Successful purging of murine plasmacytoma by mafosfamide (ASTA-Z).

Mafosfamide (ASTA-Z) is a chemotherapeutic agent currently in use for in vitro purging of tumor-bearing human BM cells prior to autologous bone marrow transplantation (ABMT). We tested the efficacy of ASTA-Z against mouse plasmacytoma cells MOPC-315 (MOPC), a model of human multiple myeloma. BALB/c mice were injected intraperitoneally with different doses of MOPC preincubated with ASTA-Z. All control mice receiving > or = 10(4) MOPC intraperitoneally (ip) died within 23 days. All recipients of ASTA-Z pretreated MOPC remained healthy for > 180 days. To simulate the clinical situation, BALB/c mice received lethal doses of 10(3) MOPC ip prior to ABMT. Subsequently, mice were treated with cyclophosphamide 200 mg/kg one day prior to syngeneic BMT with 10(7) BMC containing 10(6) MOPC; 90% of the mice receiving unpurged syngeneic BMC died within 45 days whereas all mice transplanted with ASTA-Z-treated BMC/MOPC mixtures remained disease-free for > 100 days. Our results suggest that a similar approach may be successful in patients with multiple myeloma and residual disease prior to cryopreservation of their BM for ABMT. Bone marrow purging with ASTA-Z is effective and under certain conditions could be critical for prevention of relapse following ABMT, provided that effective elimination of residual disease in the host can be achieved by the conditioning regimen prior to ABMT.

An evaluation of automatic coding of surgical pathology reports.

The use of the Systematised Nomenclature of Pathology as the basis of an indexing system to histopathological data is outlined. Two computer programs which perform the task of producing codes from free English summaries are compared. It is concluded that a simple system has a great deal to offer the pathologist who is prepared to accept a set of constraints.

Experience with the sodium sulphate-Alcian Blue stain for amyloid in cardiac pathology.

The sodium sulphate-Alcian Blue (SAB) method, which stains amyloid green, was evaluated in 220 hearts from elderly patients. The technique proved superior to the Congo red, crystal violet, and thioflavine T methods used either singly or as a battery for the demonstration of cardiac amyloid. Amyloid was easily identified under the X3 objective, even in small amounts. A few non-amyloid components stained varying shades of green but were easily distinguished on morphological grounds. No false positive or equivocal reactions occurred, and in particular elastic laminae and paravascular connective tissue were not tinctorially confused with amyloid. The SAB stain is technically simple and consistently reproducible, and no special light source is required for examination. An additional advantage in cardiac pathology is the simultaneous demonstration of any fibrosis, basophilic myofibre degeneration, tissue mast cells and mucoid degeneration of valves present.

Shape and texture analysis of liver cell nuclei in hepatomas by computer aided microscopy.

Nuclei from non-neoplastic hepatocytes and from hepatoma cells have been examined by image analysis techniques which measure size, shape, and texture. There are significant differences in these parameters between malignant and non-neoplastic cells. When values are obtained from multiple cells in a single biopsy, discriminant programs satisfactorily separate benign and malignant tissues. Techniques of image analysis offer an opportunity to quantify the nuclear changes associated with malignancy and may provide an objective basis for tumour grading systems.

Measurement of striated muscle fibre diameters using interactive computer-aided microscopy.

We have measured muscle fibre diameters using two methods of interactive computer-aided microscopy. They are simple to perform, reproducible and more convenient than manual methods of measurement. The technique is of general application to histological measurement.

Morphometric study of liver cell nuclei in hepatomas using an interactive computer technique: (i) nuclear size and shape.

Liver cells from 20 normal livers and 20 hepatomas have been studied in histological sections using an interactive computer method which measures nuclear size and shape. The variables which gave best discrimination between malignant and benign nuclei were the standard deviations of nuclear shape measurements. Though the liver is an ideal tissue for computer study it is considered that such measurements may act as a model for analysis of nuclei of other tissues.

Pulmonary oedema at necropsy: a combined pathological and radiological method of study.

Fixation of lungs at necropsy by inflation with formaldehyde vapour was used in a combined radiological and pathological study of pulmonary oedema. Pulmonary oedema was found in 79% of lungs examined. The earliest phases affect the interstitial tissue with oedematous connective tissue planes and distension of pulmonary lymphatics. These changes may be associated with reduction in the compliance of the lung. Alveolar filling is a late stage in the accumulation of oedema fluid in the lungs.

Investigation of the organelle pathology of skeletal muscle in chronic alcoholism.

The muscle abnormalities associated with chronic alcohol consumption were studied by applying histological and biochemical techniques to tissue obtained by percutaneous needle biopsy from the quadriceps muscles of 41 patients. Measurement of the fibre size showed atrophy of both type I (p less than 0.05) and type II (p less than 0.001) fibres. The degree of atrophy was more severe for type II fibres (33% reduction in median diameter) than type I (17%). Marker enzyme activities for the principal organelles were assayed. Compared with biopsy specimens from non-alcoholic controls, no differences were found in the activities of lysosomal, mitochondrial, peroxisomal, cytosolic, sarcolemmal, or sarcoplasmic reticulum enzymes, expressed per microgram DNA. A reduction in the protein to DNA ratio was evident in severely atrophic biopsies, and this was associated with a significant reduction of myofibrillary Ca2+-ATPase activity. These results suggest a selective loss of type II fibre myofibrillary protein and do not confirm earlier suggestions of specific mitochondrial damage.

Central nuclear counts in muscle fibres before and during treatment in hypothyroid myopathy.

Serial percutaneous needle muscle biopsies of vastus lateralis were studied in 10 patients who had varying degrees of hypothyroid myopathy. The biopsies were taken before and during treatment with l-thyroxine. Before treatment the most severely clinically affected patients showed type II muscle fibre atrophy and loss, together with increased central nuclear counts preferentially affecting the type II muscle fibre. Both the type II muscle fibre atrophy and loss and the increased central nuclear counts tended to return towards normal values during treatment with l-thyroxine. The severity of myopathic symptoms before and during treatment correlated with the biochemical evidence of hypothyroidism, a type II fibre atrophy, and increased central nuclear counts. The severity of myopathic signs before and during treatment was correlated with both a type II fibre atrophy and loss and increased central nuclear counts. There was no evidence that the myopathic signs before and during treatment were related to the biochemical parameters of hypothyroidism, except the level of thyroid stimulating hormone. It is suggested that sequential studies of muscle fibre percentages, diameters, and central nuclear counts may provide an additional method of assessing the response to treatment in hypothyroid and possibly other types of myopathy. When increased central nuclear counts are confined to a specific muscle fibre type, this may suggest a hitherto unsuspected specificity of muscle fibre damage.

Measurement in jejunal biopsies by computer-aided microscopy.

A method of morphometric analysis of jejunal mucosa using computer-aided microscopy is described. It is based on the technique of interactive computer graphics with definition by the observer of areas and lengths for measurement, but it also includes automatic measurements based on grey-level segmentation. The analyses are performed rapidly and efficiently. The technique is applicable to the analysis of biopsy tissue from many sites.

Immunohistochemistry in district general hospitals in England and Wales.

The findings of a survey on the use of immunohistochemistry in district general hospitals in England and Wales are reported. Immunohistochemistry is used in most district hospitals, contributes to the accuracy and objectivity of some histopathological diagnoses, and is considered to be generally useful though not without drawbacks. Its expansion is being hindered by lack of funds for reagents and staff. In a few regions attempts are being made to rationalise expenditure and coordinate development of the service. We believe that if this can be done at a regional or national level then the relatively small cost entailed will be justified by a resulting improvement in the quality of patient care.

Morphological basis of radiological band shadows on chest radiographs.

Lungs of unselected cadavers were fixed at necropsy using a formalin vapour technique. "Band shadows" were identified in the excised lungs and these were correlated with in vivo radiographs and with the morphological changes in the lung. Persistent shadows were produced by pulmonary infarction, subsegmental atelectasis, and septal fibrosis singly and in combination. Potentially transient shadows were seen in association with atelectasis and pulmonary oedema.

Immunocytochemical studies of blood group A, H, I, and i antigens in gastric mucosae of infants with normal gastric histology and of patients with gastric carcinoma and chronic benign peptic ulceration.

Indirect immunofluorescence studies of blood group A, H, I, and i antigens were performed on the gastric mucosae and tumor tissues of patients with gastric carcinoma, on the mucosae of patients with chronic benign gastric ulceration, and on the mucosae of infants who had died of causes other than gastrointestinal disease. The following findings were of particular interest: (1) Normal 'secretor' type mucosae were distinguishable from 'non-secretor' type mucosae by the uniform staining of the A or H antigens at the surface and in the pits. Normal 'non-secretor' type mucosae showed little staining of the H or A antigens but, instead, there was staining with anti-I(Ma) antibody. Staining with anti-I(Step) and anti-i(Den) did not show a clear correlation with the 'secretor'/'non-secretor' status of the normal mucosae. (2) Apparently normal areas of gastric mucosae of patients with gastric carcinoma or the normal part of gastric mucosae of patients with benign gastric ulcer frequently showed focal areas of loss or gain of the blood group antigens as is often seen in gastric carcinoma tissues. (3) In the mucosae of patients with intestinal metaplasia there was marked loss of A/H antigens in 'secretors' and I(Ma) antigen in 'non-secretors'. (4) Staining characteristics of tissues from gastric carcinoma were:(a) Focal loss of the expected A/H or I antigens was observed with much variation in staining from area to area, but only a minority showed complete loss of the expected staining. (b) A majority of the carcinomas from 'secretors' showed foci of substantial staining with anti-I(Ma) in contrast to normal 'secretor' mucosae. This is probably due to incomplete biosynthesis of A/H determinants. (c) Incompatible A-like staining by a rabbit anti-A serum was observed in one out of nine adenocarcinomas from blood group B or O persons. (d) A few cases showed substantial i antigen staining. The aberrant expression of blood group A, H, I, and i antigens in neoplastic as well as in some areas of morphologically normal mucosa of patients with benign and malignant diseases of the stomach is discussed in the context of current biochemical knowledge.

Chronic alcohol excess is associated with selective but reversible injury to type 2B muscle fibres.

Patients drinking more than 100 g alcohol/day for longer than three years develop atrophy of striated muscle fibres. This predominantly affects type 2B fibres which are dependent on anaerobic glycolytic metabolism. Atrophy of type 1 and type 2A fibres, which in addition use aerobic mitochondrial respiration, only occurs in the most severe cases and then only to a lesser degree. Abstention from alcohol reverses the changes in muscle which slowly return to normal. Selective injury to type 2B fibres indicates that search should be made for an alcohol-induced biochemical lesion affecting the anaerobic glycolytic pathways of the muscle fibre.

Density of mucosal mast cells in the lamina propria of the colon and terminal ileum of children.

Ileal and colonic biopsies from children observed to be histologically normal were fixed in Carnoy's fluid, sectioned, and stained by the chloroacetate esterase reaction. The density of mucosal mast cells was higher in the terminal ileum than the colon, but did not vary significantly within the colon.

Epstein Barr virus (EBV) encoded small RNAs: targets for detection by in situ hybridisation with oligonucleotide probes.

AIMS: To develop a rapid, sensitive, and specific non-isotopic in situ hybridisation (NISH) procedure for the detection of Epstein-Barr virus in formalin fixed, paraffin wax embedded tissues. METHODS: Two low molecular weight RNAs, designated EBER-1 and EBER-2 (Epstein-Barr encoded RNA), were used: cells latently infected with EBV secrete large amounts of EBERs. The method uses digoxigenin labelled anti-sense oligonucleotides, corresponding to sequences in EBER-1 and EBER-2. RESULTS: The use of these probes, in conjunction with high temperature microwave denaturation, ensured that the technique was considerably more sensitive than other in situ hybridisation techniques for detecting EBV. Furthermore, the hybridisation signal was morphologically distinct in that only the nucleus and not the nucleolus give a positive signal. No cross-hybridisation was observed with cells infected with other lymphotropic herpes viruses. CONCLUSION: The sensitivity, simplicity, and rapidity of this technique make it ideal for diagnostic use, and for studies investigating the role of this virus in neoplastic disease.

Blood group antigens in the normal and neoplastic bladder epithelium.

The indirect immunofluorescence technique was used to study the expression of the blood group A and H antigens in fresh-frozen and formalin-fixed paraffin-embedded sections of the bladder mucosa of 21 patients with non-neoplastic diseases of the bladder. Reliable assessment of these antigens could only be made using cryostat sections of fresh-frozen tissues. Also included in this study were tumour tissues of 5 patients with bladder cancer. The blood group A antigen was totally deleted in cryostat sections of the tumour of one patient with aggressive carcinoma and the appropriate antigens were diminished or focally deleted in the four patients with tumours that were non-invasive during one to three years of follow-up. It is concluded that sections of fresh-frozen tissues or sections processed in the absence of glycolipid-extracting solvents should be used for visualising the expression of blood group antigens in bladder tissues and assessing their value as prognosticators of the malignant potential of bladder tumours.

Comparison of prevalence of human papillomavirus antigen in biopsies from women with cervical intraepithelial neoplasia.

Human papillomavirus antigen was found in 39 (16%) of 253 colposcopic biopsies from a group of women at high risk for cervical cancer who had been examined in the early 1970s. Immunohistochemical evidence of papillomavirus infection was found in 20 (30%) of 67 cervical intraepithelial neoplasia lesions infected with wart virus from these patients. When these results were compared with results of a similar study carried out in the early 1980s there was no significant difference in the prevalence of human papillomavirus antigen in cervical intraepithelial neoplasia lesions infected with wart virus from women who had been examined over a decade apart.

Jejunal mucosal morphometry in children with and without gut symptoms and in normal adults.

Nineteen diagnostic peroral biopsy specimens from 18 children without diarrhoea, vomiting, or abdominal pain ('control' children) were compared with those taken from 23 children with diarrhoea of varying aetiology to establish the morphometric characteristics of jejunal mucosa in childhood. Comparison was also made with normal jejunal mucosa from adults. Statistical analysis of each characteristic individually showed no significant difference between the 'control' children and those with diarrhoea, but there were significant differences between the mucosae of 'control' children and those of adults; the villi tended to be shorter and the crypts longer in children. Thirty-seven per cent of specimens from the 'control' children showed a partial villous atrophy, that is, they were abnormal by adult criteria. Discriminant analysis of the features measured showed effective separation of the following groups: normal histology from partial villous atrophy in children, healthy adults from 'control' children, and normal histology in adults from normal histology in children.

Fibre type changes in striated muscle of alcoholics.

Striated muscle from patients taking more than 80 g of alcohol each day shows selective atrophy of the type II fibres which are dependent on glycogenolytic pathways. This atrophy is associated with an excess of glycogen and lipid within the fibres and may represent a selective metabolic insult.