TP53 mutation in ovarian carcinoma.

This short report describes the detection of mutations of the TP53 tumour suppressor gene in sporadic ovarian carcinomas using archival paraffin-embedded tissues and automated fluorescent DNA sequencing. TP53 mutations were detected in eight tumours. Missense mutations predominated and all were transitions. Mutations were commonest in late-stage serous tumours. In three cases, where tissue was available, the mutations were homogeneous throughout several sections of the bilateral ovarian tumours and in omental metastases. These data confirm the findings of previous investigations describing TP53 mutation in ovarian carcinoma and demonstrate that archival paraffin-embedded tissues can be used for such analyses.

Interphase cytogenetics of chromosomes 11 and 17 in fine needle aspirates of breast cancer.

The aims of this investigation were to compare quantitative with qualitative analysis of fluorescent in situ hybridization (FISH) centromere signals in interphase breast cancer cell nuclei and to evaluate the possible clinical utility of detecting numerical abnormalities of chromosomes 11 and 17 by FISH in the preoperative prediction of breast cancer histological grade. Commercial digoxigenin-labeled centromere probes to chromosomes 11 and 17 were hybridized to 69 malignant aspirates with histological follow-up. Aspirates were categorized as disomic or aneusomic for chromosomes 11 and 17 qualitatively; a subset of aspirates was also analyzed quantitatively. The quantitative and qualitative approaches resulted in almost identical categorisation. There was a significant association between the qualitative categorization of aspirates as aneusomic or disomic, the histological grade of the excised tumours (P = .0695, n = 69), and the cytological grade of the clinical aspirates (P = .006, n = 35). Although histological grade III tumors were almost invariably polysomic for one or both chromosomes, polysomy was also detected in grade I and II tumors. Qualitative FISH analysis was shown to be more sensitive than cytological grading in predicting histological grade III but was of lower specificity and was therefore not clinically useful.

Rapid electron microscopy in oncology.

The case is argued for wider use of electron microscopy as an aid to histological diagnosis in problem cases such as tumours of uncertain histogenesis. In practical terms, electron microscopy can produce results as "immediate" as many special stains in regular use. The cost of providing an effective service can be favourably compared with that of various other diagnostic aids commonly called upon in normal clinical practice. It is suggested that exploitation of this growth area of morphological pathology will enhance the attractions of the discipline of histopathology to talented potential recruits.

Value of the necropsy in perioperative deaths.

A series of 213 perioperative deaths was studied out of a total of 1451 consecutive necropsies carried out over three years. Discrepancies between the clinical and the necropsy diagnosis were assessed under four classes of discrepant diagnosis: class I, patient survival affected, treatable; class II, patient survival affected but not treatable; class III, correlated to cause of death but treatable; and class IV, incidental diagnosis which could not have been made before death. Major discrepancies of classes I and II were found in 44 (21%) and 62 (29%) cases, respectively. Minor discrepancies of classes III and IV were found in 63 (30%) and 101 (47%) cases, respectively. No discrepancies were found in 50 (23.5%) cases. These results confirm the continuing value of the necropsy in the assessment of perioperative deaths.

Myoepithelial carcinoma (malignant myoepithelioma) of the parotid gland arising in a pleomorphic adenoma.

A myoepithelial carcinoma, a rare malignant salivary gland neoplasm, arose in a pleomorphic adenoma of the parotid gland. The initial tumour was a pleomorphic adenoma with epithelial and myoepithelial elements. Subsequently the tumour recurred twice and was characterised by invasion of the mandible. Histological examination of the second recurrence showed a malignant spindle cell neoplasm with an infiltrative growth pattern and a high mitotic rate. There was involvement of local lymph nodes. The immunophenotype was characteristic of myoepithelial differentiation: tumour cells stained positively with anticytokeratin antibodies, S-100 protein, alpha smooth muscle actin, and vimentin. Electron microscopy confirmed myoepithelial differentiation, with small foci of keratinocytic phenotype. Large numbers of tumour cell nuclei were reactive with the anti-p53 antibody, DO-7, in contrast to the two previous resections. Thus malignant transformation of a pleomorphic adenoma may involve myoepithelial as well as epithelial elements. Accumulation of p53 protein, perhaps through mutational events, may have played a role in this malignant transformation.

B cell signet-ring cell lymphoma of bone marrow.

A case of signet-ring cell lymphoma affecting the bone marrow and diagnosed by bone marrow trephine biopsy is reported. Normal marrow was replaced totally by cells with large central vacuoles, many of which displaced the nucleus to the periphery of the cell, imparting a signet-ring appearance. Initially, the favoured morphological diagnosis was metastatic signet-ring adenocarcinoma, but on immunocytochemistry the tumour cells were strongly positive for CD45 (leucocyte common antigen) and the B cell marker CD20 (L26). Electron microscopy revealed electron-lucent vacuoles with no discernable internal structure. The tumour was classified as a high grade centroblastic lymphoma using the upgraded Kiel classification. Despite chemotherapeutic treatment, the patient died during an episode of septicaemic shock within two months of presentation.

Necropsies in clinical audit.

The need for specialised forms of clinical audit was highlighted by the report of the Confidential Enquiry into Perioperative Deaths (CEPOD). Necropsy rates in a Northern Ireland teaching hospital were studied with particular reference to perioperative deaths. To provide an overall context for these observations, the pattern of the necropsy services in Northern Ireland as a whole was also determined. For 600 consecutive deaths in a major teaching hospital, the overall necropsy rate was 180 (30%). In the 74 perioperative deaths in this group (as defined by the CEPOD) the necropsy rate was 26 (35%), compared with 16 out of 72 (22%) for other surgical deaths and 89 out of 386 (23%) for medical cases. More coroners' necropsies were carried out in the perioperative group. These figures are within the range of the CEPOD experience. In 1987, in the whole of Northern Ireland, there were 8859 hospital deaths, 520 (5.9%) hospital necropsies, and 516 (5.8%) coroners' necropsies, giving an overall necropsy rate of 11.7%. Outside the two major Belfast teaching hospitals, however, there were 6799 hospital deaths, 76.6% of all hospital deaths for Northern Ireland. In this group there were 180 (2.6%) hospital necropsies and 383 (5.6%) coroners' cases, the overall necropsy rate being only 8.2%. These wide variations reflect the fact that the number of pathologists in post in the peripheral areas of the province falls substantially short of levels recommended by the Royal College of Pathologists. If clinical audit along CEPOD lines is to be effective nationally, more emphasis should be placed on the value of necropsy and local deficiencies in provision will have to be identified and remedied. It is suggested that this could be achieved by combining audit provisions with budgetary incentives.

Malignant fibrothecomatous tumour of the ovary: diagnostic value of anti-inhibin immunostaining.

Malignant ovarian tumours of the fibrothecoma group are rare. The clinicopathological features of a case of ovarian malignant fibrothecoma in which there was metastatic disease in the small intestine and peritoneum at presentation are described. A number of differential diagnoses were considered but positive immunohistochemical staining of the resected ovarian and small intestinal neoplasms with anti-inhibin was of value in confirming a sex cord-stromal tumour and in excluding other lesions. The two tumours were also ultrastructurally identical. Classical malignant fibrothecomas are said to show four or more mitotic figures per 10 high power fields (HPF). Although the intestinal secondary was mitotically active, the primary ovarian tumour contained only one to two mitoses per 10 HPF, showing that formal mitotic counts are not an absolute indicator of malignant behaviour in this group of tumours.

Tartrate resistant acid phosphatase positive splenic lymphoma: a relatively benign condition occurring in a time-space cluster?

Conventional light and electron microscopic studies, together with cytochemical and immunocytochemical staining procedures, were carried out to ascertain whether the lymphomata of four elderly female patients living within 10 kilometers of each other, who presented within a short space of time with massive splenomegaly and varying cytopenia, belonged to any particular subgroup of lymphoma. In each case the lymphoma had a diffuse pattern and mature B cell phenotype. The malignant cells were of uniform cell type, slightly larger than admixed polymorphonuclear leucocytes, and showed minimal nuclear irregularity and positivity for tartrate resistant acid phosphatase (TRAP) staining. Their clinical and morphological features were compared with those of other lymphoproliferative disorders, but while sharing some features in common with each condition, this small group of patients seemed to have a unique combination of findings. The cytopenias of all four responded well after removal of the spleen and their disease has not been aggressive. It is concluded that these patients have a distinct subgroup of lymphoma, which it is important to recognise so that inappropriate use of aggressive cytotoxic drugs can be avoided.

Functional and structural changes of the human proximal small intestine after cytotoxic therapy.

The effects of cytotoxic therapy on the structure and function of the proximal jejunum were studied in six patients receiving intravenous cyclophosphamide (300 mg/m2), methotrexate (40 mg/m2), and 5-fluorouracil (600 mg/m2) as adjuvant therapy for breast cancer. Using a steady state, triple lumen tube perfusion system the absorption of water and electrolytes was measured before and 48 h after administration of the cytotoxic agents. Jejunal biopsies were obtained at each perfusion. Median (range) water absorption fell from 126 (40-142) to 84 (46-142) ml/h/30 cm, with parallel changes for electrolytes; none of the changes was significant. Brush border disaccharidases did not change at 48 h after chemotherapy, while mature enterocytes appeared normal by both light and electron microscopy. Crypt cells and immature enterocytes, however, showed focal vacuolation by light microscopy, corresponding to the occurrence of large residual bodies (secondary lysosomes) containing partially degraded fragments of damaged crypt cells. The confinement of ultrastructural changes to the immature cell population may explain the failure of this study to show a consistent change in the absorptive function of the jejunum 48 h after chemotherapy.

An unusual cause of biliary stricture in a patient with neurofibromatosis type 1.

The majority of biliary strictures occur as a consequence of iatrogenic injury to the extrahepatic biliary tract, with more than 80% following cholecystectomy. The laparoscopic era has led to heightened awareness of this problem. The occurrence of an iatrogenic stricture can be particularly devastating to both patient and surgeon. The literature highlights a number of factors involved in the aetiology of such traumatic stricture formation. We report an unusual case of a Bismuth 2 stricture of the proximal common hepatic duct,occurring in a patient with type 1 neurofibromatosis, following an iatrogenic bile duct injury that occurred during a laparoscopic cholecystectomy. Histological examination of the strictured region of bile duct removed at surgery demonstrated multiple neurofibromas of varying sizes present in the submucosa. Neurofibromatosis type 1 (von Recklinghausen disease) affects the gastrointestinal tract in up to 25% of cases, and in such cases is characterized by multiple submucosal neurofibromas. We believe this is the first reported case of a biliary stricture in a patient with neurofibromatosis type 1, which appeared to be as a consequence of neurofibromas in the submucosa of the bile duct.

Vimentin expression does not assist in predicting survival in ductal carcinoma of the breast.

AIMS: To establish the value of vimentin expression in predicting survival in patients with breast cancer. METHODS: Five-year follow-up data were obtained for 68 patients with ductal carcinoma (NOS) of the breast in whom vimentin expression had been studied in fresh frozen and formalin-fixed, paraffin-embedded tissue. The predictive value on survival of tumour size, growth fraction (as assessed using the Ki67 monoclonal antibody), oestrogen receptor status and Bloom and Richardson grade of the primary tumour, and the presence or absence of lymph node metastases in axillary samples, were also studied. RESULTS: Twenty-two patients died of their disease within 5 years of diagnosis. Vimentin expression either on frozen or paraffin sections did not provide a statistically significant prediction of survival. On univariate analysis tumour grade, size and the presence of lymph node metastases provided prognostic information. Only lymph node status was of independent prognostic importance on multivariate analysis. CONCLUSIONS: Whilst these results confirm the value of established prognostic factors, they do not support the use of vimentin expression in either fresh or fixed tissue for the prediction of survival in ductal carcinoma (NOS) of the breast.

Scanning electron microscopy of mouse intestinal mucosa after cobalt 60 and D-T neutron irradiation.

The stem-cell population of the intestinal crypt is an important model system in experimental radiobiology. Standardized techniques have been developed to allow quantitation of the response of crypt cells to radiation injury following doses of 0-2 krad of D-T neutrons or 60Co gamma rays. These techniques rely on the identification of regenerating crypt cells three-and-a-half days after irradiation. The results are expressed as the number of regenerating crypts per circumference of small intestine, as determined by conventional histological examination; the more profound the injury, the smaller the crypt count. The practical relevance of crypt-counting techniques to clinical radiotherapy is limited by their relative insensitivity; the dose levels commonly used in fractionated radiotherapy produce no detectable response. Scanning electron microscopy of the mucosal surface provides a more sensitive measure of radiation injury. The earliest detectable changes occur at the level of 300 rad of gamma radiation, well below the threshold of the crypt-counting technique. At around 1,000 rad, where the first drop in crypt counts occurs, there are well-marked morphological changes which become more severe with increasing dose levels. Some differences have been observed between the morphological effects of gamma and neutron irradiation at points of radiobiological equivalence in terms of crypt counts (using an RBE value of about 2). The changes observed may reflect more than the disruption of epithelial cell kinetics. Mucosal morphology is the total expression of many different biological parameters of which the regenerative ability of the crypt cells is only one. The surface microanatomy of the gut may be the most sensitive indicator of radiation injury which is conveniently available for study.

C-erbB-2 oncogene product expression depends on tumour type and is related to oestrogen receptor and lymph node status in human breast carcinoma.

Although over-expression of the C-erbB-2 oncogene product can be detected on formalin-fixed, paraffin-embedded sections from tumours of the human breast, fixation results in a considerable loss of immunostaining for this protein. In this study, C-erbB-2 expression was studied immunohistochemically on frozen sections in 63 infiltrating carcinomas, 3 phyllodes tumours, 15 fibroadenomas and 16 cases of fibrocystic disease. The value of C-erbB-2 expression as a prognostic indicator in ductal carcinomas was examined by seeking an association with known prognostic indicators. C-erbB-2 over-expression was confined to ductal carcinomas of the breast and was positively correlated with tumour size, grade, oestrogen receptor status and the presence of metastatic deposits in axillary lymph nodes. Only the association with lymph node and oestrogen receptor status were statistically significant. No evidence of a correlation between C-erbB-2 expression and growth fraction (as assessed with the Ki-67 antibody) was demonstrated in the cases studied. Formalin fixation was associated with the loss of immunoreactivity in all cases studied. 75% of cases which were positive on frozen sections were negative in paraffin sections. These results highlight the significance of the method of fixation used in studying C-erbB-2 expression. They confirm the association between C-erbB-2 expression and known prognostic indicators.

Familial expansile osteolysis. A new dysplasia.

We report 40 cases in one family of an autosomal dominant bone dysplasia, which, though similar in some aspects to Paget's disease, seems unique in some features and in its natural history. The disease shows both general and focal skeletal changes, the latter being mainly in the limbs with an onset from the second decade. Progressive osteoclastic resorption is accompanied by medullary expansion which leads to pain, severe deformity and a tendency to pathological fracture. The serum alkaline phosphatase and urinary hydroxyproline are variably elevated, while other biochemical indices are normal. Most patients had an associated deafness of early onset and loss of dentition. No previous description of this disease has been found in the literature.

Mechanism of actinomycin D-induced resistance in Ridgway osteogenic sarcoma: an ultrastructural study.

The ultrastructural appearances of actinomycin D treated sensitive and resistant sublines of Ridgway osteogenic sarcoma (ROS) have been correlated with the suggested mechanism of drug-induced resistance. The resistant subline (designated ROS/ADX/G2) was developed by repeated suboptimal treatment of tumor bearing animals and passage of the fastest growing tumors. In the present experiments, animals bearing sensitive and resistant tumors were given a single intraperitoneal injection of actinomycin D (0.3 microgram/g) and examined at 1, 6 and 24 h after injection. The principal effect of actinomycin D treatment in both cell lines was the development of nucleolar segregation. This change, however, followed a different time-scale in each case, appearing more prominently at an early stage and returning more quickly to normal in the actinomycin D resistant cell line. These findings can be interpreted as being in agreement with the suggestion that reduced drug retention or an increased rate of detoxification provides the mechanism of acquired resistance.

Gastric paraganglioma: an immunohistological and ultrastructural case study.

This is a report of the immunohistochemical and ultrastructural features of a case of paraganglioma of the stomach in a 61-year-old woman who presented with melaena. The typical 'Zellballen' arrangement of epithelial cells with granular cytoplasm was present and the tumour was highly vascular. The polygonal epithelial cells stained strongly for neuron specific enolase and the intervening elongated sustentacular or support cells were positive for S100 protein and vimentin. Many dense core granules were present within the epithelial cells while the elongated support cells contained plentiful diffuse intermediate filaments. No hybrid cells or ganglion cells were present in the tumour. This is the sixth recorded case of gastric paraganglioma and the first to be studied in detail by immunostaining and electron microscopy.

The declining autopsy rate and clinicians' attitudes.

The autopsy rate has been declining worldwide for decades. This study determined the overall and differential autopsy rates for the Royal Victoria Hospital, Belfast for the years 1997-1999 inclusive. Trends were examined by comparison with previously collected data for the years 1990, 1991 and 1993. Reasons for the decline in autopsy rates as perceived by hospital clinicians were assessed by means of a questionnaire. Over the last decade, there has been a steady decline in the overall autopsy rate from 30.4% in 1990 to 18.4% in 1999. This is due to a decrease in the hospital autopsy rate from 21.6% in 1990 to 7.9% in 1999. The coroner's autopsy rate has remained comparatively unchanged at around 11%. The decline in the overall and hospital autopsy rates involves all of the principal bedholding directorates, but is most dramatic in medicine, surgery and intensive care, where hospital autopsy rates are currently 7% or less. The main reasons for this decline as perceived by clinicians are difficulty in obtaining consent from relatives and advances in modern diagnostic techniques. The findings of this enquiry are in keeping with trends elsewhere, despite repeated studies which clearly demonstrate the continuing value of the autopsy in clinical practice. Recent publicity concerning the retention of organs can only have an adverse affect. Pathologists and clinicians who value the autopsy must become actively engaged in both public and medical education. Renewed emphasis must be placed on the importance of the autopsy in teaching, training and clinically relevant research, and as a means of medical audit.

A preliminary study of scanning electron microscopic changes of the duodenum during healing of duodenal ulcers.

Using the scanning electron microscope, changes in the rat duodenum have been studied during the healing of gastric secretagogue-induced duodenal ulcers on the 1st, 3rd, 7th and 10th days post-infustion. A montage technique with fine resolution has allowed both the entire area of the duodenum and the cellular organization in individual villi in proximity to the ulcers to be investigated. There was a considerable range of appearances noted at any one time point due to the variations in the size and depth of the original area of ulceration. On day 1, there was acute destruction of the mucosal surface with distortion of the villi at the ulcer margin. By the 3rd day, some specimens already showed full epithelialization over the defects whereas others up to day 10 revealed persistent areas of ulceration. Occasional specimens showed areas suggestive of gastric surface metaplasia on the villous surface adjacent to these ulcers.

The ultrastructure of some gastrointestinal lesions in experimental animals and man.

This has been a brief and necessarily selective review, covering only a few of the numerous experimental and diagnostic uses of electron microscopy in the field of gastroenterology. The roles of experimentalist and diagnostician have emerged in a kind of counterpoint. We have identified the contrasting themes of the controllable laboratory experiment and the uncontrollable experiment of disease; of the three-dimensional image of the surface scanning technique and the two-dimensional world of the thin section. There is harmony, also, in our common concern for morphology and our shared interest in any structural change. Altered morphology, whether in tissue architecture or cellular organisation, may offer a key to the better understanding of altered function. In the future, both the experimental and the diagnostic electron microscopist will come to rely more on correlative procedures, such as the re-processing of specimens for a second look with a different technique. Functional dividends are promised to the morphologist by advances in detector technology and associated techniques such as analytical microscopy. It remains to be seen whether medical benefits will accrue, in terms of a more precise diagnosis or a more effective prognosis in individual cases of human gastrointestinal disease.