Oligotide anti-ischemic effect is related to in vitro inhibition of leukocyte-endothelial cell adhesion.
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3T3 Cells↗
Biomedical subjects
F Pellegatta
Publications and source records attributed to F Pellegatta.
Oxidized LDL induce hsp70 expression in human smooth muscle cells.
Heat shock protein 70 (hsp70) has been detected in atherosclerotic lesions, in which endothelial cells and smooth muscle cells are involved. In a previous report we showed that Ox-LDL, a causal factor in atherosclerosis, could induce hsp70 expression in cultured human endothelial cells [Zhu et al. B.B.R.C. 1994, 200:389]. Here, with immunofluorescence and immunoblotting techniques, we show that Ox-LDL are capable of inducing hsp70 expression also in human smooth muscle cells, and that this induction is dependent on cell density and on the concentration of Ox-LDL. The induced expression of hsp70 was higher in human umbilical vein smooth muscle cells than in a human smooth muscle cell line. Conversely, Ox-LDL was cytotoxic to both types of cells, more so to the human smooth muscle cell line. These observations indicate that Ox-LDL may be a stress responsible for hsp70 expression in atherosclerotic plaques and the presence of hsp70 in plaques may be a useful marker for continuous oxidative damage in the arterial wall.
Actins↗
Oxidized-LDL induce the expression of heat shock protein 70 in human endothelial cells.
Heat shock proteins are detectable in human atherosclerotic plaques, especially in endothelial cells. In this report we show by immunofluorescence that incubation "in vitro" with OxLDL is a stress capable of inducing the expression of heat shock protein 70 in both the EAhy-926 cell line and human umbilical vein endothelial cells (HUVEC). This induction was parallel to the cytotoxicity of oxidized LDL as determined by [3H]adenine release. When cells were confluent, however, both effects were greatly reduced. We speculate that induction of hsp70 is related to the cytotoxicity of oxidized LDL and that the detection of heat shock proteins in human atherosclerotic plaques is a further indication for the presence "in vivo" of oxidized LDL. These observations may be relevant to the understanding of endothelial response to injury in proatherosclerotic events.
Adenine↗
Inducible nitric oxide synthase modulates fibronectin production in the EA.hy926 cell line and cultured human umbilical vein endothelial cells.
We wished to test the hypothesis of a connection existing between inducible nitric oxide (NO) synthesis and production of extracellular matrix proteins in endothelial cells (EC). We recently reported that the inducible-NO pathway contributes to cytokine-induced enhancement of tumor cell (TC) adhesion to cultured vascular endothelium, independent of changes in E-selectin expression on endothelial cells (EC). We now show that inducible NO-synthase is involved in enhancing fibronectin production by EC. Indeed, fibronectin synthesis and secretion increased both in the EA.hy926 EC line and in human umbilical vein EC (HUVEC) after prolonged exposure to tumor necrosis factor-alpha (TNF-alpha) or interferon-gamma (IFN-gamma). This effect was reversed by the reported inhibitor of NO synthase N omega-nitro-L-arginine methyl ester (L-NAME 10(-5) M). The two cytokines exerted no additive effect, suggesting that they trigger a common metabolic pathway. NO production by cytokine-stimulated EC was dependent on the inducible NO-pathway, as demonstrated by studies of EC-dependent inhibition of platelet aggregation. This inhibition was also evident in calcium-free medium and was reversed by L-NAME and by two inhibitors of protein synthesis that are reported to block the inducible-NO synthase, such as dexamethasone (Dex 10(-7) M) and cycloheximide (Chx 10(-6) M). We conclude that modulation of the inducible NO-synthase may regulate matrix protein production by vascular endothelium during inflammation.
Amino Acid Oxidoreductases↗
Platelet calcium homeostasis is abnormal in patients with severe arteriosclerosis.
To evaluate platelet calcium homeostasis in a typical thrombosis-prone clinical condition, 14 patients with severe arteriosclerosis and 11 healthy control subjects were studied. Platelet intracellular free calcium concentration ([Ca2+]i) was evaluated by means of the fluorescent probe fura 2 under resting conditions and after challenge with 0.05, 0.1, and 0.5 U/mL thrombin (final concentrations). Three different concentrations of extracellular ionized calcium ([Ca2+]e) were used: 1 mmol/L, 1 mumol/L, and < 1 nmol/L. Resting platelet [Ca2+]i was significantly higher (P < .001) in patients than in control subjects. After addition of 0.05 U/mL thrombin, the relative increase of [Ca2+]i was lower in patients than in control subjects in each of the three [Ca2+]e conditions (P = .05 at 1 mmol/L, P = .02 at 1 mumol/L, and P = .04 at < 1 nmol/L). After addition of 0.1 U/mL thrombin, the relative increase of [Ca2+]i was lower in patients than in control subjects under two [Ca2+]e conditions, 1 mumol/L and < 1 nmol/L (P = .04 and P = .03 respectively). With 0.5 U/mL thrombin, a trend toward lower values in patients than in control subjects was observed, reaching statistical significance (P = .03) only at < 1 nmol/L [Ca2+]e. These results suggest that calcium homeostasis is abnormal in platelets from patients with severe arteriosclerosis and probably reflects a chronic activation.
Adult↗
Deranged platelet calcium homeostasis in poorly controlled IDDM patients.
OBJECTIVE: Platelet hyperfunction frequently occurs in IDDM. As in many other cellular systems, cytosolic free Ca plays a key role in platelet activation. RESEARCH DESIGN AND METHODS: We measured cytosolic free Ca concentration ([Ca2+]i) by means of the fluorescent probe fura-2 in 60 IDDM patients (mean age 30.8 yr, range 18-50 yr) and in 31 age-matched healthy control subjects. Platelets were studied in both resting conditions and after stimulation with thrombin at 0.05, 0.1, and 0.5 U/ml. RESULTS: No differences were noted between control subjects and diabetic patients, as a whole. Patients with a poor metabolic control (HbA1c > 8%) had significantly (P < 0.01 and P < 0.03) higher [Ca2+]i in resting platelets. The presence or absence of retinopathy did not modify resting platelet [Ca2+]i. After stimulation with thrombin, a significantly (P < 0.009) higher rise of platelet [Ca2+]i was observed only in those patients who were both free from complications and had good metabolic control. A highly significant (P < 0.001) correlation was found between resting [Ca2+]i and both blood cholesterol and HbA1c in the diabetic patients. Platelets from 10 young healthy subjects also were studied after in vitro incubation with various glucose concentrations (from 1.68 to 56 mM): resting and thrombin-stimulated platelet [Ca2+]i and thrombin-induced aggregation were not modified. CONCLUSIONS: These data confirm that platelet hyperfunction is present in IDDM patients who have unsatisfactory metabolic control, and give evidence that such an activation involves Ca homeostasis. Acute variations of blood glucose concentration are probably not influent, in this respect.
Adult↗
Involvement of nitric oxide in tumor cell adhesion to cytokine-activated endothelial cells.
The aim of the present experiments was to test the possible involvement of nitric oxide (NO) in cytokine-induced enhancement of tumor cell (TC) adhesion to endothelial cells (ECs). Exposure of EA hyb 926 cells to TNF (500 U/ml) plus IFN (100 U/ml) for 24 h significantly enhanced their adhesivity for the 51Cr-labeled GLC1 (small cell lung carcinoma) TCs. Conversely, exposure of TCs to cytokines did not result in an increased adhesion of these cells to ECs. TC-stimulated adhesion to EA hyb 926 was abrogated by the glucocorticoid dexamethasone (Dex, 10(-7) M), the NO synthase inhibitors N omega-nitro-L-arginine methyl ester (L-NAME, 10(-5) M) and NG-monomethyl-L-arginine (L-NMMA, 10(-5) M) and the protein synthesis inhibitor cycloheximide (Cex, 10(-6) M). Furthermore, GLC1-stimulated adhesion to EA hyb 926 was reversed following removal of L-arginine from the medium or pretreatment with the guanylate cyclase inhibitor methylene blue. TC-stimulated adhesion was also prevented when TCs were pretreated with the monoclonal antibody CD15 directed against the endothelial-leukocyte adhesion molecule (ELAM-1) ligand or following exposure of ECs to anti-ELAM-1 monoclonal antibody. Although suppressing TC-stimulated adhesion, L-NMMA failed to modify significantly cytokine-induced ELAM-1 expression in EA hyb 926. These results (a) provide evidence for the NO-inducible pathway contributing to cytokine-induced enhancement of tumor cell adhesion to the vascular endothelium and (b) demonstrate the involvement of the ELAM-1/CD15 adhesion system in tumor cell-stimulated adhesion to ECs.
Arginine↗