Image analysis cytometry of dysplastic nevi.

Computerized image analysis was used to assess nuclear atypia in 24 dysplastic nevi (DN), 19 CN (CN), and five thin melanomas. DN were selected for the study using architectural criteria alone. Feulgen-stained, 6-um sections were analyzed with a microTICAS cytometer. At least 100 nuclei were measured in each case. The standard deviation of nuclear area, mean nuclear roundness, standard deviation of nuclear roundness, mean ploidy, and standard deviation of ploidy were found to be significantly greater for DN than for CN. DNA histograms from DN showed an increased fraction above 2N, suggesting that DN are more proliferative than CN. No DN were aneuploid. All melanomas were aneuploid, and differed significantly from DN in mean nuclear area, standard deviation of nuclear area, mean ploidy, and standard deviation of ploidy. There were no significant differences between the junctional and intradermal populations of compound DN in any of the measured parameters, except that the intradermal nuclei were significantly rounder than the junctional nuclei. There were no significant differences between DN from patients with only a single DN and DN from patients with at least two dysplastic nevi.

Evaluation of brush cytology as an independent technique for detection of esophageal carcinoma.

In this study the accuracy of indirect brush cytology for detection of esophageal carcinoma is evaluated against current standard methods of diagnosis and is compared with the known accuracy rate of endoscopically directed brush cytology. A standard endoscopic nylon brush placed inside a nasogastric tube was used in 203 patients with various esophageal problems. Correct diagnosis was made in 78% of cancers, 95% of potentially premalignant cases, and 100% of cases of normal esophageal mucosa with both indirect and directed brushing procedures. The technique meets most requirements of a new screening procedure as being simple, safe (no complications), relatively inexpensive, and acceptable to patients (98% compliance). Currently it is employed to monitor high-risk esophageal conditions and post-treatment courses of patients with pharyngoesophageal tumors for local recurrence or a new primary lesion in the esophagus.

Automated image analysis in clinical pathology.

The applications of automated cell image analyses are primarily in cytopathology, hematology, and cellular immunology. This paper presents data evaluation technics and statistical methods for automated cell analysis.

DNA ploidy profiles as prognostic indicators in CIN lesions.

The prognostic significance of DNA ploidy measurements in cervical tissues was examined. Microphotometric measurements of 302 Feulgen-stained tissue sections (91 normal squamous epithelia, 14 condylomata, 29 cervical intraepithelial neoplasia (CIN) I, 78 CIN II, and 90 CIN III) were performed with a personal computer (PC)-based video microphotometry system. Analysis of these data shows that the DNA profile provides significant prognostic information: CINs with a polyploid DNA profile are more likely to return to normal than are those exhibiting an aneuploid pattern. Of 211 abnormal cases, 38% had polyploid DNA profiles and 62% were aneuploid. Eight-six percent of the cases that regressed were polyploid and 14% were aneuploid. Of the 130 aneuploid DNA cases, 95% remained static or progressed and only 5% regressed. Of these nonregressing aneuploid lesions, 90 remained static and 34 progressed, whereas within the nonregressing polyploid group 37 remained static and only 6 progressed. This result holds across diagnostic categories. Several other ploidy-related descriptors also showed prognostic significance (including mean ploidy, the 5N exceeding rate and 2N deviation index, and discriminant functions derived from order statistic analysis of the cumulative DNA histograms), but not to the degree or with the consistency of expression as the DNA profile categorization. These results indicate that important information about the prognosis of CIN lesions may be obtained by the DNA profile on Feulgen-stained tissue specimens. The data were acquired by a cytophotometry system of relatively modest cost consisting of readily available hardware components.

Differentiation by TICAS analysis of cell populations of tracheal aspirates from hamsters with squamous-cell carcinoma.

Individual cells from the tracheal aspirates of hamsters exposed to benzo-a-pyrene were scanned at .5 mum in three colors. Features relating to size, shape, and color were extracted and calculated by computer. The single cells were then classified by these features into separate populations with varying degrees of atypia, extending up to frank cancer cells. A high degree of accuracy was attained in classification by these methods.

Follow-up study of male and female offspring of DES-exposed mothers.

This follow-up study presents the effects of DES on the genital tract of male and female offspring of mothers who were part of a double-blind, placebo-controlled investigation during 1951 and 1952 aimed at determining the effect of DES on pregnancy. Epididymal cysts, hypotrophic testes, and capsular induration were the more common genital lesions found in 25% of 163 DES-exposed males as compared to 6% in 168 control males. Semen analysis data on 39 subjects of the DES-exposed group and 25 subjects of the control group showed that 26% of the DES-exposed group produced an ejaculate volume under 1.5 ml; no such cases were observed in the control group. The average values for sperm density ant total motile spermatozoa per ejaculate, although in the normal range, were more than two times lower in the DES-exposed group as compared to the controls. A quality score of greater than 10 ("severely pathologic semen") was found in 28% of the DES-exposed group as compared to 0 in the control group. An association of pathologic semen quality with physical abnormalities was found only in the DES-exposed group. Two cases of azoospermia, one without genital abnormalities on physical examination and one with bilateral hypotrophic testes were observed so far in the DES-exposed group. Eighteen percent of 229 DES-exposed female patients had irregular menstrual cycles (oligomenorrhea) as compared to 10% of 136 controls. The history of pregnancy revealed a lower incidence of pregnancy in the DES-exposed group (18%) than in the control group (33%). Circumferential ridges of the vagina and cervix were seen in 40% of 229 DES-exposed females but in none of 136 controls. Colposcopic findings in the vagina revealed adenosis in 66.8% of the DES-exposed females and in 3.6% of the control group. Dysplastic lesions were more prevalent in the vagina and cervix of the DES-exposed subjects. No cases of cancer were observed in either the male or female offspring.

Dysplasia during pregnancy: a cytologic follow-up study.

To clarify the characteristics of dysplasia in the pregnant state, 525 of 703 dysplasias in pregnant women recorded in the laboratory computer at the University of Chicago Lying-in Hospital during a ten-year period were selected for study through medical records. These materials were basically diagnosed by cytologic techniques. The regression rates of moderate and marked dysplasia within a six-month period after delivery seemed to be much higher than those of dysplasia in the general population. The progression rates of dysplasia to carcinoma in situ during pregnancy and after delivery were almost the same as those of dysplasia in the nonpregnant state, whereas the progression rate of dysplasia to invasive carcinoma after delivery (0.4%) was almost half that in the nonpregnant state (1%). With reference to the latent period during which dysplasia progressed to malignancy, dysplastic lesions during pregnancy, as compared to those in nongravid women, had a higher potential for progression. The number of dysplastic cells decreased with the course of pregnancy. The rates of metaplastic and keratinizing types of dysplasia were remarkably higher than those in the general population, and the rates of those types of dysplasias increased with the course of pregnancy.

Human papillomavirus standardization and DNA cytophotometry in cervical intraepithelial neoplasia.

Human papillomavirus (HPV) standardization and DNA cytophotometry were carried out in 29 cases of cervical intraepithelial neoplasia (CIN) grades I-III. A prognostically unfavorable DNA distribution pattern with an aneuploid stemline was found in 14 of the 16 dysplasias with HPV types 16 and 18, while in 11 of 13 dysplasias with HPV types 6 and 11 there was a favorable DNA distribution with a euploid-polyploid stemline. Among 178 colposcopically, cytologically and histologically confirmed cervical lesions, there was a statistically significant incidence of HPV 16/18 infections in severe dysplasias and carcinomas, while HPV 6/11 was found predominantly in mild cervical lesions. It seems that CIN can be divided into high- and low-risk lesions not only by the degree of severity and the DNA distribution pattern but also by HPV typing.

A quality-control procedure on cervical lesions for the comparison of cytology and histology.

A new method for the quality control of cytologic and histologic diagnoses of cervical lesions is based on the automated high-resolution scanning, image processing and computer analysis of cytometric data by the TICAS system. It determines and then compares optical-density-based ploidy patterns of cells in cytologic smears and the corresponding histologic sections, with the results available both as computer-graphic displays and printouts. Examples of the former appear for an "agreement case," in which the cytologic and histologic patterns corresponded, and a noncorrespondence (nonrepresentative) case, in which the tissue sample had been nonrepresentative of the lesion sampled by cytology. Computergraphic examples concern one case of condyloma and one of tissue repair, in both of which both the cytologic and histologic diagnoses had been overcalled. A further example shows the method's use in monitoring response to therapy. The DNA ploidy patterns on which this method is based can give diagnostic and prognostic clues when morphology alone may be equivocal or insufficient. The utility of ploidy pattern determinations of material from other body sites is also well established. With the use of microprocessors, the system described could be made inexpensive and operationally simple for the routine quality control of many cytopathologic studies as well as for the clinical follow-up of patients.

Follow-up study of male and female offspring of DES-treated mothers a preliminary report.

This is a follow-up study of male and female offspring of mothers who were part of a double-blind placebo controlled investigation during the years 1951-1952, originally aimed at determining the usefulness of DES administration in maintaining pregnancy. So far, 84 DES-exposed females, 43 female controls, 42 DES-exposed males and 37 male controls have been examined. Circumferential ridges of the vagina and cervix were seen in 39% of the DES-exposed females but in none of the controls. Colposcopy revealed vaginal epitheleal changes in 78% of the DES-exposed females 2% of the female controls. Cytology proved to be reliable as a screening test for vaginal epithelial changes in the DES-exposed female. Urine cytology was negative for tumor cells in all patients. The main abnormal finding in the DES-exposed males was that cysts in the epididymis were detected in 10%. No cases of cancer were observed in either the male or female offspring.

PIP: The original study conducted 22 years earlier at the Chicago Lying-in Hospital attempted to determine the value of diethylstilbestrol (DES) in maintaining pregnancy. The number completing the course of therapy was 840 in the DES group; there were 860 in a control group. Increasing doses were given beginning during the 7th week of pregnancy. The present study was to determine the level of risk of cancer and other anomalies in the female and male offspring of mothers who participated in the study. So far, 84 DES-exposed females, 43 female controls, 43 DES-exposed males, and 37 male controls have been examined. No cases of cancer have been found. The average age was 22 years. For female patients the medical history, a general physical examination, a gynecological examination, a colposcopic study, and laboratory tests were made. Laboratory tests consisted of cervical, endocervical, and 4 vaginal wall Pap smears, urine cytology, and follicle stimulating hormone and luteinizing hormone determinations. Biopsies were performed when indicated. Progesterone and total estrogens were determined only in patients with irregular menstrual cycles. In male patients, a general physical examination, urologic studies, and laboratory work-up were done. Medical records of all the newborn infants were surveyed and pediatric records examined. No cases of congenital malformations were recorded. Minor differences in menstrual histories and in ability to conceive were noted. The differences appeared mainly at vaginal examinations. Circumferential ridges in the vagina and cervix were seen in 39% of the exposed females but in none of the controls. Erythroplakia of the cervix was seen in 67% of the exposed and in 53% of the controls. Colposcopic findings in the vagina revealed vaginal epithelial changes in 78% of the DES-exposed females and 2% of female controls. Iodine negative areas in the vagina were noted in 78% of the exposed females compared with 2% of the unexposed females. Iodine negative areas on the cervix were seen in 74% of the exposed and 58% of the unexposed. All dysplastic lesions were confirmed by histology. The cytology was negative in all. In the males abnormal findings were noted mainly in the DES-exposed group. An undersized penis was noted in 2, small testes in 2, varicocele in 1, and epididymal cysts in 4. Urine cy tology and prostatic fluid cytology did not reveal unusual findings. A more detailed analysis of findings will follow when material is larger and older.

Endometrial cancer: rising incidence, detection and treatment.

PIP: This editorial consists of summaries of the discussions on incidence, pathogenesis, prognosis and patient follow-up, and transcripts of the discussions on detection and treatment of endometrial carcinoma, from a symposium held in Carefree, Arizona. 75% of the cancers occur in postmenopausal women; average age is 52 years, but is decreasing. Endometrial carcinoma rose from 20.3 to 46.3% of all uterine cancers in Cleveland University Hospitals from 1941-1970. Older patients are often diabetic, overweight, nulliparous, with anovulatory or familial history; young women frequently resemble mild Stein-Levinthal syndrome. Clinically, 20% of patients are assymptomatic, others may have softer or larger uterus, larger ovaries, irregular postmenopausal bleeding, or lengthy onset of menopause. The Gravlee jet wash is indicated for high risk patients and those about to take estrogen. Endometrial carcinoma first affects epithelium, then endometrial stroma, then upper myometrium, lower myometrium, then other organs, perhaps via lymphatics, vagina, tubes, but ascites is uncommon. Generally, U.S. physicians use intrauterine radium followed by surgery, British use surgery first, and Swedish use radiation only. Cases must be treated individually, e.g. surgery only for minimal cancer, radium and surgery for more serious cases, and preoperative external radiation also for advanced disease. Although radiation lessens chance of implantation during surgical trauma, insertion of intrauterine radium enhances spread of tumor cells. Injectable progestins sometimes control metastatic disease, although they require 8 weeks to act. Progestins may help those with late recurrence, squamous metaplasia, or who are under 50 years of age. Estrogens are rarely effective. Prognois for terminal patients often includes subjective improvement, bowel obstruction, lung complications, hemorrhage. Radiation side effects and menopausal symptoms are often problems for cured patients. In young cured patients the endometrium should be suppressed with progestins or oral contraceptives.^ieng

Accuracy of three sampling technics for the diagnosis of endometrial cancer and hyperplasias.

Intrauterine aspiration (Vakutage), endocervical aspiration cytology and vaginal, ectocervical and endocervical (VCE) smears were used as outpatient procedures for diagnosis of endometrial cancer and hyperplasias in 840 patients. The results of the three technics were correlated with D&C or hysterectomy specimens. Ninety-three percent of the malignant lesions (82 adenocarcinomas, 5 mixed mesodermal tumors, 5 mixed adenosquamous carcinomas and 5 metastatic adenocarcinomas) were diagnosed by the Vakutage sample but only 67% by the VCE smears and 68% by endocervical aspiration cytology. The diagnostic accuracy of Vakutage in 50 cases of cystic hyperplasia was 88% (as compared to 14% for VCE smears and endocervical aspiration cytology) and 89% in 90 cases of adenomatous hyperplasias (as compared to 20% for the other two technics). Endometrial polyps were diagnosed exclusively from the Vakutage tissue sample in 83% of the 42 cases.

Bronchial metaplasia as a benign or premalignant lesion. I. Cytologic and ultrastructural discrimination between acute carcinogen effects and toxin-induced changes.

The respiratory tracts of hamsters were exposed to either of two different chemical compounds contained in cigarette smoke, the carcinogenic chemical benzo(a)pyrene or the noncarcinogenic chemical formaldehyde. At various times during or shortly after the exposure, the acute cytologic, histologic and ultrastructural responses to the two agents were compared. Both agents induced microscopically similar squamous metaplastic changes of the tracheobronchial epithelium. However, there were several cytologic and ultrastructural differences in the appearance of the chromatin, nucleoli and cytoplasm that could be used to separate carcinogen-induced from noncarcinogen-induced acute metaplastic changes. Furthermore, after cessation of the exposures, squamous metaplastic changes induced by the noncarcinogenic agent formaldehyde regressed to normal, while the acute carcinogen-induced changes were replaced by other, more persistent metaplastic changes.

Automated hierarchic decision structures for multiple category cell classification by TICAS.

TICAS-PROBE, a computerized system has been developed to generate hierarchic decision structures with little human intervention. It has been tested with several tasks, and the results are comparable, if not better, than those produced by human designers. Additional methods of generating the individual elements will be included in the near future.

Dimensionality reducing displays in cell image analysis.

The paper discusses the problem of representation of multivariate cell data sets in two dimensions such that the essence of the situation as represented by the multivariate feature space is preserved. A corresponding projectional technique has been developed and illustrated on a set of five different cell types of ectocervical cells, including normal and carcinoma cells.

Chromatin appearance in intermediate cells from patients with uterine cancer.

This paper presents the differences found by the computer analysis of digitized images of visually apparently normal intermediate cells from patients with normal cytology and patients with uterine cancer. There exist differences in the absolute amount of nuclear staining, the average density of staining, cell shape and size and also differences in spectral contrasts of the nuclear chromatin. Visual cytologic detection of these changes became more conspicuous when the photomicrographs were arranged in order of their discriminant function scores.

Clinical applications for an inexpensive, microcomputer-based DNA-cytometry system.

Some of the clinical capabilities of an inexpensive, microcomputer-based DNA-cytometry system are described. A variety of applications is illustrated: ploidy assessment, which provides significant information for the management of patients with tumors from almost any organ site; quality control of cytologic and histologic diagnoses; determination of tumor heterogeneity; monitoring of cell profile changes during therapy; and quantitation of immunocytochemical staining.

Stereotaxic fine needle aspiration cytology of clinically occult malignant and premalignant breast lesions.

Stereotaxic fine needle aspiration (FNA) cytology was used to study clinically occult (nonpalpable) breast lesions in 114 consecutive patients with mammographically suspicious findings prior to excisional biopsy. The aspirate contained insufficient material for cytologic evaluation in 15 cases (13.2%), which were histologically diagnosed as benign (7 cases), atypical hyperplasia (7 cases) or carcinoma in situ (1 case). The cytologic findings indicated a benign lesion in 77 cases (67.5%), which were histologically diagnosed as benign (71 cases) or atypical ductal hyperplasia (6 cases). The cytologic sample showed atypia in eight cases (7.0%), which were histologically diagnosed as severe atypical ductal hyperplasia (three cases), carcinoma in situ (one case) or proliferative fibrocystic disease (four cases). In the eight cases (7.0%) cytologically interpreted as probably malignant, histology confirmed six invasive carcinomas, one carcinoma in situ and one fibrocystic disease. Of six cases (4.4%) cytologically reported as malignant, five were histologically diagnosed as invasive carcinoma and one as carcinoma in situ. Overall, stereotaxic FNA cytology reported as malignant or probably malignant 14 of the 15 cases with a histologic confirmation of malignancy, for a sensitivity of 93.3%. Cytology correctly identified 78 of the 83 histologically negative cases, for a specificity of 94.0%. The 16 cases histologically diagnosed as ductal hyperplasia, which carries a high risk for subsequent malignancy, were studied in detail in an effort to define histologic and cytologic criteria for this entity. Using selected histologic criteria, 11 of these cases were graded as showing mild-to-moderate atypical hyperplasia and 5 as showing severe atypical hyperplasia. Three of the latter cases were similarly identified by an analogous cytologic grading; the other two cases had insufficient cytologic samples. The total results in this series of 114 cases support the use of stereotaxic FNA cytology in the diagnosis of these nonpalpable breast lesions, examples of which are illustrated. In particular, it may help to raise the low specificity yielded by mammography alone, which would represent a significant advance for the patient in terms of the accuracy, expediency and reduced cost of diagnosing these lesions.

Correlation of light and scanning electron microscopy a new method for exfoliative cytology.

A method for preparing locator slides for light and scanning electron microscopy has been developed. These slides make possible correlation of light and scanning electron microscopy of individual cells. Some examples of the value of this method are given.

The TICAS-RTCIP real time cell identification processor.

A preliminary model of a real time processing system, the TICAS-RTCIP (for Taxonomic Intra-Cellular Analytic System--Real Time Cell Identification Processor) has been disigned and is being tested under operational conditions, proving the substantial advantages of employing a network of microprocessors operating in parallel. The system is designed only to demonstrate the feasibility, while more extensive development and biologic testing will be required to assess general applicability to automated uterine cytology.