bluuebunny/crossref_metadata_2025 huggingface dataset:
p53 gene point mutations in relation to p53 nuclear protein accumulation in colorectal cancers
Authors: Costa Aurora , Marasca Roberto , Valentinis Barbara , Savarino Marco , Faranda Alessandra , Silvestrini Rosella , Torelli Giuseppe
Published: June 2005

It is known that structural alterations of the p53 tumour suppressor gene cause malignant transformation and tumour progression in colorectal mucosa. In this study, 38 colorectal cancers were analysed for mutations detected in the p53 gene by single‐strand conformational polymorphism and DNA sequence analysis, and the results were compared with p53 protein expression detected by immunohistochemistry. A very strict association (P<0·0001) was found between genetic alterations and protein accumulation, as detected by the PAb 1801 monoclonal antibody. p53 expression and gene mutations were more frequent in rectal than in colonic cancers. No relation was observed with Dukes' stage, even though most of the mutations were at exon 7 in Dukes' A–B cancers and almost all mutations at exon 8 were observed in Dukes' C–D cancers. DNA ploidy was not generally associated with p53 protein expression or gene mutations. However, 83 per cent of cases with exon 5 and 6 mutations were diploid or near‐diploid and 71 per cent of cases with mutations at exons 7 and 8 were aneuploid. Tumours with p53 gene mutations at exon 5 had a higher median [3H]thymidine labelling index (17 per cent) than those with mutations at exons 6, 7, and 8 (11·8 per cent).
An evaluation of the cell cycle‐associated monoclonal antibody Ki‐S1 as a prognostic factor in primary invasive adenocarcinoma of the breast
Authors: Morris E. S. , Elston C. W. , Bell J. A. , Galea M. , Blamey R. W. , Ellis I. O.
Published: June 2005

Immunohistochemical staining with the novel monoclonal antibody Ki‐S1, believed to recognize a cell cycle‐associated antigen, was investigated in 110 cases of invasive carcinoma of the breast. Immunoreactivity indices were compared with disease‐free interval (DFI), overall survival, and a series of other prognostic indicators. Significant positive correlations were found between the percentage of strongly positive immunoreactive nuclei and tumour size, histological grade and type, vascular invasion, and mitotic count. A significant negative correlation was found with age. No significant correlation was found with either DFI or overall survival. Although a correlation with mitotic count does imply that the Ki‐S1 antigen is cell cycle‐associated to some extent, Ki‐S1 does not appear to be a useful prognostic factor in human breast carcinoma.
Lectin‐binding properties of the antral and body surface mucosa in the human stomach—are the differences relevant for Helicobacter pylori affinity?
Authors: Baczako Karl , Kühl Philippa , Malfertheiner Peter
Published: June 2005

Biopsy specimens from histologically normal human gastric mucosa were examined histochemically with the use of lectins. Significant differences were found in the glycoconjugate patterns of the surface mucous cells between the body and the corresponding antrum. Two lectins (Ulex europaeus agglutinin I and Ricinus communis agglutinin I) specific for fucose and galactose residues stained the apical membrane and the intracellular (perinuclear) region of the antral mucosa more strongly than those of the body (P<0·04). Body and antral biopsies from patients with Helicobacter pylori‐positive chronic active gastritis were also studied, but the lectin patterns of Helicobacter pylori‐infected body and antrum did not differ. On comparing these chronically infected gastric biopsies with normal antral mucosa, however, significant changes could be seen with the fucose‐binding lectin as well as with two other lectins (succinylated concanavalin A and peanut agglutinin).

These differences could have been induced directly by bacterial virulence factors or could be due to non‐specific reactions to the inflammatory process.
Endometrial extramedullary haemopoiesis
Authors: Creagh Teresa M. , Bain Barbara J. , Evans David J. , Reid Cecil D. L. , Young Robert H. , Flanagan Adrienne M.
Published: June 2005

Four cases of endometrial extramedullary haemopoiesis are reported, all with associated haematological disease. The diagnoses of a myeloproliferative disorder and thalassaemia trait were made as a consequence of the histological observations and subsequent haematological investigations in two cases. The third case occurred in a patient with an established diagnosis of chronic myeloid leukaemia. The diagnosis of extramedullary haemopoiesis in the final case was made on autopsy material from a patient with multiple myeloma. The endometrium from five other women with known myelofibrosis was examined but extramedullary haemopoiesis was not found. Endometrium from 32 fetuses did not contain haemopoietic elements, excluding the likelihood of the endometrium being a common site for extramedullary haemopoiesis in development. Endometrial extramedullary haemopoiesis is an uncommon finding, but it is worthy of note, as it may herald the presence of an underlying haematological abnormality.
Non‐isotopic in situ hybridization at the ultrastructural level
Authors: Morey Adrienne L.
Published: June 2005

Non‐isotopic in situ hybridization techniques are becoming increasingly widely used at the ultrastructural level, permitting rapid localization of nucleic acid targets with a high degree of resolution. Technical considerations dictate that the great specificity of the method cannot be matched by a similar degree of sensitivity; the value of non‐isotopic ultrastructural in situ hybridization lies in its unique ability to localize nucleic acid targets in relation to submicroscopic cellular structures. This article presents an overview of non‐isotopic ultrastructural hybridization methods and applications.
The phenotypic changes in tumour infiltrating lymphocytes and tumour cells following intra‐arterial infusion of interleukin‐2 in patients with squamous cell carcinoma
Authors: Salter Janine , Maclennan Kenneth A. , Moore Jaqualyn , Dadian Grace , Riches Pamela G. , Gore Martin E.
Published: June 2005

This study evaluates the morphological and phenotypic changes that occur in squamous cell carcinoma of the head and neck when local infusions of interleukin‐2 (IL‐2) are given. Twelve patients were treated with a range of doses of IL‐2 (3 × 103 to 3 × 106 international units/day) by continuous intra‐arterial infusion for 10 days. Biopsies of the tumour were taken pre‐ and 48 h post‐therapy, snap‐frozen, cut, and examined histologically and immunocytochemically. Local infusions of IL‐2 increase the numbers of antigen‐presenting Langerhans cells (CDIa‐positive) and infiltrating lymphocytes, predominantly of the CD3 and CD4 (T‐helper) phenotypes. Locally infused IL‐2 results in the expression of MHC (major histocompatibility complex) class II antigens on the surface of the tumour cells, capillary and post‐capillary endothelial cells, and peri‐tumoural macrophages. Intratumoural NK (natural killer) cells and CD8‐positive (T‐cytotoxic) infiltrating lymphocytes were not increased by this therapy and CD25 (IL‐2 receptor) was only increased in those patients treated at the lower does levels. The system of intra‐arterial cytokine infusion into head and neck tumours developed in this study is a useful model to examine the biological effects of cytokines, since in vivo they are mainly produced and act locally, Furthermore, the infused tumours are easily accessible to biopsy. The results from studies such as this may influence the design of tumour‐targeted cytokine gene therapy programmes.
ICAM‐1 and VCAM‐1 expression in accelerated cardiac allograft arteriopathy and myocardial rejection are influenced differently by cyclosporine a and tumour necrosis factor‐α blockade
Authors: Molossi Silvana , Clausell Nadine , Sett Suvro , Rabinovitch Marlene
Published: June 2005

Expression of the vascular cell adhesion molecules intercellular adhesion molecule‐1 (ICAM‐1) and vascular cell adhesion molecule‐1 (VCAM‐1) occurs in allograft myocardium and in coronary arteries, promoting adhesion and transendothelial migration of inflammatory cells. We therefore investigated, in cholesterol‐fed rabbits 9–10 days following heterotopic cardiac transplantation, whether the reduction of both myocardial rejection and graft arteriopathy with cyclosporine A (CsA) or graft arteriopathy alone with tumour necrosis factor‐α soluble receptor (TNFsr) was associated with suppression of ICAM‐1 and VCAM‐1 expression. Host hearts showed negative immunostaining for these adhesion molecules, whereas donor specimens from untreated (control) rabbits showed moderate immunostaining for ICAM‐1 and weaker immunostaining for VCAM‐1 in the coronary arteries, myocardium (cardiac myocytes), and perivenular regions. The selective reduction of the coronary arteriopathy with TNFsr was associated with somewhat reduced expression of these adhesion molecules in the arteries, whereas CsA also suppressed myocardial rejection and markedly decreased both vascular and myocyte expression of ICAM‐1 and VCAM‐1.
Specific accumulation of exogenous fibronectin in experimental glomerulosclerosis
Authors: Bergijk Eline C. , Baelde Hans J. , De Heer Emile , Killen Paul D. , Bruijn Jan Anthonie
Published: June 2005

The prognosis of patients showing glomerulosclerosis as a complication of an immunologically mediated kidney disease is poor. To improve the diagnosis and treatment of these patients, it is important to understand the processes involved in the development of glomerulosclerosis. In this study, we investigated the molecular composition of experimental end‐stage glomerular sclerotic lesions and their pathogenesis in chronic graft‐versus‐host disease (GvHD) in the mouse and chronic serum sickness in the rat. Accumulation studies were performed to determine the degree of specific trapping of constituents from the circulation. Two different models were investigated to determine whether differences in disease initiation resulted in different compositions of the glomerulosclerotic lesions. In both models, glomeulosclerosis was preceded by expansion of the mesangial matrix and thickening of the glomerular basement membrane (GBM). The end‐stage sclerotic lesions consisted mainly of fibronectin, which appeared to displace the other extracellular matrix (ECM) components peripherally in the mesangial matrix and GBM. The abundance of fibronectin in the lesions was not reflected in the mRNA levels for this component. Indeed, antibodies directed against the cellular form of fibronectin did not stain positive in the end‐stage lesions. These findings, together with accumulation studies, suggest that specific accumulation rather than de novo synthesis of fibronectin plays a major role in the development of experimental glomerulosclerosis, which appears to be independent of the pathway of induction.
DNA content and expression of tumour markers in germ cells adjacent to germ cell tumours in childhood: Probably A different origin for infantile and adolescent germ cell tumours
Authors: Jørgensen Niels , Müller Jørn , Giwercman Aleksander , Visfeldt Jakob , Møller Henrik , Skakkebæk Niels E.
Published: June 2005

The origin of testicular germ cell tumours occurring during childhood is poorly understood. In adults, the classical seminomas and non‐seminomas originate from carcinoma in situ of the testis, which can usually also be detected in seminiferous tubules adjacent to the tumours. In order to contribute with information regarding a possible association between carcinoma in situ and the childhood group of germ cell tumours, we investigated seminiferous tubules adjacent to 13 infantile yolk sac tumours, five infantile teratomas, and six adolescent germ cell tumours of various types, using morphological evaluation, immunohistochemical staining with markers for carcinoma in situ cells, and densitometric DNA measurement of the germ cells. We detected clear differences between the germ cell populations adjacent to adolescent and infantile germ cell tumours. The former were associated with both normal germ cells and carcinoma in situ cells. The presence of carcinoma in situ cells strongly suggested that the adolescent tumours arose from carcinoma in situ cells, like germ cell tumours occurring in adult men. Although we were in doubt in two cases, the infantile germ cell tumours were in general not associated with carcinoma in situ cells. The aetiology of infantile yolk sac tumours and teratomas may therefore be fundamentally different from that of adolescent and adult germ cell tumours. The origin of yolk sac tumours and teratomas remains to be elucidated.
The effect of interleukin‐6 and soluble interleukin‐6 receptor protein on the bone resorptive activity of human osteoclasts generated in vitro
Authors: Flanagan Adrienne M. , Stow Mark D. , Williams Richard
Published: June 2005

The role of interleukin‐6 (IL‐6) in the regulation of bone resorption is and has not been studied using human tissue in vitro. This study exploits a recently described in vitro model, whereby osteoclasts, defined as cells that resorb bone, can be generated from human bone marrow, and investigated the effect of IL‐6 and its soluble receptor on bone resorption, in the presence of 1,25‐dihydroxyvitamin D3[1,25(OH)2 vitamin D3]. Human bone marrow was cultured to form a confluent stroma, sedimented onto devitalized bone slices, and recharged with non‐adherent bone marrow cells. 1,25(OH)2 vitamin D3 increased bone resorption, whereas IL‐6 failed to induce a similar stimulatory effect. Both IL‐6 at 100 ng/ml and soluble IL‐6 receptor protein in the absence of exogenous IL‐6 inhibited the stimulatory effect of 1,25(OH)2 vitamin D3. Bone resorption was never observed when non‐adherent haemopoietic cells were cultured in the absence of stroma but in the presence of IL‐6, which indicates that IL‐6 cannot replace the stromal factor(s) required for the formation of cells capable of resorbing bone. These results suggest that IL‐6 at high concentrations is not a critical cytokine in stimulating osteoclastic bone resorption.
Apoptosis in the human liver during allograft rejection and end‐stage liver disease
Authors: Afford Simon C. , Hubscher Stefan , Strain Alastair J. , Adams David H. , Neuberger James M.
Published: June 2005

The contribution of apoptosis (programmed cell death) to cellular damage in human liver disease is unknown. Using the in situ DNA end labelling method (ISEL), evidence was sought of programmed cell death (PCD) in liver tissue from patients with various liver diseases. In particular, the study aimed to determine whether PCD is involved in either the loss of interlobular bile ducts (vanishing bile duct syndrome—VBDS) or the perivenular hepatocyte drop‐out, both of which are characteristic of irreversible graft rejection. Large numbers of apoptotic hepatocytes were found in pervenular areas in tissues taken from patients with chronic graft rejection. Significant hepatocyte apoptosis, was not seen in long‐term stable allografts, primary biliary cirrhosis, cholestasis, paracetamol‐induced fulminant hepatic failure, or fulminant hepatic failure of indeterminate origin (non‐A, non‐B, non‐C hepatitis). Bile ducts rarely stained positively, but mononuclear cells present in the post‐transplant tissues were frequently positive, showing nuclear or cytoplasmic staining. The presence of cytoplasmic staining suggested that some mononuclear cells had ingested apoptotic DNA from other cellular sources. PCD may thus contribute to the perivenular hepatocyte loss in chronic rejection. The absence of ductular epithelial cell staining suggests that PCD is not involved significantly in the bile duct loss of VBDS. Furthermore, apoptosis of monomuclear cells implies that PCD may be involved in regulating the inflammatory cell infiltration of graft rejection.
Subcellular localization of pyruvate dehydrogenase dihydrolipoamide acetyltransferase in human intrahepatic biliary epithelial cells
Authors: Joplin Ruth , Wallace Lorraine L. , Johnson Gerald D. , Lindsay J. Gordon , Yeaman Stephen J. , Palmer Jeremy M. , Strain Alastair J. , Neuberger James M.
Published: June 2005

In previous histological studies, biliary epithelial cells (BEC) in the liver of patients with primary biliary cirrhosis (PBC), but not controls, reacted strongly with antibodies specific for the major autoantigen associated with PBC, the E2 component of pyruvate dehydrogenase complex (PDC‐E2). In this study we have used transmission electron microscopy (TEM) to document the precise subscellular localization of PDC‐E2 in BEC. Two antibodies which recognize PDC‐E2 were used: affinity‐purified anti‐PDC‐E2 raised in rabbits; and human antibody from the serum of patients with PBC, affinity‐purified against human heart PDC. The intracellular localization of antibody binding was determined by laser scanning confocal microscopy and TEM. Both antibodies bound to the inner membrane of mitochondria in BEC isolated from both patients with PBC and controls, but binding to the external aspect of the plasma membrane was observed only in BEC from patients with PBC. Surface antigen expression in PBC may make BEC immunological targets.
The abnormal isoform of the prion protein accumulates in late‐endosome‐like organelles in scrapie‐infected mouse brain
Authors: Arnold Jane E. , Tipler Carron , Laszlo Lajos , Hope James , Landon Michael , Mayer R. John
Published: June 2005

The prion encephalopathies are characterized by accumulation in the brain of the abnormal form PrPsc of a normal host gene product PrPc. The mechanism and site of formation of PrPsc from PrPc are currently unknown. In this study, ME7 scrapie‐infected mouse brain was used to show, both biochemically and by double‐labelled immunogold electron microscopy, that proteinase K‐resistant PrPsc is enriched in subcellular structures which contain the cation‐independent mannose 6‐phosphate receptor, ubiquitin‐protein conjugates, β‐glucuronidase, and cathepsin B, termed late endosome‐like organelles. The glycosylinositol phospholipid membrane‐anchored PrPc will enter such compartments for normal degradation and the organelles may therfore act as chambers for the conversion of PrPc into infectious PrPsc in this murine model of scrapie.
Distribution of cyclooxygenase isoforms in murine chronic granulomatous inflammation. Implications for future anti‐inflammatory therapy
Authors: Appleton Ian , Tomlinson Annette , Mitchell Jane A. , Willoughby Derek A.
Published: June 2005

Inhibition of the enzyme cyclooxygenase (COX) is the basis for the mechanism of action of non‐steroidal anti‐inflammatory drugs (NSAIDs). COX exists as a constitutive (COX‐1) and a mitogen‐inducible (COX‐2) isoform. The relative contribution of COX‐1 and COX‐2 to inflammation is unknown. This study investigated COX activity and the distribution of COX‐1 and COX‐2 during the development of a murine air pouch model of chronic granulomatous inflammation. COX activity progressively rose and was maximal at day 14. Of the COX metabolites measured, PGE2 was the greatest >6‐keto PGF1α>TXB2>PGF2α. By day 7, COX‐2‐labelled fibroblast‐ and macrophage‐like cells were observed and their number and distribution increased with time. At all time points, endothelial cells of venules in the loose connective tissue of the dermis showed immunoreactivity for COX‐2. After day 14, labelling of capillaries in the granuloma was also observed. This study is the first to show that COX‐2 is the predominant COX isoform in all stages of the inflammatory response. These results suggest that selective inhibition of COX‐2 may prove more beneficial, with fewer gastric and renal side‐effects, than existing NSAID therapy for the treatment of chronic inflammatory diseases.
Analysis of the BCL‐6 gene configuration in diffuse B‐cell non‐Hodgkin's lymphomas and Hodgkin's disease
Authors: Pescarmona Edoardo , Lo Coco Francesco , Pacchiarotti Alberto , Rapanotti Maria Cristina , Cimino Giuseppe , Di Paolo Barbara , Baroni Carlo D.
Published: June 2005

BCL‐6 is a novel proto‐oncogene that codes for a zinc‐finger protein sharing homologies with many transcription factors. It has recently been shown that BCL‐6 is involved in chromosome band 3q27 aberrations in non‐Hodgkin's lymphomas (NHLs) and BCL‐6 rearrangements have been detected in 34–45 per cent of diffuse large cell lymphomas with B immunophenotype. We have studied the BCL‐6 gene configuration by Southern blot analysis in 60 cases of B‐cell NHL and in 17 cases of Hodgkin's disease (HD). BCL‐6 was rearranged in 15/46 (32·6 per cent) diffuse B‐large cell lymphomas, mainly with centroblastic morphology, and in 2/11 (18·2 per cent) follicular (centroblastic‐centrocytic) lymphomas. Conversely, all cases of HD, including four cases of lymphocyte predominant, nodular type (nodular paragranuloma), had a germline configuration. These findings confirm that BCL‐6 is rearranged in a significant percentage of diffuse B‐large cell lymphomas, suggesting that this proto‐oncogene might have a pathogenetic role in this subset of NHLs, but our preliminary analysis suggests that BCL‐6 lesions are not involved in the pathogenesis of HD. However, further investigations using more sensitive techniques are required to confirm these findings.
MDM2 expression in lymphoid cells and reactive and neoplastic lymphoid tissue. Comparative study with p53 expression
Authors: Martinez Juan C. , Mateo Marisol , Sá;nchez‐Beato Margarita , Villuendas Raquel , Orradre Juan L. , Algara Patrocinio , Sáanchez‐Verde Lydia , García Pedro , López Carmen , Martínez Pedro , Piris Miguel A.
Published: June 2005

MDM2 and p53 immunohistochemical protein expression was analysed in lymphocytes and in reactive and neoplastic lymphoid tissue. Phytohaemagglutinin (PHA)‐stimulated lymphocytes displayed MDM2 and p53 co‐expression. In 8 of 8 tonsils, 24 of 24 Hodgkin's disease (HD), and 10 of 24 high‐grade non‐Hodgkin's lymphoma (HG‐NHL) specimens, MDM2 paralleled p53 nuclear expression in non‐tumour and tumour cells. The number of positive cells was greater and the staining intensity was stronger for p53 than for MDM2. In another nine of the 24 HG‐NHL cases studied, dissociated expression was observed, with high p53 expression and very low or absent MDM2 expression. In five cases, both MDM2 and p53 were negative. The eight low‐grade NHL (LG‐NHL) cases were also MDM2‐ and p53‐negative. MDM2 and p53 expression in PHA‐activated lymphocytes and reactive lymphoid tissue is probably an expression of opposing biological signals regulating cell proliferation. Parallel MDM2 and p53 expression in all HD and in 10 out of 24 HG/NHL cases may indicate that this growth suppressive pathway is maintained in those cases. However, dissociated MDM2/p53 expression (nine cases) and the absence of expression of both proteins (five cases) may represent examples of deregulation of this growth control pathway. These findings are in agreement with previous in vitro studies in cell lines regarding the role of MDM2/p53 interaction in the regulation of growth control and extend this observation to normal lymphocytes and reactive lymphoid tissue, suggesting a possible role for MDM2 deregulation in lymphomagenesis.
CD25‐negative hairy cell leukaemia: Intracytoplasmic detection of Tac antigen and interferon‐induced surface expression
Authors: Tison Tiziana , Dazzi Francesco , Giacon Claudia , Vianello Fabrizio , Valente Maria Luisa , Radossi Paolo , Sgarabotto Dino , Girolami Antonio , Pennelli Natale , Poletti Alessandro
Published: June 2005

Hairy cell leukaemia (HCL) is a chronic lymphoproliferative disease of B‐cell lineage. One of the peculiar immunophenotypic markers is the strong expression of the p55 chain of the interleukin‐2 receptor (IL2R), recognized by anti‐CD25 (or anti‐Tac) monoclonal antibody. However, it is known that in rare cases CD25 may not be detectable, even when variant forms of HCL are excluded. The possibility has not been investigated that in these situations CD25 is present in the cytoplasm of the neoplastic cells. This paper describes a case in which the clinical, histological, and electron microscopic features were consistent with a typical HCL. Immunophenotype analysis showed the whole spectrum of markers of HCL, except for the expression of IL2R. The soluble form of the molecule was, however, increased in the patient's serum. Cytospin staining of the neoplastic B cells with anti‐CD25 clearly demonstrated the presence of IL2R in the cytoplasm of hairy cells. When the cells were cultivated in vitro in the presence of interferon‐α2b, CD25 was detectable at the membrane level. These findings suggest that at least some cases of CD25‐negative HCL may express cytoplasmic IL2R.
p53 immunoreactivity and mutation of the p53 gene in smooth muscle tumours of the uterine corpus
Authors: Jeffers Michael D. , Farquharson Maura A. , Richmond James A. , McNicol Ann Marie
Published: June 2005

Mutation and overexpression of p53 have been described in uterine malignant mixed Müllerian tumours and in endometrial adenocarcinoma, where it has been associated with a poor prognosis. This study examines p53 expression and mutation of the p53 gene in benign and malignant smooth muscle tumours of the uterine corpus. p53 expression was evaluated by immunohistochemistry in formalin‐fixed, paraffin‐embedded tissue from 23 leiomyo‐sarcomas, 10 tumours of uncertain malignant potential (TUMPs), and 18 leiomyomas. Single‐stranded conformational polymorphism, (SSCP) analysis of exons 5–8 of the p53 gene was performed on 13 leiomyosarcomas, nine TUMPs, and eight leiomyomas. With microwave antigen retrieval, p53 immunoreactivity was seen in 13/23 leiomyosarcomas, 6/10 TUMPs, and 1/18 leiomyomas. Normal myometrium was unstained. In the absence of microwave treatment, staining was abolished in three leiomyosarcomas, all immunoreactive TUMPs, and the single positive leiomyoma. SSCP analysis revealed mutation in three leiomyosarcomas. There was one mutation in exon 5 in a case with positive immunohistochemistry. Two cases with negative staining showed mutation, one in exon 7 and one in exon 8. Mutation was present in exon 7 in 4/9 and in exon 6 in 1/9 TUMPs. All of these cases showed positive immunohistochemistry. There was no significant difference in outcome between cases with and without positive immunohistochemistry. p53 expression is seen in a significant proportion of uterine leiomyosarcomas. Microwave antigen retrieval increases the proportion of positive cases and also results in positive staining in TUMPs. Mutation of the p53 gene occurs in only a minority of leiomyosarcomas and in a significant proportion of TUMPs. Positive immunohistochemistry does not, however, correlate with the presence of mutation and other factors may be responsible for p53 detection in many cases.
Ultrastructure of rat pulmonary arterioles after neonatal exposure to hypoxia and subsequent relief and treatment with monocrotaline
Authors: King Andrew P. , Smith Paul , Heath Donald
Published: June 2005

A group of rats was born in and spent the first 4 weeks of life at a simulated altitude of 3550 m. Two animals were killed immediately afterwards and the remaining 16 were allowed to recover for various times up to a maximum of 12 weeks at sea‐level atmospheric pressure. On ultrastructural examination, the pulmonary arterioles of hypoxic rats showed muscularization, the new layer of mature smooth muscle cells containing abundant organelles and myofilaments. These cells were bounded by prominent elastic laminae. During the recovery period, the medial layer became progressively thinned, but the cells still retained some characteristics of smooth muscle by 12 weeks' recovery. When a similar group of ten hypoxic rats was allowed to recover for 12 weeks before being given monocrotaline, there was early enlargement of the residual smooth muscle cells in the media of pulmonary arterioles and within 5 weeks there was again a thick layer of medial smooth muscle. This was in contrast to the sparse, weakly muscularized arterioles seen in eight similarly treated rats born under normoxic conditions. The relevance is discussed of these findings to the rare occurrence of primary pulmonary hypertension in people who were born at high altitude but returned to sea‐level during childhood.
Fractal patterns for cells in culture
Authors: Vilela Marcelo J. , Martins Marcelo L. , Boschetti Sandro R.
Published: June 2005

The present study aims to understand the growth of human malignant tumours in vitro, using the geometry of fractals as a method of analysis. The fractal dimensions of HN‐5 and MDCK cell growth patterns have been measured. The first results may suggest the possibility of distinct growth processes characterized by different (time‐dependent) effective fractal dimensions for MDCK and HN‐5 cells. If this is true, the fractal dimension may yet prove to be a useful discriminant for comparing different diagnostic categories.
Non‐Hodgkin's lymphoma, coeliac disease, and Epstein–Barr virus: A study of 13 cases of enteropathy‐associated T‐ and B‐cell lymphoma
Authors: Ilyas M. , Niedobitek G. , Agathanggelou A. , Barry R. E. , Read A. E. , Tierney R. , Young L. S. , Rooney N.
Published: June 2005

A group of 166 patients with coeliac disease was followed for a period of up to 25 years. During this time, 17 patients developed intestinal tumours that were diagnosed as lymphoma, of which 15 cases were available for review. Eleven of the lymphomas were of T‐cell type (enteropathy‐associated T‐cell lymphoma, EATL) and two were of B‐cell type. Two cases were reclassified as undifferentiated carcinoma. The interval between the diagnosis of enteropathy and the onset of lymphoma varied from less than 2 months in four patients to more than 5 years in seven. Seven of the T‐cell and both B‐cell lymphomas were investigated for the presence of Epstein–Barr virus (EBV) by in situ hybridization (ISH) using probes against Epstein–Barr virus‐encoded RNAs (EBERs) and by immuno‐histochemistry with EBV‐specific monoclonal antibodies. All EATL cases were negative, suggesting that EBV is not an important factor in these cases. In one of the B‐cell cases, EBV was detectable by ISH and immunohistochemistry in most tumour cells in the mesenteric lymph nodes, but not in any of the tumour cells in the primary ileal tumour, indicating that in this case EBV infection was a late event in the neoplastic process. These results show that lymphoma may develop any time after the onset of coeliac disease and that in our cases of EATL, EBV was not an important factor. In some cases of EBV‐related neoplasia, virus infection may be a late event.
Determination of transforming growth factor beta1 mRNA expression in breast carcinomas by in situ hybridization
Authors: Walker Rosemary A. , Gallacher Barbara
Published: June 2005

The expression of transforming growth factor beta1 (TGF‐β1) mRNA has been determined in 16 breast carcinomas using in situ hybridization and compared with TGF‐β protein as detected by antibodies against TGF‐βT1 and TGF‐β1 plus TGF‐β2. Digoxigenin‐labelled riboprobes were used, with alkaline phosphatase and immunogold silver detection systems. TGF‐β1 mRNA was only detected in carcinomas in which TGF‐β1 protein was found (9 of 16 cases) and not in those with prominent reactivity for TGF‐β2. RNA preservation was poor in two other cases in which TGF‐β1 protein had been detected. In general, those tumours with greater numbers of cells labelled for TGF‐β1 mRNA had prominent reactivity for TGF‐β1 protein. The mRNA was localized to cancer cells with no labelling of stromal cells, although in a small number of cases scanty staining for TGF‐β1 protein had been observed in stromal cells. The incidence of detection of TGF‐β1 mRNA is lower than the published data from Northern analysis studies of breast carcinomas, suggesting that only higher levels of TGF‐β1 mRNA expression are being detected by in situ hybridization. However, this approach has provided useful information about the cellular sites of expression of TGF‐β1 in breast carcinomas.
The intracellular polymerase chain reaction for small CMV genomic sequences within heavily infected cellular sections
Authors: Ray R. A. , Smith M. , Sim R. , Nystrom M. , Pounder R. E. , Wakefield A. J.
Published: June 2005

The indirect intracellular polymerase chain reaction (in situ PCR) combines the potential sensitivity of the polymerase chain reaction (PCR) with the high specificity and morphological preservation of in situ hybridization (ISH). This study describes a method for the amplification of small, specific cytomegalovirus (CMV) genomic sequences [100 base pairs (bp)] within large formalin‐fixed, paraffin‐embedded tissue sections. A heat‐resistant glue surrounds the section, creating a well which contains a relatively large volume of isotonic reaction solution without a covering mineral oil layer; this optimizes morphological preservation, permits the evaluation of large sections, and allows both denaturation steps and up to 40 cycles of in situ PCR to be performed, whilst progressively concentrating the reaction solution by evaporation during thermal cycling. ISH was performed using a non‐isotopic DNA probe with specificity for CMV, with or without prior in situ PCR amplification, both for samples on slides (fibroblasts and lung) and in suspension (fibroblasts). Samples on slides were evaluated by both blinded studies and image analysis, comparing the intensity of signal (P<0·003) and the numbers of positive cells detected (P<0·007), with or without intracellular amplification. Cells in suspension were analysed by blinded studies on cytospins and by gel electrophoresis of cell lysates. Successful intracellular amplification was achieved in this high copy model.
A megakaryocyte analysis of the bone marrow in patients with myelodysplastic syndrome, myeloproliferative disorder and allied disorders
Authors: Ohshima Koichi , Kikuchi Masahiro , Takeshita Morishige
Published: June 2005

A morphometric analysis was performed on aspirate clots of bone marrow to identify the presence of atypical megakaryocytes after immunohistological staining with a monoclonal antibody against Factor VIII. This study included cases of myelodysplastic syndrome (MDS), myeloproliferative disorder (MPD), aplastic anaemia (AA), idiopathic thrombocytopenic purpura (ITP), chronic myelogenous leukaemia (CML), and control cases free from any haematological disease. Quantitative and qualitative abnormalities of megakaryocytes were assessed using an image analyser and a personal computer to perform a morphometric analysis of the number (per mm2), arrangement (μm), nuclear size (μm2), cell size (μm2), nuclear size/cell size (N/C) ratio, and nuclear contour index (NCI). Micromononuclear megakaryocytes were detected in MDS, while in MPD, large over‐mature magakaryocytes were observed to increase in number. In AA, the megakaryocytes decreased dramatically in number without showing any morphological abnormality. In CML, the megakaryocytes also increased without any remarkable morphological abnormality. In ITP, only the NCI increased. The above results show that MDS is characterized by the presence of atypical micromegakaryocytes, while MPD is characterized by atypical large over‐mature megakaryocytes.
Pathogenesis of segmental glomerular changes at the tubular origin, as in the glomerular tip lesion
Authors: Howie A. J. , Lee S. J. , Sparke J.
Published: June 2005

Segmental abnormalities at the glomerulo‐tubular junction, or tip changes, are a common and early feature in several experimental and human renal disorders. Scanning and transmission electron microscopic study of an experimental model, Lewis rats given anti‐glomerular basement membrane antibodies, showed that at 5 days there were monocytes/macrophages in glomerular capillary loops next to the tubular origin, with abnormalities of visceral epithelium, including prolapse of groups of these cells into the tubular origin. At 12 days, there was contact and adhesion between glomerular capillary loops and Bowman's capsule. Transmission electron microscopic study of human renal biopsies showed that the human tip changes resembled the later lesions seen in rats, with intracapillary foam cells confirmed by immunohistological study to be monocytes/macrophages. These findings show that abnormalities of the glomerular tuft precede adhesion to Bowman's capsule and that the earliest changes recognized by light microscopy in human renal biopsies are relatively late in their natural history, meaning at least several days old. Prolapse of visceral epithelial cells into the tubular origin may explain the characteristic site of tip changes.
Expression of p53 protein has no independent prognostic value in breast cancer
Authors: Pietiläinen Timo , Lipponen Pertti , Aaltomaa Sirpa , Eskelinen Matti , Kosma Veli‐Matti , Syrjänen Kari
Published: June 2005

A series of 392 female breast carcinomas was analysed immunohistochemically for expression of p53 protein with special emphasis on the role of p53 as an independent prognostic factor. Altogether, 54·8 per cent of the carcinomas expressed p53 protein, with the mean [standard error (SE)] fraction of positive nuclei being 17·1 per cent (1·2 per cent). Expression of p53 protein was independent of tumour metastasis at diagnosis, axillary lymph node status, tumour diameter, histological type, tubule formation, proportion of intraductal growth, margin formation, necrosis, DNA ploidy, and S‐phase fraction. A high fraction of p53‐positive nuclei was significantly related to patient age under 70 years, high grade, severe nuclear pleomorphism, dense infiltration of tumour by lymphocytes, high mitotic index, and high apoptotic index (for all,P<0·05). Impaired survival probability in the entire cohort (P=0·05) and in the axillary lymph node‐positive (ANP) tumours (P=0·015) was associated with a fraction of p53‐positive nuclei less than 25 per cent, while in the axillary lymph node‐negative (ANN) tumours, expression of p53 had no prognostic value. In multivariate analysis, independent prognostic predictors included axillary lymph node status, tumour diameter, and mitotic index. In the ANN tumours, tumour diameter, fraction of p53‐positive nuclei, and tumour grade were independent prognostic factors, whereas in the ANP tumours, diameter and mitotic index were the two independent prognostic factors. The results suggest that abnormal expression of p53 protein is only a weak independent prognostic factor in female breast cancer.
In situ detection of Epstein–Barr virus in non‐small cell lung carcinomas
Authors: Wong Maria P. , Chung L. P. , Yuen S. T. , Leung S. Y. , Chan S. Y. , Wang Elaine , Fu K. H.
Published: June 2005

Epstein–Barr virus (EBV) is strongly associated with nasopharyngeal carcinoma and lymphoepithelioma‐like carcinomas (LELC) of foregut‐derived organs. Recently this group of EBV‐associated carcinomas has been expanded by the identification of the virus in conventional adenocarcinomas of the stomach. In situ hybridization (ISH) using a sensitive digoxigenin‐labelled EBER RNA probe was performed on 167 consecutive unselected primary non‐small cell lung carcinomas, to determine the frequency of EBV association in these tumours. Nine cases (5·4 per cent) showed strong EBER signals in the tumour cell nuclei. By immunohistochemistry, four of the EBER‐positive tumours showed patchy expression of the viral latent membrane protein (LMP‐1) and none showed any expression of the EBV nuclear antigen 2 (EBNA2). Morphologically, all the positive tumours were LELC, whereas no conventional type of non‐small cell lung carcinoma showed EBV association. The LELC presented a morphological spectrum from undifferentiated to squamoid or glandular differentiation. The patients showed a male to female ratio of 8:1. The mean age at presentation was 48 years. Smoking was not a risk factor. All patients were alive at follow‐up periods of 23–52 months. Southern blot analysis performed on eight of the nine positive tumours showed a clonal episomal form of EBV, suggesting the clonal expansion of an infected tumour cell early in oncogenesis. These characteristics of the EBV‐associated lung tumours justify their consideration as a distinct clinicopathological entity.
Expression of bcl‐2 oncoprotein in renal cell tumours
Authors: Paraf François , Gogusev Jean , Chrétien Yves , Droz Dominique
Published: June 2005

Expression of bcl‐2 is associated with inhibition of apoptosis and extension of cell survival. The importance of apoptosis in relation to the development and progression of renal cell neoplasia remains undefined so far. In order to determine the expression of bcl‐2 oncoprotein in normal and neoplastic renal cells, 37 renal tumours were investigated by immunolabelling, including 13 clear cell carcinomas, ten tubulopapillary carcinomas, four chromophobic renal cell carcinomas, and ten oncocytomas. Twenty‐six samples of adjacent normal renal tissue served as controls. bcl‐2 expression was correlated with cell proliferation activity as estimated by Ki67 antigen expression, and p53 protein expression in the tumour samples. The results demonstrate that in the normal kidney, positive bcl‐2 immunostaining was present in glomerular parietal epithelial cells, in distal tubular cells, and in sparse proximal tubule cells. Renal cell tumours showed heterogeneous bcl‐2 expression according to the tumour cell type. While the majority of carcinomas of clear cell type were usually negative or contained sparsely distributed positive cells, all tubulopapillary carcinomas were consistently positive for bcl‐2. In oncocytomas and chromophobic carcinomas, there was a low percentage of bcl‐2 immunoreactive tumour cells; some nuclear bcl‐2 positivity was detected in one chromophobic tumour. These findings indicate variable bcl‐2 oncoprotein expression in different types of renal cell tumours, with the highest level of expression in tubulopapillary carcinomas. No clear relationship was found between nuclear grade, cell proliferation activity, and level of bcl‐2 expression. p53 protein was detected in only one tubulopapillary carcinoma.
Expression of vascular endothelial growth factor in renal vascular disease and renal allografts
Authors: Gröne Hermann‐Josef , Simon Mathias , Gröne Elisabeth F.
Published: June 2005

Vascular endothelial growth factor (VEGF) is a dimeric glycoprotein that exerts a proliferative effect specifically on endothelial cells. VEGF can increase vascular permeability and collagenase activity, is chemotactic for monocytes, and may dilate blood vessels. It can be induced by phorbol ester and cAMP in both mesenchymal and epithelial cells. In vitro cell culture experiments suggest that VEGF is upregulated by oxygen deprivation. In this study we tested whether in vivo acute and/or chronic reduction of renal blood flow by vascular obstruction would result in increased expression of VEGF mRNA and protein. Three normal kidneys, five human kidneys with narrowing of preglomerular vessels by vascular rejection or by vasculitis, and eight kidneys with nephrosclerosis and/or diabetic nephropathy were examined. In situ hybridization with 35S‐labelled riboprobes showed a pronounced expression of VEGF mRNA in acutely hypoxic proximal and distal tubules of both the cortex and medulla; VEGF protein was demonstrated in the epithelia of these tubules by immunohistochemistry. In kidneys with chronically reduced blood flow, the majority of atrophic tubules were negative for VEGF mRNA and protein, although interstitial cells expressed VEGF mRNA. In arcuate arteries showing intimal and adventitial fibrosis, some medial smooth muscle cells were positive for VEGF mRNA. In glomeruli with segmental sclerosis, viable podocytes showed a prominent signal for VEGF mRNA. Mesangial cells did not express VEGF in the cases studied. It is possible that hypoxia itself led to the upregulation of VEGF in tubular epithelia and vascular smooth muscle cells. The vasodilatory and permeability‐promoting effects of the endothelial growth factor produced by damaged tubular epithelia may constitute a mechanism to alleviate a decrease in blood flow and substrate availability and to re‐establish vascular integrity.
Thymic epithelial neoplasms in transgenic mice expressing SV40 T antigen under the control of an erythroid‐specific enhancer
Authors: Teitz Tal , Chang Judy C. , Kan Yuet Wai , Yen T. S. Benedict
Published: June 2005

The β‐globin locus control region is known to be a powerful erythroid‐specific enhancer. In an attempt to produce immortalized erythroid percursor cells, transgenic mice have been generated with the simian virus (SV) 40 T antigen gene under the control of the locus control region. As previously reported, transgenic mice did not develop erythroleukaemia, but rather succumbed to insulinomas and poorly differentiated rhabdomyosarcomas. This paper describes additional mice containing this transgene that developed thymomas of the mixed epithelial/lymphocytic type, in which only the epithelial component expressed the T antigen. Epithelial cell lines have been established from these tumours. This system may be useful in future studies on the pathogenesis of thymomas and the function of thymic epithelial cells.
Inhibition of gastric cell proliferation by acetaldehyde
Authors: Matysiak‐Budnik T. , Kärkkäinen P. , Methuen T. , Roine R. P. , Salaspuro M.
Published: June 2005

Helicobacter pylori possesses alcohol dehydrogenase activity and is capable of producing acetaldehyde from ethanol in vitro. Acetaldehyde is a toxic and reactive compound and has been shown to inhibit the proliferation of many different cell lines in vitro. To study the effects of acetaldehyde on the proliferation of gastric epithelial cells in vivo, we employed an immunohistochemical method after labelling proliferating cells with 5′‐bromo‐2′‐deoxyuridine in rats receiving acetaldehyde intragastrically. Chronic (16 weeks) exposure of gastric mucosa to acetaldehyde given to rats in their drinking water in concentrations of 10 or 20 mM resulted in significant (P<0·05) inhibition of gastric epithelial cell proliferation, expressed as 332 ± 36, 348 ± 8, and 695 ± 15 proliferating cells per ten high‐power ( × 400) fields in the groups drinking 10 mM acetaldehyde, 20 mM acetaldehyde, and in controls respectively. In an acute study, significant inhibition of proliferation was observed after as few as 4 days of exposure to acetaldehyde, but only when a higher dose (50 mM) of acetaldehyde was given (438 ± 44 versus 615 ± 19 in controls, P<0·05). The inhibition of gastric cell renewal by acetaldehyde may play a role in the pathogenesis of ethanol‐ and/or H. pylori‐associated gastric diseases by inhibiting normal gastric mucosal protection and repair.
Protein extraction and western blotting from methacarn‐fixed tissue
Authors: Lalani El‐Nasir , Golding Matthew , Hudson Mark , Chieffi Gabriella , Stamp Gordon , Anilkumar T. V. , Sarraf Catherine , Alison Malcolm R.
Published: June 2005

Polyacrylamide gel electrophoresis (PAGE) of proteins with subsequent western blotting has become a routine technique for the analysis of proteins from both cultured cells and fresh whole tissue. We have developed a method to extract proteins from methacarn‐fixed tissue which renders them suitable for SDS‐PAGE and western blotting. With a panel of antibodies to specific intermediate filaments, transforming growth factor‐α (TGF‐α), and albumin, immunohistochemistry was performed in parallel with western blotting on sections cut from methacarn‐fixed samples of normal rat liver and liver from rats treated under a regime which induces oval cell proliferation. Immunohistochemistry enabled the determination of changes in tissue distribution and abundance of the target proteins, which was mirrored by the corresponding western blot data. This technique can be especially effective when used in conjunction with immunohistochemistry. Tissue samples are easy to prepare, avoiding the precautions which need to be taken when handling fresh tissue (Abstract:J Pathol1994;173S: No. 41).