-Coexistence of neural proliferations with early epithelial neoplasias and benign ectopias of the uterine cervix-. / Koexistenz nervaler Proliferate mit frühen epithelialen Neoplasien und benignen Ektopien der Cervix uteri.

OBJECTIVE: The study wanted to test the hypothesis that stromal systems can influence the development of the epithelium. METHOD: The stroma of specimens obtained by conizations of the cervix was histologically investigated. RESULTS: Neural proliferations such as neuromas, neurofibromas and vascular neurofibromatoses were found. They were common in the presence of epithelial lesions. CONCLUSION: Thus, the frequency of neural proliferations correlated with the nature of the epithelial abnormality. This association argues against the conventional concept that the growth of cervical intraepithelial neoplasias is absolutely autonomous.

Quantitative measurements of the changes in protein thiols in cervical intraepithelial neoplasia and in carcinoma of the human uterine cervix provide evidence for the existence of a biochemical field effect.

Quantitative micromethods have been used for measuring reactive protein thiols (PSHr), total reactive protein sulfur (TRPS), total protein thiols (PSHt), and protein disulfides (PDS) in fixed frozen sections of human uterine cervix. PSHr and TRPS were stained using 2,2'-dihydroxy-6,6'-dinaphthyl disulfide; PSHt and PDS were stained using mercurochrome methods. Microspectrophotometric measurements were made on the stained sections using a microdensitometer with associated data processing; the results obtained for areas of epithelium and stroma were converted to absorbance values per micron 2. Samples of uterine cervix that were diagnosed as containing cervical intraepithelial neoplasia (CIN) I-III or carcinoma were examined and compared with samples of normal uterine cervix. Measurements were made not only on identified lesions but also on apparently normal tissue obtained from the same cervix. Epithelial/stroma ratios (E/S) were calculated for PSHr, TRPS, PSHt and PSHt + PDS; in addition, the double ratios of PSHr/TRPS and PSHt/PSHt + PDS were also calculated for E/S. The mean E/S values for PSHr and PSHt were significantly different for all types of lesion compared with control samples. The E/S ratios for apparently normal tissue obtained from cervices with CIN or carcinoma were also significantly different compared with corresponding control values, indicating a field effect. There was a considerable degree of overlap between individual values in the control groups versus those obtained with each type of lesion. The corresponding mean E/S values for TRPS and for PSHt + PDS in the samples containing lesions were not significantly different from control means except for the group containing CaCx. However, the mean values for the double ratios (PSHr/TRPS and PSHt/PSHt + PDS) were significantly different in the groups containing lesions compared with the controls. Moreover, apparently normal tissue obtained from cervices containing CIN or carcinoma had different mean values compared with the controls, confirming the existence of a field effect. The degree of overlap of individual values in the lesion groups compared with the control values was much less with double ratio values than previously noted for single ratio values. In consequence, the double ratio measurements clearly discriminated CIN I + II and CIN-III from controls. Our data show that CIN is associated with marked changes in tissue protein thiols and disulfides and that these differences extend to neighboring apparently normal tissue indicative of a field effect.

Histophotometry of protein thiols and disulphides in tissue samples from the human uterine cervix and the skin reveals a "field effect" as well as an "extended field effect" of malignant tumours.

Fresh frozen and fixed serial sections were stained with 2,2'-dihydroxy-6,6'-dinaphthyldisulfide (DDD) and Fast blue B for reactive protein thiols (PSHr) and total reactive protein sulfur (TRPS). The mean optical densities of PSHr and TRPS determined histophotometrically at a distinct part of a tissue were related to each other (PSHr: TRPS). If this quotient has been determined, e.g. for normal epithelium and the adjacent stroma, both quotients can be related to each other by a double quotient (Q PSHr: TRPS). With the aid of the double quotient highly significant differences could be found between tumours and normal tissue from patients without tumour of the human uterine cervix. Similar differences exist between normal skin from healthy patients and skin tumours. Q PSHr: TRPS revealed similar differences to exist between normal tissue of patients without tumour and apparently normal tissue in the neighbourhood of tumours of the uterine cervix and of skin ("field effect" of tumours). Histophotometric investigations on abdominal skin (and skin of breast) showed highly significant differences between normal skin of patients without tumour and patients with various kinds of tumours of the uterine cervix, ovaries, liver and breast ("extended field effect" of tumours).

Histophotometric quantification of the field effect and the extended field effect of tumors.

Histophotometric investigations have been made on samples of human skin. Fresh frozen serial sections were fixed and stained for either reactive protein thiols (PSHr) or total reactive protein sulphur (TRPS) using modifications of the DDD-Fast blue B-method. In addition, total protein thiols (PSHt) were stained with the Mercurochromcyanide-method, and proteins were stained using a modified amido-black procedure. Significant differences were found between the different tumours investigated and normal tissue, and also between apparently normal tissue adjacent to the tumours and normal tissue from patients without tumour. To reveal such tumour-related changes of apparently normal tissue, termed the field effect of tumours, a double quotient had to be calculated from the PSHr-and TRPS-values determined from both epithelium (epidermis) and connective tissue. In addition, abdominal skin was investigated from patients without tumour and patients with tumours of the female genital tract, liver or breast. With the aid of the double quotient procedure, highly significant differences were found between normal abdominal skin of patients without tumours versus similar samples taken from patients with tumours. The tumour-related changes found with abdominal skin distant from the tumours have been termed the extended field effect of tumours. These general tumour-related changes, independent of the size, state or degree of malignancy of the distant tumour, could be shown to be due to changes in abdominal dermis.

Quantitative cytospectrophotometric studies on protein thiols and reactive protein disulphides in samples of normal human uterine cervix and on samples obtained from patients with dysplasia or carcinoma-in-situ.

Quantitative microspectophotometric studies have been made on sections of human cervix after staining for reactive protein thiol-groups (PSHr), and the sum of protein thiols with so-called reactive protein disulphides (together abbreviated as TRPS). Measurements were made on normal epithelium, apparently normal epithelium adjacent to a pathological lesion, dysplastic epithelium, carcinoma-in-situ, and adjoining stroma. The numbers of cases studied were: normal healthy controls (53); patients with dysplasias (34) and patients with carcinoma-in-situ (29). In the normal control sections the ratio of PSHr in epithelium:stroma was approximately 2.7 and this ratio was strongly decreased in dysplasias (1.6) and carcinoma-in-situ (1.5); the 3 populations of values had sufficient overlap to prevent this measurement being an effective discriminator. No significant variations were observed with TRPS-values except with changes in the stroma adjacent to apparently normal epithelium. However, the ratio of PSHr:TRPS was effectively discriminatory when this double-staining ratio was calculated for epithelial values:stromal values. These results are discussed in relation to the importance of thiol-groups in cell division and cancer, and the biological implications of similar changes observed in neighbouring apparently normal epithelium.

Protein thiols in normal and neoplastic human uterine cervix.

Protein-thiol groups that react with dihydroxydinaphthyl disulphide during a 7 h incubation (so-called reactive protein thiols, PSHr) have been quantitatively measured on sections of human uterine cervix by microcytospectrophotometry. Measurements were made on areas (1 micron 2) of epithelium and adjoining stroma in samples of normal cervix, and in samples obtained from patients with dysplasia, carcinoma-in-situ and invasive cancer. The ratio of PSHr in epithelium to stroma is substantially reduced in the pathological conditions compared with normal and in apparently normal adjacent areas. Such changes in PSHr are discussed in relation to the redox balance of the tissue, and free radical disturbances previously described.

[Neural proliferation as a concomitant finding in neoplasias and benign ectopias of the uterine cervix]. / Nervale Wucherungen als Begleitbefunde von Neoplasien und benignen Ektopien der Cervix uteri.

Histological examinations of the uterine cervix showed neural proliferations structurally resembling microscopically small neuromas, neurofibromas and vascular neurofibromatoses. They were found regularly (83%) with existing epithelial neoplasias, and much less frequently (in 7 of 14 cases) with glandular ectopias with focally developed benign metaplastic squamous epithelium. These pictures were not found in normal cases with unchanged cylindrical epithelium in the cervical canal and unchanged squamous epithelium on the endocervix. The neural structures thus prove to be findings accompanying epithelial alterations. Their frequency is proportional to the extent of epithelial change. Along with other findings, the neural proliferations are taken as an argument against the usual concept of the absolute autonomy of cancerous epithelium.

Histogenesis of spontaneous regression of cervical intraepithelial neoplasias.

It is justifiable to assume that a spontaneous regression of cervical intraepithelial neoplasia is in progress when pathologic epithelium is replaced by normal epithelium. Very rare histologic pictures of this process are shown. Dysplastic squamous epithelium or intraepithelial carcinomas become detached from their bases and are pushed up by replacement tissue in the form of normal squamous or columnar epithelium.

[Colposcopic findings and their histologic correlates]. / Kolposkopische Befunde und ihre histologischen Korrelate.

An evaluation was made of 457 colposcopically guided punch biopsies from patients with suspicious areas in the ectocervical mucous membrane showing leukoplakia, punctations , mosaic structures or atypical transformation zones. Intraepithelial cervical neoplasias could be demonstrated histologically in 56 cases (12.3%). Irrespective of the nature of the epithelial findings, the histological pictures correlated with colposcopic findings in 388 cases (84.9%) as far as epithelial structure, abnormal epithelial-stromal relations, keratoses and stroma alterations are concerned. The correctness of the colposcopic findings can be certified by precise formulation of these changes in the histological report. This additional information is offered to encourage more extensive use of the colposcope . Colposcopy, used routinely and independent of cytology, provides satisfactory quality control for cytology.

[Corresponding behaviour of the epithelium and stroma of the cervix uteri]. / Korrespondierende Verhaltensweisen von Epithel und Stroma der Cervix uteri.

Different types of epithelium in the human cervix are regularly found within circumscribed areas and obviously respect predetermined borderlines. The determination theory may explain these findings and could in consequence lead to the hypotheses of stromal regulation of the epithelium. Corresponding morphological changes of epithelium and associated stroma are evaluated as another relevant aspect. More or less obvious borderlines between two morphologically different environments are found in the stroma underlying epithelium with distinct borders. Such stroma changes were seen to be correlated to benign and neoplastic alterations of both squamous and cylindrical epithelium.

[Possibilities and limits of the quantitative demonstration of protein thiols with the DDD-reagent in cells of cervical epithelium (author's transl)]. / Möglichkeiten und Grenzen der quantitativen Bestimmung von Protein-Thiolen mit dem DDD-Reagens in Cervix-Epithelien.

From six patients, all with invasive carcinoma of the cervix uteri, two groups fo smear preparations, each consisting of eight samples, were taken: The first, group A, before and the second, group B, after mechanical removal of the discharge which was present in abundant quantities. All samples were treated for 24 h with DDD to reach the fast-reaching SH-groups, coupled with Fast Blue B and measured cytophotometrically at 560 nm. Without any exception, the neoplastic cells of group A showed significantly lower extinction values both of the nucleus and of the total cell, if compared to group B. The mean extinction difference amounts to 55% and is highly significant and some evidence is given that the viability of the cells plays an important role. For this reason, the quantitative evaluation of the DDD Fast Blue B does not seem to be useful for cytologic routine investigations.

[Combined use of colposcopy and cytology]. / Zur Frage des kombinierten Einsatzes von Kolposkopie und Zytologie.

The widespread view of the nearly absolute accuracy of cervical cytology in the diagnosis of severe dysplasia, carcinoma in situ and small invasive carcinoma is not correct. The rate of cytologically missed cases computed from the first smear only is more than 10%. Using cytology and colposcopy simultaneously in each single case, the errors of one method will be greatly compensated by the other. Presupposition for the compensation, however, is the independent indication for biopsy. In other words, biopsy must be done based on cytological findings, even when colposcopy is unsuspicious. On the other hand a colposcopically suspected and cytologically unsuspected case must also be biopsied. For the gynecological clinic as well as for the gynecological practice, the additional effort resulting from the combined method is easily realizable. For large screening programs, however, the colposcope can be used only as an instrument for target punch biopsies, indicated by pathologic smears.

[Quantification of proteinthiols in morphologically normal basal cells and pathological squamous cells of the cervix uteri]. / Quantifizierung der Proteinthiole in morphologisch normalen Basalzellen und pathologischen Platten-Epithelzellen der Cervix Uteri.

Smears taken from eight probands with carcinoma in situ or invasive carcinoma of the cervix uteri have been stained with DDD and Fast Blue B. The extinctions microspectrometrically measured at 560 nm are directly proportional to the quantity of protein-SH-groups. The extinctions of the total cell (Eges) and of the cell nucleus (EK) are measured in 67 basal cells (BAS), 78 dysplatic cells (DYS), 122 undifferentiated cancer cells (UNIF) and 89 differentiated cancer cells (POLY). From BAS through DYS and UNIF to POLY EK increases by a total of 176%. In all four cell types investigated, linear correlations between EK and Eges have been found to occur with a probability of over 99%. The straight lines ascertained represent a relation between EK and Eges which is obviously very characteristic for each cell type, and it becomes apparent that the measuring points corresponding to each single cell are in each instance so close to the straight line that in most cases a differentiation of the three pathological cell types is possible even without a morphological criterion. The straight lines corresponding to BAS, DYS and UNIF start from a common origin, whereas the straight line corresponding to POLY branches off from the UNIF line only. This is in accordance with the formal genesis of pathological variants observed in the cervical squamous epithelium or in differentiated carcinomas of the squamous epithelium respectively.

Quantification of protein thiols in morphologically intact cells of the cervical epithelium. II. Effects of cell differentiation.

Protein-SH-groups (PSH) were determined quantitatively in basal (B), parabasal (P), intermediary (I) and superficial (S) cells of normal portio epithelium in order to obtain information about the correlation between PSH and cell differentiation. It was found that total PSH increase from B to S by 67 per cent, the main step occuring from B to P. Nuclear PSH drop continuously from B to S by 80 per cent. Thus, cytoplasmic PSH must raise (by 400%). The area of the increases strongly from B to S (by more than 1000%). Thus, differentiation is accompained by a drastic "dilution" of total PSH and PSH per area unit (1 micron2) both of nuclear and cytoplasm drop considerably. The findings about nuclear PSH are interpreted in terms of the loss of proliferative activity during differentiation, about cytoplamic PSH as expression of decreasing activities of PSH depending metabolic processes.

Microphotometrical quantification of protein thiols in morphologically intact cells of the cervical epithelium. I. Basic results with superficial cells of healthy women.

The investigations are based on Esterbauer's results according to which the staining of protein thiols (PSH) with DDD-Fast Blue B (Barrnett and Seligman) is of a quantitative order. Consequently, the PSH of the nucleus and the cytoplasm of superficial cells of the ectocervical epithelium of healthy women are determined to 0.3 and 4.1 X 10(-14) moles, respectively. After it has been found that the mean absorption spectrum between 310 and 238 nm of the pure proteins, the extinction at 278 nm of a cytoplasmic area (300 to 400 micron2) of unstained cells is taken as a first measure for the cytoplasmic protein content which is calculated to approximately 5 X 10(-18) moles per micron2 whereas the PSH content results to 1.9 X 10(-17) moles per micron2. From this value results a very plausible number of 3.6 SH per 10(5) g of cytoplasmic proteins.