bluuebunny/crossref_metadata_2025 huggingface dataset:
Can p53 staining be used to identify patients with aggressive superficial bladder cancer?
Authors: Masters John RW , Vani Urvi D , Grigor Kenneth M , Griffiths Gareth O , Crook Angela , Parmar Mahesh KB , Knowles Margaret A
Published: April 2003

Approximately 10% of patients with superficial bladder cancer (pTa/pT1) recur with life‐threatening muscle‐invasive disease. Identification of these patients has been a major goal of bladder cancer research. In 1994, it was suggested that p53 immunostaining could identify the cancers that would progress and it was proposed that tumours that stain for p53 should be treated aggressively with radiotherapy or cystectomy. Despite the hundreds of studies published since on the relationship between p53 and progression in superficial bladder cancer, the clinical utility of p53 immunostaining has not been resolved because of limitations concerning the numbers of patients and the length of follow‐up. This study set out to overcome these limitations by using tissue from a large multicentre trial that recruited 502 patients with a median follow‐up of 10 years. Each of 34 patients that had progressed with ≥ pT2 disease or had distant metastases or had died from bladder cancer was compared with one or two matched controls. Sections were stained with a mouse monoclonal antibody to p53, pAb1801. In agreement with many of the earlier studies, p53 immunostaining had prognostic significance. The adjusted hazard ratio for time to progression for the pAb1801‐positive versus negative group was 2.5, with 95% confidence intervals of 1.05–5.98 (p = 0.039). The other major risk factor that is associated with progression of superficial bladder cancer is pT1G3 disease. Of the 42 pT1G3 cancers, 14 (33%) progressed. The proportion of cancers with p53 staining that progressed was similar to the proportion of pT1G3 cancers that progressed, but neither the sensitivity nor the specificity of association of p53 staining with progression is sufficient to recommend cystectomy in individual patients. Copyright © 2003 John Wiley & Sons, Ltd.
Malignant phyllodes tumours show stromal overexpression of c‐myc and c‐kit
Authors: Sawyer Elinor J , Poulsom Richard , Hunt F Toby , Jeffery Rosemary , Elia George , Ellis Ian O , Ellis Paul , Tomlinson Ian PM , Hanby Andrew M
Published: April 2003

Phyllodes tumours are fibroepithelial neoplasms of the breast, the stroma of which can undergo malignant progression to sarcoma. The frequency of malignant lesions varies in different series from 5% to 30%. The aim of this study was to elucidate potential molecular mechanisms in the progression to malignancy in phyllodes tumours. c‐myc and c‐kit were studied at the protein, RNA(c‐myc only) and DNA level. We chose to study c‐myc as we have previously shown that Wnt signalling is important in benign, but not malignant, phyllodes tumours. If c‐myc is constitutively activated in malignant tumours, this may provide an explanation for why the Wnt pathway is no longer important in these tumours. c‐kit is a membrane‐bound tyrosine kinase receptor and overexpression is characteristic of gastrointestinal stromal tumours. A previous report suggested that this may also be the case in malignant phyllodes tumours, and we wished to confirm this. We assessed expression of c‐myc and c‐kit in 30 phyllodes tumours (10 malignant) using in situ hybridization (c‐myc) and immunohistochemistry (c‐myc and c‐kit). 9/10 malignant tumours showed c‐myc expression in the stroma, compared to 7/20 benign tumours (p = 0.006, Fisher's exact test). Stromal c‐kit expression was found in 5/10 malignant tumours, compared to 1/20 benign tumours (p = 0.008, Fisher's exact test). One tumour had high‐level amplification of c‐myc, but we found no evidence of mutations of c‐kit. We hypothesize that the overexpression of c‐myc may drive stromal proliferation in malignant phyllodes tumours, and that c‐kit overexpression contributes to the growth of these lesions. c‐kit may also be a new therapeutic target in these tumours. Copyright © 2003 John Wiley & Sons, Ltd.
Cholesterol storage and tau pathology in Niemann–Pick type C disease in the brain
Authors: Distl Roland , Treiber‐Held Stephanie , Albert Frank , Meske Volker , Harzer Klaus , Ohm Thomas G
Published: April 2003

Niemann–Pick type C disease is an inherited neurovisceral storage disorder with intracellular accumulation of cholesterol. In affected brains, many ballooned neurons are seen. Considerable nerve cell loss of unknown pathogenesis leads to neurological deterioration and dementia. Chemical examination of brains has failed to demonstrate increased levels of cholesterol. Using filipin fluorometry of neuronal cells in tissue slices, we found massive accumulation of cholesterol in neurons in four out of five human Niemann–Pick type C cases including adult patients. Neurofibrillary tangles composed of aggregates of the otherwise highly soluble protein tau were present in three Niemann–Pick type C cases and were also immunologically identical to those associated with Alzheimer's disease. However, only a thin slab of spinal cord or a tiny piece of isocortex was available for examination in the two cases without tangles. In a further semi‐quantitative analysis of 576 neurons, we determined higher cholesterol content in tangle‐bearing neurons than in adjacent tangle‐free neurons. The association of cholesterol accumulation with neurofibrillary degeneration in Niemann–Pick type C disease and Alzheimer's disease awakens interest in the role of impaired cholesterol metabolism in the development of neurofibrillary tangles in both diseases. Copyright © 2003 John Wiley & Sons, Ltd.
Sustained interleukin‐6 signalling leads to the development of lymphoid organ‐like structures in the lung
Authors: Goya Sho , Matsuoka Hiroto , Mori Masahide , Morishita Hiroshi , Kida Hiroshi , Kobashi Yoichiro , Kato Terufumi , Taguchi Yoshio , Osaki Tadashi , Tachibana Isao , Nishimoto Norihiro , Yoshizaki Kazuyuki , Kawase Ichiro , Hayashi Seiji
Published: April 2003

A variety of pathological changes are seen in lymphoproliferative disorders of the lung but the histogenesis of these abnormalities is not yet fully understood. We previously showed that adenovirus vector‐mediated transient expression of both the human interleukin‐6 (IL‐6) and IL‐6 receptor (IL‐6R) genes, but not the IL‐6 gene alone, in the rat lung induced lymphocytic alveolitis. In the present study, we explored the lung pathology of human IL‐6 and IL‐6R double transgenic mice to elucidate the effects of prolonged IL‐6 signalling on the lung. The transgenic animals developed mononuclear cell accumulation in peribronchovascular regions, but little infiltration into alveolar spaces. Immunohistochemical analysis revealed that the cellular accumulations contained not only mixtures of inflammatory cells but also lymphoid tissue‐like structures. As the expression of CXCL13/BLC, the indispensable chemokine for lymphoid organogenesis, was recognized in the B cell follicles of the pulmonary lesions, we speculate that this chemokine plays an inductive role in the development of the lymphoid tissue‐like structures. These structures were distinguished from bronchus‐associated lymphoid tissues (BALTs) by their location and by the lack of lymphoepithelium, which is a characteristic of BALT. These findings imply that IL‐6 signalling may play a role in the pathogenesis of lymphoproliferative disorders of the lung. Copyright © 2003 John Wiley & Sons, Ltd.
Overexpression of the mitotic checkpoint genes BUB1, BUBR1, and BUB3 in gastric cancer—association with tumour cell proliferation
Authors: Grabsch Heike , Takeno Shinsuke , Parsons Wendy J , Pomjanski Natalja , Boecking Alfred , Gabbert Helmut Erich , Mueller Wolfram
Published: April 2003

The mitotic spindle assembly checkpoint modulates the timing of anaphase initiation in response to improper alignment of chromosomes at the metaphase plate. The BUB gene family encodes proteins which are part of a large multi‐protein kinetochore complex and which are believed to be key components of the checkpoint regulatory pathway. Failure of this surveillance system can lead to genomic instability and could be responsible for the increased incidence of aneuploidy in gastric cancer. Since mutations of BUB genes have not been identified in gastric cancer to date, altered BUB expression levels may significantly impair mitotic checkpoint function. To explore this possibility, the expression levels of BUB1, BUBR1, and BUB3 were determined in 43 gastric carcinomas and corresponding normal gastric mucosa by reverse transcription‐polymerase chain reaction (RT‐PCR). Gene expression levels were compared with histopathological parameters and DNA ploidy, as well as with proliferative activity, measured by Ki‐67 mRNA expression. To the authors' knowledge, this is the first study to investigate the expression levels of mitotic checkpoint genes together with DNA ploidy in gastric cancer. BUB1 was overexpressed in 84%, BUBR1 in 68%, and BUB3 in 79% of gastric cancers. This study also revealed that all three genes were simultaneously overexpressed in 61% of the tumours and that there was a statistically significant positive correlation between overexpression of BUB1, BUBR1 or BUB3 and Ki‐67 expression (p < 0.001). Eighty‐one per cent of the tumours were classified as aneuploid. However, no correlation was found between ploidy and BUB transcript expression levels. These results suggest that inactivation of the mitotic checkpoint genes BUB1, BUBR1, and BUB3 by epigenetic silencing does not seem to play a role in gastric carcinogenesis. The strong correlation of BUB expression level and tumour cell proliferation suggests that BUB overexpression is a proliferation‐dependent phenomenon in gastric cancer. However, overexpression due to lack of normal BUB protein function or due to a yet unknown additional BUB function has to be considered. Copyright © 2003 John Wiley & Sons, Ltd.
Concerted promoter hypermethylation of hMLH1, p16INK4A, and E‐cadherin in gastric carcinomas with microsatellite instability
Authors: Kim Hyunki , Kim Yun Hee , Kim Sung Eun , Kim Nam‐Gyun , Noh Sung Hoon , Kim Hoguen
Published: April 2003

In most sporadic gastric carcinomas, microsatellite instability (MSI) originates from inactivation of the hMLH1 gene by promoter hypermethylation. However, the methylation patterns of other genes and their consequences in high MSI (MSI‐H) gastric carcinomas are not well characterized. To address the aberrant promoter methylation profiles of MSI‐H gastric carcinomas, promoter methylation of six genes (hMLH1, p16INK4A, E‐cadherin, Rb, RASSF1A, and VHL) and CpG island methylator phenotype (CIMP) were explored in 36 MSI‐H gastric carcinomas and the results were compared with those of 43 microsatellite‐stable (MSS) gastric carcinomas. Frequent promoter hypermethylation was found in hMLH1, p16INK4A, and E‐cadherin and the frequency was significantly higher in MSI‐H gastric carcinomas. Promoter hypermethylation of hMLH1, E‐cadherin, and p16INK4A was found in 89%, 78%, and 33% of MSI‐H gastric carcinomas and in 16%, 32%, and 11% of MSS carcinomas, respectively (p = 0.01). Selective absent or decreased expression of the gene product related to the hypermethylated promoter was found for hMLH1 and p16INK4A in MSI‐H carcinoma, whereas the expression of E‐cadherin was generally decreased both in the MSI‐H and in the MSS carcinomas. MSI‐H gastric carcinomas were also related to the high CIMP (CIMP‐H, three or more of the five loci examined showing methylation). Twenty‐two (61%) MSI‐H gastric carcinomas were CIMP‐H, compared with only seven (16%) MSS carcinomas (p = 0.001). These findings indicate that hMLH1 is one of the frequent methylation targets in CIMP‐H gastric carcinomas and that inactivation of hMLH1 through promoter hypermethylation results in tumours following the MSI pathway. Copyright © 2003 John Wiley & Sons, Ltd.
The role of E‐cadherin in low‐grade ductal breast tumourigenesis
Authors: Roylance Rebecca , Droufakou Stavroula , Gorman Patricia , Gillett Cheryl , Hart Ian R , Hanby Andrew , Tomlinson Ian
Published: April 2003

Grade I invasive ductal breast carcinomas have a specific pattern of genetic aberrations, namely gain of 1q and loss of 16q. This pattern is very similar to the changes seen in invasive lobular breast carcinomas (ILCs). The gene on 16q involved in ILC is known to be E‐cadherin (CDH1). This study has investigated whether the same gene is responsible for grade I invasive ductal carcinoma (IDC), using allele imbalance analysis, mutation screening, and immunohistochemistry (IHC). The data suggest that despite the shared pattern of genetic aberrations seen in grade I IDC and ILC, CDH1 is not the target gene in low‐grade ductal tumourigenesis. Copyright © 2003 John Wiley & Sons, Ltd.
Promoter methylation of TGFβ receptor I and mutation of TGFβ receptor II are frequent events in MSI sporadic gastric carcinomas
Authors: Pinto Mafalda , Oliveira Carla , Cirnes Luis , Carlos Machado José , Ramires Maria , Nogueira Ana , Carneiro Fátima , Seruca Raquel
Published: April 2003

Transforming growth factor beta (TGFβ) is a potent inhibitor of cell growth, whose action is transduced through interaction between type I (RI) and type II (RII) receptors. Abnormal expression of these receptors has been identified in several human cancers and was found to be associated with resistance to TGFβ. TGFβ RII mutations occur in many types of malignancy. TGFβ RI hypermethylation has been suggested as a cause of abnormal or absent expression of this receptor in cancer. This study has analysed the methylation status of the promoter region of the TGFβ RI gene using a methylation‐sensitive enzyme followed by polymerase chain reaction (PCR), and TGFβ RII mutations (BAT‐RII and a GT3) in order to determine the frequency of alteration of the TGFβ receptors in a series of 40 sporadic gastric carcinomas (SGCs), 25 of which showed microsatellite instability (MSI) and 15 of which were microsatellite stable (MSS). Methylation in the promoter region of the TGFβ RI gene was detected in 20 of the 40 (50%) SGCs (64% of the MSI cases and 26.7% of the MSS); 17 of the 40 (42.5%) cases had mutations in the BAT‐RII region of the TGFβ RII gene (68% in the MSI cases; 0% in the MSS). In total, 25 of the 40 (62.5%) SGCs had alterations in at least one of the TGFβ receptors (84% of the cases in the MSI group, in contrast with 16% of the MSS cases) (p = 0.0003). The clinicopathological features of the cases were also studied and significant associations were found between the presence of alterations in TGFβ receptors and the age of the patients (p = 0.046), size (p = 0.011), and proliferative rate of the tumours (p = 0.048). In conclusion, alterations in the receptors of TGFβ (TGFβ RI promoter hypermethylation and TGFβ RII mutations) are frequent events in MSI SGC and are associated with large size and high proliferative activity of the tumours, in keeping with loss of the growth inhibitory effects of TGFβ in this setting. Copyright © 2003 John Wiley & Sons, Ltd.
Age‐related pseudocapillarization of the human liver
Authors: McLean Allan J , Cogger Victoria C , Chong Guan C , Warren Alessandra , Markus Astrid MA , Dahlstrom Jane E , Le Couteur David G
Published: April 2003

Age‐related changes in liver function are important because they may promote susceptibility to adverse drug reactions, neurotoxicity, atherosclerosis, and other important diseases in older people. Age‐related changes in the rat hepatic sinusoidal endothelium, termed pseudocapillarization, have been described recently and these may contribute to hepatic impairment. The present study has examined surgical and post‐mortem specimens with immunohistochemistry and transmission electron microscopy to determine whether pseudocapillarization also occurs in older humans. The age of the subject, independent of systemic disease or hepatic pathology in surgical and post‐mortem samples of human liver, was associated with increased peri‐sinusoidal expression of von Willebrand's factor, collagen I, collagen IV, and staining with Masson's trichrome. Electron microscopy revealed significant age‐related thickening of the sinusoidal endothelium (young 165 ± 17 nm, middle age 222 ± 11 nm, older 289 ± 9 nm, p < 0.001) with loss of fenestrations (young 7.7 ± 0.7 per 10 µm, middle age 3.6 ± 0.5 per 10 µm, older 1.5 ± 0.4 per 10 µm, p < 0.001), and age‐related deposition of basal lamina and collagen. In conclusion, ageing in humans is associated with morphological changes in the sinusoidal endothelium and space of Disse which are presumptively related to the ageing process and potentially represent an important link between the ageing process and disease susceptibility. Copyright © 2003 John Wiley & Sons, Ltd.
ALK‐negative systemic anaplastic large cell lymphoma: differential diagnostic and prognostic aspects–a review
Authors: ten Berge Rosita L , Oudejans Joost J , Ossenkoppele Gert J , Meijer Chris JLM
Published: April 2003

Anaplastic large cell lymphoma (ALCL) can be divided into two major groups. The first is a spectrum of CD30+ T‐cell lymphoproliferative disorders including primary cutaneous ALCL and lymphomatoid papulosis, usually affecting older patients but characterized by an excellent prognosis. The second is systemic nodal ALCL, which on the basis of genetic and immunophenotypic features combined with clinical parameters can be divided into two subgroups: anaplastic lymphoma kinase (ALK)‐positive and ALK‐negative systemic ALCL. ALK expression, usually the result of a t(2;5) translocation, correlates with the expression of other markers such as EMA and a cytotoxic phenotype, and is strongly related to younger age groups, lower international prognostic index (IPI) risk groups, and a good prognosis. ALK‐negative ALCL, however, shows a more heterogeneous immunophenotype and clinical behaviour, and prognostic parameters are needed to determine treatment strategies in individual patients. Besides clinical parameters included in the IPI, recent studies have pointed out several biological prognosticators of potential value, such as the percentage of tumour‐infiltrating activated cytotoxic T‐lymphocytes. The expression of proteins involved in the execution or regulation of apoptosis, such as activated caspase 3, Bcl‐2, and PI9, was also found to be strongly related to clinical outcome. These studies indicate that inhibition of the apoptosis cascade in particular is an important mechanism that can explain the poor clinical outcome in therapy refractory ALCL. Functional studies are required to investigate whether disruption of one or more of the apoptosis pathways is the major factor in the fatal outcome of the disease and whether apoptosis resistance based on inhibition of one pathway can be overcome by activating another pathway that is still intact. Copyright © 2003 John Wiley & Sons, Ltd.
Evidence for cell‐specific changes with age in expression of oestrogen receptor (ER) α and β in bone fractures from men and women
Authors: Batra Gaurav S , Hainey Linda , Freemont Anthony J , Andrew Glynne , Saunders Philippa TK , Hoyland Judith A , Braidman Isobel P
Published: April 2003

Oestrogen is recognized as important for maintaining bone mass in men and women. Oestrogen receptor (ER) α and the recently described ER‐β are both expressed in bone cells, but have different affinities for oestrogen agonists and plant oestrogens, which could be important in developing treatments for bone loss in both men and women. It is unclear, however, which isoform predominates in bone; cell type and age may influence their relative expression. The present study has compared ER‐α and ER‐β expression in serial sections of human fracture callus from males (n = 19, age range 5–72 years) and females (n = 15, age range 3–86 years) by indirect immunoperoxidase. Fracture callus was used as it can be readily obtained from individuals over a wide age range and contains a variety of bone cells. Antibody specificity was confirmed by western blotting and comparison of immunoreactivity in sections of breast tumour and benign prostate hyperplasia. No gender difference in ER expression was found in bone from individuals less than 40 years old. Proliferative chondrocytes were positive for both isoforms, but few larger hypertrophic cells were immunoreactive. ER‐α and ER‐β were co‐expressed in osteoclasts, suggesting that oestrogen may act directly on these cells. Osteoblasts, osteocytes, and mesenchymal cells also expressed both isoforms. In women over 40 years of age, however, relatively fewer biopsies contained osteocytes positive for ER‐α and ER‐β. Likewise, the proportions of osteoblasts and mesenchymal cells expressing ER‐β were reduced but ER‐α remained unaffected. In contrast, in men over 40 years, only the proportion of biopsies containing ER‐β‐positive mesenchymal cells was lower. In these older men and women, ER‐α and ER‐β expression was retained by the small proliferative chondrocytes. These results demonstrate that gender, age, and cell type are important determinants of ER isoform expression in skeletal cells. Copyright © 2003 John Wiley & Sons, Ltd.
The evolution of crescentic nephritis and alveolar haemorrhage following induction of autoimmunity to glomerular basement membrane in an experimental model of Goodpasture's disease
Authors: Reynolds John , Moss Jill , Duda Mark A , Smith Jennifer , Karkar Ayman M , Macherla Vamshi , Shore Ian , Evans David J , Woodrow David F , Pusey Charles D
Published: April 2003

Goodpasture's, or anti‐glomerular basement membrane (GBM), disease presents with rapidly progressive glomerulonephritis and lung haemorrhage, and is caused by autoimmunity to the NC1 domain of the α3 chain of type IV collagen (α3(IV)NC1). This study examines the development of crescentic nephritis and alveolar haemorrhage in a model of Goodpasture's disease, experimental autoimmune glomerulonephritis (EAG), induced in WKY rats by immunization with rat GBM in adjuvant. An increase in circulating anti‐GBM antibodies and albuminuria was observed by week 2, which increased further by weeks 3 and 4, while a decrease in creatinine clearance was observed by week 2, which decreased further by weeks 3 and 4. The kidneys of animals with EAG showed linear deposits of IgG on the GBM and a transient glomerular infiltration by CD4+ T cells at week 2. By week 3 there were large deposits of fibrin in Bowman's space, and glomerular infiltration by CD8+ T cells and macrophages, accompanied by focal necrotizing glomerulonephritis with crescent formation. Ultrastructural studies showed glomerular endothelial cell swelling and epithelial cell foot process effacement at week 2. As the lesion progressed, capillary loops became occluded and the mesangium became expanded by mononuclear cells. By week 3 there was detachment of the endothelium from the GBM, and accumulation of fibrin beneath the disrupted endothelial cells and in Bowman's space. Occasional breaks were observed in the continuity of the basement membrane, and cytoplasmic projections from infiltrating mononuclear cells could be seen crossing the capillary wall between the lumen and the crescent. The lungs of animals with EAG showed patchy binding of IgG to the alveolar basement membrane (ABM) at week 2, and infiltration of the interstitium by CD8+ T cells and macrophages by weeks 3 and 4, accompanied by both interstitial and alveolar haemorrhage. Ultrastructural studies showed focal mononuclear cell infiltrates in alveolar walls at week 2. Occasional breaks were observed in the basement membrane and adjacent endothelium by weeks 3 and 4, together with accumulation of surfactant and erythrocytes within the alveolar spaces. This study defines for the first time the relationship between the immunological and pathological events during the evolution of EAG, and provides the basis for further work on the pathogenesis of Goodpasture's disease. Copyright © 2003 John Wiley & Sons, Ltd.
The pattern of expression of protease‐activated receptors (PARs) during early trophoblast development
Authors: Even‐Ram Sharona Cohen , Grisaru‐Granovsky Sorina , Pruss Diana , Maoz Miriam , Salah Zaidoun , Yong‐Jun Yin , Bar‐Shavit Rachel
Published: April 2003

Human fetal development depends on the ability of the embryo to gain access to the maternal circulation. Thus, specialized stem cells of the newly formed placenta, trophoblast, invade the uterus and its arterial network to establish an efficient feto‐maternal molecular exchange. To accomplish this task, trophoblast differentiation during the first trimester of pregnancy involves cell proliferation, invasion, and extracellular matrix (ECM) remodelling. Trophoblast invasion shares many features with tumour cell invasion, with the distinction that it is strictly spatially and temporally controlled. We have previously demonstrated that PAR1, the first member of the protease‐activated receptor (PAR) family, plays a central role in tumour cell invasion. In the present study we have examined the pattern of expression of PAR1 and other PAR family candidates during early human placental development. We show that PAR1 and PAR3 are highly and spatially expressed between the 7th and 10th weeks of gestation but not at the 12th week and thereafter. Likewise, high expression levels of PAR1 and PAR3 were observed in the cytotrophoblast cells of complete hydatidiform mole as compared to minimal levels in normal age‐matched placenta. Together, our data suggest the involvement of PAR1 and PAR3 in restricted and unrestricted pathological trophoblast invasion. Copyright © 2003 John Wiley & Sons, Ltd.
Spatial and temporal distribution of intracellular free cholesterol in brains of a Niemann–Pick type C mouse model showing hyperphosphorylated tau protein. Implications for Alzheimer's disease
Authors: Treiber‐Held Stephanie , Distl Roland , Meske Volker , Albert Frank , Ohm Thomas G
Published: April 2003

Niemann–Pick type C (NPC) disease is a fatal hereditary neurovisceral disorder with diagnostically relevant intracellular accumulation of cholesterol in non‐brain tissue, for example the spleen and fibroblasts. In the brain, many ballooned neurons are seen. Using filipin microfluorodensitometry, significant accumulations of free cholesterol in specified neurons have been described in NPC patients. The present study demonstrates spatial and temporal accumulation of free cholesterol in the brains of homozygous NPC (−npc/−npc) mice, a widely acknowledged mouse model, and in primarily cultured neurons therefrom. Intraneuronal storage of free cholesterol was already prominent at a pre‐clinical stage in various grey matter areas of the murine cerebral cortex. Hippocampal areas showed differential development of the pathological distribution of free cholesterol. The pyramidal cells in the CA3 sector of Ammon's horn were affected much earlier than in CA1. Some of the deeper cerebral nuclei were affected only slightly, even at the final stage. Neurons (E15–E17) cultured in a cholesterol‐free medium also showed massive accumulation of intracellular free cholesterol. In addition, brains from the murine NPC model for Alzheimer's disease (AD)‐like changes in the microtubule‐associated protein tau were tested using the Gallyas silver technique and AT8‐immunolabelling, since both human diseases are accompanied by intraneuronal tangles made up of tau protein aggregations. Although the analysis failed to show classical silver‐stainable tangles of the AD type in the NPC mice, tau protein phosphorylated at epitopes considered to represent early stages of AD was found. This further strengthens the concept that an alteration in cholesterol metabolism may play an important role in AD. The NPC mouse model may thus serve as a tool to analyse the role of cholesterol in initial changes of tau that eventually lead to the formation of tangles in both NPC and AD. Copyright © 2003 John Wiley & Sons, Ltd.
Epithelial carcinogenesis: challenging monoclonality
Authors: Going James J
Published: April 2003

How carcinomas begin remains unclear, but experimental data do not entirely exclude the participation of more than one clone of neoplastic cells, even in relatively advanced epithelial tumour growth. Microdissection and new PCR clonality assays exploiting X‐linked polymorphisms, some of which (including XIST) are expressed in RNA, create investigational opportunities complementary to other molecular analyses, but a reliable in situ assay of X‐inactivation remains desirable. The necessity for stringent controls in clonality analysis is emphasized. While it may be possible to reconcile ‘pluriclonal’ (oligoclonal or polyclonal) carcinogenesis with widely accepted paradigms of genetic/epigenetic change and clonal selection in epithelial neoplasia, it deserves critical study as novel carcinogenic mechanisms would be implied. Copyright © 2003 John Wiley & Sons, Ltd.
Pathobiology of autochthonous prostate cancer in a pre‐clinical transgenic mouse model
Authors: Kaplan‐Lefko Paula J. , Chen Tsuey‐Ming , Ittmann Michael M. , Barrios Roberto J. , Ayala Gustavo E. , Huss Wendy J. , Maddison Lisette A. , Foster Barbara A. , Greenberg Norman M.
Published: April 2003

Background

Animal models that closely mimic clinical disease can be exploited to hasten the pace of translational research. To this end, we have defined windows of opportunity in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model of prostate cancer as a paradigm for designing pre‐clinical trials.

Methods

The incidence of cancer, metastasis, and distribution of pathology were examined as a function of time in TRAMP mice. The expression of various markers of differentiation were characterized.

Results

The TRAMP model develops progressive, multifocal, and heterogeneous disease. Each lobe of the prostate progressed at a different rate. Cytokeratin 8, E‐cadherin, and androgen receptor (AR) were expressed during cancer progression but levels were reduced or absent in late stage disease. A distinct epithelial to neuroendocrine (ENT) shift was observed to be a stochastic event related to prostate cancer progression in TRAMP.

Conclusions

This study will serve as the basis for the rational design of pre‐clinical studies with genetically engineered mouse models. Prostate 55: 219–237, 2003. © 2003 Wiley‐Liss, Inc.
Growth inhibition and differentiation in human prostate carcinoma cells induced by the vitamin D analog 1α,24‐dihydroxyvitamin D2
Authors: Bauer Joshua A. , Thompson Todd A. , Church Dawn R. , Ariazi Eric A. , Wilding George
Published: April 2003

Background

Vitamin D has been suggested as a chemopreventive and therapeutic modality for prostate cancer. However, hypercalcemic toxicity has limited the use of 1α,25‐dihydroxyvitamin D3 (1,25‐(OH)2D3) in clinical trials, prompting the search for analogs of vitamin D with less toxicity while retaining efficacy as a modality for cancer intervention. In this study, the less hypercalcemic vitamin D analog 1α,24‐dihydroxyvitamin D2 (1,24‐(OH)2D2) was examined for its effects on cellular growth inhibition and differentiation induction in the LNCaP human prostate carcinoma cell line.

Methods

LNCaP cell growth was determined by quantifying DNA levels. Protein levels were determined using the ELISA method and immunoblotting. Levels of mRNA were determined using real‐time quantitative reverse transcriptase PCR.

Results

LNCaP growth was decreased 50% by exposure to 0.01 nM 1,24‐(OH)2D2 after 96 hr in the presence of a growth stimulatory 0.1 nM dose of the androgen R1881. Prostate specific antigen (PSA) levels were increased 3.5‐fold with 10 nM 1,24‐(OH)2D2 treatment compared to a 1.9‐fold increase in PSA levels found with 10 nM 1,25‐(OH)2D3 under low androgen conditions. Neither 1,24‐(OH)2D2 nor 1,25‐(OH)2D3 affected the expression of cytokeratin 18 protein levels. Treatment with 10 nM 1,24‐(OH)2D2 alone produced a 1.3‐fold increase in AR mRNA and a 2.2‐fold increase in AR protein levels after 96 hr. Surprisingly, the addition of 1.0 nM R1881 alone or in combination with 10 nM 1,24‐(OH)2D2 produced an approximately 60% decrease in AR mRNA, whereas AR protein levels were increased 1.6‐fold.

Conclusions

Overall, 1,24‐(OH)2D2 was found to be at least as effective as 1,25‐(OH)2D3 at inhibiting growth and inducing differentiation markers in LNCaP prostate carcinoma cells and may thus prove useful in prostate cancer treatment. Prostate 55: 159–167, 2003. © 2003 Wiley‐Liss, Inc.
Variation in chromogranin A serum levels during intermittent versus continuous androgen deprivation therapy for prostate adenocarcinoma
Authors: Sciarra Alessandro , Monti Salvatore , Gentile Vincenzo , Mariotti Gianna , Cardi Antonio , Voria Giuseppe , Lucera Rossana , Di Silverio Franco
Published: April 2003

Objectives

It has been hypothesized that continuous androgen‐suppression therapy produces hyperactivation of neuroendocrine (NE) cells and an increase in chromogranin A (CgA) in prostate carcinoma (PC). The aim of this study was to verify whether the intermittent administration of androgen deprivation (IAD) reduces the risk of CgA increase in PC cases treated with complete androgen deprivation (CAD).

Materials and Methods

We analyzed changes in serum CgA levels in patients with PC who successfully responded to the first 24 months of IAD versus continuous CAD therapy. Two different populations were analyzed: Type 1 = pT3pN0M0 prostate cancers with biochemical (PSA) progression after RRP; Type 2 = metastatic PC directly submitted to CAD. Cases in Type 1 and Type 2 population were randomly assigned to IAD versus continuous CAD therapy. Forty cases each in Type 1 and Type 2 population were included in the analysis. At 1, 3, 6, 12, 18, 24 months of IAD versus continuous therapy, serum levels of CgA compared to PSA levels were analyzed.

Results

In population Type 1 and Type 2, in the group of cases continuously treated with CAD (Group 2), there was a significant trend to increase for CgA levels from baseline to 24 months of therapy. On the contrary, no significant variations were found in cases treated with IAD (Group 1). Either in population Type 1 or Type 2, at 12‐ and 24‐month follow‐up, mean and median serum levels of CgA were significantly (P < 0.005) lower in Group 1 than in Group 2.

Conclusions

The present study represents the first evidence in the literature that the intermittent administration of CAD therapy significantly reduces the increase in serum CgA levels during CAD therapy. Prostate 55: 168–179, 2003. © 2003 Wiley‐Liss, Inc.
Intraepithelial and stromal lymphocytes in the normal human prostate
Authors: Bostwick David G. , de la Roza Gustavo , Dundore Paul , Corica Federico A. , Iczkowski Kenneth A.
Published: April 2003

BACKGROUND

The distribution and significance of lymphocytes in the normal human prostate is uncertain. It may be important as a determinant of the immune response, playing a role in hyperplasia and carcinogenesis.

METHODS

We studied 28 normal autopsy prostates from patients between 1 and 64 years of age. Cases with nodular hyperplasia, intraepithelial neoplasia, cancer, and florid prostatitis were excluded. Each prostate was routinely processed to paraffin and immunohistochemically stained for CD20, CD45, and CD45RO. Three frozen samples obtained from these cases were stained for CD8 and CD4. One low power field (25× magnification, 50 mm2) was randomly chosen for examination from each anatomic zone of the prostate, namely the peripheral, transition, and central zone. Lymphocytes were visually counted in the stroma and epithelium of each zone. Lymphocyte counts were converted to number of cells per square millimeter based on the area of the slide per field and the percentage of field that was glandular.

RESULTS

Stromal lymphocytes were more numerous than epithelial lymphocytes (mean, 7.32 and 2.04 cells/mm2, respectively). They were mostly T cells (stromal lymphocytes: 93% CD45RO +; epithelial lymphocytes: 99% CD45RO +). Helper/inducer (CD4 +) T cells predominated in the stroma, whereas those in epithelium were chiefly cytotoxic/suppressor (CD8 +). No difference was found in the number of stromal and epithelial T or B cells according to patient age, race, or location within the prostate (peripheral, central, and transition zones) (all P > 0.05), although the number of lymphocytes in the peripheral zone tended to exceed that in the other zones after correcting for gland:stroma ratio.

CONCLUSIONS

The immune response in the prostate is primarily cell‐mediated. Cell distribution is constant according to patient age, race, and anatomic zonal location within the prostate. The greatest concentration is in the stroma with a small but significant number of intraepithelial cells. The inverted CD4/CD8 ratio in the intraepithelial compartment suggests that cytotoxic/suppressor T cells are the first line of defense against luminal foreign agents reaching the prostate through retrograde flow. Prostate 55: 187–193, 2003. © 2003 Wiley‐Liss, Inc.
Reduced circulating androgen bioactivity in patients with prostate cancer
Authors: Raivio Taneli , Santti Henrikki , Schatzl Georg , Gsur Andrea , Haidinger Gerald , Palvimo Jorma J. , Jänne Olli A. , Madersbacher Stephan
Published: April 2003

Background

Previous studies on immunoreactive androgen levels in serum have revealed equivocal associations with the risk of prostate cancer (CaP). The aim of this study was to compare serum biological androgen activity between men with newly diagnosed CaP and age‐matched men with benign prostatic hyperplasia (BPH).

Methods

Caucasian men with newly diagnosed, untreated CaP (n = 101) and age‐matched patients with BPH (n = 103) were investigated. Serum androgen bioactivity (ABA) levels were measured using a recently developed recombinant cell bioassay.

Results

In comparison to men with BPH, CaP patients with Gleason score ≥8 (n = 16) had lower serum ABA (P < 0.05), and patients with Gleason score ≤5 (P < 0.05) or ≥8 (P = 0.07) displayed suppressed ABA levels in relation to serum testosterone. As the entire group, men with CaP (n = 101) had significantly lower serum ABA than age‐matched men with BPH (n = 103): median 3.0 nM (range, 0.8–6.4 nM) versus 3.2 nM (range, 0.8–7.9 nM) testosterone equivalents, respectively (P < 0.005). By contrast, serum immunoreactive testosterone and SHBG concentrations and free androgen indices did not differ significantly between the two groups.

Conclusions

Patients with CaP have lower serum ABA than controls with BPH, and men with low or high Gleason score display suppressed circulating ABA‐to‐testosterone ratio. These features may reflect interaction between variables such as the degree of tumor differentiation and tumor volume with androgen metabolism. Prostate 55: 194–198, 2003. © 2003 Wiley‐Liss, Inc.
Hypermethylation of GSTP1, CD44, and E‐cadherin genes in prostate cancer among US Blacks and Whites
Authors: Woodson Karen , Hayes Richard , Wideroff Louise , Villaruz Liza , Tangrea Joseph
Published: April 2003

Background

In the US, the incidence and mortality of prostate cancer is about twofold higher among US Blacks compared to Whites, suggesting racial differences in prostate tumor occurrence and aggressiveness. The reason for these racial differences is unknown. Epigenetic events such as promoter‐region gene hypermethylation may be influenced by environmental exposures and have been implicated in prostate carcinogenesis (by the silencing of tumor suppressors and other regulatory genes).

Methods

Using real‐time methylation‐sensitive PCR, we assessed differences in DNA hypermethylation of GSTP1, CD44, and E‐cadherin (three genes thought to be important in the progression of prostate cancer) in archival tumor tissue of black (n = 47) and white men (n = 64).

Results

We found a high prevalence of GSTP1 hypermethylation overall (84%) but no differences by race (89 and 83% in black vs. white men, respectively), tumor stage, or grade. Although CD44 hypermethylation was less prevalent overall (found in 32% of tumors), we observed a 1.7‐fold higher frequency among black men (43 vs. 25% in black vs. white men, P = 0.05) and a correlation with tumor grade (CD44 was hypermethylated in 10, 42, and 52% of well, moderate, and poorly differentiated tumors, respectively, P = 0.003) but not disease stage. The E‐cadherin gene was not hypermethylated in any of the tumors. In summary, of the three genes examined, only CD44 hypermethylation differed by race and correlated with tumor grade, independent of race.

Conclusions

These preliminary findings suggest that differences in gene promoter hypermethylation may potentially underlie racial differences in prostate cancer pathogenesis and should be explored in larger studies. Prostate 55: 199–205, 2003. © 2003 Wiley‐Liss, Inc.
Expression of ERα and ERβ in prostate cancer
Authors: Linja Marika J. , Savinainen Kimmo J. , Tammela Teuvo L.J. , Isola Jorma J. , Visakorpi Tapio
Published: April 2003

BACKGROUND

It has been suggested that estrogens and their receptors (ERs) may be involved in the development and progression of prostate cancer. To elucidate the significance of these receptors, expression of both ERα and ERβ was measured in benign and malignant prostate tumors, as well as in cell lines.

METHODS

Expression of ERα and ERβ was measured in prostate hyperplasia (BPH, n = 7), androgen‐dependent (n = 30) as well as hormone‐refractory (n = 12) prostate carcinomas, and in four prostate cancer cell lines (LNCaP, DU145, PC‐3, and 22Rv1) using real‐time quantitative RT‐PCR.

RESULTS

Only low‐level expression of ERα was found in all tumor types and cell lines. The level of expression was similar to that observed in breast carcinomas found to be negative for ERα by immunohistochemistry. All cell lines showed low, but detectable, levels of ERβ expression. The mean expression of ERβ in the hormone‐refractory carcinomas was about half that seen in BPH or the androgen‐dependent carcinomas; however, the difference was not statistically significant.

CONCLUSIONS

The data suggest it is unlikely that alterations in the expression of either ER are commonly involved in the progression of prostate cancer. Prostate 55: 180–186, 2003. © 2003 Wiley‐Liss, Inc.
Intermediate basal cells of the prostate: In vitro and in vivo characterization
Authors: Garraway Levi A. , Lin Douglas , Signoretti Sabina , Waltregny David , Dilks James , Bhattacharya Nandita , Loda Massimo
Published: April 2003

Background

Progenitor cells within the prostate basal layer may play important roles in differentiation and carcinogenesis; however, prostate stem cell populations remain uncharacterized.

Methods

Immunohistochemical and immunoblot analyses were used to characterize prostate epithelial cells (PrEC), a commercially available prostate basal cell isolate.

Results

Proliferating PrECs exhibited immunophenotypic characteristics most consistent with basal cells, but during senescence PrECs up‐regulated androgen receptor (AR) mRNA, p27, and low‐molecular‐weight cytokeratin (LMWCK) expression, suggestive of partial differentiation. PrECs also stained strongly for involucrin, which marked a subset of intermediate prostate basal cells in vivo. Basal hyperplasia consisting of involucrin‐positive cells was prevalent in prostate tissue from androgen‐ablated patients, and formed epithelial clusters flanked by involucrin‐negative basal and luminal monolayers. Cultivation of PrECs on matrigel together with androgen‐treated stromal conditioned media resulted in dense aggregates, with a peripheral rim of basal‐like cells expressing p63 and basal cytokeratins.

Conclusions

PrEC represents an epithelial population whose basal characteristics are modified in response to matrigel, stromal factors, and senescence, consistent with a transient amplifying population. These cells may derive from a previously unrecognized, involucrin‐positive subset present in vivo. Prostate 55: 206–218, 2003. © 2003 Wiley‐Liss, Inc.
Algorithms for computational solvent mapping of proteins
Authors: Kortvelyesi Tamas , Dennis Sheldon , Silberstein Michael , Brown Lawrence , Vajda Sandor
Published: April 2003

Computational mapping methods place molecular probes (small molecules or functional groups) on a protein surface to identify the most favorable binding positions by calculating an interaction potential. We have developed a novel computational mapping program called CS‐Map (computational solvent mapping of proteins), which differs from earlier mapping methods in three respects: (i) it initially moves the ligands on the protein surface toward regions with favorable electrostatics and desolvation, (ii) the final scoring potential accounts for desolvation, and (iii) the docked ligand positions are clustered, and the clusters are ranked on the basis of their average free energies. To understand the relative importance of these factors, we developed alternative algorithms that use the DOCK and GRAMM programs for the initial search. Because of the availability of experimental solvent mapping data, lysozyme and thermolysin are considered as test proteins. Both DOCK and GRAMM speed up the initial search, and the combined algorithms yield acceptable mapping results. However, the DOCK‐based approaches place the consensus site farther from its experimentally determined position than CS‐Map, primarily because of the lack of a solvation term in the initial search. The GRAMM‐based program also finds the correct consensus site for thermolysin. We conclude that good sampling is the most important requirement for successful mapping, but accounting for desolvation and clustering of ligand positions also help to reduce the number of false positives. Proteins 2003;51:340–351. © 2003 Wiley‐Liss, Inc.
Characterization of carbonic anhydrases from Riftia pachyptila, a symbiotic invertebrate from deep‐sea hydrothermal vents
Authors: De Cian Marie‐Cécile , Bailly Xavier , Morales Julia , Strub Jean‐Marc , Van Dorsselaer Alain , Lallier François H.
Published: April 2003

The symbiotic hydrothermal vent tubeworm Riftia pachyptila needs to supply its internal bacterial symbionts with carbon dioxide, their inorganic carbon source. Our aim in this study was to characterize the carbonic anhydrase (CA) involved in CO2 transport and conversion at various steps in the plume and the symbiotic tissue, the trophosome. A complete 1209 kb cDNA has been sequenced from the trophosome and identified as a putative α‐CA based on BLAST analysis and the similarities of total deduced amino‐acid sequence with those from the GenBank database. In the plume, the putative CA sequence obtained from cDNA library screening was 90% identical to the trophosome CA, except in the first 77 nucleotides downstream from the initiation site identified on trophosome CA. A phylogenetic analysis showed that the annelidan Riftia CA (CARp) emerges clustered with invertebrate CAs, the arthropodan Drosophila CA and the cnidarian Anthopleura CA. This invertebrate cluster appeared as a sister group of the cluster comprising mitochondrial and cytosolic isoforms in vertebrates: CAV, CAI II and III, and CAVII. However, amino acid sequence alignment showed that Riftia CA was closer to cytosolic CA than to mitochondrial CA. Combined biochemical approaches revealed two cytosolic CAs with different molecular weights and pI's in the plume and the trophosome, and the occurrence of a membrane‐bound CA isoform in addition to the cytosolic one in the trophosome. The physiologic roles of cytosolic CA in both tissues and supplementary membrane‐bound CA isoform in the trophosome in the optimization of CO2 transport and conversion are discussed. Proteins 2003;51:327–339. © 2003 Wiley‐Liss, Inc.