Kinetics of cell replication of the uterine cervix. II. Parabasal cell production in atypical epithelium.

The rate of cell division during a 6-hour period of time was determined in cervices from 31 mice, 16 having normal epithelium, 8 with atypical epithelium and 7 with invasive carcinoma. Cervical atypias and carcinoma were induced by local application of benzo(a)pyrene (BP) and mitosis was arrested with vincrotal number of cells in all layers as well as the proportion of dividing cells in the parabasal layer were significantly greater in atypias and invasin cell production in that layer.

Ultrastructure of normal epithelial cells in Papanicolaou-stained cervical smears. An application of a modified open-face embedding technique for transmission electron microscopy.

The results of an open-face embedding technique applied to Papanicolaou-stained uterine cervical smears are presented. By this technique, cells are smeared on a plastic sheet, fixed in 1.5% buffered glutaraldehyde and stained by the Papanicolaou method; areas of special interest are then selected by light microscopy for electron microscopic observations. Thus, a comparison is possible between the light microscopic characteristics of cells in smear preparations and their ultrastructural counterparts. The good ultrastructural preservation allowed detailed study of nuclei and cytoplasm. The endocervical cells had intact nuclei with nuclear envelope, Golgi apparatus, well-preserved mitochondria and two types of secretory material. The superficial and intermediate squamous cells showed loss of desmosomes; dense, fine cytoplasmic fibrils; small, fatty droplets; and degenerated nuclei with loss of the nuclear envelope. Compared with ultrastructural studies on cervical tissues, the intermediate cells in cervical smears were more degenerated, presumably because predominantly degenerated intermediate cells exfoliate. This study indicates that the light microscopic impression of the nuclear appearance is misleading: in the routine smear the nuclei of superficial and intermediate cells may look "intact" and "well preserved" whereas ultrastructurally the nuclei are degenerated.

A comparison of thiomersal and 50% alcohol as preservatives in urinary cytology.

The efficacy of 50% ethyl alcohol and of thiomersal as preservatives in urinary cytology were compared. In both methods over 80% of the cells were sufficiently well preserved after three days to allow cytomorphological evaluation, and over 50% on the seventh day. In the specimens without preservative, only 54% and 28% were intact after the same time intervals. In contrast with 50% ethyl alchohol, thiomersal is a more effective bactericide, it does not increase the volume of the sample, it is cheaper, and it does not affect the cytomorphology.

Effect of various routine cytopreparatory techniques on normal urothelial cells and their nuclei.

Nuclear and cytoplasmic sizes of cells in permanent, stained smear preparations differ from those in unfixed unstained cells. In air-dried MGG-stained smears the area of the nucleus is 50% larger and that of the cytoplasm 30% larger. In wet-fixed Papanicolaou-stained smears the nucleus is 10--30% and the cytoplasm is 15--55% smaller. The shrinkage in the wet fixation method is dependent on the concentration of the ethyl alcohol applied. The staining method has relatively little influence on nuclear and cytoplasmic size. The three-dimensional appearance of the smeared, stained cells is also dependent on the cytopreparatory technique applied:in the methods with air-drying the nuclei and cells are flat and in the wet fixation method more spherical. In the methods with air-drying the nuclear:cytoplasmic ratio is larger than that seen with the wet fixation methods.

Cell yield obtained with various cytopreparatory techniques for urinary cytology.

Information concerning the main pitfalls of cytopreparatory techniques, e.g., cell loss, is of paramount importance for general acceptance of the routine use of urinary cytology. Therefore, we studied the cell yield obtained with five Papanicolaou and two May-Grünwald-Giemsa (MGG) smear techniques commonly used in diagnostic cytology laboratories, and compared the results with those obtained with the Millipore filter technique. All smear methods using wet fixation gave very low cell harvests (2-26%), and all methods using air drying gave high cell yield (55-95%), the spray fixation method giving intermediate results (about 40%). Since the morphometric studies showed that there was no preferential cell loss, the cells remaining on the slides can be considered representative of the original cell population. However, if the cell concentration of the urinary sample is extremely low, the Millipore filter and the albumin MGG methods, in both of which cell loss is minimal, are recommended. The results obtained with the spray fixation Papanicolaou method are quantitatively acceptable and qualitatively excellent.

The identification of atypical reserve cells in smears of patients with premalignant and malignant changes in the squamous and glandular epithelium of the uterine cervix.

Atypical reserve cells can be recognized in cervical smears by their characteristic exfoliation pattern and the fragility of their cytoplasm. The frequency of cases with such cells in 30,000 cervical smears is given, as well as that in 440 cases of dysplasia, epidermoid carcinoma in situ and invasive carcinoma. The presence of atypical reserve cells was found to increase in frequency with the increasing severity of the lesions. The bipotential character of these cells is discussed in relation to the possible progression to carcinoma in situ of the adenomatous type in two of the 42 cases of pure atypical reserve cell hyperplasia, and the presence of these cells in the majority of the 43 cases of adenocarcinoma of the uterine cervix in the present series.

Selection of material for training cytotechnologists.

Out of 450 specially selected cases (gynecological and non-gynecological material) presented to 30 students during their educational program lasting 12 months, 20 cases caused the most problems. These 20 cases, in effect "selected by the students", were again presented as "unknowns" at the end of the course. The mean percentage of error improved from 64 per cent to 26 per cent. Furthermore, we gave the ten gynecological "unknowns" to ten cytotechnologists with at least five years' experience and their diagnoses were compared with those of the students at the end of the course. The majority of the diagnoses were similar with the exception of two cases: in one case the experienced cytotechnologists overlooked the scarce adenocarcinoma cells more often. In the other case follicular cervicitis was correctly diagnosed by the trained cytotechnologists but frequently misinterpreted by the students.

Cytological changes due to urinary calculi: a consideration of the relationship between calculi and the development of urothelial carcinoma.

Urinary calculi can induce urothelial cellular abnormalities comparable with those of malignancy; this was found in 11 out of 62 lithiasis cases. Severe cellular changes, comparable with those of carcinoma in situ, may be seen in the epithelium adjacent to a calculus. Squamous metaplasia was frequently observed in cases with staghorn stones in the renal pelvis. The abrasive effect of the calculus may result in many multinucleated cells in the sediment. When the calculi are removed the cytological atypia and the observed multinucleation disappeared, and none of these 62 patients developed urothelial carcinoma. A possible relationship was found between a long-term clinical history of lithiasis and the development of cancer of the upper urinary tract in a study of 92 cases of carcinoma of the ureter and renal pelvis.

The efficacy of urinary cytology in the detection of urothelial tumours. Sensitivity and specificity of urinary cytology.

The sensitivity and specificity of urinary cytology in the detection of urothelial tumours using voided urine and applying simple smear preparation and staining techniques have been assessed. Of the 2704 patients under investigation 207 had urothelial tumours. The first urine analysis was positive in 66% of the patients with urothelial carcinoma; an additional 23% of the patients showed positive cytology in repeat smears, resulting in a sensitivity of 89%. The efficacy of urinary cytology depends on the tumour type: for grade 2 tumours, 79% were cytologically positive, 92% of grade 3 and 98% of the grade 4 tumours. The diagnostic efficacy in cases of carcinoma-in-situ, squamous cell carcinoma and adenocarcinoma was comparable with that of grade 4 carcinomas. Grade 0-1 tumours did not result in positive cytology. In eleven cases of lithiasis and in two cases of cyclophosphamide therapy the cytological diagnosis was positive but no neoplasm could be established histologically; these represent true false positive diagnoses. Thus, the false positive rate, 13 out of 165, was 7.88% and the false negative rate, 61 out of 207, was 29.4% when grades 0-1 were included, but 8.75% (14 our of 160) when grades 0-1 were excluded.

The effect of calculi on transitional epithelium. A clinical and cytological study.

An analysis has been made of the 9 false-positive cytologic diagnoses in 4000 samples of urine from 1500 patients. They could be traced to 23 patients who had stones in their urinary tract. After removal of the stones the urinary cytology became negative. Thorough clinical examination excluded malignancy. The histological findings in one of these patients offers a possible explanation for the exfoliation of malignant cells and gives support ot the hypothesis that stones can cause reversible changes of the urothelium with the appearance of a malignancy. A casual relationship between stones in the urinary tract and the development of urothelial cancer could not be established.