Distribution and significance of the smooth muscle component in polyps of the large intestine.

Immunocytochemical staining for alpha smooth muscle actin specifically reveals the distribution of the muscularis mucosae and pericryptal intramucosal smooth muscle fibers in the normal mucosa and polyps of the large intestine. In the latter, the muscular component is hyperplastic, especially in pedunculated polyps, which display a thick "muscular zone" at the top of the stalk. The diagnostic (and prognostic) significance of the distribution pattern of the muscular component in polyps, and in "early invasion" cases, is discussed.

Significance of the smooth muscle cell component in Peutz-Jeghers and juvenile polyps.

We have previously reported the presence of smooth muscle cells (SMCs) in the lamina propria of both tubular and villous adenomatous polyps. In the current study, we investigated the presence and distribution of SMCs with immunohistochemical techniques using anti-alpha smooth muscle actin and anti-desmin antibodies on a series of hamartomatous polyps. In five Peutz-Jeghers polyps, large tree-like branches of SMCs were observed, while in 13 juvenile polyps, rare elongated SMCs were found in the lamina propria, partially surrounding cystically dilated glands. Apart from the quantitative differences, the SMC distribution was similar in the two types of hamartomatous polyps and not dissimilar from the pattern described in neoplastic polyps. These findings make the category "juvenile polyp" less clear and its differentiation from other histologic types of polyp less sharp. Thus, the presence or absence of SMCs cannot form the basis of the differential diagnosis between adenomatous and hamartomatous polyps.

Squamous cell carcinoma with prominent myxoid stroma.

Three cases of a squamous cell carcinoma with a prominent myxoid stroma are reported. One case in a 70-year-old woman had presented as a lump in the breast, the other two presented as polypoid lesions of the larynx and cervix uteri in a 65-year-old man and 61-year-old woman, respectively. The carcinomatous component was immersed in abundant extracellular mucosubstances. In addition to occasional squamous pearls, it displayed immunocytochemical evidence of high-weight keratin present in the neoplastic cells and, in one case, desmosomal attachment under electron microscopy. The extracellular mucosubstances proved to be similar to those seen in connective tissue. The differential diagnosis with histologically similar lesions has been taken into consideration, and it has been suggested that this newly described entity showing abundant myxoid stroma has to be distinguished from numerous benign and malignant myxoid soft tissue tumors.

Mucin histochemical analysis in the interpretation of Barrett's esophagus. Results of a multicenter study. The Operative Group for the Study of Esophageal Precancer.

A multicentric study of Barrett's esophagus (BE) was started in November 1987 to evaluate (1) the prevalence of BE among subjects undergoing upper gastrointestinal endoscopic examination; (2) the pathologic features of BE; and (3) the correlation between BE and early malignant changes. In 157 of 330 patients who underwent multiple standardized biopsies, BE was confirmed with histologic evaluation. Specialized intestinal-type BE was observed in 84 patients. By applying Alcian blue (pH 2.5)-periodate oxidation-Schiff, high-iron diamine-Alcian blue (pH 2.5), and periodate borohydride-saponification-periodate oxidation-Schiff techniques, the intestinal type of BE was subclassified into colonic and ileal types, both complete and incomplete. Fifty cases had incomplete colonic metaplasia with sulphomucins in the columnar cells and 64 had complete colonic intestinal metaplasia, 49 of them containing O-acetylated sialomucins in the goblet cells. These patients are being included in a short-term follow-up. Dysplasia (six low grade, two high grade) was observed in eight patients in areas of intestinal colonic-type epithelium; in these patients, a complete loss of O-acetylated sialomucins in the dysplastic areas and a remarkable reduction of these mucins in the surrounding tissue were observed. The reduction of O-acetylated sialomucins might indicate relative tissue immaturity, which could represent an early sign of neoplastic dedifferentiation. Therefore, the relevance of O-acetylated sialomucin content in BE, first demonstrated in intestinal type, is now evident, although its biologic importance is being studied.

The incidental finding of abnormal cervical histology in postmenopausal patients.

A report is supplied on 216 samples of cervical tissue incidentally found in 684 endometrial specimens collected during hysteroscopic examination of postmenopausal women with uterine bleeding and a recent negative Pap smear. We found 43 (19.9%) specimens including cervical tissue with some histologic sign of pathology. Twenty-five (11.6%) had histologic features suggestive for human papillomavirus (HPV) infection, while 18 (8.3%) had cervical intraepithelial neoplasia (CIN). Of the 18 CIN cases, 9 were CIN I, 6 CIN II and 3 CIN III. Altogether, the prevalence of dysplasia in postmenopausal women with recently referred normal cervical cytology was impressive. A significant number of dysplastic lesions (14 out of 18, 77.7%) did not present any histologic sign of HPV. Also, none of the histologic diagnoses of sub-clinical HPV infection was confirmed by the in situ hybridization. Considering the significant prevalence of high grade lesions (9 CIN II and III, 4.2% of the analysed samples) found in this randomly selected patient population, our data strongly suggest the need for a regular follow-up of the transformation zone in all postmenopausal women. Although in women of postmenopausal age some low grade lesions seem to have only a reactive-reparative significance, a more accurate screening procedure, taking into account the peculiar modifications of the menopausal uterine cervix, is advisable.

Endometrial response in sequential cyclic therapy assessed with associated hysteroscopy and histology.

A morphologic study was performed on the endometrium in 37 asymptomatic postmenopausal women under effects of cyclically administered oestrogens. Eighty-seven postmenopausal women were taken as control group. Medroxyprogesterone acetate (MPA), 10 mg daily, was administered in association with two types of oestrogen replacement therapy: conjugated equine oestrogens 0.625 mg (CEE) or transdermal 17 beta-oestradiol 0.05 mg (E2-TTS). Endometrial biopsies were taken under hysteroscopic control before treatment and on days 8-12 of combined therapy at the 6th month. Follow-up at 12 and 18 months was only performed in 8 and 5 patients, respectively, with transdermal 17 beta-oestradiol treatment. Various types of endometrial response were identified from atrophy to hyperplasia and secretory patterns. No atypical hyperplasia was found. All cases of simple or complex hyperplasia showed a regression after increased MPA dosage treatment (20 mg). This work is aimed at investigating the endometrial response during sequential cyclic therapy by using morphologic criteria based on hysteroscopy and histology. A large number of patients with hyperplasia can be detected with target biopsy under hysteroscopy, thus playing an important role in the management of patients during replacement therapy in research protocols.

Histochemical study of mucinous ovarian cystomas of intestinal and mixed types: O-acetylated sialomucin content in relation to biologic behavior.

The subgroup of mucinous ovarian cystomas of intestinal type containing site 8 O-acetylated sialomucins and its relation to malignant behavior were studied. Of 75 mucinous cystomas of the ovary from the files of the Institute of Pathological Anatomy and Histology (University of Genoa), 54 were endocervical, 19 were mixed, and 2 were intestinal. As regards their histologic pattern, the 54 mucinous endocervical cystomas were benign, whereas the 19 mixed type group included 8 benign, 5 borderline and 6 malignant; the 2 intestinal type tumors were borderline. According to histochemical investigations, the frequency of tumors with goblet cells containing site 8 O-acetylated sialomucins (positive to PB/KOH/PAS) was 47% in the mixed type cystomas and 100% in the intestinal type cystomas. Twenty-five percent of benign mixed cystomas, 60% of borderline mixed cystomas, and 67% of mixed cystadenocarcinomas were positive to PB/KOH/PAS. One hundred percent of the borderline cystomas of intestinal type were positive. Our results confirm that among mucinous ovarian cystomas, those containing intestinal type epithelium are the most likely to fall within the borderline or malignant categories. Moreover, of all ovarian cystomas of intestinal type, those characterized by site 8 O-acetylated sialomucins seem to be more aggressive and, in general, to have a more malignant behavior than the mucinous cystic tumors of müllerian or endocervical type.

Patterns of mucous secretion in normal and pathological conditions of the endocervix.

20 specimens of normal endocervix, 30 of chronic cervicitis, 20 of endocervical polyps and 10 of adenocarcinomas were histochemically investigated in order to assess their mucin pattern. Diastase periodic acid-Schiff (D-PAS), Alcian blue pH 2.5 (AB), High iron-diamine (HID) and HID followed by Alcian blue pH 2.5 (HID-AB) were used. Both the superficial and glandular epithelium of the normal endocervix contained abundant amounts of neutral mucins. Generally sialomucins were scarce and predominant over sulphomucins: the latter were in some cases absent in the proliferative phase of the menstrual cycle and increased in the secretory phase. In chronic cervicitis a slight amount of sulphomucins in the residual glandular epithelium, especially in areas presenting severe inflammation, pseudoerosion, or both was observed. Nabothian cysts mainly contained sulphomucins. In the endocervical polyps, the mucin pattern was various. In adenocarcinomas mucin secretion was usually scanty and mixed with a predominance of sialomucins. Therefore resulted: the mucin pattern of the normal endocervical epithelium is related to the menstrual cycle; the histochemical evaluation of mucous secretion, presently doesn't seem to be helpful in differentiating endocervical from endometrial adenocarcinomas.

Blood free endometrial sampling: a useful tool in routine diagnosis.

The presence of clots or of a great amount of blood in the specimen from curettage or endometrial aspiration may lead to considerable problems for the pathologist. The application of a lysis buffer/ethanol solution before applying conventional fixation techniques may solve this problem. This new technique was tested on a series of endometrial samples. No disadvantages have been recorded, quite the contrary, optimum specimens were obtained for histochemical and immunohistochemical studies.

Patterns of intestinal metaplasia in gastric biopsies. A comparison of different histochemical classifications.

A total of 217 cases of gastric intestinal metaplasia (IM), out of 491 cases of consecutive multiple gastric biopsies, were examined using various histochemical methods. Three classifications were adopted (1) Jass and Filipe's (1979), (2) Segura and Montero's (1983), and (3) our own distinguishing two types of small intestine IM (incomplete and complete) and two types of colonic IM (incomplete and complete). Our classification revealed (1) a group of 'pure' IM cases (44%), (2) a group of 'combined' IM cases (15%), either small intestinal or colonic, expressing a maturating line, and (3) a conspicuous group of mixed 'mosaic' IM cases (41%) consisting of all possible combinations between the basic IM types. No significant correlations were found between each IM type and associated diseases (i.e. gastritis, stump gastritis, ulcer, adenoma and adenocarcinoma). However, a suggestive association between colonic IM and malignancy was observed. Our results confirmed the heterogeneity of IM and indicated two differentiation cell lines (ileal and colonic), thus providing additional evidence of impaired maturation and aberrant differentiation along various cell lines, supporting Leblond's (1976) unifying theory on the origin of intestinal epithelial cells.

Assessment of proliferating cell nuclear antigen expression in dysplastic intestinal type Barrett's esophagus. Gruppo Operativo per lo Studio delle Precancerosi Esofagee.

In the management of patients with Barrett's Esophagus (BE) the definition of a sub-group of patients at high risk for malignant transformation of the metaplastic epithelium is important. Undoubtedly, the histological evaluation of dysplasia is the most important and objective parameter to identify a malignant transformation of BE. In order to obtain additional data a group of cell proliferation markers was applied in dysplastic lesions. We studied PCNA in a selected group of patients with BE. One hundred six biopsies were examined, referring to fifty five patients with BE, who were followed up for a period of at least one year (1-5 years). All patients showed one or more biopsies positive for dysplasia. PCNA was detected immunohistochemically on formalin fixed and paraffin embedded biopsies, and positive cells were evaluated in a crude percent count. Our results showed a remarkable number of PCNA positive cells in high grade dysplasia and a variable amount of PCNA positive cells in the others groups. In addition, there was an overlap in the number of positive cases between low grade dysplasia (30%), indefinite (33%) and negative (34%). Therefore, the assessment of S-phase cells correlated with the degree of dysplasia seems to be of limited practical use. Cell proliferation is probably affected by many factors and mainly inflammation.

Prevalence and prognostic significance of endocrine cells in colorectal adenocarcinomas.

Sporadic differentiation of endocrine cells inside adenocarcinomas of the gut (combined tumors) has been observed mainly in the stomach and less frequently in the colon. Since one or more lines of endocrine cells could be present, a wide-spectrum marker of neuroendocrine cells seems to be useful in defining the prevalence of an endocrine component in colorectal carcinomas. One hundred and fifty-eight cases of unselected colorectal adenocarcinomas were immunohistochemically evaluated on formalin fixed and paraffin embedded material, employing streptavidin-biotin-peroxidase-complex (SABC) with anti chromogranin antibodies. All samples were obtained in peripheral areas of neoplastic proliferation without necrosis or haemorrhage. Semiserial sections from each block were cut and stained with H.E., Grimelius, Masson Fontana and SABC anti chromogranin. Chromogranin positive cells were found in 22 of 158 cases (14%). In 18 cases (11.39%), scattered cells (one single cell or a few cells in each section) were located between neoplastic epithelial cells; in 4 cases (2.53%), clusters of chromogranin positive cells were found. Twelve tumors were located in the sigma-rectum, eight in the right colon and two in the left colon. According to tumor grading, five were G1 and seventeen were G2. According to tumor staging following Dukes' classification, eleven were stage B, and eleven stage C. In conclusion, no statistically significant correlations resulted between the adenocarcinoma with endocrine component and tumor location (p = 0.1 n.s.), tumor grading (p = 0.2 n.s.), and tumor staging (p = 0.1 n.s.).(ABSTRACT TRUNCATED AT 250 WORDS)

Immunohistochemical investigations of the human palatine tonsil from surgical specimens using polyclonal and monoclonal antikeratin antibodies.

The keratinization process of tonsillary epithelium in patients with different age, who underwent surgery because of chronic inflammatory processes, has been studied by employing low (40 KD), medium (52-56-58 KD), high (56-64 and 68 KD) as well as broad spectrum antikeratin antibodies. The findings thus obtained outline the various keratin patterns of the tonsil in the outer covering and, even more, in the crypt labyrinth wall. These findings have been compared with those obtained by histochemical reactions for acid phosphatase and non-specific esterase activities (Favrz et al. 1986 II). In crypt epithelium keratinization, 56-64 KD keratins are involved in cells where also high enzymic activities are taking place, namely 1. in those elements delimiting the lumen as well as in those belonging to the reticulum mesh which are nearest to it and host lymphoid elements active in immunity reactions; 2. in elements at the bottom of small and large crypts, where excavation continues into the lymphoid tissue and interfollicular spaces. 68 KD keratin, although present in lower amounts, is synthesized almost everywhere, but only irregularly. It marks surface epithelium surrounding some dermal papillae, some meshes of the crypt reticulum, the "gallery" bottom as well as elements of interfollicular proliferation developing in the lymphoid tissue like arborescence. The path of congested blood vessels in the crypt wall towards the surface is marked by keratinized epitheliocytes. 56-64 KD keratin is present in the most superficial elements and is generally accompanied by the wall growing thinner, its blistering and breaking. The most frequent pathologic alterations of the epithelium (like cellular hypertrophy) involve variations in cytoplasm keratin pattern, in still un-flattened polyhedral elements of the intermediate layers.

O-acylated sialic acid variants in mucinous tumours of the ovary.

O-acylated sialic acid variants (site 8) can be demonstrated histochemically by the PB/KOH/PAS method. They are secreted by goblet cells of the lower gastrointestinal tract, by colorectal adenocarcinomas, and by their metastases. Since the metastases are positive only when the primary tumour is positive, O-acylated sialomucins can be considered to be specific markers of colorectal adenocarcinomas if identified in metastases of a tumour of unknown origin. In our histochemical study we evaluated 29 mucinous cystomas of the ovary (23 benign and 6 malignant). We found that six cases were positive to PB/KOH/PAS. The positivity was observed in a limited number of cells and only in areas which presented an intestinal type epithelium. It was also more evident in malignant cystomas than in benign ones. We therefore think that the PB/KOH/PAS positivity can not only be considered a marker of colorectal adenocarcinomas, but also of all neoplasms which originate from an intestinal epithelium or appear to an "intestinal type epithelium".

O-acetylated sialic acid variants in intestinal glandular metaplasia of the urinary tract.

Two cases of chronic cystitis, two of chronic ureteritis, one of chronic pyelitis and two transitional cell carcinomas of the urinary tract showed glandular metaplasia or focal mucin production. Positive staining for O-acetylated sialic acid variants (sites 7 and 8) detected by the PB-KOH-PAS method was found in the two cases of cystitis, in one of chronic ureteritis and in one of chronic pyelitis. Positive staining was observed only in areas showing an intestinal type of metaplasia and in which goblet cells and argyrophil cells were also identified. These findings taken together with earlier studies on ovarian cystadenomas lead the authors to believe that O-acetylated variants are markers of an intestinal type of epithelium. Evaluation of O-acetylated sialomucins may be useful in further studies on the histogenesis of primary adenocarcinomas of the bladder.

Immunohistochemical study of carcinoembryonic antigen, epithelial membrane antigen, and secretory immunoglobulin system in the large bowel adenoma-carcinoma sequence.

To identify parameters for the malignant potential of large bowel adenomas, we used the avidin-biotin-peroxidase complex (ABC) technique. Patterns of carcinoembryonic antigen, epithelial membrane antigen, IgAs, and secretory component in 31 tubular, tubulovillous, and villous adenomas with different grading of dysplasia (mild, moderate, severe) and with early cancer were studied. All markers showed different degrees of staining intensity and various cellular localizations. We found that these variations might be related to dysplasia grading. Adenoma size did not influence the marker patterns. IgAs seemed to be the more selective among the markers studied. Our results suggested a dysplasia-carcinoma sequence instead of an adenoma-carcinoma sequence concept.

Human papillomavirus in cervical adenocarcinoma. An in situ hybridization study.

Twenty cervical adenocarcinomas (CACs) in women aged 22 to 71 were investigated by in situ hybridization (ISH) with 6, 11, 16, 18, 31, 35 and 51 HPV biotinylated probes. Two cases, one adenocarcinoma and one adenosquamous carcinoma (in women aged 28 and 40 respectively) showed focal nuclear positivity to 16 HPV Probe in some neoplastic glands. We used ISH, rather than other hybridization techniques, in order to exclude a positivity to viral DNA, due to adjacent squamous epithelium, either normal or metaplastic, and in squamous foci within adenosquamous tumors. Reviewing the literature, we found 33 out of 98 CACs positive to viral DNA by ISH (33.6%). In spite of the differences found from author to author, a relationship between adenocarcinomas of the uterine cervix and HPV infection seems to be possible, as was demonstrated for CIN and invasive cervical carcinomas. These data could explain why the incidence of this neoplasia has tended to increase over the last few years, mainly in younger patients.

Studies on the mechanism of the carcinogenic activity of amitrole.

Amitrole, a widely used herbicide found to produce thyroid and liver tumors in rodents and classified as possibly carcinogenic to humans, was investigated to acquire further information about its mechanism of action. A 20-hr exposure to amitrole concentrations ranging from 5.6 to 18 mM did not induce DNA fragmentation, as measured by the alkaline elution technique, in primary cultures of human thyroid follicular cells and of human liver cells. Under the same experimental conditions a minimal frequency of DNA breaks was detected in primary cultures of rat hepatocytes, but this event was presumably the unspecific consequence of a cytotoxic effect. In rats given amitrole with drinking water for 12 successive days at a daily dose of approximately 200 mg/kg, plasma levels of triiodothyronine and thyroxine displayed a progressive reduction, and a concurrent increase of both the mitotic index and frequency of S-phase cells revealing a clear-cut follicular cell hyperplasia was observed. In a group of these rats euthanized after 8 days of treatment any evidence of DNA fragmentation was absent in both thyroid and liver cells. Taken as a whole these results provide further evidence that the mechanism of amitrole carcinogenic activity is most likely nongenotoxic but due to hormone imbalance.