Ultrastructural studies of semen abnormalities and Herpesvirus associated with cultured testicular cells from domestic turkeys.

Abnormal cells and macrophages found in white and yellow turkey semen were studied by electron microscopy. Yellow semen contained many abnormal cells, most of which were large and round or smaller and ellipsoidal. It was concluded that they were aberrant spermatids, with differentiation being more complete in the smaller cells. Only a few cells of the smaller type were detected in normal white semen. Macrophages were occasionally seen in white semen but were numerous in yellow semen. Phagocytic vacuoles of these cells contained structural elements of spermatozoa and abnormal spermatids. Virus particles were not detected in any of the seminal cells observed. Ultrastructure studies of cultured testicular cells obtained from several of the turkeys examined showed the presence of intranuclear Herpesvirus particles in germinal cells. Macrophages from the testicular cultures seldom were seen with intranuclear Herpesvirus, although these cells commonly were found with Herpesvirus particles and cellular debris contained within phagocytic vacuoles.     

Histochemical evidence for lysosomal storage of acid glycosaminoglycans in splenic cells of rats treated with tilorone

Tilorone, an agent with antiviral and antitumor activities, has previously been reported to produce clear cytoplasmic vacuoles in many cell types of the rat. The present study on rat spleen was planned to investigate the ultrastructural and histochemical features of the tilorone-induced vacuoles occurring in sinus endothelium, trabecular smooth muscle cells, and macrophages of the red pulp. Evidence was obtained that the vacuoles represent lysosomes overloaded with acid glycosaminoglycans (aGAG). The main purpose of the present study was to overcome the technical difficulties of preserving the intralysosomal storage materials which were highly water-soluble and non-fixable by aldehyde fixatives. Preservation, at least for the light microscopical level, was achieved by freeze drying and by means of cationic dyes which served also to characterize the storage materials on the basis of their acidities. Tissue slices were used to determine the critical MgCl2 concentration necessary to abolish Alcian blue staining; cartilage and mast cells served as references. For the storage material in sinus endothelium, the critical MgCl2 concentration was found to be greater than 0.7 M, as compared to greater than 0.5 M for cartilage and greater than 0.9 M for mast cells. The storage materials in trabecular cells and macrophages were slightly less acidic than cartilaginous matrix and more heterogeneous than that in sinus endothelium. Ultrastructurally, positive staining with high iron diamine (HID) confirmed the presence of aGAG within the tilorone-induced vacuoles.     

Histochemical evidence for lysosomal storage of acid glycosaminoglycans in splenic cells of rats treated with tilorone

Summary Tilorone, an agent with antiviral and antitumor activities, has previously been reported to produce clear cytoplasmic vacuoles in many cell types of the rat. The present study on rat spleen was planned to investigate the ultrastructural and histochemical features of the tilorone-induced vacuoles occurring in sinus endothelium, trabecular smooth muscle cells, and macrophages of the red pulp. Evidence was obtained that the vacuoles represent lysosomes overloaded with acid glycosaminoglycans (aGAG). The main purpose of the present study was to overcome the technical difficulties of preserving the intralysosomal storage materials which were highly water-soluble and non-fixable by aldehyde fixatives. Preservation, at least for the light microscopical level, was achieved by freeze drying and by means of cationic dyes which served also to characterize the storage materials on the basis of their acidities. Tissue slices were used to determine the critical MgCl₂ concentration necessary to abolish Alcian blue staining; cartilage and mast cells served as references. For the storage material in sinus endothelium, the critical MgCl₂ concentration was found to be >0.7 M, as compared to >0.5 M for cartilage and >0.9 M for mast cells. The storage materials in trabecular cells and macrophages were slightly less acidic than cartilagineous matrix and more heterogeneous than that in sinus endothelium. Ultrastructurally, positive staining with high iron diamine (HID) confirmed the presence of aGAG within the tilorone-induced vacuoles.     

Infectious HIV-1 assembles in late endosomes in primary macrophages

Although human immunodeficiency virus type 1 (HIV-1) is generally thought to assemble at the plasma membrane of infected cells, virions have been observed in intracellular compartments in macrophages. Here, we investigated virus assembly in HIV-1-infected primary human monocyte-derived macrophages (MDM). Electron microscopy of cryosections showed virus particles, identified by their morphology and positive labeling with antibodies to the viral p17, p24, and envelope proteins, in intracellular vacuoles. Immunolabeling demonstrated that these compartments contained the late endosomal marker CD63, which was enriched on vesicles within these structures and incorporated into the envelope of budding virions. The virus-containing vacuoles were also labeled with antibodies against LAMP-1, CD81, and CD82, which were also incorporated into the viral envelope. To assess the cellular source of infectious viruses derived from MDM, virus-containing media from infected cells were precipitated with specific antibodies. Only antibodies against antigens found in late endosomes precipitated infectious virus, whereas antibodies against proteins located primarily on the cell surface did not. Our data indicate that most of the infectious HIV produced by primary macrophages is assembled on late endocytic membranes and acquires antigens characteristic of this compartment. This notion has significant implications for understanding the biology of HIV and its cell-cell transmission.     

AUTOPHAGIC VACUOLES PRODUCED IN VITRO : I. Studies on Cultured Macrophages Exposed to Chloroquine

Mouse macrophages exposed to 30 µg/ml of chloroquine in vitro develop autophagic vacuoles containing various cytoplasmic components and acid phosphatase. The early toxic vacuoles appear in the perinuclear region within 15 min; on electron microscopy, they show irregular shape, amorphous moderately dense content, apparent double membranes, and in some instances curved thin tubular extensions with a central, dark linear element. Cytoplasmic structures are probably transported into the vacuoles by invagination of the vacuolar membrane. After exposure to chloroquine for 1–4 hr, macrophages display large vacuoles containing degraded cytoplasmic structures, membranous whorls, and amorphous material. When chloroquine is removed by changing the culture medium after 4 hr, the cells survive and 24 hr later they exhibit no abnormality except for large cytoplasmic dense bodies packed with membrane lamellae. During recovery chloroquine disappears from the cells. 24 hr after exposure to chloroquine the macrophages have accumulated less hydrolases than control cells.     

Electron Microscopic Study of the Phagocytosis Process in Lung

Diluted India ink was instilled into the nasal cavity of mice and the lungs of some animals were fixed with osmium tetroxide at various intervals after one instillation. The lungs of other animals were fixed after 4, 7, 9, 16, or 18 daily instillations. The India ink was found to be phagocytized almost exclusively by the free alveolar macrophages. A few particles are occasionally seen within thin portions of alveolar epithelium, within the "small" alveolar epithelial cells, or within occasional leukocytes in the lumina of alveoli. The particles are ingested by an invagination process of the plasma membrane resulting in the formation of intracellular vesicles and vacuoles. Ultimately large amounts of India ink accumulate in the cell, occupying substantial portions of the cytoplasm. The surfaces of phagocytizing macrophages show signs of intense motility. Their cytoplasm contains numerous particles, resembling Palade particles, and a large amount of rough surfaced endoplasmic reticulum. These structures are interpreted as indicative of protein synthesis. At the level of resolution achieved in this study the membranes of this reticulum appear as single dense "lines." On the other hand, the plasma membrane and the limiting membranes of vesicles and of vacuoles often exhibit the double-line structure typical of unit membranes (Robertson, 37). The inclusion bodies appear to be the product of phagocytosis. It is believed that some of them derive from the vacuoles mentioned above, and that they correspond to similar structures seen in phase contrast cinemicrographs of culture cells. Their matrix represents phagocytized material. Certain structures within this matrix are considered as secondary and some of these structures possess an ordered form probably indicative of the presence of lipid. The possible origin and the fate of alveolar macrophages are briefly discussed.     

鸡中枢淋巴器官肥大细胞的组织化学与形态学

对哺乳动物的,特别是啮齿动物和人类肥大细胞已有了比较深入的研究, 但关于家禽肥大细胞的研究很少.本研究旨在阐明鸡中枢淋巴器官中肥大细胞的组织化学与形态学特征.本研究证实Carnoy 氏液是鸡肥大细胞的优良的固定液,而中性缓冲福尔马林(NBF) 却阻断了大多数肥大细胞的着染力.甲苯胺蓝和阿尔新蓝是鸡肥大细胞的良好的染料,但阿尔新蓝能使更多的肥大细胞着染,虽然其也可使杯状细胞着染.作者的一种新的染色法, 长时间阿尔新蓝染色(LAB-S)可用于NBF固定的组织中肥大细胞的染色,因为其着染的细胞数与Carnoy 氏液固定甲苯胺蓝染色的细胞数无显著差异(P<0.001).在胸腺髓质中见有大量的肥大细胞,而胸腺皮质仅可见个别肥大细胞位于血管周围及小叶间结缔组织中.腔上囊的皮质与髓质中很少见有肥大细胞.肥大细胞有血管周围分布的倾向,但一个有趣的发现是血管内偶尔也有个别肥大细胞.电镜下可见肥大细胞的胞浆颗粒内充满无定形的颗粒状基质,但其电子密度有的较高,有的较低.少数胞浆颗粒内有旋涡状及网状亚微结构.但未见有人类肥大细胞胞浆颗粒内特征性的晶格状和卷轴状的亚微结构,也未见到在绵羊肥大细胞中描述过的特殊亚微结构.     

The Thymic Microenvironment of the Common Sole,Solea solea

Abstract The thymus of the sole Solea solea contained lymphoblasts and thymocytes within a network of pale and dark epithelial cells. The pale cells were characterized by tonofilaments and desmosomes and some embraced rodlet cells within their cytoplasmic processes. The dark epithelial cells had numerous electron-dense inclusions and electron-lucent vacuoles. Lymphocytes were closely associated with the plasma membrane of both types of epithelial cells and with macrophages. Breakdown of effete lymphocytes appeared to be the main function of the macrophages. Some macrophages were multinucleated. Those containing melanin granules associated with phagosomes were classified as melanomacrophages. Pigment cells including melanophores and guanophores were present along the connective tissue trabeculae and surrounding the blood vessels. A few plasma cells and mucous cells were present.     

Involvement of smooth muscle cells in collagen degradation in the postpartum uterus

Ultrastructural study of gravid and postpartum involuting human uteri revealed a number of cells containing collagen fibrils in their cytoplasm. In gravid uteri these cells could be identified as macrophages and fibroblasts; in the postpartum uteri smooth muscle cells (SMC) were also found, containing cytoplasmic collagenous vacuoles. The morphology of intracellular collagen in SMC was similar to that observed in macrophages: fragments of banded collagen fibrils with a diameter corresponding to that of extracellular collagen were located within structures considered to be phagosomes. Limiting membranes were always smooth, most often in apposition to the fibrils that were single or packed in small groups; some cytoplasmic vacuoles contained banded elongated profiles barely discernable as collagen. The collagen fibrils within SMC of the involuting human uterus are regarded as a morphological manifestation of heterogenic enclosure of collagen fibrils and their intracellular degradation. It seems that in the postpartum uterus, where a substantial amount of collagen needs to be removed rapidly, both macrophages and SMC are involved in the process of collagen phagocytosis and degradation. These data suggest that SMC may be involved in the cellular mechanism for collagen breakdown in remodelling SMC-containing tissues like the uterus and the vascular wall.     

Effects of leupeptin on endocytosis and membrane recycling in rat visceral yolk-sac endoderm

The effect of exposure to leupeptin (25 micrograms/ml for 24 h) on the endocytotic activity and the membrane flow of apical cell membranes was studied in endodermal cells of cultured rat visceral yolk sacs by applying a double-labelling method using concanavalin-A ferritin (Con-A Fer) and horseradish peroxidase (HRP). Control and leupeptin-treated yolk sacs were labelled with Con-A Fer at 4 degrees C and then incubated with HRP for 5, 15 or 60 min at 37 degrees C. In controls, HRP reaction product was detected after 5 min in many of the apical vacuoles as well as a few lysosomes; after 15 min, reaction product was observed in all apical vacuoles and in lysosomes of various sizes. These HRP-positive structures usually contained a variable amount of membrane-bound Fer. After 60 min, all apical vacuoles and almost all lysosomes exhibited HRP reactions, but only some of these structures contained Fer particles. At this time, many apical canaliculi (which are involved in membrane recycling) exhibited positive HRP reactions and sometimes also contained Fer particles. In leupeptin-treated cells, HRP reaction product and variable amounts of membrane-bound Fer particles were found in apical vacuoles after 5 min; after 15 min, both labels were also observed in some small lysosomes, and after 60 min, they were found in all apical vacuoles as well as some small and middle-sized lysosomes. Significantly fewer labelled apical vacuoles, lysosomes and apical canaliculi were present after leupeptin treatment than in controls at corresponding times. At all times examined, the giant lysosomes found in leupeptin-treated cells did not exhibit any labeling.(ABSTRACT TRUNCATED AT 250 WORDS)     

THE USE OF SILVER NITRATE AS A VITAL STAIN, AND ITS DISTRIBUTION IN SEVERAL MAMMALIAN TISSUES AS STUDIED WITH THE ELECTRON MICROSCOPE

After chronic administration of a dilute solution of silver nitrate in drinking water to rats, mice, and guinea pigs, granular deposits of metallic silver were detected in electron micrographs of the kidney, liver, thyroid, and pancreas. The silver deposits were in the form of extremely dense, angular particles with sharp outlines. They varied from aggregates a few microns in diameter down to granules at the limit of resolution of the electron microscope. The principal sites of deposition were (1) basement membranes, especially those of the renal glomeruli, proximal convoluted tubules, and various glands, and those associated with vascular endothelium, and (2) the cytoplasm of fixed and free macrophages. Both in Kupffer cells lining hepatic sinusoids and in the wandering macrophages of other tissues, the silver was segregated in discrete vacuoles. In addition, granular deposits were observed in occasional vesicular structures in the proximal convoluted tubules of the kidney, the hepatic cells, and the pancreatic acinar cell. These structures, in favorable preparations, contained an outer double layered membrane and internal folds similar to those of mitochondria, from which they appear to have been derived. The significance of these findings in heavy metal poisoning and in cellular physiology is briefly discussed.     

The fine structure of the cells of the mouse peritoneum

Summary The cells of the peritoneum of the mouse have been examined with the electron microscope both by studying the gastro-splenic omentum and by washing the cells out of the peritoneal cavity. They comprise mesothelial cells, mast cells, lymphocytes and macrophages. The mesothelial cells were probably nearly all degenerate. The mast cells released granules which were phagocytosed by the other cells. The lymphocytes were either classical small lymphocytes, or rather larger cells similar to previously described immunoblasts. The macrophages varied considerably in size. Some were probably derived from the covering cells of the milk spots. They contained varying numbers of dense bodies, with the structure of lysosomes. A series of appearances was seen which suggested that these were synthesized in the granular endoplasmic reticulum. A gradation of structure was seen between lymphocytes and small macrophages.The gastro-splenic omentum consisted of two layers of mesothelium, in places fenestrated. The milk spots which were scattered throughout this structure were covered by cells similar to macrophages, and had a core of lymphoid cells in which ran a small blood vessel. The most notable difference between the mesothelial cells and the macrophages was the presence of many small caveolae at the surface of the mesothelial cells, and of larger vacuoles and indentations at the surface of the macrophages. Acknowledgements. I am grateful to Professor R. Barer for much advice and criticism, to Dr. G. A. Meek for guidance on electron microscopy, and to Miss M. Tune and Mr. M. Turton for photographic assistance.This work was supported by a grant from the Medical Research Council and by grants to the Department from the S.R.C., Nuffield Foundation and Unilever Limited.     

Cytoplasmic assembly and accumulation of human immunodeficiency virus types 1 and 2 in recombinant human colony-stimulating factor-1-treated human monocytes: an ultrastructural study.

Recombinant human colony-stimulating factor-1-treated human peripheral blood-derived monocytes-macrophages are efficient host cells for recovery of the human immunodeficiency virus (HIV) from blood leukocytes of patients with acquired immunodeficiency syndrome. These cells can be maintained as viable monolayers for intervals exceeding 3 months. Infection with HIV resulted in virus-induced cytopathic effects, accompanied by relatively high levels of released progeny virus, followed by a prolonged low-level release of virus from morphologically normal cells. In both acutely and chronically infected monocytes, viral particles were seen budding into and accumulating within cytoplasmic vacuoles. The number of intravacuolar virions far exceeded those associated with the plasma membrane, especially in the chronic phase, and were concentrated in the perinuclear Golgi zone. In many instances, the vacuoles were identified as Golgi elements. Fusion of virus-laden vacuoles with primary lysosomes were rare. The pattern of cytoplasmic assembly of virus was observed with both HIV types 1 and 2 and in brain macrophages of an individual with acquired immunodeficiency syndrome encephalopathy. Immunoglobulin-coated gold beads added to acutely infected cultures were segregated from the vacuoles containing virus; relatively few beads and viral particles colocalized. The assembly of HIV virions within vacuoles of macrophages is in contrast to the exclusive surface assembly of HIV by T lymphocytes. Intracytoplasmic virus hidden from immune surveillance in monocytes-macrophages may explain, in part, the persistence of HIV in the infected human host.     

Analysis of pure and mixed murine mast cell colonies

Mast cells have been proposed to originate from diverse sources, including connective tissues, macrophages, T lymphocytes, and hemopoietic cells. Evidence for a hemopoietic origin of mast cells includes the presence of mast cell precursors in spleen colonies and the presence of mast cells in hemopoietic colonies in culture. Here we report a detailed analysis of mouse spleen mixed hemopoietic colonies containing mast cells. All of the colonies in cultures plated at low cell densities were individually removed for analysis by May-Grunwald-Giemsa staining on day 15 of culture. Examination of five dishes which contained a total of 82 colonies showed 16 pure mast cell colonies and 36 mixed mast cell colonies. Sixteen different combinations of cell types were seen and were not distinguishable from each other in situ. The most diverse type of mixed colony contained macrophages (m), neutrophils (n), eosinophils (e), mast cells (Mast), megakaryocytes (M), erythroid cells (E), and blast cells. The clonal origin of mixed mast cell colonies was established by the replating of single cells obtained from blast cell colonies. Individual cells were removed with a micromanipulator, replated, and allowed to grow for 15 days. Cytospin preparations of 10 such colonies showed diverse combinations of cell lineages which were seen in the different types of mixed mast cell colonies described above. Replating studies of mixed mast cell colonies were carried out and a high incidence of replating was seen. Approximately one half of these colonies formed only mast cell colonies upon replating. Further studies showed that pure mast cell colonies could be serially replated four to five times. The replating efficiency of cells in the primary mast cell colonies varied over a wide range (2.5–44%) with an average replating efficiency of 13%. The data also revealed that cells containing metachromatic granules possess significant proliferative capacity. From these studies of pure and mixed mast cell colonies, we concluded (1) that mast cells are in wide variety of types of mixed colonies and that the in situ identification of mixed colonies is unreliable, (2) that mast cells are derived from pluripotent hemopoietic stem cells, and (3) that mast cells with metachromatic granules can have a high proliferating ability.     

Mast cells, macrophages, and crown-like structures distinguish subcutaneous from visceral fat in mice

Obesity is accompanied by adipocyte death and accumulation of macrophages and mast cells in expanding adipose tissues. Considering the differences in biological behavior of fat found in different anatomical locations, we explored the distribution of mast cells, solitary macrophages, and crown-like structures (CLS), the surrogates for dead adipocytes, in subcutaneous and abdominal visceral fat of lean and diet-induced obese C57BL/6 mice. In fat depots of lean mice, mast cells were far less prevalent than solitary macrophages. Subcutaneous fat contained more mast cells, but fewer solitary macrophages and CLS, than visceral fat. Whereas no significant change in mast cell density of subcutaneous fat was observed, obesity was accompanied by a substantial increase in mast cells in visceral fat. CLS became prevalent in visceral fat of obese mice, and the distribution paralleled mast cells. Adipose tissue mast cells contained and released preformed TNF-α, the cytokine implicated in the pathogenesis of obesity-linked insulin resistance. In summary, subcutaneous fat differed from visceral fat by immune cell composition and a lower prevalence of CLS both in lean and obese mice. The increase in mast cells in visceral fat of obese mice suggests their role in the pathogenesis of obesity and insulin resistance.     

APPEARANCE AND DISTRIBUTION OF FERRITIN IN MOUSE PERITONEAL MACROPHAGES IN VITRO AFTER UPTAKE OF HETEROLOGOUS ERYTHROCYTES

Mouse peritoneal macrophages have been studied in vitro after ingestion of treated rat, rabbit, or sheep erythrocytes. Under light microscopy, phagocytic vacuoles persist up to 24 h. Macrophages lose benzidine reactivity about 5 h after red cell ingestion, and they become prussian blue positive at 2 days. Ultrastructural studies show little or no ferritin in control macrophages not fed erythrocytes. In contrast, after red cell ingestion, ferritin is widely distributed in the cytoplasmic matrix and in some cytoplasmic granules by 48 h. The Golgi complex, pinocytic vacuoles, endoplasmic reticulum, nuclei, and mitochondria do not contain ferritin. Between 2 and 4 days, ferritin in cytoplasmic granules increases, concomitant with decrease in the ferritin in the cytoplasmic matrix. Evidence is presented suggesting that ferritin in the cytoplasmic matrix is translocated into cytoplasmic granules by autophagy. Polyacrylamide gel studies on macrophages after uptake of red blood cells labeled with radioiron confirm that macrophages produce radiolabeled ferritin by 4 days.     

Histochemical and Morphological Characteristics of Canine Cardiac Mast Cells

Cardiac mast cells have been recently isolated and characterized in humans, however canine cardiac mast cells have not been investigated. The objective of this study is to describe the histological and morphological characteristics of canine cardiac mast cells and examine the potential usefulness of canine models in investigating the role of mast cells in cardiovascular pathology. Canine cardiac mast cells could be easily identified by staining with Toluidine Blue or FITC-avidin. Using Toluidine Blue staining, we demonstrated fewer mast cells in formalin-fixed samples than in specimens fixed in Carnoy's, thus identifying a formalin-sensitive mast cell population in the canine heart. Mast cells were equally distributed in atria and ventricles with approximately 50% showing a perivascular location. Using enzyme-histochemical techniques, we detected tryptase and chymase activity in canine cardiac mast cells. Ultrastructural studies identified mast cells as granular cells with an eccentric non-segmented nucleus. Immunohistochemistry with the macrophage specific antibody AM-3K demonstrated that resident cardiac macrophages were 1.9 times more numerous than mast cells, also showing a predominantly perivascular (60%) location. Perivascular macrophages were more often periarteriolar, whereas perivascular mast cells were more often located along small veins and capillaries. Due to their ability to release cytokines and growth factors and their strategic perivascular location, resident cardiac inflammatory cells, such as mast cells and macrophages, may be important in pathological processes causing myocardial inflammation and fibrosis. Furthermore, mast cell-derived chymase, an important angiotensin II-forming enzyme may have a significant role in regulating the cardiac renin-angiotensin system.     

Electron microscopy of Listeria monocytogenes-infected mouse spleen

Armstrong, B. A. (The University of Kansas, Lawrence), and C. P. Sword. Electron microscopy of Listeria monocytogenes-infected mouse spleen. J. Bacteriol. 91:1346-1355. 1966.-Mouse spleen infected with Listeria monocytogenes was observed during the acute phase of infection; 72 hr after infection, organisms were usually found within phagocytic vacuoles in the cytoplasm of macrophages. These vacuoles, which resembled phagosomes, often contained several organisms as well as varying amounts of amorphous electron-dense material, ferritin-like particles, membrane fragments, and vesicles of varying density. Breakdown of vacuolar membranes appeared to be accompanied by damage to the host cell cytoplasm. Nuclear membrane damage was occasionally observed when phagocytic vacuoles were close to the nucleus.     

ULTRASTRUCTURAL STUDIES PLASMA MEMBRANE RELATED SECONDARY VACUOLES IN CULTURED CELLS

The plasma membrane of cultured cells of several plant species was observed to possess invaginations, or secondary vacuoles, of variable size in the adjacent cytoplasm. These structures, which occurred in cells at different phases in vacuolation, were very numerous in thin sections of some cells but fewer in others. In vacuolated cells enlarged secondary vacuoles protrude into the primary vacuole but are delimited from the tonoplast by an intermembrane zone of variable width. The plasma membrane at the orifice of an invagination may fuse and detach the secondary vacuole from the membrane to form in the cytoplasm a structure bounded by a single membrane. Complex accumulations of membranes consisting of spherical, tubular, and laminar structures, possibly containing cytoplasm, may develop within secondary vacuoles. Contents of many of these vacuoles arise from folds along its limiting membrane which pinch off into the interior of the secondary vacuole. A fibrous substance, possibly derived from the wall, is present in some secondary vacuoles. Observed folding of the plasma membrane and measurements of membrane width of various organelles and cytomembranes support an interpretation that endocytosis occurs in cultured cells.     

Phagocytic and motile properties of endothelial cells measured magnetometrically: effects of endotoxin

Endothelial cells lining the vasculature share some properties with macrophages and neutrophils in that they can take up material from the blood and are known to migrate, particularly during wound healing. We observed that endothelial cells isolated from bovine pulmonary arteries ingested magnetic iron oxide particles during culture in vitro. Using a non-optical, magnetometric method, we examined motions of magnetic-particle containing intracellular vacuoles. We demonstrated that these organelles move within endothelial cells, but at a slower rate than phagosomes within macrophages. Magnetometry was used to show that incubation with endotoxin (10 micrograms/ml) for 4 hr resulted in a decrease in cytoplasmic movement; yet the fluidity of the cytoplasm was increased, as measured by intracellular particle response to forced motion. We conclude that intracellular magnetic probe particles can detect vesicular motion in endothelial cells, and that endotoxin exposure can affect endothelial cells directly, altering their physical properties; these alterations precede ultrastructural evidence of cell death.