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Biomedical subjects

D J Silverman

Publications and source records attributed to D J Silverman.

At least 37 records · Page 2Linked to original sources
PubMed1992

Penetration of host cells by Rickettsia rickettsii appears to be mediated by a phospholipase of rickettsial origin.

Internalization of obligate intracellular bacteria belonging to the genus Rickettsia by eukaryotic cells requires participation of both the parasitized host and the microorganism. The term "induced phagocytosis" has been used specifically to describe the entry of Rickettsia prowazekii, although a similar mechanism is likely for R. rickettsii. A role for a phospholipase in the internalization process has been proposed for both of these organisms, with the strongest supporting evidence provided for R. prowazekii. Despite general acceptance of the notion that phospholipase activity is involved in the internalization process of these bacteria, the origin of the enzyme is not known. The results of the study presented here, which used R. rickettsii and Vero cells, suggest that a rickettsial phospholipase, rather than a host cell phospholipase, mediates internalization of the organism. This conclusion is based upon results which show that pretreatment of R. rickettsii, but not of host cells, with a specific chemical inhibitor of phospholipase, and also antiserum to this enzyme, significantly reduces uptake of the organism and its ability to cause plaque formation.

Acetophenones↗
PubMed1991

Rickettsia rickettsii infection of cultured endothelial cells induces release of large von Willebrand factor multimers from Weibel-Palade bodies.

The clinical manifestations of Rocky Mountain spotted fever (RMSF) result from Rickettsia rickettsii (R rickettsii) infection of endothelial cells and are mediated by pathologic changes localized to the vessel, including in situ thrombosis and tissue ischemia. This study uses in vitro infection of cultured human umbilical vein endothelial cells with R rickettsii to test the hypothesis that such infection induces von Willebrand factor (vWF) release from Weibel-Palade bodies, a process that could contribute to thrombotic changes. At 24 hours postinfection, there was an increase in metabolically prelabeled large multimers of vWF in the culture medium, with a concomitant decrease of these forms in the cell lysate samples. This release reaction was specific for the large multimer pool of vWF, localized to Weibel-Palade bodies, because no change in the distribution of dimeric forms between cells and culture medium was detected. Double-label immunofluorescence staining showed an inverse correlation between the number of R rickettsii and the number of Weibel-Palade bodies in infected cells. Cell lysis was minimal at 24 hours postinfection, as no detectable intracellular precursor forms (molecular weight 260,000) of vWF were released into the culture medium, there was no decrease in cell viability as measured by trypan blue exclusion, and no increase in 51Cr-release into the culture medium was observed when compared with uninfected controls. Release was likely a direct effect of the intracellular presence of the organism, rather than due to a noxious soluble factor such as endotoxin, because culture medium conditioned by infected endothelial cells was ineffective at inducing release in uninfected endothelial cell cultures. In summary, in vitro infection of endothelial cells by R rickettsii induces release of Weibel-Palade body contents, a process that may contribute to the pathogenesis of RMSF.

Cell Survival↗
PubMed1991

Some contributions of electron microscopy to the study of the rickettsiae.

Electron microscopy has provided valuable insights into the study of rickettsiae as intracellular parasites from several important perspectives. This tool has allowed researchers to delineate the fine structural features of these organisms and to show that they truly resemble free-living bacteria. Furthermore, it has been shown that there are subtle, but distinct differences in the outer envelope structure of some members of the genus Rickettsia that may explain reported differences in tinctorial properties and in their sensitivity to certain antibiotics. With Coxiella burnetii, electron microscopy has helped significantly in the characterization of the pleomorphic nature of the organism including formation of terminal bodies that resemble endospores of gram-positive bacteria. Electron microsxopy has also helped to define the relationship of the rickettsiae to their host cells. For example, ultrastructural analysis can reveal whether organisms exist free within the cytoplasm or nucleus (members of the genus Rickettsia), or whether they are bound by a phagosomal or phagolysosomal membrane (Ehrlichia and Coxiella). Finally, although all rickettsiae eventually destroy their host cell, it has been shown through transmission electron microscopy that this destruction might be mediated by different mechanisms that are specific for different rickettsial species.

Animals↗
PubMed1991

Heparin protects human endothelial cells infected by Rickettsia rickettsii.

Routine culture of endothelial cells currently includes the use of heparin, which significantly reduces cell doubling time and increases cell population size. Heparin protects cultured arterial endothelial cells from damage by toxic oxygen metabolites produced by the action of xanthine and xanthine oxidase. Because of our hypothesis implicating free radicals in cell injury caused by Rickettsia rickettsii, we have carried out a series of experiments to examine the effects of heparin on injury to endothelial cells infected by this microorganism. These studies showed that heparin does not inhibit replication of R. rickettsii in the cytoplasm of endothelial cells. Furthermore, heparin appears to exhibit a protective effect on the infected host cell as measured by (i) reduced plaque size, (ii) increased longevity of the cell monolayer, (iii) reduction in the amount of lactic dehydrogenase released from infected cells, and (iv) reduction in the levels of intracellular peroxides formed in infected cells. Electron microscopic studies also show a significant reduction in dilatation of the rough-surfaced endoplasmic reticulum of the infected cells in the presence of heparin. These observations appear to lend additional support to involvement of an oxidative mechanism in human endothelial cell injury caused by R. rickettsii.

Antioxidants↗
PubMed1990

Experimental infection of human vascular endothelial cells by pathogenic and nonpathogenic hantaviruses.

To investigate whether the hantaviruses replicate in endothelial cells, we inoculated human umbilical vein endothelial cells with several pathogenic and nonpathogenic strains of hantavirus. Intracytoplasmic, virus-specific granular fluorescence was detected initially at three days postinoculation and nearly 100% of cells contained viral antigen at 10 days. Cytopathic effect or inclusion bodies were not observed. The in vitro demonstration of the susceptibility of endothelial cells to hantaviruses corroborates in vivo findings, and suggests that endothelial cells may serve as target cells in hemorrhagic fever with renal syndrome.

Animals↗
PubMed1990

A non-pharmacological treatment of vascular headache during pregnancy.

This study investigates the use of biofeedback, relaxation and psychotherapy on five patients with severe, vascular headaches that occurred during the course of pregnancy. The subjects received between four and twelve sessions of treatment overall. The subjects all showed a marked reduction or complete cessation of headaches during treatment, the term of pregnancy, and during a follow-up evaluation months after the birth of the child. Possible alternate explanations for improvement are discussed along with the study's limitations. This preliminary investigation strongly suggests that psychological treatment may be a particularly useful intervention for management of headaches that occur in pregnant women.

Adult↗
PubMed1988

Potential for free radical-induced lipid peroxidation as a cause of endothelial cell injury in Rocky Mountain spotted fever.

Cells infected by Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever, display unusual intracellular morphological changes characterized by dilatation of the membranes of the endoplasmic reticulum and outer nuclear envelope. These changes are consistent with those that might be expected to occur following peroxidation of membrane lipids initiated by oxygen radical species, such as the hydroxyl radical or a variety of organic radicals. Using a fluorescent probe, we have found significantly increased levels of peroxides in human endothelial cells infected by R. rickettsii. Studies with desferrioxamine, an iron chelator effective in preventing formation of the hydroxyl radical from hydrogen peroxide and the superoxide free radical, reduced peroxide levels in infected cells to those found in uninfected cells. This observation suggests that the increased peroxides in infected cells may be lipid peroxides, degradation products of free radical attack on polyenoic fatty acids. The potential for lipid peroxidation as an important mechanism in endothelial cell injury caused by R. rickettsii is discussed.

Deferoxamine↗
PubMed1986

Associative recall properties of the trion model of cortical organization.

We developed a cooperative model of the cortical column incorporating an idealized subunit, the trion (which represents a localized group of neurons), and a discrete time step for firing. We found that networks composed of a small number of trions (with symmetric interactions) supported up to thousands of quasi-stable, periodic firing patterns (MPs) which could be selected out with only small changes in interaction strengths using a Hebb-type algorithm. Here we report a study of the associative recall properties showing striking features: By considering all possible initial firing patterns (for a given set of network connections), we find 1) It takes on the average only 2-5 time steps to recall an MP. 2) Many of the MPs can be individually accessed by thousands of different initial patterns. The variety of examples presented illustrate the rich, general nature of the model.

Action Potentials↗
PubMed1986

Adherence of platelets to human endothelial cells infected by Rickettsia rickettsii.

Rickettsia rickettsii, the etiologic agent of Rocky Mountain spotted fever, causes widespread vasculitis due to extensive injury of endothelial cells lining small blood vessels. Injury to these cells can lead to denudation of the endothelial lining and exposure of the thrombogenic subendothelium. Modification of the cell surface, either through minor trauma or as a result of altered cell surface biochemistry, could induce changes reflected in increased platelet reactivity. Because membrane modification is likely when R. rickettsii enters endothelial cells, the possibility that infection by this organism caused increased reactivity of platelets with the endothelial cell surface was examined. A fourfold increase in adherence of platelets to cultured endothelial cells from the human umbilical vein infected by R. rickettsii compared with uninfected cells was found. These studies suggest that the adherence of platelets to the surface of Rickettsia-infected endothelial cells can contribute to reduction in the number of circulating platelets in blood during human infection.

Blood Platelets↗
PubMed1985

Model of cortical organization embodying a basis for a theory of information processing and memory recall.

Motivated by V. B. Mountcastle's organizational principle for neocortical function, and by M. E. Fisher's model of physical spin systems, we introduce a cooperative model of the cortical column incorporating an idealized substructure, the trion, which represents a localized group of neurons. Computer studies reveal that typical networks composed of a small number of trions (with symmetric interactions) exhibit striking behavior--e.g., hundreds to thousands of quasistable, periodic firing patterns, any of which can be selected out and enhanced with only small changes in interaction strengths by using a Hebb-type algorithm.

Animals↗
PubMed1984

Infection of human vascular endothelial cells by Rickettsia rickettsii.

Rocky Mountain spotted fever is caused by Rickettsia rickettsii, an obligate intracellular bacterial parasite. The organism primarily attacks endothelial cells and occasionally attacks smooth-muscle cells of small blood vessels. An effective means of examining host-parasite interaction in Rocky Mountain spotted fever would be to use an in vitro model system with a host-cell type that is similar in structure and function to the putative target cell in human infections. Because human umbilical-vein endothelial cells in culture retain many, if not all, of their characteristic properties in vivo and because they also share many properties of capillary endothelium, the use of this endothelial cell system is appropriate in the study of the interaction between R rickettsii and the cell that is principally parasitized in humans. Uptake by umbilical-vein endothelial-cell cultures of R rickettsii is dose dependent. The organism replicates in both the nucleus and cytoplasm of infected cells and exhibits early cell-to-cell spread without detectable host-cell injury.

Cells, Cultured↗
PubMed1984

Rickettsia rickettsii-induced cellular injury of human vascular endothelium in vitro.

The endothelial cell is the putative primary target cell in humans infected with Rickettsia rickettsii, the etiological agent of Rocky Mountain spotted fever. Although the clinical manifestations of infection by this organism are well documented, the mechanism of injury to the endothelial cell is not understood. The ability to culture human endothelial cells in vitro provides a unique system with which to study this host-parasite interaction directly. Human vascular endothelial cells derived from the umbilical vein, when infected by R. rickettsii, became severely damaged within a few days postinfection. The primary lesion observed at the ultrastructural level appeared to occur at intracellular membranes, specifically, the rough-surfaced endoplasmic reticulum. Widespread dilatation of these membranes eventually led to the creation of large intracellular cisternae and the apparent circumscription of rickettsiae and cellular organelles by the rough-surfaced endoplasmic reticulum. Small membrane-bound fragments of host cytosol created by dilating membranes also were present within the cisternae. Within 5 to 6 days postinfection, cells lost their osmoregulatory control and lysed. Some possible mechanisms of cell injury directed at the level of intracellular membranes are discussed.

Animals↗
PubMed1982

Penicillin-induced unstable intracellular formation of spheroplasts by rickettsiae.

Penicillin G (greater than or equal to 20 micrograms/ml) is rapidly rickettsiacidal for intracellular Rickettsia prowazekii. Light and electron microscopic examinations revealed that penicillin G in culture medium induced a predictable transformation into typical enlarging spheroplasts deficient in the internal, putative peptidoglycan layer of the outer membrane. Under certain conditions, spheroplasts ruptured to discharge contents into host cell cytoplasm and to leave empty shells of defective outer membrane and diffuse amorphous intracytoplasmic antigen. Host cell destruction often accompanied spheroplast rupture. Penicillin G (100 micrograms/ml) caused similar spheroplast formation by Rickettsia rickettsii, but 1,000 micrograms/ml caused neither growth inhibition nor spheroplast formation in Rickettsia tsutsugamushi. The clinical and epidemiological significance of a practical rickettsiacidal drug for the treatment of louse-borne typhus fever is discussed. Practical pharmacologic considerations preclude the use of penicillin for the treatment of typhus or spotted fever.

Animals↗
PubMed1981

Some characteristics of heavy and light bands of Rickettsia prowazekii on Renografin gradients.

Suspensions of partially purified Rickettsia prowazekii yielded two bands of organisms when centrifuged to equilibrium in Renografin density gradients. Rickettsiae from the lower, heavy band were defective in their infective and metabolic activities, as compared to organisms from the light band. The greater density in Renografin of heavy-banding organisms was due to their lack of permeability barrier to it, as evidenced by the absence of plasmolysis in hypertonic Renografin. In contrast, light-banding rickettsiae were able to exclude Renografin, since they were plasmolyzed in it. The proportion of heavy-banding organisms in a rickettsial suspension was influenced by the growth phase they were in when harvested from infected yolk sacs, as well as by the conditions and media to which they subsequently were exposed. We have concluded that these defective forms arise from the degeneration of light-banding rickettsiae. This separation of two functional classes of rickettsiae in Renografin density gradients has been exploited (i) to increase the uniformity of the suspensions by removing many noninfectious particles and (ii) to determine rapidly the integrity of certain properties of the cytoplasmic membrane of organisms exposed to a variety of conditions.

Cell Membrane↗
PubMed1980

In vitro studies of Rickettsia-host cell interactions: ultrastructural study of Rickettsia prowazekii-infected chicken embryo fibroblasts.

Secondary chicken embryo fibroblasts infected in suspension with the Breinl strain of Rickettsia prowazekii and grown in monolayer culture were examined by both transmission and scanning electron microscopy at specific intervals after infection to study the effects of prolonged intracellular growth on the fine structure of the host cell and the rickettsiae. Cytopathological changes in the infected host cells were not apparent until late in the intracellular growth cycle when the cells began to rupture as a result of a large rickettsial burden. The only recognizable changes in heavily infected cells before lysis were the condensation of the intercristal matrix of some mitochondria and the apparent dissociation of ribosomes from the rough-surfaced endoplasmic reticulum. Although the effects of intracellular growth of rickettsiae on the fine structure of the host cell were rather unremarkable when compared with those imposed by Rickettsia rickettsii in a similar cell system, noticeable morphological changes in the rickettsiae were recognized during the intracellular growth cycle. These changes first became apparent about 40 h postinfection and consisted primarily of an increased electron density of the rickettsiae, the appearance of numerous vacuoles in the rickettsial cytoplasm, and a slight reduction in size of the rickettsiae. Changes of this nature may reflect transitional phases of growth characteristically seen in free-living bacterial cell systems.

Animals↗
PubMed1979

In vitro studies of rickettsia-host cell interactions: ultrastructural changes induced by Rickettsia rickettsii infection of chicken embryo fibroblasts.

Secondary chicken embryo fibroblasts infected with the Sheila Smith strain of Rickettsia rickettsii and grown in monolayer culture undergo rapid morphological alterations. Transmission electron microscopic examination of cells at intervals after infection showed several progressive host cell lesions, including widespread dilatation of the rough endoplasmic reticulum and outer nuclear envelope and the accumulation of electron-dense material within the cisternae of intracellular membranes. Dilatation of the rough endoplasmic reticulum is a common, early reversible manifestation of other forms of cell injury. However, the severity of the damage to the host cell resulting from the progressive distention of intracellular membranes and the subsequent formation of small segments of membrane-bound host cytoplasm within the cisternae of these membranes is unknown. Early in the infection cycle, the rickettsiae were found free in the host cell cytoplasm, within invaginations of the nuclear envelope, occasionally free in the space between the outer and inner nuclear membranes, and in the host nucleoplasm, but not within cisternae formed by swollen endoplasmic reticulum. As a consequence of intracisternal swelling and fusion of intracellular membranes later in the infection cycle, the majority of the rickettsiae were found surrounded by host cytoplasm bound by host-derived internal membranes and appeared to persist in this state until cell lysis. The overall cytopathological changes in cells infected with R. richettsii appear dramatic and, from other studies in our laboratory, are significantly different from those observed in cells infected with Rickettsia prowazekii.

Animals↗