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J Ustinov

Publications and source records attributed to J Ustinov.

13 recordsLinked to original sources
PubMed1992

Towards understanding the pathophysiology of chronic rejection.

Chronic allograft rejection is the major reason why allografts are lost. While only 2%-3% of all allografts are lost during the first year to irreversible acute rejection, approximately 6%-7% are lost during each subsequent year to chronic rejection. The major manifestation of chronic rejection in all organs is persistent perivascular inflammation and allograft arteriosclerosis. Bearing this in mind, we have developed a model to investigate the pathophysiology of allograft arteriosclerosis using aortic transplantations between inbred rat strains. The results obtained thus far indicate that several different inflammatory cascades are operative within the vascular wall during allograft arteriosclerosis. The relative importance of these different cascades, and particularly the role of growth factors as final effectors, has not yet been defined. Attempts to suppress allograft arteriosclerosis under experimental conditions have already met with some success: under conditions where no immunosuppression is provided we have been able to delay the process by at least 3 months, though we have not been able to block it indefinitely. It may be expected, however, that once the inflammatory cascades leading to smooth muscle cell replication in the allograft media and their influx into the intima are better defined, more specific approaches to the inhibition of allograft arteriosclerosis will be developed.

Animals↗
PubMed1992

Chronic rejection of rat aortic allografts. III. Synthesis of major eicosanoids by vascular wall components and effect of inhibition of the thromboxane cascade.

We have previously demonstrated that rat aortic allografts from the DA (RT1a) to the WF (RT1v) strain develop chronic arteriosclerotic changes in the vascular wall after a short spontaneously reversible acute rejection episode. These changes, which are lacking in syngeneic DA-to-DA control grafts, are virtually identical with those observed in human allografts during chronic rejection. In this study we have investigated whether eicosanoids are involved in the generation of arteriosclerotic changes. Incubation of aortic wall rings in vitro and immunochemical assays demonstrated that the arteriosclerotic allografts synthesize significantly more thromboxane B2 (TxB2) but not 6-ketoprostaglandin F1 alpha (6-keto-PGF1 alpha) or leukotriene B4. The increased synthesis of TxB2 in the allografts persisted for at least 5 months after transplantation. Separate incubation of the two major components of the vascular wall, after microdissection of the intima and (media plus) adventitia, demonstrated that most of the synthesis of TxB2 during chronic rejection was due to the outer layer of aorta, presumably the inflammatory cells of the adventitia. In contrast, most of the 6-keto-PGF1 alpha was synthesized by the inner layer of the aorta, presumably the endothelial cells and the smooth muscle cells of the intima. Administration of 1 mg.kg-1 x day-1 of a specific TxA2 receptor inhibitor, GR32191B, to the recipient rat reduced the proliferative response of inflammatory cells in the adventitia by 30%, as detected by in vivo tritiated-thymidine (3H-TdR) labeling and autoradiography, but did not reduce the proliferative response of smooth muscle cells in the media and intima.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Carbohydrate synthesis inhibitors decrease interleukin 1-stimulated lymphocyte binding to endothelial cells.

Lymphocyte extravasation (homing) is initiated when lymphocyte adheres to endothelial cells. All know protein structures involved in the lymphocyte binding located on the endothelial surface are heavily glycosylated. We asked therefore whether these carbohydrate motifs had a role in the lymphocyte homing. The relative importance of the N-linked chains on biological effects mediated by glycoproteins can be studied with specific inhibitors of carbohydrate processing, i.e. 1-deoxynojirimycin (DN), castonospermine (CAST), 1-deoxymannojirimycin (DMN) and swainsonine (SW), which produces different kinds of blocked carbohydrate chains. N-linked carbohydrate chains are modified in the Golgi apparatus and in the final glycoprotein they can be either of high-mannose-, hybrid- or complex-type motifs, having the same core structure but different terminal structures. We show here that when all N-linked carbohydrates were cleaved off from the cell surface glycoproteins by treating endothelial cells with N-glycosidase F interleukin 1-induced lymphocyte binding was reduced almost to non-stimulated control values. Treatment of endothelial cells with CAST led to generation of glycoproteins carrying high-mannose-type oligosaccharides, which are glucose capped with three glucose molecules on the chain. CAST treatment resulted in an 85% decrease in lymphocyte binding compared to interleukin 1-induced levels. DMN treatment, resulting in accumulation of high-mannose type oligosaccharides without any terminal glucoses on the cell surface, caused a similar inhibition of lymphocyte binding to that induced by CAST treatment. SW treatment, leading to accumulation of hybrid-type glycoproteins, decreased only slightly the lymphocyte binding. These results suggest that carbohydrates indeed have a role in lymphocyte binding to endothelial cells.

1-Deoxynojirimycin↗
PubMed1990

Protein kinase C-regulated production of prostacyclin by rat endothelium is increased in the presence of lipoxin A4.

Prostacyclin is generated by cultured rat endothelial cells. Compound blocking activity of protein kinase C and cyclic nucleotide-dependent protein kinases (H7) and compound blocking interaction between Ca2+ and calmodulin (W7) diminish generation of prostacyclin in rat endothelial cells. These compounds give a synergistic effect when they are introduced to the endothelial cell cultures simultaneously. Compound HA1004, an inhibitor of cAMP- and cGMP-dependent protein kinases has no effect on prostacyclin generation. Lipoxin A4, a potent direct stimulator of protein kinase C, rapidly induces prostacyclin generation in rat endothelium in a dose- and time-dependent fashion. Lipoxin A4-induced generation of prostacyclin can be inhibited by H7 and W7 but not by HA1004. Lipoxin B4 has no significant effect on prostacyclin generation in rat endothelium. In conclusion, our results demonstrate that generation of prostacyclin by rat endothelial cells is regulated via a pathway involving protein kinase C and Ca2+.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1990

Interleukin 1-induced lymphocyte binding to endothelial cells. Role of cAMP as a second messenger.

Interleukin 1 (IL 1) is a potent protein mediator of inflammation. Among other things it increases the number of lymphocytes adhering to endothelial cell monolayers. We analyzed the signal transduction during IL 1-induced lymphocyte binding. Dibutyryl cyclic AMP, which is a cAMP analog able to penetrate into the cytosol, increased lymphocyte binding to the same extent as IL 1. Direct activation of adenylate cyclase by forskolin enhanced also lymphocyte binding. IL 1 increased the level of cytosolic cAMP in a time- and dose-dependent manner measured with radioimmunoassay. 2',5'-Dideoxyadenosine, which is an inhibitor of adenylate cyclase, decreased both the IL 1-induced lymphocyte binding to endothelial cells and elevation in cytosolic cAMP levels. Lymphocyte binding increased with cytosolic cAMP levels in accordance with elevation of IL 1 concentration. These results suggest that cAMP is essential in signal transduction during IL 1-induced lymphocyte binding to cultured endothelial cell monolayers.

Animals↗
PubMed1990

Interferon-gamma augments hydrolysis of LTA4 to LTB4 by endothelial cells.

LTB4 is a potent mediator of inflammation acting at local sites of inflammation. LTB4 increases the lymphocyte binding to and penetration through the endothelium. In this paper we demonstrate that while endothelial cells were unable to metabolize LTB4 from arachidonic acid they were able to hydrolyse LTA4 into LTB4 in a granulocyte-endothelial co-culture assay. This hydrolysis is markedly increased if endothelial cells were pretreated with IFN-gamma prior to the assay. The IFN-gamma induced effect was shown to be time- and dose-dependent. The ability of endothelial cells to hydrolyse LTA4 to LTB4 may provide an answer how LTB4 can be produced in large quantities by nonheamatopoetic cells (i.e. by endothelial cells) at sites of acute inflammation.

Animals↗
PubMed1990

Signal transduction during in vitro lymphocyte homing.

Lymphocyte binding to endothelium is a necessary prerequisite for lymphocyte homing through endothelium. This is mediated by the binding of ligands on endothelial cells to lymphocyte surface homing receptors. We show in this paper that the intracellular second messenger pathways involved in interferon gamma-induced intercellular adhesion molecule 1 upregulation on endothelial cells are protein kinase C and calcium dependent. Lymphocyte binding to endothelial cells is enhanced by both platelet activating factor and interleukin 1 alpha. Platelet activating factor added to endothelial cultures increases lymphocyte binding within 10 min and operates via protein kinase C but not via cAMP. On the other hand interleukin 1 alpha increases binding within 4 hr and operates via cAMP but not via protein kinase C. These results imply that different mediators of inflammation can activate different signal transduction pathways but lead to similar increases in lymphocyte binding.

Animals↗
PubMed1990

Properties of B cells and Thy-1-antigen-expressing cells infiltrating rat renal allografts.

We have examined (1) the frequency of B cells secreting antibodies against donor major histocompatibility complex (MHC) antigens and (2) the properties of Thy-1-antigen-expressing leukocytes in rats rejecting renal allografts. Our results show that B cells secreting antibodies are present in the inflammatory cell population at the frequency of 1:850. Among them only 1 out of 2-150 is engaged in production of antibodies directed to the graft MHC antigens, depending on the method of assay. This suggests that despite the observed significant production of nonspecific immunoglobulin in situ, only a minority of the B-cell population is specifically committed to the graft MHC antigens. This finding is concordant with the described previously low frequencies of the T cells specifically directed toward the graft MHC antigen. The role of the "immunologically noncommitted" cells in graft rejection is unknown. We have found that a substantial part (up to 60%) of inflammatory cells invading a rat kidney allograft express the Thy-1 antigen. This suggests that they might be immature (progenitor?) cells and, therefore, unable to respond to the graft antigens. Progenitor-like properties of these cells have been confirmed by their ability to reconstitute lethally irradiated syngeneic rat. Finally, these immature cells are of lymphoid, not of myeloid, linkage, because they do not proliferate in the presence of GM-CSF.

Animals↗
PubMed1990

Activation of protein kinase C is crucial in the regulation of ICAM-1 expression on endothelial cells by interferon-gamma.

ICAM-1 (CD54) is expressed on endothelial cells and serves as an important ligand for the white cell adhesion molecule CD11a/CD18 (LFA-1). Many studies have demonstrated that increased numbers of white cells binding to endothelial cells correlate with the level of ICAM-1 expression on endothelial cells. Several cytokines, including IFN-gamma, increase ICAM-1 expression in cultured human endothelial cells. We have analysed the second intracellular messenger pathways involved in IFN-gamma-induced up-regulation of ICAM-1 expression in endothelial cells. IFN-gamma induced a rapid activation of phospholipase C, leading to a breakdown of phosphoinositoldiphosphate (PIP2) into diacyglycerol (DAG) and inositoltriphosphate (IP3). DAG is a natural activator of the protein kinase C pathway. We were able to show that the effect induced by IFN-gamma could be inhibited by a protein kinase C inhibitor, H7, in a dose-dependent manner and mimicked by PMA, which stimulates protein kinase C. IFN-gamma induced a 5-fold translocation (activation) of protein kinase C from the cytosol into the endothelial cell membrane. Elevation of the IP3 levels led to activation of the calcium-dependent pathway. An inhibitor of calcium calmodulin, W7, decreased the IFN-gamma induced ICAM-1 expression, and addition of calcium ionophore to endothelial cells could replace IFN-gamma in the up-regulation of ICAM-1. Finally, IFN-gamma caused a significant increase in the calcium flux of endothelial cells. cAMP and cGMP had no effect on the regulation of ICAM-1 expression on cultured human endothelial cells.

Calcium↗
PubMed1990

Signal transduction during platelet-activating factor-induced lymphocyte binding to endothelial cells.

Platelet-activating factor (PAF) is a lipid mediator of inflammation. PAF pretreatment of cultured endothelial cells leads to an increase in lymphocyte binding. We have analysed the intracellular signal transduction during this PAF-induced effect. The protein kinase C activator, phorbol 12-myristate, 13-acetate, mimicked PAF in the binding assay. Concomitantly, the protein kinase C (PKC) inhibitor H7 down-regulated the PAF-induced binding to nearly control level. Also dibutyryl-cAMP treatment of endothelial cells increased lymphocyte binding, but the protein kinase A inhibitor HA1004 did not alter the PAF-induced binding. Furthermore, PAF did not increase the level of cytosolic cAMP in the endothelial cells. Other second messengers, cGMP and Ca2+, had no effect on lymphocyte binding. These findings suggest that protein kinase C, but not other signal transduction pathway, is essential in the PAF-induced lymphocyte binding to endothelial cells.

Animals↗