Epithelial and muscle antigens in benign cystic teratomas of ovary.

Thirty beign cystic teratomas of the ovary were studied by immunofluroesecence, using specific human antisera for (a) squamous epithelial intercellular areas (pemphigus sera), (b) squamous epithelial basement membranes (bullous pemphigoid sera), (c) smooth muscle (chronic active hepatitis sera, and (d) striated muscle (thymona sera). Squamous epithelial intercellular staining was seen in all specimens and was present in the basal cell layers as well as in the prickle cell layers, suggesting a slow regeneration rate for the squamous epithelial lining of these tumors. Well-formed basement membranes were identified at the periphery of all squamous epithelial structures. Smooth muscle was found in 73% of the neoplasms, and the immunoflurorescent findings correlated with light microscopy. No skeletal muscle components were identified. Within the limits of the antibodies used and the antigens detected, these resutlts support the concept that benign ovarian cystic teratomas are composed of mature, heterotopic tissue elements.

Pulmonary hemorrhage in systemic lupus erythematosus.

Massive pulmonary hemorrhage confirmed by pathologic examination occurred in three of 12 patients with systemic lupus erythematosus (SLE) complicated by lung disease. Two patients died. Hemorrhage was clinically unrecognized in two cases. The third patient survived after an open lung biopsy and an increase in prednisone therapy. In the absence of hemoptysis, awareness of the possibility of pulmonary hemorrhage in patients with SLE and a high index of suspicion are necessary for early recognition of this uncommon but potentially fatal complication.

Immunofluorescent detection of angiotensin-converting enzyme (ACE) in Gaucher cells.

The cellular localization of the elevated angiotensin-converting enzyme (ACE) in the spleen of a patient with Gaucher's disease was examined by immunofluorescence using an antibody prepared against highly purified human lung enzyme. Intense ACE-specific cytoplasmic fluorescence was observed in Gaucher cells, but not in various controls, indicating the localization of large quantities of enzyme in these cells. These results and the demonstrated capability of mononuclear phagocytes for marked induction of this enzyme suggest the possibility that induction of active synthesis of ACE in Gaucher cells may be responsible for the elevated enzyme levels in the serum and spleen of patients with Gaucher's disease.

Quantitation of the immunocytochemical assay for estrogen receptor protein (ER-ICA) in human breast cancer by television imaging.

A Quantimet 720D Image Analysis System has been programmed for light microscopic evaluation of the nuclear estrogen receptor distribution in frozen sections of human breast cancer stained by the peroxidase-antiperoxidase method using monoclonal antibodies to estrogen receptor protein (ER). This method provides precise criteria for distinguishing ER-positive and -negative cells and a sensitive and reproducible means for densitometric quantification of the staining patterns. Although imaging sequence and graphic analysis are automated by computer programs, light pen interaction provides supervision of feature selection. Imaging of the immunocytochemical assay (ER-ICA) in 50 patients revealed marked heterogeneity of nuclear estrogen receptor concentration varying over a nearly 100 fold concentration range. Various ER concentration patterns were evident: (I) distributions with a single peak (CV = 5%) present at various concentration levels; (II) bimodal distributions, revealing co-existent ER-positive and ER-negative subpopulations; (III) multimodal distributions with a number of resolvable concentration peaks; and (IV) highly skewed distributions with or without discernible peaks, frequently extending over the entire concentration range. Statistical methods of de-convolution were applied to determine the frequency and ER concentration characteristics of component subpopulations in the mosaic cases and for resolving the proportion of ER-positive and -negative cells. An approach for evaluating nuclear ER content in conjunction with ER concentration patterns in individual patients revealed whether spread in the ER concentration distribution resulted from differences in nuclear ER content or from variability in nuclear volume distribution.

Histochemical analyses of steroid hormone receptors in breast and prostatic carcinoma.

Histochemical analyses estrogen (ER) and progesterone (PgR) receptors in breast cancer were statistically correlated with results of dextran-coated charcoal (DDC) and sucrose gradient assays. Correlated for ER was 91% of 363 cases, and for PgR 88% of 255 specimens. Breast cancer ER/PgR positivity by histochemistry correlated with a favorable clinical response to endocrine therapies in 72% of 25 cases, while ER/PgR negativity correlated with a lack of response in 96% of 22 cases with Stage IV disease. Nuclear ER/PgR correlated with a poor response to therapy in 8 of 12 patients. An in vitro technique to detect nuclear translocation of ER revealed two groups of ER positive cases, with 11 of 17 exhibiting translocation and 6 not displaying translocation. In prostatic carcinoma, 72% of 65 men were positive for ER and/or androgen receptor. Comparison of specimens obtained without and with electrocautery revealed a preponderance of nuclear binding in the latter, suggesting heat-induced nuclear translocation of receptor. coumestrol, a naturally fluorescent, entirely unaltered estrogen was also used for histochemical detection of ER. Results correlated with ER by DCC in 87% of 61 breast cancers. Coumestrol was additionally used to visually observe receptor and nuclear translocation of ER in intact whole cells in culture.

Immunofluorescence of soft-tissue tumors with anti-smooth-muscle and anti-skeletal-muscle antibodies.

One hundred thirteen soft-tissue tumors were studied by immunofluorescence using anti-smooth-muscle (SMA) and anti-skeletal-muscle antibodies (MGA). Of the classic leiomyomas examined, 94% manifested bright fluorescence with SMA, whereas cellular leiomyomas and leiomyosarcomas failed to stain. Of the rhabdomyosarcomas studied, 21% showed fluorescent cross-striations or fluorescent granules after incubation with MGA. None of 48 control tumors fluoresced with either antibody. No tumor vessel seen showed smooth-muscle fluorescence in its walls. Although of little diagnostic value, immunofluorescence utilizing specific human antibodies is a valuable tool with which to study some of the antigenic components of these neoplasms.

Immunohistologic diagnosis of sarcoidosis. Detection of angiotensin-converting enzyme in sarcoid granulomas.

Angiotensin-converting enzyme (ACE) was detected by indirect immunofluorescence examination. ACE was present in the cytoplasm of epithelioid cells in granulomas from 38 of 39 patients who had clinical sarcoidosis but not in any of 37 nonsarcoid granulomas, normal lymph nodes, or splenic controls. Intensity of staining with ACE antiserum was inversely proportional to the degree of fibrosis and was significantly less intense in fibrotic granulomas. The detection of ACE appears to be a distinct aid in establishing a definitive tissue diagnosis of sarcoidosis.

An improved histochemical method for detection of estrogen receptors in mammary cancer. Comparison with biochemical assay.

Unselected, consecutive surgical specimens from 120 women with cancer of the breast were subjected to histochemical assay for the presence of estrogen receptor. A fluoresceinated bovine serum albumin--estradiol conjugate was used that linked estradiol at position 17 and contained 5 mol fluorescein and 4 mol estradiol per mole albumin. Simultaneous competitive binding studies with excess unlabeled estradiol, diethylstilbestrol, and the antiestrogen nitromifene citrate were regularly performed. Results were compared to those obtained by the dextran-coated charcoal receptor assay. Three specimens were necrotic, two others thawed, and two lacked sufficient protein for biochemical analysis. One specimen did not contain tumor, and 11 others showed a predominant nuclear staining pattern. Nuclear receptor was not assayed biochemically. Comparison of results in the remaining 101 cases showed agreement in 92%. The precedure is uncomplicated, economical, and could be performed and interpreted in any pathology laboratory.

Acute pulmonary complications in systemic lupus erythematosus. Immunofluorescence and light microscopic study.

Lung tissue obtained from eight consecutive patients with systemic lupus erythematosus complicated by severe, acute pulmonary disease was studied by both light and immunofluorescence microscopy. Light microscopic examination disclosed interstitial pneumonia in four cases, cytomegalovirus pneumonitis in one case, bronchiolitis and peribronchiolitis in one case, pulmonary infarction in one case and focal atelectasis in the remaining case. Direct immunofluorescence examination revealed focally bound immunoglobulins or complement (C3) within pleural and/or pneumocyte nuclei in each specimen. Immunohistologic studies in these cases may thus suggest a diagnosis of systemic lupus erythematosus with acute pulmonary complications, despite the lack of specificity of the pathologic changes seen by light microscopy.

Glomerulonephritis due to Staphylococcus aureus antigen.

Staphylococcus aureus antigen was identified within the glomeruli of a patient with acute bacterial endocarditis and diffuse glomerulonephritis. Routine immunofluorescence had revealed only granular deposits of complement (C3). C3 activator and C4 were not present. Direct immunofluorescence studies with a specific anti-Staphylococcus aureus conjugate were positive. Electron microscopy showed subepithelial and intramembranous electron-dense deposits. Eluates of the kidney did not contain anti-S. aureus antibody. The absence of immunoglobin suggests that the toxic action of S. aureus antigens may activate complement and cause glomerular injury and that immune complexes are not essential for the production of glomerulonephritis in this entity.

An immunofluorescent study of basement membranes in squamous cell carcinoma of the cervix, vagina, and vulva.

Basement membranes of 33 samples of in situ or invasive squamous cell carcinomas of the cervix uteri, vagina, and vulva were studied by defined immunofluorescence technics. Pooled serum from patients with bullous pemphigoid, containing specific antibody to squamous epithelial basement membrane, was utilized. Essentially normal basement membranes were found in all cases of in situ carcinoma and in 18 cases of invasive carcinoma. Basement membranes appeared poorly formed in three specimens with invasion and were absent in the remaining three. These findings support the premise that penetration of the basement membrane is not a valid criterion for distinguishing in situ from invasive squamous cell carcinoma.

A morphological comparison of the regional distribution of methadone in human and rat brain as demonstrated by immunofluorescence.

The brains of ten narcotic addicts who had died from an overdose of methadone, and the brains of ten rats, given methadone for one month, were examined by the immunofluorescent technic. Positive neuronal fluorescence was seen primarily in the limbic systems of both species as well as in closely associated areas. In the human, Purkinje cell fluorescence was seen in the cerebellum, whereas only stellate cell staining was observed in rat cerebellum. Neurons of the hippocampal denate gyrus often fluoresced in man while only pyramidal cell staining was seen in this region of rat brain. The method should prove to be of value in the detection and tracing of narcotic drugs and may be helpful in investigations of drug tolerance and dependence.

Lysosomes in Brenner's tumor simulating secretory argentaffin granules.

Argentaffin cells were searched for in the epithelial nests of 30 Brenner's tumors, with the Fontana-Masson stain for the screening. Although these cells were found in five tumors, ultrastructural examination of one case, in which the argentaffin cells were multiple, identified the cytoplasmic granules as lysosomes and not as amine precursor uptake and decarboxylation (APUD)-type granules. We conclude that the epithelial component of Brenner's tumors consists of urothelium only and does not include the cells containing argentaffin APUD-type granules.

Ocular immunopathology in murine lupus.

Immunoglobulin deposits and DNA were identified by use of immunofluorescence microscopy within the ciliary process and choroid of a majority of (NZB X W)F1 mice with murine lupus. The number of animals with such deposits increased from 57% at 36 weeks of age to 90% at one year of age. Ultrastructural examination showed electron-dense deposits between the basement laminae of the endothelium and pigmented epithelium of the ciliary process, accompanied by alterations in the epithelial basement membranes. Similar electron-dense deposits were also identified in the vicinity of the lamina choroidocapillaris. The titer of antibody to DNA did not correlate with the presence of ocular deposits, suggesting that time is an important factor in the development of ocular immunopathology in these animals.

Disseminated toxoplasmosis. Case report and review of the literature.

A 40-year-old man had disseminated toxoplasmosis preceded by miliary tuberculosis and associated with skin anergy. In previously reported cases of disseminated toxoplasmosis, the three organs most commonly involved were brain, heart, and lungs. The histopathologic picture of toxoplasmal lymphadenopathy was not present in our case or in previously reported cases. Therefore, we believe that the diagnosis of disseminated toxoplasmosis should not be rejected simply because of the likely absence of toxoplasmal lymphadenopathy in biopsy material. The thymus in our case was morphologically abnormal and showed features suggestive of an autoimmune process.

Histochemistry of steroid receptors in breast cancer: an overview.

Immunofluorescence and a new histochemical technique were employed to assay 226 breast cancer specimens for estrogen receptor. Results showed an overall correlation of 91 percent when compared to those of biochemical assays. The histochemical technique is rapid, easy to perform and reveals the same parameters as does immunofluorescence without the need for antiserum. Tumor cell receptor heterogeneity and location of receptor in cytoplasm or nucleus is readily defined by both methods. These histologic tests should prove to be useful in extending the availability of estrogen receptor analysis to all patients with breast cancer.

Histochemistry of steroid receptors in prostatic diseases.

Tissue obtained from 55 men with prostatic disease was assayed for estrogen and androgen receptors by a newly developed histochemical technique. The material studied consisted of 45 specimens of benign nodular prostatic hyperplasia and 10 specimens of prostatic adenocarcinoma. The results obtained were compared to those of parallel biochemical assays in 17 cases and successfully correlated in 85 percent. The new procedure is rapid, inexpensive and accurate, allowing for the detection of receptor in cytoplasm and/or nucleus and evaluation of receptor heterogeneity. The histochemical method may offer an alternate to biochemical assay of prostatic tissue as contamination with steroid binding globulins does not appear to be a problem at this time.

Histochemical determination of estrogen and progesterone binding in fine needle aspirates of breast cancer. Correlation with conventional biochemical assays.

Estrogen and progesterone binding sites were assayed in patients with primary, recurrent and metastatic mammary carcinoma using a new histochemical technique. This histochemical assay employed fluorescein-labeled conjugates of bovine serum albumin covalently bound to estradiol or progesterone. In a double-blind study in which the results of histochemistry were correlated with those obtained by a conventional biochemical assay, there was 83% correlation for estrogen binding in 41 cases and 87% correlation for progesterone binding in 31 cases.