A.J. Robertson

Observer variability in histopathological reporting of transitional cell carcinoma and epithelial dysplasia in bladders.

Sections from 90 urinary bladder biopsy specimens were examined by 11 consultant histopathologists with varying experience to determine the appropriateness of existing pathology terminology. Analysis with kappa statistics showed fair to good agreement in the grading and staging of transitional cell carcinoma. There was also reasonable agreement in the diagnosis of high grade dysplasia in random biopsy specimens from the urothelium adjacent to the neoplasm, but very poor agreement for lesser degrees of dysplasia. It is concluded that the present classification of bladder carcinomata is reliable and that pathologists can determine stage with a high degree of reproducibility and grade with a fair degree of reproducibility.

Observer variability in histopathological reporting of malignant bronchial biopsy specimens.

AIMS--To evaluate the ability of histopathologists to classify lung carcinomas on bronchial biopsy material using the current World Health Organisation (WHO) classification. METHODS--Eleven histopathologists each reviewed 100 randomly selected bronchial biopsy specimens which had originally been reported as showing lung carcinoma. A single haematoxylin and eosin stained section from each case was circulated and a standard proforma completed. These were analysed using kappa statistics. RESULTS--The histopathologists were excellent at distinguishing between small cell and non-small-cell carcinoma kappa = 0.86), but not so good at subclassifying the non-small cell carcinoma group kappa = 0.25). CONCLUSIONS--The clinically important distinction between small cell and non-small cell carcinoma of the lung is reliably made by competent histopathologists even on limited material.

Observer variability in histopathological reporting of non-small cell lung carcinoma on bronchial biopsy specimens.

AIMS: To evaluate the ability of histopathologists to sub-classify non-small cell lung carcinomas on bronchial biopsy material using the current World Health Organisation (WHO) classification. METHODS: Twelve histopathologists each reviewed 100 randomly selected bronchial biopsy specimens which had originally been reported as showing non-small cell lung carcinoma. For each case, two sections were circulated, one stained by haematoxylin and eosin and the other by a standard method for mucin (alcian blue/periodic acid Schiff). The participants were allowed to indicate their degree of confidence in their classification of each case. A standard proforma was completed and the results were analysed using kappa statistics. RESULTS: Where the participants were confident in their classification, they were actually quite good at sub-classifying the non-small cell carcinoma sections (kappa = 0.71, standard error = 0.058). Overall, however, the results were only fair (kappa = 0.39, standard error = 0.034). CONCLUSIONS: The majority of non-small cell lung carcinomas can be correctly categorised on adequate bronchial biopsy material. Where a confident diagnosis was made, both squamous carcinoma (kappa = 0.73) and adenocarcinoma (kappa = 0.83) were well recognised.

Observer variability in the histopathological reporting of abnormal rectal biopsy specimens.

AIMS--To study the consistency of reporting of abnormal rectal biopsy specimens, especially in the differentiation of inflammatory bowel disease from other causes of abnormality. METHODS--Sixty rectal biopsy specimens were identified from patients presenting with bloody diarrhoea. These were then circulated to the 11 consultant pathologists in the study who filled in a proforma with a list of 12 diagnostic categories and 22 features. RESULTS--Forty one of the 60 cases were examples of inflammatory bowel disease. In 33 of these cases nine or more pathologists had made the diagnosis. Further categorisation into ulcerative colitis and Crohns disease showed better recognition of ulcerative colitis. In the 19 cases of non-inflammatory bowel disease recognition of pseudomembranous colitis and solitary rectal ulcer syndrome was good, but the results were poorer in the case of infective colitis. CONCLUSION--The findings suggest that a group of consultant pathologists can differentiate between inflammatory bowel disease and other causes of an abnormal rectal biopsy specimen and can also recognise pseudomembranous colitis and solitary rectal ulcer syndrome satisfactorily.

A new method of testing for mitogen-induced lymphocyte stimulation: measurement of the percentage of growing cells and of some aspects of their cell kinetics with an electronic particle counter.

Human lymphocytes were cultured with or without added PHA for periods not exceeding one day so that none of the cells had entered the first mitotic division. The size distribution of the cells was measured with an electronic particle counter and the results were collected in a multichannel analyser. An iterative stochastic model was developed to estimate the proportion of responding cells and their growth characteristics. This mathematical model was based on a few simple assumptions about the pattern of cell growth and was considered to fulfill basic requirements of plausibility and parsimony. The technique described in this paper makes measurement of the absolute percentage of responding cells and their average growth rate in mitogen-induced lymphocyte stimulation tests possible in routine diagnostic laboratories for the first time.

Dose-related depression of pha-induced stimulation of human lymphocytes by hydrocortisone

Abstract Hydrocortisone inhibits PHA-induced lymphocyte stimulation by reducing the number of cells entering G1-phase and by slowing the volume growth rate during the first 24 h of culture: the kinetics of cellular recruitment is unmodified. This inhibition has a linear log-dose response curve and there is marked intersubject variation in normal adults. Pre-incubation of the cells with hydrocortisone greatly potentiates its inhibitory effect: when added after the start of PHA stimulation, hydrocortisone becomes progressively less effective until it has lost most of its activity by 5 h, even though the cells do not show any microscopic evidence of having entered the G1-phase at that time. The experiments have elucidated the mode of action of hydrocortisone inhibition on the early stages of PHA-induced lymphocyte stimulation and shown an unexpectedly large intersubject variation in responsiveness that might have therapeutic significance. The methods that have been employed could be adapted for clinical monitoring of the immunosuppressive effect of glucocorticoids in individual patients and for screening synthetic steroids for glucocorticoid activity.

A simple method for determining the extent of cellular contamination in peripheral blood lymphocyte preparations

A method is described for calculating, from the size distribution of the cells, the extent of cellular contamination of lymphocyte concentrates prepared from venous blood. The precision of this method was checked by direct comparison with differential leucocyte counts obtained by direct microscopy on 29 samples chosen because they showed a wide range of intensity of contamination. By virtue of its technical simplicity, the method has proved useful in a diagnostic service laboratory for checking the purity of lymphocyte preparations either before performing lymphocyte function tests or after a period of 24 h in tissue culture.

Serum Inhibitory Factor in Lepromatous Leprosy: Its Effect on the Pre-S-Phase Cell-Cycle Kinetics of Mitogen-stimulated Normal Human Lymphocytes

The sera of ten Egyptian men with long‐standing lepromatous leprosy (LL) (mean duration 17.4 years) that had failed to respond in dapsone treatment were shown to inhibit mitogen stimulation responses of normal human lymphocytes. When first tested, the sera partly inhibited the response to phytohaemagglutinin (PHA) and pokeweed mitogen and virtually abolished that to concanavalin A (Con A): after repeated freezing and thawing, the Con A inhibition had disappeared, whereas the PHA response was still partly Inhibited. The inhibitory serum factor(s) had similar actions on lymphocytes from each of six normal donors. Although the sera varied in potency, they showed similar dose response curves when tested against lymphocytes from a single donor. The principal action of the sera was to reduce the number of cells responding to mitogen, without modifying the kinetics of recruitment or rate of volume growth during G1‐phase in those cells that were unaffected by the inhibitory substances(s). Study of PHA dose‐response curves and of the effect of delayed addition of LL serum suggested that the serum factor(s) act by diminishing the responsiveness of the cells, rather than by reducing the concentrations of free mitogen or by blocking cell membrane mitogen receptors The serum from one apparently healthy attendant, who had nursed leprosy patients for 30 years but who did not have leprosy or other chronic infective disease, inhibited completely stimulation by all three mitogens in a manner different from that of LL sera. Serum from the other 13 control patients did not modify the response of normal lymphocytes to stimulation by any of the three mitogens studied. It was concluded that the inhibitory factor(s) in the scrum of patients, with LL were a consequence of the disease and not of the environment in which the patients lived. Microscopy confirmed that the techniques used for recovery of the cultured cells did not introduce bias into the volume spectroscopy measurements.

Development of asynchrony in growth of normal human lymphocytes during first day of culture after PHA stimulation

Abstract The growth in volume of human peripheral blood lymphocytes after PHA stimulation was measured with an electronic particle counter. The percentage of growing cells and average values describing growth during the elapsed period of culture were estimated by fitting, to the observed data, volume distributions derived from a mathematical model based on simple, plausible and parsimonious assumptions. Most of the responding cells were recruited between 9 and 18 h after the start of culture. The averaged growth rate of the stimulated cells did not change materially between 18 h and the end of the experiment (36 h). Progressive recruitment over the first 18 h of culture is probably the main cause of the asynchronous growth encountered when such cultures are sampled during continuous replication with T c previously estimated at 18–30 h. This conclusion was supported by computer simulation experiments.

Mechanisms of phytohaemagglutinin (PHA) stimulation of normal human lymphocytes: ‘trigger’, ‘push’ or both?

Abstract. The growth in volume of human peripheral blood lymphocytes after stimulation with various concentrations of PHA was measured with an electronic particle counter. The percentage of growing cells and averaged values describing their growth rates during the elapsed period of culture were estimated by fitting to the observed data the volume distributions derived from a mathematical model. With sub‐optimal doses, the percentage of cells stimulated, and their incremental growth rate, increased with increasing dose of PHA, but the time‐course of recruitment into the G1‐phase was similar with all PHA concentrations studied. The results provide strong support for the ‘trigger’ hypothesis that there is a distribution of stimulation thresholds within the lymphocyte population: consequently, increasing mitogen concentration will be expected to result in increased numbers of responding cells within the suboptimal concentration range.