Cytological analysis of the urn cell complex of Sipunculus nudus before and after serum-induced secretion.

This study analyzes the cytology of the urn cell complex (UCC) of Sipunculus nudus, an invertebrate cell model for humorally regulated mucus secretion. An unstimulated UCC is composed of a vesicle cell and a ciliated cell joined together by desmosomes. Another cell population (third-type cells) is frequently associated with ciliated cells. Vesicle cells are thin, have few mitochondria or lipid droplets, and enclose a bubble-like cavity containing microfibrillar material. Ciliated cells contain several rows of cilia that are anchored by prominent rootlets and propel UCCs forward. Five to six concentric bundles of microfilaments are distributed along the outer convexity of ciliated cells and may have a role in the plasticity of the UCC. Many fibrillar deposits that lack a demonstrable limiting membrane are distributed around intracytoplasmic vacuoles facing the mouth-like opening of the UCC. These deposits are reactive to periodic acid-Schiff and resistant to diastase. After stimulation with serum, they appear to migrate through the ciliated cell's plasma membrane, contributing to the formation of a secretory tail. Discharge of secretory material is not observed in third-type cells, which instead contain lysosome-like granules and autophagic-like vacuoles and become displaced distalward by the emerging tail of the UCC. This study indicates that formation and elongation of the UCC secretory tail are functions of ciliated cells.

Ultrastructure of an arterial lesion induced in rats by fenoldopam mesylate, a dopaminergic vasodilator.

Fenoldopam mesylate (FM) is a dopaminergic vasodilator with demonstrated efficacy and a favourable safety profile in hypertensive and congestive heart failure patients. FM produced a novel arterial lesion in renal and splanchnic arteries of rats, but not dogs or monkeys. The studies reported here were undertaken to investigate the ultrastructure of the arterial lesion induced in rats by FM in an attempt to shed light on its pathogenesis. Rats were infused intravenously with FM, either 50 micrograms/kg/min for 1 or 4 h, or 5 or 100 micrograms/kg/min for 24 h. Control rats were infused for 4 or 24 h with vehicle alone. Perfusion-fixed tissue from the stomach and pancreas of control and drug-treated rats was examined by transmission electron microscopy. No arterial lesions were seen in rats infused with the drug for 1 or 4 h, or in control rats. All drug-treated rats infused with 5 or 100 micrograms/kg/min of FM for 24 h had lesions in subserosal gastric arteries and interlobular pancreatic arteries. In areas of mild arterial damage, medial smooth muscle cells contained intracytoplasmic pseudovacuoles, autophagic vacuoles, and electron-dense, myofilamentous inclusions. More severe lesions were characterized by overt medial necrosis and haemorrhage. The endothelium of affected arteries was invariably intact, except in areas of severe medial damage. The internal elastic lamina and connective tissue elements within the arterial wall were unaffected. These findings suggest that medial smooth muscle cells are the primary site of damage caused by fenoldopam mesylate in splanchnic arteries of the rat. This iatrogenic arterial lesion could provide an interesting model to study the response of medial smooth muscle to pharmacologically mediated injury.

Rabbit endocervical epithelium: morphometric analysis of secretory cell populations.

In this report we quantitated ultrastructural changes in two cytologically distinct secretory cell populations from the rabbit endocervix. Type I and type II cells from estrous animals differ only in the presence of one or more empty cytoplasmic vacuoles in type II cells. Comparing type II cells from 5-day pseudopregnant (PSP) rabbits with type II cells from estrous controls, there is no increase (P greater than .05) in the average vacuole volume. When type I and type II cells from PSP animals are compared to cells from estrous controls, there is a decrease (P less than .01) in the average cell volume, a decrease (P less than .01) in the average nuclear volume, and a decrease (P less than .01) in the average granule volume. This reduction in the granule content of secretory endocervical cells was correlated with a dramatic decrease in protein glycosylation into the microsomal fraction. Serum estradiol concentrations for estrous (13.7 +/- 1.0 pg/ml) and PSP (18.1 +/- 1.5 pg/ml) animals were comparable. However, the 36-fold increase in serum progesterone concentrations for PSP (12.04 +/- 1.7 ng/ml) animals compared to estrous (0.33 +/- 0.1 ng/ml) animals may be responsible for the decrease in protein glycosylation.

Gold-induced immune thrombocytopenia in the dog.

In two seven-year studies with gold compounds in dogs of both sexes, thrombocytopenia was observed after 45 to 72 months of dosing in three of 14 and two of 14 dogs in high-dose groups that received 2.4 to 3.6 mg/kg of auranofin per day orally or 0.5 to 2.0 mg/kg of gold sodium thiomalate intramuscularly once every three days, respectively. An immune basis for the disorder was suggested by the apparent consumptive nature of the thrombocytopenia (increased bone marrow megakaryocytes and large peripheral blood platelets), the response to corticosteroid therapy and the demonstration of increased platelet-associated immunoglobulin. The latter was demonstrated with a solid phase radioimmunoassay and by electron microscopy using a staphylococcal protein A-colloidal gold conjugate. Platelet-associated immunoglobulin decreased as the platelet counts rose, and in one dog monitored over periods of steroid-induced remissions and subsequent relapses, the amount of platelet-associated immunoglobulin G correlated inversely with the platelet count (r = 0.82). These findings suggest that the long-term administration of gold compounds in dogs is associated with a dose-dependent incidence of thrombocytopenia, which is immune-mediated and similar to that associated with parenteral chrysotherapy in man. The application of tests for platelet-associated immunoglobulin to canine patients with immune thrombocytopenia should be useful in the diagnosis of the disorder in clinical practice.

Changes in the macrophage content of lung metastases at different stages in tumor growth.

The macrophage content of experimental B16 melanoma metastases at different stages of their growth has been quantified with the use of morphometry in conjunction with a recently developed histochemical method for selectively staining intratumoral macrophages. Data are presented from analyses of 954 sections of 155 individual lung metastases, showing that the macrophage content of individual B16 melanoma lung metastases not only varies significantly but also falls dramatically once metastases contain more than 700 tumor cells. In addition to providing new information on host response reactions of micrometastases, these experiments also indicate that conclusions on intratumoral macrophages derived from studies of large primary tumors and metastases in advanced stages of growth may have little or no relevance to events in micrometastases.

Ultrastructural immunocytochemical localization of lysozyme in the mucociliary epithelium of the rabbit endocervix in different hormonal states.

The distribution of lysozyme in the endocervix of estrous, pseudopregnant, and ovariectomized rabbits was studied using two different immunocytochemical techniques--the unlabeled antibody enzyme method of Sternberger et al. (1970) and the peroxidase-labeled antibody method of Taylor and Burns (1974). With both procedures, a fine immunostaining precipitate was seen over the entire area of basal mucous granules, while immunodeposits were coarser and mostly located in the outer zone of central and apical granules. A nonspecific staining was noted when tissues were reacted with peroxidase-antiperoxidase complex alone. This troublesome artifact was abolished by preincubating tissues with human IgA. This step did not affect the specific immunostaining for lysozyme yet nonspecific staining was absent from specificity and method controls carried out for both immunocytochemical procedures. The presence of high levels of lysozyme in the endocervical epithelium of estrous rabbits was also confirmed in enzymatically isolated endocervical epithelia using the lysoplate method of Osserman and Lawlor (1966). Mucous granules and immunostainable intracellular lysozyme were abundant during estrus, decreased during early pseudopregnancy, and were absent after long-term ovariectomy. However, they were restored by the administration of estradiol (5 micrograms/12 hours/10 days) to ovariectomized animals. These data indicate a common hormonal regulation and secretory mechanism for endocervical mucous glycoproteins and lysozyme.

Enrichment of lysozyme-containing cells from the rabbit endocervix by unit gravity sedimentation.

Lysozyme is a bacteriolytic enzyme component of the secretory granules of endocervical mucous cells. In order to study the subcellular distribution of this enzyme in specific cell populations, endocervical cells from estrous and 5-day pseudopregnant rabbits were separated by unit gravity sedimentation. The application of this technique to pronase-dispersed endocervical cells from estrous rabbits resulted in the isolation and enrichment of two mucous cell types that were distinguished morphologically into type I and type II cell populations. Lysozyme was identified in both cell types, using an unlabeled antibody enzyme method, and the degree of staining paralleled the number of mucous granules. In the absence of estrogen dominance in 5-day pseudopregnant rabbits, there was a 50% reduction in the number of mucous cells with a concomitant reduction in both the number of secretory granules per cell and the intracellular concentration of lysozyme. In the absence of ovarian steroid hormones, i.e., 15-16 weeks after ovariectomy, endocervical cells were devoid of secretory granules and lysozyme staining was negative. Enriched populations of endocervical cells represent a potential experimental model for studying the hormonal role in the regulation of lysozyme synthesis by specific cell populations.

Isolation and characterization of rabbit endocervical cells.

Three cytologically distinct cell populations were identified, in addition to ciliated cells, when a unit gravity sedimentation procedure was applied to pronase-dispersed rabbit endocervical cells. Two of these cell populations contained histochemically distinguishable (periodic acid- Schiff [PAS]) mucoproteins and were designated vacuolated and granular PAS-positive cells. The third, designated as vacuolated PAS-negative, did not contain secretory granules. Cell integrity was confirmed by trypan blue dye exclusion, [(3)H]leucine incorporation, and ultrastructural analysis. To demonstrate hormonal modulation of endocervical cell morphology, cell distribution profiles were compared from animals in different hormonal states. In the absence of estrogen dominance, PAS- positive cells from 5-d pseudopregnant rabbits were reduced 50 percent, while vacuolated PAS-negative cells increased fourfold as compared with estrous cell populations. The PAS-positive cells sedimented toward the top of the gradient where the bovine serum albumin concentrations were lower, consistent with a reduction in the number of secretory granules. In the sustained absence of ovarian steroid hormones, the number of PAS-positive mucous cells from ovariectomized rabbits was reduced to only 4 percent of the total endocervical cell population. The biosynthetic capacity of isolated endocervical cells was determined by incubating the three nonciliated cell populations from estrous and 5-d pseudopregnant rabbits for 36 h with the mucin precursor, [(14)C]N-acetyl- D-glucosamine. Only PAS-positive cells incorporated significant amounts of labeled precursor. This study indicates that steroid hormones influence cervical secretions by modulating the type of endocervical cells.