Effect of steroids on brain lipocortin immunoreactivity.

LCT-1, LCT-2 and LCT-5 were assessed in uninjured rats and rats subjected to a cortical freezing injury or middle cerebral artery (MCA) occlusion. Apart from animals receiving no treatment, other uninjured or injured animals received methylprednisolone (2 or 30 mg/kg) or the 21-aminosteroid U-74389F (10 mg/kg) one day and 2 hours before killing. The animals were killed by decapitation 1 hour after the freezing injury or the MCA occlusion and the area containing the lesion was removed and frozen in Freon. Frozen sections were treated with rabbit polyclonal anti-LCT antibody; binding of antibody was visualized by horseradish peroxidase-conjugated swine antirabbit antibody. Without steroid pretreatment, in the uninjured brain LCT immunoreactivity was absent in the greater part of the brain, except in sporadic microglia. In steroid-pretreated animals and in the freezing lesion of both pretreated and untreated animals there was extensive immunostaining; in the freezing lesion it may be due to passage of systemic LCT across the impaired blood-brain barrier in the lesion. The cellular elements showing immunostaining were meningeal cells, neurons, ependyma, choroid plexus, oligodendroglia and capillary endothelium. It implies that also in the brain the steroid effect is consistent with LCT formation.

The effect of steroid treatment on lipocortin immunoreactivity of rat brain.

Lipocortin-1, lipocortin-2 and lipocortin-5 were immunohistochemically assessed in rats. Apart from animals receiving no treatment, other animals received pretreatment with methylprednisolone, or the 21-aminosteroid U-74389F. Whereas Hpocortin immunoreactivity was absent in the greater part of the brain in animals not pretreated with steroid (except in sporadic microglial cells and choroid plexus), there was obvious immunostaining of parenchymatous elements in steroid pretreated animals. In the steroid pretreated animals lipocortin immunoreactivity of the brain tissue may indicate local formation of lipocortin under the influence of steroids that had entered the tissue. The cellular elements which showed immunostaining included meningeal cells, neurones, ependyma, oligodendroglia and capillary endotheHum.

Direct visualisation and quantification of cellular cytotoxicity using two colour flourescence.

A fluorescence method is described for the evaluation of cell death induced by cellular cytolytic activity. A green fluorescent membrane dye, D275, was used to label various target cell lines and propidium iodide (PI) uptake was used to assay cell death. Natural killer (NK), lymphokine activated killer (LAK) as well as targeted T cell mediated cellular cytotoxicity were quantified using the fluorescence method and compared to results obtained with the 51chromium (51Cr) release assay. A good correlation was found after an assay period of 4-8 h indicating that the fluorescence method is a reliable alternative to the 51Cr release assay.

Distribution of endotracheally instilled surfactant protein SP-C in lung-lavaged rabbits.

In lung-lavaged surfactant-deficient rabbits (n = 6) requiring artificial ventilation, porcine surfactant was instilled endotracheally. This resulted in improvement of lung function so that the animals could be weaned off artificial ventilation. The animals were killed 4 1/2 h after surfactant administration and the porcine surfactant protein was localized in the lung with a MAb. We found surfactant protein in all lobes of the lung but the distribution was not homogeneous. Surfactant protein C was found in less than 15% of the alveolar spaces and in less than 1% of the bronchi.

Immunochemistry and immunogenicity of low molecular weight human salivary mucin.

The purposes of this study were to examine the immunogenicity of the low molecular weight human salivary mucin (MG2) and determine its distribution within major and minor human salivary glands. Anti-MG2 sera were produced in Balb/c mice by a variety of immunization schedules. Chromatographically or electrophoretically purified MG2 and partially purified mucin chromatographic fractions exposed to mild denaturing conditions were not immunogenic. Only MG2 without prior exposure to urea or guanidine was able to elicit an immune response. A murine anti-MG2 monoclonal antibody (clone 1/F9) was produced and its monospecificity confirmed by immuno-dot blotting and SDS-PAGE Western transfer. Clone 1/F9 (IgG1; kappa) was of moderate affinity and was directed to a Pronase- and TPCK trypsin-sensitive but periodate-resistant epitope which was not blood group- or sialic acid-specific. Immunocytochemical studies of frozen tissue sections with clone 1/F9 using both indirect and direct methods revealed that MG2 was more heterogeneously distributed within submandibular than labial glands and was not found in parotid or palatine glands. The use of a polyclonal rabbit anti-MG2 reagent in either frozen or paraffin-embedded tissues gave the same immunocytochemical results as those obtained with the monoclonal antibody.

Acquired factor VIII inhibitor with squamous cell cancer of the epiglottis.

A 73-year-old woman developed an acquired factor VIII inhibitor in association with squamous cell carcinoma of the epiglottis. The inhibitor was an IgG antibody that reacted with factor VIII in vitro and in vivo. Intravenous gamma-globulin therapy was successful in reducing the inhibitor so that curative surgery could be undertaken. With surgical resection of the tumor the inhibitor did not recur. The relevance of this type of coagulation disorder to the surgical management of the patient's head and neck cancer is discussed.

Immunochemistry of high molecular-weight human salivary mucin.

The purpose of this study was to determine the distribution of mucin glycoprotein 1 (MG1) within submandibular, parotid, labial and palatine salivary tissues. Formalin-fixed and frozen tissue sections were examined histochemically with PAS, Alcian blue and Meyer's mucicarmine, and immunocytochemically with an anti-mucin glycoprotein 1 monoclonal antibody (clone 3/E8). Clone 3/E8 was produced in Balb/c mice using mucin-enriched chromatographic fractions from submandibular-sublingual saliva. The monospecificity of 3/E8 was confirmed by immuno-dot blotting and SDS-PAGE/electrophoretic transfer. Clone 3/E8 (IgG1; kappa) was of moderate affinity, and was directed to a carbohydrate-containing, TPCK-trypsin-insensitive and pronase-insensitive epitope on this mucin, which was not blood-group specific. The location of mucin glycoprotein 1 was determined by both indirect (peroxidase-antiperoxidase) and direct methods. Mucin glycoprotein 1 was localized within all labial acini examined, but was not found within parotid tissues. Histochemical methods stained all submandibular, palatine and labial acini, but immunocytochemistry with monoclonal antibody revealed heterogeneous staining with clone 3/E8 in submandibular and palatine tissues. These findings suggest the presence of mucin glycoprotein 1-specific acinar cell subpopulations within human submandibular and palatine salivary tissues.

Detection of small cell lung cancer metastases in bone marrow aspirates using monoclonal antibody directed against neuroendocrine differentiation antigen.

To detect metastases in the bone marrow of patients with small cell lung cancer, immunofluorescence with a monoclonal antibody detecting a membrane antigen (MOC-1) associated with small cell lung cancer was performed on 53 bone marrow aspirates from 30 patients. In 19 (63%) patients MOC-1 reactive cells were detected. Simultaneous histopathological examination of the bone marrow biopsy specimens detected tumour cells in only six (20%). The method is more sensitive than conventional histochemical staining of bone marrow aspirate and may eventually be able to show additional subgroups, such as patients with limited disease who might benefit from adjuvant radiotherapy or surgery.

Immunoperoxidase staining on frozen tissue sections as a first screening assay in the preparation of monoclonal antibodies directed against small cell carcinoma of the lung.

To prepare monoclonal antibodies against small cell carcinoma of the lung (SCCL), we have used an SCCL-derived cell line as immunogen. A first screening of hybridoma supernatants was performed on frozen tissue sections of a biopsy with histologically proven SCCL involvement. Screening on tissue sections is a valuable technique, especially for the isolation of monoclonal antibodies directed against tumor antigens. A drawback of this procedure is that it is laborious. To circumvent this, we have reduced the number of supernatants to be screened by increasing the number of seeded hybridomas per well. Although this resulted in the growth of 5-20 hybridomas per well, among which there was always at least one that secreted antibodies against 'common antigens', clones that secreted specific antibodies could still be revealed by testing supernatants which were preabsorbed with thrombocytes. This has resulted in the isolation of 7 monoclonal antibodies directed against SCCL associated antigens.

Primary hyperparathyroidism and laryngeal carcinoma: a cause of associated hypercalcemia.

Two cases of laryngeal carcinoma are presented which, upon initial evaluation, were noted to have asymptomatic hypercalcemia with no evidence of bone metastases. Exploration of the parathyroid glands at the time of definitive surgery revealed parathyroid pathology in both cases. Calcium levels returned to normal postoperatively in both cases. We conclude that in patients with head and neck cancer with hypercalcemia and no evidence of bone metastases, parathyroid pathology should be considered. Exploration of the parathyroid glands at the time of definitive surgery or as a separate procedure may prove rewarding.

Granulomatous laryngitis of unknown etiology.

A 21-year-old man was admitted for odynophagia and hoarseness of four months duration. He smoked one and a half packs of cigarettes a day and occasionally inhaled marijuana. Indirect laryngoscopy revealed a massive swelling of the entire epiglottis, aryepiglottic folds, and arytenoids. The histopathologic diagnosis of chronic but active nonspecific inflammation was made. Combined antibiotics and steroid therapy gave temporary relief. He was readmitted several months later with progressive shortness of breath, dysphagia, and hoarseness. Biopsy of the epiglottic tissues showed multiple noncaseating epithelioid granulomatous lesions consistent with sarcoidosis. All pertinent laboratory tests failed to establish a definitive diagnosis. The patient eventually underwent supraglottic laryngectomy. He has been symptom-free for 20 months following surgery.

Olfactory neuroblastoma (esthesioneuroblastoma): a light and ultrastructural study of two cases.

Olfactory neuroblastoma is a malignant neoplasm with a varied biological behavior. Its clinical course is unpredictable and there is no correlation between its microscopic features and biological behavior. The present study deals with light and ultrastructural characteristics of two cases of olfactory neuroblastoma of the nasal cavity. In one patient, the definitive diagnosis was established on the basis of ultrastructural features of the lesion. The most consistent fine structural findings were the presence of intracytoplasmic densecored neurosecretory granules, "true" and "pseudo-" rosettes, and the neuritic processes emanating from the tumor cells. On the basis of their biochemical, histochemical, and ultrastructural characteristics, olfactory neuroblastomas are similar to neuroblastomas arising from the adrenals or sympathetic nervous system. These findings, therefore, support the hypothesis that olfactory neuroblastomas are most likely of neural crest origin and thus belong to a group of neoplasms collectively known as "apudomas" or neurocristomas. The literature review strongly favors combined surgery and postradiation as the most effective treatment of olfactory neuroblastoma.

A nonfunctioning paraganglioma of vagus nerve: an ultrastructural study.

The clinical, histological, and ultrastructural aspects of a cervical paraganglioma of the vagus nerve, in a 66-year-old white man, have been discussed in detail. Ultrastructurally, the tumor chief cells contained characteristic membrane-bound and dense-cored neurosecretory granules which ranged in size from 85 millimicron to 190 millimicron. Unlike earlier ultrastructural reports, the present study showed the presence of sustentacular or supporting cells. These cells were smaller, darker, polymorphic, and were commonly located at the periphery of a single or group of chief cells. Furthermore, unlike earlier reports on vagal paragangliomas, nonmyelinated nerve fibers and an occasional axon were identified in the present fine structure study.

Chondroblastic osteosarcoma of the larynx.

Osteosarcoma of the larynx is a rare lesion. Acceptable cases in the literature are difficult to find. We report a case in which total laryngectomy was carried out for a large, obstructing mass that arose from the cricoid cartilage. We present the histologic criteria for the diagnosis of chondroblastic osteosarcoma.