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S J Galli

Publications and source records attributed to S J Galli.

At least 163 records · Page 9Linked to original sources
PubMed1987

Neovascularization and coronary atherosclerotic plaque: cinematographic localization and quantitative histologic analysis.

A new technique was developed for analyzing the neovascularization associated with coronary artery atherosclerosis: cinematography during silicone polymer injection of the coronary arteries of fixed and cleared human hearts, followed by histologic analysis in routine and 1-micron-thick, Epon-embedded sections. Twenty-two hearts obtained at autopsy were studied. On the basis of cinematographic findings, individual regions of the coronary arteries were classified as negative, positive, or abundantly positive for neovascularization. Positive and abundantly positive areas, which invariably occurred in segments exhibiting changes of atherosclerosis, contained numerous small vessels in the adventitia and outer media (4.7 +/- 1.5 and 9.8 +/- 1.3 [SE] vessel profiles/artery cross-section in positive and abundantly positive areas, versus 1.0 +/- 0.6 in negative regions). Abundantly positive areas, which occurred in coronary artery segments demonstrating the most extensive atherosclerotic change, contained numerous small vessels in the inner media or in the plaque itself. Some of these microvessels were in close proximity to mast cells, which represent potentially rich sources of mediators affecting vascular tone and permeability. Vessels were not observed in the inner media or in atherosclerotic plaque in areas designated either positive or negative by cinematography. These findings show how our approach can be used both to define the three-dimensional, in situ configuration of coronary artery neovascularization and to characterize the histology of this process in detail. They also confirm previous work indicating that areas of coronary arteries involved by atherosclerosis frequently exhibit extensive neovascularization.

Adult↗
PubMed1987

Identification, purification, and characterization of a mast cell-associated cytolytic factor related to tumor necrosis factor.

The role of mast cells and mast-cell-derived factors in natural cytotoxic reactions was investigated. Cultured and freshly isolated murine mast cells are shown to be cytotoxic to WEHI-164 and YAC-1 targets in 18-hr viability assays but not in 4-hr assays. Here, we describe a cytotoxic factor in murine mast cells that is immunologically related to tumor necrosis factor (TNF). This TNF-like factor lyses WEHI-164 cells with a slow time course requiring 16-20 hr for the lytic reaction to complete. Antibodies specific for human and murine TNF and human lymphotoxin partially block mast cell lysis of WEHI-164 cells. These antibodies react on immunoblots with one major mast cell protein band of 50 kDa. Immunoblot analysis shows this factor in cloned and uncloned cultured mouse mast cells and in mature "connective tissue-type" mast cells freshly purified from rat or mouse peritoneal cavities. The amount of this factor is greatly enhanced in cells that have been stimulated with a combination of phorbol ester/concanavalin A or bacterial lipopolysaccharide. Subcellular fractionation analysis of mast cells with Percoll gradients reveals two pools of TNF-related cytotoxic activity that are associated with free cytosolic material and granule fractions. In contrast to cytotoxic T lymphocytes and natural killer cells, granule-enriched fractions of mast cells do not contain any hemolytic activity. The localization of the TNF-like molecule in mast cell granules may play a strategical role in the rapid delivery of this mediator to the target cell membrane following cell surface stimulation and degranulation.

Animals↗
PubMed1987

Beige mouse mast cells generated in vitro: ultrastructural analysis of maturation induced by sodium butyrate and of IgE-mediated, antigen-dependent degranulation.

We derived growth factor-dependent mast cell lines from C57BL/6-bgJ/bgJ ('beige') mouse bone marrow cells using techniques previously described for deriving mast cell lines from normal mice. According to examination by transmission electron microscopy, the cytoplasmic granules of cultured mast cells derived from beige mice were larger in size and fewer in number than those in cultured mast cells derived from normal C57BL/6 mice. Mast cells derived from beige mice underwent maturation when exposed to sodium butyrate, as judged by increased content of electron-dense material in the cytoplasmic granules. Cultured mast cells derived from beige mice also underwent IgE-mediated, antigen-dependent anaphylactic degranulation which was similar in its ultrastructural features to that described for cultured mast cells from normal mice. However, in mast cells not stimulated with IgE and antigen, fusion between individual cytoplasmic granules was observed more commonly in mast cells derived from beige mice than in normal mast cells. This might mean that these events are more common in C57BL/6-bgJ/bgJ mast cells, and/or that their resolution is slower and thus more easily captured by electron microscopy.

Animals↗
PubMed1987

Analysis of mast cell function in biological responses not involving IgE.

Mast cells are critical for the expression of certain IgE-mediated responses, but the precise contributions of mast cells to biological processes not involving IgE are obscure. We have employed genetically mast cell-deficient WBB6F1-W/Wv and WCB6F1-S1/S1d mice to investigate the roles of mast cells in several different biological responses. This work strongly suggests that mast cells are not required for the elicitation of contact sensitivity (CS) responses, suppressor T cell-dependent tolerance to CS, reserpine-induced inhibition of CS responses, or bleomycin-induced pulmonary fibrosis. By contrast, mast cells appear to contribute to the acute gastric injury induced by ethanol and the acute inflammation of the skin induced by croton oil.

Animals↗
PubMed1987

Ultrastructural cytochemical and autoradiographic demonstration of nonspecific esterase(s) in guinea pig basophils.

We used ultrastructural autoradiographic and cytochemical methods to localize esterase activities in unstimulated guinea pig basophils and in basophils undergoing degranulation or recovery from degranulation. We used tritium-labeled diisopropylfluorophosphate (DFP) as a probe for serine enzymes and localized this probe by ultrastructural autoradiography to cytoplasmic granules of immature or mature unstimulated basophils, as well as to granules released by degranulating basophils. Ultrastructural cytochemistry using alpha naphthyl acetate (ANA) as substrate localized nonspecific esterase activity to extruded granules, either within the interiors of degranulation sacs or within granules completely separated from degranulating basophils. Extruded granules retained their esterase activity for as long as 24 hr after antigen-induced degranulation. The plasma membranes of unstimulated or degranulating basophils, as well as of basophils recovering from degranulation, displayed prominent cell surface ANA esterase ectoenzyme activity. Lipid bodies, organelles present in the cytoplasm of both control and recovering basophils, were also alpha naphthyl acetate esterase (ANAE)-positive. Thus, cytochemical and autoradiographic techniques localized esterase and/or [3H]-DFP-binding activities to cytoplasmic granules, lipid bodies, and cell surface of basophils, and these enzyme activities persisted during both degranulation and recovery from degranulation.

Animals↗
PubMed1987

Nonspecific esterase activity expressed in Weibel-Palade bodies of cloned guinea pig aortic endothelial cells.

We studied the localization of nonspecific esterase activities in cloned guinea pig aortic endothelial cells using ultrastructural cytochemistry. Weibel-Palade bodies (WPB), which are known to contain von Willebrand protein, were positive for esterase, defining a heretofore unrecognized activity of these organelles. Esterase activity was also found localized to the external surface of the plasma membrane, to cytoplasmic lipid bodies, and to the outer (cytoplasm-facing) surface of certain membrane-bound cytoplasmic vacuoles. Localization of esterase activity to these four discrete sites probably reflects the presence of a number of endothelial cell enzymes capable of hydrolyzing alpha-naphthyl acetate or butyrate. The physiological substrate and biological function of these enzyme activities are not presently understood.

Animals↗
PubMed1987

Rapid microwave fixation of rat mast cells. I. Localization of granule chymase with an ultrastructural postembedding immunogold technique.

We defined the ultrastructural localization of chymase in rat peritoneal mast cells using standard aldehyde fixation and a newly described microwave fixation method (Login GR, Dvorak AM: Microwave energy fixation for electron microscopy. Am J Pathol 120: 230, 1985; Login GR, Stavinoha WB, Dvorak AM: Ultrafast microwave energy fixation for electron microscopy. J Histochem Cytochem 34:381, 1986) and postembedding immunogold labeling. Thin sections were exposed first to goat IgG anti-rat chymase and second to gold-conjugated rabbit Ig directed against goat IgG. By transmission electron microscopy, gold particles were localized to the matrix of cytoplasmic granules. Control sections treated with nonimmune sera did not exhibit labeling of mast cells. Thin sections treated simultaneously with purified rat mast cell chymase and anti-chymase antibody in competition studies, showed a marked reduction in granule staining. These findings demonstrate that a microwave fixation method can be used to rapidly fix cell suspensions for postembedding immunocytochemical studies.

Animals↗
PubMed1987

New approaches for the analysis of mast cell maturation, heterogeneity, and function.

Mast cells are regarded as potentially critical participants in a wide variety of important biological processes. Yet it has been difficult to ascertain the precise contribution of mast cells to many of these reactions. In part, this reflects recognition that different populations of mast cells may vary in phenotype and therefore may express different functions. Another problem has been the lack of suitable model systems for identifying and quantitating the mast cell's unique roles in biological processes in intact animals. This review will outline the development of promising new approaches for analyzing mouse mast cell differentiation and phenotypic heterogeneity and for studying the roles of mast cells in vivo.

Animals↗
PubMed1987

Development of large numbers of mast cells at sites of idiopathic chronic dermatitis in genetically mast cell-deficient WBB6F1-W/Wv mice.

The normal skin and other tissues of adult mast cell-deficient WBB6F1-W/Wv or WCB6F1-Sl/Sld mice contain less than 1.0% the number of mast cells present in the corresponding tissues of the congenic normal (+/+) mice. As a result, genetically mast cell-deficient WBB6F1-W/Wv or WCB6F1-Sl/Sld mice are widely used for studies of mast cell differentiation and function. We found that mast cells developed at sites of idiopathic chronic dermatitis in WBB6F1-W/Wv mice and that the number of mast cells present in the skin of WBB6F1-W/Wv mice was proportional to the severity of the dermatitis (in ear skin, there were 33 +/- 4 mast cells/mm2 of dermis at sites of severe dermatitis v 9 +/- 3 at sites of mild dermatitis, 0.8 +/- 0.3 in skin without dermatitis, and 100 +/- 7 in the normal skin of congenic WBB6F1-+/+ mice; in back skin, the corresponding values were 2.0 +/- 0.6, 1.1 +/- 0.9, 0.025 +/- 0.025, and 26.2 +/- 3.2). The development of mast cells was a local, not systemic, consequence of the dermatitis. Thus, WBB6F1-W/Wv mice with severe dermatitis lacked mast cells in skin not showing signs of dermatitis and also in the peritoneal cavity, stomach, cecum, and tongue. Idiopathic chronic dermatitis was not associated with the local development of mast cells in WCB6F1-Sl/Sld mice, a mutant whose mast cell deficiency is due to a mechanism distinct from that of WBB6F1-W/Wv mice. These findings may have implications for understanding the nature of the mast cell deficiency in WBB6F1-W/Wv and WCB6F1-Sl/Sld mice and for the use of these mutants to analyze mast cell differentiation and function.

Animals↗
PubMed1987

Ethanol-induced acute gastric injury in mast cell-deficient and congenic normal mice. Evidence that mast cells can augment the area of damage.

The authors used stereomicroscopy and planimetry to measure the area of glandular stomach mucosa acutely injured by oral ethanol in mast cell-deficient and congenic normal (+/+) mice, and examined the damaged areas in 1-mu sections. Ethanol caused degranulation and/or disruption of gastric mucosal mast cells, and, at certain concentrations of ethanol, mast cell-deficient WBB6F1-W/Wv or WCB6F1-Sl/Sld mice developed significantly less (43-90% less) acute gastric injury than either congenic +/+ mice or WBB6F1-W/Wv mice whose mast cells were restored by bone marrow transplantation from WBB6F1-+/+ mice. Nevertheless, ethanol produced detectable, and in some cases substantial, gastric injury even in the complete absence of mast cells. Thus, ethanol can produce some damage to the gastric mucosa independently of mast cells. But these data suggest that under certain circumstances mast cells can augment the area of acute gastric injury induced by ethanol.

Alcoholic Intoxication↗
PubMed1987

Defective cytoplasmic granule formation. I. Abnormalities affecting tissue mast cells and pancreatic acinar cells of beige mice.

Beige mice (C57BL/6-bgJ/bgJ) express the Chediak-Higashi syndrome, a genetically determined constellation of morphologic and functional abnormalities affecting cells that synthesize cytoplasmic granules; a similar disorder also occurs in humans and several other mammalian species. We used a computer-assisted morphometric approach to identify and quantitate the effect of the beige mutation on the structure of mast cell or pancreatic acinar cell cytoplasmic granules. Beige and control mouse mast cell or pancreatic acinar cell granules exhibited periodic, multimodal distributions of equivalent volumes in which the modes fell at volumes that were integral multiples of the volume of the "unit granule," whose volume (the "unit volume" or v1) was defined by the first mode in the granule equivalent volume distribution. But the modal frequency of the C57BL/6-bgJ/bgJ mast cell granule equivalent volume distribution fell at v1, a pattern consistent with a haphazard pattern of "unit granule" fusion, whereas the corresponding modal frequency for the control mast cell granules fell at v3, a pattern consistent with a "unit addition" model of granule fusion. In addition, the unit volume of beige mouse mast cell granules was 18 times that of control mouse mast cell granules. By contrast, the unit volume of beige mouse pancreatic acinar cell granules was only slightly (23%) greater than that of control cells. C57BL/6-bgJ/bgJ and control cells did not differ significantly in total cell or nuclear volume, or in the aggregate volume of their cytoplasmic granules. However, C57BL/6-bgJ/bgJ mast cells or pancreatic acinar cells contained significantly fewer granules than did their normal counterparts. These findings are consistent with the hypothesis that the beige mutation affects the formation of unit granules and also alters the pattern of aggregation and fusion of unit granules. The data also identify quantitative differences in the expression of the beige mutation in mast cells and pancreatic acinar cells.

Animals↗
PubMed1987

Antigen-induced, IgE-mediated degranulation of cloned immature mast cells derived from normal mice.

Cloned, immature mast cells derived from normal mice were passively sensitized with mouse monoclonal IgE antibodies with specificity for DNP, and then stimulated to degranulate with DNP35-HSA. Cells were fixed for transmission electron microscopy or recovered for quantitation of histamine release at various intervals up to 30 minutes after antigen challenge. The cloned mast cells rapidly extruded the contents of their immature granules (dense progranular material and membrane-bound vesicles) to the exterior via multiple openings in the plasma membrane. Degranulation was associated with striking activation of the cell surface, characterized initially by elongation of surface processes, as well as by close approximation of strands of rough endoplasmic reticulum to the cell surface and by the development of coated pits. At later times after stimulation, degranulated mast cells had released nearly all of their granules and exhibited angular surfaces lacking elongated processes. These findings demonstrate for the first time that cloned, immature mast cells, like their mature counterparts, can undergo classic morphologic release reactions involving exocytosis of granules.

Animals↗
PubMed1986

125I-fibrin deposition in contact sensitivity reactions in the mouse. Sensitivity of the assay for quantitating reactions after active or passive sensitization.

We investigated the clotting associated with delayed hypersensitivity (DH) responses in the mouse by sensitizing the animals to the contactant oxazolone (Ox), and then administering 125I-guinea pig fibrinogen i.v. 10 to 30 min before antigen challenge 5 days later. Early (4 to 8 hr) contact sensitivity (CS) responses in immunized mice were barely detectable by three conventional measures of CS, but the total 125I-cpm in ears challenged with hapten was 3.6 to 4.5 X that in control ears challenged with vehicle alone; moreover, the amount of urea-insoluble cpm (cross-linked 125I-fibrin-associated cpm) in the reactions to Ox was 6.5-fold to 8.2-fold that present in the control reactions. In 24 hr reactions that were near peak intensity by measurements of ear swelling, ear weight ratios, and ratios of 125I-5-iodo-2-deoxyuridine-labeled leukocyte infiltration, the cpm in antigen-challenged ears exceeded that in control ears by 13-fold to 53-fold. In addition, antigen-challenged ears contained 27 to 300 X the urea-insoluble cpm present in control ears. 125I-Fibrin deposition was not a specific characteristic of CS reactions, because a small amount of urea-insoluble reactivity was also detected in some reactions to Ox in native mice. Nevertheless, the assay was exquisitely sensitive and readily detected quantitative differences between the immunologically specific and nonspecific reactions at very early intervals after challenge or with suboptimal doses of antigen. Furthermore, it was more sensitive than conventional tests of CS in detecting the reactions elicited in mice that had been passively sensitized to Ox by adoptive transfer of immune lymph node cells. Finally, we showed that the assay gave similar results when we tested CS reactions elicited in mast cell deficient WBB6F1-W/Wv and littermate normal (+/+) mice, demonstrating yet another similarity in the phenotype of DH reactions elicited in the presence or absence of mast cells.

Animals↗
PubMed1986

Cellular and vascular manifestations of cell-mediated immunity.

The studies reviewed here emphasize both the complexity and the heterogeneity of cell-mediated immunity. In addition to the round cell infiltrate of the classic descriptions, cell-mediated immunity includes reactions that feature many types of inflammatory cells, that exert profound effects on the blood microvasculature, and that initiate extravascular clotting and, possibly, angiogenesis. The common denominator of all of these reactions is a subset or subsets of sensitized T lymphocytes that, on exposure to specific antigen, recruit and collaborate in other ways with one or more populations of circulating bone marrow-derived cells. Although the reactions generally resemble chronic inflammation by virtue of the lymphocytes and monocytes present, cell-mediated immunity may also take the guise of acute or subacute inflammation when neutrophils or eosinophils predominate and an entirely different morphologic pattern when basophils predominate, as in CBH. Tissue mast cells undergo changes (activation, proliferation) that are generally observed at later stages of delayed hypersensitivity, but no convincing evidence has been presented that these cells play an essential role in the elicitation of cell-mediated immunity. The concept that an essential prerequisite for the elicitation of delayed hypersensitivity is the mast cell-dependent generation of microvascular gaps, favoring inflammatory cell diapedesis, is clearly incorrect. First, lymphocytes fail to traverse certain of the vessels that exhibit such gaps (i.e., those of the SCV) in delayed hypersensitivity reactions in humans. Second, there is no diminution in the cellular infiltration associated with cell-mediated immunity reactions in mast cell-deficient mice. Cell-mediated immunity does not consist of an inflammatory cell infiltrate alone. The local microvasculature is rendered hyperpermeable to varying extents, with resulting extravasation of plasma proteins, including fibrinogen. The majority of extravasated fibrinogen is clotted to cross-linked fibrin, presumably as the result of the actions of procoagulants associated with fixed connective tissue cells and perhaps also because of the activity of infiltrating cells, such as monocytes/macrophages. Clotted fibrin forms a water-trapping gel, which accounts for the induration seen in many delayed hypersensitivity reactions. The microvasculature may also be affected in other ways. Endothelial cells may undergo hypertrophy and cell division or, alternatively, may exhibit profound and progressive injury.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1986

Anticoagulantly active heparin-like molecules from mast cell-deficient mice.

To assess the contribution of mast cells to the maintenance of blood fluidity, the hindlimb vasculature of mast cell-deficient mice (W/Wv) and littermates containing normal levels of mast cells (+/+), were perfused with purified human thrombin and antithrombin. Enzyme-inhibitor complex generation within the vasculature was enhanced to a comparable extent for W/Wv and +/+ mice over the uncatalyzed rate, that level of complex produced within a similar time interval in the absence of heparin. Perfusion of purified Flavobacterium heparinase prior to infusion of the hemostatic components, or perfusion of antithrombin modified at the heparin-binding domain, reduced W/Wv and +/+ hindlimb thrombin-antithrombin complex formation to the uncatalyzed rate. To further define the cellular source of the vascular-associated heparin-like molecules, endothelial cells isolated from epididymal fat pads of W/Wv and +/+ mice were grown in vitro. The acceleration of thrombin-antithrombin interactions in the presence of endothelial cell-derived glycosaminoglycans was similar for W/Wv and +/+ mice, was abolished with purified bacterial heparinase, and was expressed to only a minor extent when utilizing modified antithrombin. The biologically active mucopolysaccharides appear to be present on the cell surface.

Adipose Tissue↗
PubMed1986

Inbred guinea pig aortic endothelial cell clones. Model for studying the vascular endothelium under totally isologous conditions.

Aortic endothelial cell clones were established from adult inbred strain 2 guinea pigs by using a simple method requiring neither feeder layers nor growth factors other than isologous serum. One clone has been maintained in continuous culture for 26 passages over a 9-month period. Cloned endothelial cells grew in a cobblestone pattern at confluence, formed intercellular junctions, and expressed factor-VIII-related antigen by immunofluorescence microscopy. The cloned cells also contained high levels of angiotensin-converting enzyme activity, although the level of enzyme activity expressed by different cloned cell lines varied. Cloned endothelial cells contained numerous Weibel-Palade bodies which appeared larger and less electron-dense than those of strain 2 guinea pig aortic endothelial cells in vivo. These clones may serve as an in vitro model to investigate the formation and function of this organelle. Cloned populations of strain 2 guinea pig vascular endothelial cells maintained in medium supplemented only with strain 2 guinea pig serum also provide an immunologically isologous system ideal for investigating interactions between the vascular endothelium and immunocompetent leukocytes in vitro.

Animals↗
PubMed1985

Reevaluation of reserpine-induced suppression of contact sensitivity. Evidence that reserpine interferes with T lymphocyte function independently of an effect on mast cells.

It has been suggested that reserpine blocks expression of delayed hypersensitivity (DH) by depleting tissue mast cells of serotonin (5-HT), thereby preventing a T cell-dependent release of mast cell 5-HT necessary to localize and to amplify the DH response. However, reserpine blocks expression of DH in mast cell-deficient mice. We therefore decided to reevaluate the mechanism by which reserpine abrogates expression of cellular immunity, and investigated whether the drug might interfere with T cell activity in vitro or in vivo. At concentrations as low as 4 microM, reserpine profoundly suppressed baseline or antigen-augmented levels of [3H]thymidine incorporation by immune lymph node cells obtained from mice sensitized to the contactant oxazolone [I-LNC(Ox)]. This effect was observed both with I-LNC derived from normal mice and with I-LNC derived from congenitally mast cell-deficient W/Wv mice, cell preparations that lacked detectable mast cells, histamine, and 5-HT. Furthermore, treatment of I-LNC with reserpine (20 microM) for 1 h in vitro virtually abolished the ability of these cells to transfer CS to naive mice. This was not a cytolytic effect, as the viability of the I-LNC treated with reserpine was not affected, and washing of the reserpine-treated I-LNC before transfer fully restored their ability to orchestrate a CS response. The action of the drug was not mediated by an effect on mast cells, since the experiment could be performed using mast cell-deficient W/Wv mice as both donors and recipients of I-LNC. In addition, the effect was specific for the treated cells: mice that received reserpine-treated I-LNC(Ox) intravenously together with untreated I-LNC(DNFB) did not develop CS to Ox but responded normally to DNFB; and local intradermal injection of reserpine-treated I-LNC(Ox) which failed to transfer reactivity to Ox, did not interfere with the development of CS to DNFB at the same site. Finally, cotransfer experiments indicated that the effect of reserpine on the transfer of CS was not due to activation of suppressor cells. Our findings strongly suggest that whatever effects reserpine might have on immunologically nonspecific host cells, the drug's effects on sensitized T cells are sufficient to explain its ability to block cell-mediated immune responses in vivo.

Animals↗