia. Moreover, the significantly lower AI of villous adenomas, known to develop into large tumours when compared with tubular adenomas, suggests that reduced apoptoses may lead to a shift in tissue kinetics towards expansive growth.
Immunohistochemical detection of p53 protein in archival tissues from squamous cell carcinomas of the oral cavity using wet autoclave antigen retrieval
Authors: Piffko J. , Bankfalvi A. , Öfner D. , Joos U. , Böcker W. , Schmid K. W.
Published: June 2005

Using a panel of four anti‐p53 antibodies (CM‐1, PAb 1801, DO7, and PAb 240), the phenotypic characteristics of p53 protein overexpression were investigated in 22 routinely formalin‐fixed and paraffin‐embedded squamous cell carcinomas of the oral cavity. Conventional immunohistochemical staining without pretreatment for antigen retrieval was compared with wet autoclaving (AC) and microwave irradiation (MW). Applying AC or MW, p53 immunoreactivity was demonstrated in 9/22 cases with PAb 1801 and DO7 antibodies and in 8/22 cases with the CM‐1 antibody. Immunostaining with the PAb 240 antibody after AC pretreatment showed p53 positivity in 8/22 tumours, but in only 4/22 following MW pretreatment. Immunostaining without pretreatment revealed only 2/22 p53‐positive cases using CM‐1, 4/22 with PAb 1801, and 3/22 with DO7. Without pretreatment, the PAb 240 antibody was consistently negative. Wet autoclave pretreatment was significantly superior for all the antibodies used in this study when compared with conventional immunohistochemistry without pretreatment (P<0·005). It yielded better results than microwave irradiation for antigen unmasking in relation to the PAB 240 antibody (P<0·05). Wet autoclave pretreatment is recommended as a reliable and highly reproducible method for p53 antigen retrieval in routinely processed archival material. The advantages over microwave pretreatment include simple handling and good preservation of morphology.
Numerical aberrations of chromosomes 1 and 7 in renal cell carcinomas as detected by interphase cytogenetics
Authors: Beck J. L. M. , Hopman A. H. N. , Feitz W. F. J. , Schalken J. , Schaafssma H. E. , Van De Kaa C. A. , Ramaekers F. C. S. , Hanselaar A. G. J. M. , De Wilde P. C. M.
Published: June 2005

Alcohol‐fixed single cell suspensions of 37 renal cell carcinomas (RCCs) were assessed by both flow cytometry (FCM) and the fluorescence in situ hybridization (FISH) technique, using chromosome 1‐ and chromosome 7‐specific centromere DNA probes. DNA diploidy or near‐diploidy was observed in 30 of the 37 RCCs and only 12 of these (near‐)diploid tumours were disomic for both chromosomes 1 and 7. Numerical aberrations of chromosome 1 and/or chromosome 7 were present in 18 of the 30 (near‐)diploid RCCs and five of these cases showed monosomy for chromosome 1 in more than 50 per cent of the tumour cells. A double target FISH, with a centromeric and a telomeric specific probe for 1p36, excluded misinterpretation on the basis of clustering of 1q12, and suggested a complete loss of chromosome 1. All these five (near‐)diploid RCCs with monosomy for chromosome 1 were eosinophilic chromophilic cell carcinomas, according to the Thoenes classification of RCC. This observation is of special interest, because it was recently concluded from cytogenetic studies that the diagnosis of chromophilic renal cell carcinoma must be considered as obsolete. Monosomy for chromosome 1 seems to be a non‐random numerical aberration of (near‐)diploid eosinophilic chromophilic cell carcinomas, and a gain of one or more chromosomes 1 appeared to be a common phenomenon in RCCs, especially in the DNA aneuploid tumours. As these chromosomal abnormalities were not found in the earlier classical cytogenetic studies, we conclude that in situ hybridization techniques are required in addition to chromosome banding techniques to obtain a complete characterization of the chromosome imbalances in RCCs.
Diagnosis of Ewing's sarcoma and related tumours by detection of chromosome 22q12 translocations using fluorescence in situ hybridization on tumour touch imprints
Authors: McManus Aidan P. , Gusterson Barry A. , Pinkerton C. Ross , Shipley Janet M.
Published: June 2005

It is increasingly recognized that the identification of t(11;22)(q24;q12) is a useful aid in the accurate diagnosis of Ewing's sarcoma and related tumours. However, cytogenetic studies have a low success rate and adequate tumour is not always available. This study describes the use of fluorescence in situ hybridization (FISH) to detect translocations at 22q12, the site of the EWS gene involved in t(11;22)(q24;q12), on tumour touch imprints made from true cut core‐needle biopsy and frozen tumour. Of the seven tumours analysed, five diagnosed as Ewing's sarcoma or primitive neuroectodermal tumour demonstrated chromosome translocation at 22q12. This is a rapid and reliable method to detect a diagnostically relevant chromosome translocation using minimal amounts of fresh or frozen tumour.
Distribution of sialosyl Tn and Tn antigens within normal and malignant colorectal epithelium
Authors: Jass Jeremy R. , Allison Louise J. , Edgar Stephen G.
Published: June 2005

Expression of the core blood group structures sialosyl Tn (STn) and Tn is regarded as a colorectal cancer‐specific change reflecting truncated synthesis of the oligosaccharide component of goblet cell mucin. The distribution of STn and Tn in normal and malignant epithelium has been studied in detail by a combination of mucin‐, lectin‐, and immunohistochemistry with and without pretreatment with potassium hydroxide (KOH), neuraminidase, and KOH–neuraminidase. When O‐acetylated sialic acid (neuraminidase‐resistant) is converted by saponification to non‐O‐acetylated sialic acid (neuraminidase‐sensitive), normal colorectal goblet cells (mainly of the lower two‐thirds of crypts) are immunoreactive with the monoclonal antibody TKH2 (specific for STn). This immunoreactivity is abolished by the interposition of neuraminidase, but goblet cells then become immunoreactive with Hb‐Tn1 (specific for Tn). While colorectal cancer mucin expresses STn, expression of Tn is not seen in either goblet cell mucin or extracellular material showing the morphological and histochemical characteristics of secretory mucin. Tn expression in cancers is mainly limited to the Golgi zone and in a proportion of cases to cytoplasm and apical membrane (glycocalyx) of columnar cells and inspissated material within lumina. The material reacting with Hb‐Tn1 may be upregulated, membrane‐associated MUC1 glycoprotein rather than MUC2 or MUC4 goblet cell mucin. The presence of STn and cryptic Tn within normal colorectal goblet cells and the absence of Tn expression within colorectal cancer secretory mucin contradicts the generally accepted concept of cancer‐specific incomplete glycoprotein synthesis within these neoplasms.
Altered expression and function of E‐cadherin in cervical intraepithelial neoplasia and invasive squamous cell carcinoma
Authors: Vessey Carina J. , Wilding Jonathan , Folarin Najeem , Hirano Shinji , Takeichi Masatoshi , Soutter Pat , Stamp Gordon W. H. , Pignatelli Massimo
Published: June 2005

HECD‐1 monoclonal antibody has been used to localize E‐cadherin, a calcium‐dependent cell–cell adhesion molecule, in microwave‐treated, paraffin‐embedded sections from 53 cases of cervical intraepithelial neoplasia (CIN) (11 CIN I, 22 CIN II, and 20 CIN III), 16 invasive cervical squamous cell carcinomas, and seven metastases. In normal cervix, E‐cadherin was expressed on the cell membrane of basal and parabasal cells. Cytoplasmic staining was present in occasional basal cells only. In CIN, the presence and localization of cytoplasmic E‐cadherin were found to be significantly correlated with the grade of the CIN lesion. In squamous cell carcinomas, reduced membranous and increased cytoplasmic staining was seen with worsening differentiation. Loss of membranous E‐cadherin expression was also detected in 4/7 metastatic deposits. E‐cadherin expression (120 kD form on Western blotting) was seen in human cervical carcinoma cell lines (HT3, ME180, C41, Caski) that maintained the ability to aggregate in a homotypic adhesion assay and showed a typical epithelial morphology. E‐cadherin‐negative cell lines (Hela, SiHa, C33A) did not show adhesion. HOG‐1 was the only E‐cadherin‐negative cell line which showed a significant degree of cell–cell aggregation. These data indicate that loss of membranous E‐cadherin expression may represent one of the abnormalities underlying loss of cell polarity and differentiation which characterize CIN and invasive cervical cancer.
Platelet‐derived endothelial cell growth factor/thymidine phosphorylase expression in normal tissues: An immunohistochemical study
Authors: Fox Stephen B. , Moghaddam Amir , Westwood Mark , Turley Helen , Bicknell Roy , Gatter Kevin C. , Harris Adrian L.
Published: June 2005

Angiogenesis is the formation of new blood vessels from the existing vascular bed. It is a complex multi‐step process controlled by a number of angiogenic factors. One such factor is platelet‐derived endothelial cell growth factor (PD‐ECGF), recently shown to be thymidine phosphorylase (TP), which is angiogenic in several in vivo assays and tumour systems. PD‐ECGF/TP catalyes the reversible phosphorylation of thymidine to deoxyribose‐1‐ phosphate and thymine. Since PD‐ECGF/TP has an important role in cellular metabolism and in angiogenesis and its expression has been only partially characterized, we raised a monoclonal antibody against recombinant PD‐ECGF/TP and used an immunohistochemical approach to examine the expression of PD‐ECGF/TP in a comprehensive range of normal human tissues. The clone P‐GF44. C, which recognized recombinant PD‐ECGF‐TP and cell lysates transfected with a plasmid expressing PD‐ECGF/TP cDNA on Western blotting, was selected for its ability to stain routinely processed tissue. Staining was observed in both the cytoplasm and/or the nucleus. Immunoreactivity was strongly expressed by macrophages, stromal cells, glial cells, and some epithelia. Gastrointestinal epithelium, smooth muscle, adrenal, lung, and testis were negative. Although endothelial cell expression was observed, there was no correlation with sites of new vessel growth. This pattern of expression suggests tight PD‐ECGF/TP regulation and that cellular thymidine pools may serve to control its different functions. Thus, in the nucleus it might modulate the pool for DNA synthesis, whilst in the cytoplasm it could control other effects through different enzyme systems. The high expression present in macrophages and skin might be important for total body thymidine homeostasis.
Absence of in situ hybridization evidence for latent ‐ or lytic‐phase Epstein‐Barr virus infection of preinvasive squamous lesions of the cervix
Authors: Payne S. , Kernohan N. M. , Walker F.
Published: June 2005

To investigate whether Epstein‐Barr virus (EBV) infection of the uterine cervix plays a significant role in cervical carcinogenesis, 30 preinvasive squamous lesions were subjected to in situ hybridization for (EBER−1,−2, and BHLF1) EBV transcripts which are expressed in latent and lytic infection, respectively. Twenty cases were known to contain EBV sequences by previous polymerase chain reaction (PCR) analysis. Irrespective of EBV PCR status or histological grade, none of the 30 cases demonstrated EBV transcripts in squamous epithelial cells. Two cases showed very occasional EBER‐positive stromal cells, most probably representing resident cervical lymphocytes. These findings suggest that EBV plays no part in early cervical carcinogenesis.
p53 protein expression and nuclear DNA content in breast intraductal proliferations
Authors: Schmitt Fernando C. , Leal Conceição , Lopes Carlos
Published: June 2005

Immunohistochemical analysis of the p53 gene protein and cytometric assessment of nuclear DNA were performed in a series of 51 cases of intraductal breast proliferation. The series included 22 cases of intraductal hyperplasia without atypia, 6 cases of intraductal hyperplasia with atypia, and 23 cases of pure intraductal carcinoma. Expression of p53 protein was detected in one case of intraductal hyperplasia without atypia (4·5 per cent), one case of intraductal hyperplasia with atypia (16·6 per cent) and six cases of intraductal carcinoma (26·0 per cent). No significant correlation was observed between p53 expression and histological subtype of intraductal carcinoma. Aneuploidy was demonstrated in two cases of intraductal hyperplasia with atypia (33·3 per cent) and in 18 cases of intraductal carcinoma (78·2 per cent). All cases of intraductal hyperplasia without atypia were euploid. No significant association was observed between p53 protein expression and ploidy in intraductal hyperplasia. The only case of intraductal hyperplasia without atypia positive for p53 was euploid, whereas the only p53‐positive case of intraductal hyperplasia with atypia was aneuploid. Among the intraductal carcinomas, only the aneuploid cases showed positivity for p53, regardless of histological subtype. The results suggest that some of the changes observed in invasive breast carcinoma, such as p53 expression and aneuploidy, are already present in breast intraductal proliferation, especially in areas with atypia and in intraductal carcinoma. The expression of p53 in breast intraductal proliferation may reflect the acquisition of p53 gene mutations in cells unable adequately to repair DNA damage, with genomic instability which would lead to clonal expansion and putative evolution to invasive disease.
CD44v6 expression in non‐Hodgkin's lymphoma: An association with low histological grade and poor prognosis
Authors: Ristamäki Raija , Joensuu Heikki , Söderström Karl‐Ove , Jalkanen Sirpa
Published: June 2005

The family of CD44 glycoproteins has diverse functions in cell–cell and cell–matrix interactions. The standard form of CD44 is of importance in the dissemination of lymphoma, whereas the clinical significance of the variant exon v6‐containing forms of CD44 (CD44v6) is not known. The expression of different forms of CD44 was investigated by using antibodies against the constant part of CD44 (CD44c) and CD44v6 in 56 primary and 17 recurrent non‐Hodgkin's lymphomas and correlated with several clinicopathological parameters and with prognosis. Fifty‐seven per cent of the primary non‐Hodgkin's lymphomas expressed CD44v6 and 73 per cent expressed the constant epitope. Expression of both CD44c and CD44v6 was associated with low histological grade of malignancy. CD44c expression was associated with a low cellular proliferation rate as assessed by DNA flow cytometry. Of several factors tested, high expression of the variant from v6 was the only factor that was associated with unfavourable recurrence‐free survival (P=0·04). We conclude that CD44v6 is associated with a low histological grade, but, on the other hand, with an unfavourable outcome, which suggests that the combination of CD44v6 and histological grading may form a particularly strong prognostic parameter in non‐Hodgkin's lymphoma.
VWF release and platelet aggregation in human melanoma after perfusion with TNFα
Authors: Renard Nathalie , Nooijen Peet T. G. A. , Schalkwijk Lia , De Waal Robert M. W. , Eggermont Alexander M. M. , Liénard Danielle , Kroon Bin B. R. , Lejeune Ferdy J. , Ruiter Dirk J.
Published: June 2005

Twenty‐nine stage IIIA/B melanoma patients treated by isolated limb perfusion (ILP) with a high dose of recombinant human tumour necrosis factor alpha (rHuTNFα), interferon γ (IFNγ), and melphalan were histologically documented with emphasis on therapy‐induced changes of the tumour vasculature. Sequential biopsies were taken at various intervals before and after the treatment to compare the morphological changes. In order to visualize microvascular changes, immunostaining was performed for von Willebrand factor (VWF), type IV collagen, α‐smooth muscle actin, endothelial antigen PAL‐E, tissue factor, CD41 (thrombocyte marker), and fibrin. In biopsies prior to perfusion, necrosis, haemorrhage, and fibrin thrombi were not found. Within 3 h following triple combination therapy, a change in the distribution of VWF staining occurred, from a discrete endothelial pattern in the untreated lesions to a fuzzy perivascular and subepidermal pattern in the treated lesions. Within 24 h, this was accompanied by intravascular thrombocyte aggregation and erythrostasis, in the absence of tissue factor and fibrin deposits. These findings indicate that the thrombocyte aggregation observed is not caused by local procoagulant activity, but is rather the result of the therapy‐associated vascular damage or haemostasis. Although it is difficult to derive the dynamics of this process from static images, we assume that TNFα induced endothelial cell damage, leading to VWF release. Released VWF may play a role in the adhesion between thrombocytes and the damaged endothelium or the denuded subendothelium. As a consequence, the blood flow is impaired, leading to congestion and oedema, compatible with an early stage of haemorrhagic infarction.
TP53 mutations in prostatic cancer. Analysis of pre‐ and post‐treatment archival formalin‐fixed tumour tissue
Authors: Berner Aasmund , Geitvik Gry , Karlsen Frank , Fosså Sophie D. , Nesland Jahn M. , BØRresen Anne‐Lise
Published: June 2005

The TP53 gene mutation pattern in prostatic cancer was examined in relation to progression and survival, using archival formalin‐fixed pre‐and post‐treatment tumour specimens from 84 prostatic cancer patients. Thirty‐four had hormone‐sensitive tumours and 50 were hormone‐resistant. Six of the 34 (18 per cent) therapy‐responding tumours and 19 of the 50 (38 per cent) hormone‐resistant tumours showed p53 protein accumulation in the post‐treatment specimen. Both pre‐ and post‐treatment specimens from these 25 patients were analysed for mutation of the conserved regions of the TP53 gene (exons 5–8), using constant denaturant gel electrophoresis (CDGE) followed by DNA sequencing. In the post‐treatment samples, mutations were detected in three of the six patients with hormone‐responsive tumours and in 11 of the 19 patients with hormone‐resistant tumours. The three (100 per cent) patients with therapy‐responsive tumours with mutations and nine of the 11 (82 per cent) patients with therapy‐resistant tumours with mutations died of the disease. Thirteen of the 14 mutations in the post‐treatment specimens were transitions, 11 occurring at CpG dinucleotides in which codon 273 was involved in ten. A significantly higher proportion of tumours with mutations were poorly differentiated compared with tumours without mutation (P<0·04). Our findings indicate that TP53 mutation is a late event in tumour development of the prostate gland and that codon 273 might be a ‘hotspot’ for mutation in the progression of the disease.
A novel model for human interstitial lung disease: Hapten‐driven lung fibrosis in rodents
Authors: Roberts S. N. , Howie S. E. M. , Wallace W. A. H. , Brown D. M. , Lamb D. , Ramage E. A. , Donaldson K.
Published: June 2005

A novel model is described of chronic pulmonary fibrosis in rodents. The condition is induced by a single intratracheal instillation of a well‐characterized fluorescent haptenic antigen, fluorescein isothiocyanate (FITC), into non‐immune animals. This results in an acute inflammatory response involving a granulocytic infiltrate, which disappears over a week and is replaced by a chronic mononuclear infiltrate in which T lymphocytes predominate. Over several months, a chronic patchy fibrosis is accompanied by a sustained mononuclear interstitial infiltrate localized at sites of persistent FITC deposition. Where no FITC is present, the lung tissues are apparently normal. An immune response is mounted, as measured by the appearance of specific anti‐FITC serum antibodies. This model has relevance to the pathogenesis of some forms of human interstitial lung disease.