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An oligomer targeted against protein kinase C alpha prevents interleukin-1 alpha induction of cyclooxygenase expression in human endothelial cells
No full textWe have previously demonstrated that interleukin-1 alpha (IL-1), a potent polypeptide mediator of immune and inflammatory responses, induces the expression of cyclooxygenase (cox) in human endothelial cells. The mechanism by which binding of IL-1 to its receptor stimulates gene expression remains unclear. Since phorbol 12-myristate 13-acetate, which directly binds and activates protein kinase C (PKC), induced cox expression, we examined the role of PKC as an intracellular mediator of IL-1 activity in human endothelial cells. IL-1 induced the translocation of PKC from the cytosol to the membrane. H7, a selective inhibitor of PKC, as well as an antisense oligomer blocking PKC alpha translation suppressed IL-1 induction of cox mRNA. These findings establish that PKC plays a role in the signal transduction pathway leading to the induction of cox in human endothelial cells
Different molecular pathways regulate cellular senescence in human fibroblasts and in human endothelial cells
No full textSenescence stimulates U937-endothelial cell interactions
No full textProgressive pathophysiologic modifications of endothelial cells are associated with aging. In vitro, endothelial cell senescence is accompanied by the failure to proliferate as well as by perturbations in gene expression. Here we show that (i) senescence enhances monoblastoid U937 cell adhesion to the endothelial monolayer; (ii) the enhanced interaction between senescent endothelial cells and U937 cells is mediated, at least in part, by the overexpression of ICAM-1; and (iii) LPS and interleukin 1 alpha, but not tumor necrosis factor alpha, are unable to stimulate the adhesion of U937 to senescent endothelial cells. Since monocyte adhesion to the endothelium is an early event in atherosclerosis, the altered adhesive properties observed in senescent cells could give insights into the formation of atherosclerotic lesions
The mitogenic signaling pathway but not the plasminogen activator-inducing pathway of basic fibroblast growth factor is mediated through protein kinase C in fetal bovine aortic endothelial cells
No full textBasic fibroblast growth factor (bFGF) induces cell proliferation and plasminogen activator (PA) activity in transformed fetal bovine aortic endothelial (FBAE) GM 7373 cells. A similar response is observed after treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA). In these cells, bFGF and TPA cause activation of protein kinase C (PKC), as demonstrated by the induction of the phosphorylation of an 87-kD PKC substrate in intact cells and by the increase in membrane-associated PKC activity. Activation of PKC by bFGF or TPA is inhibited in cells made PKC-deficient by pretreatment with high concentrations of TPA. The mitogenic activity of bFGF or of TPA is completely inhibited in PKC-deficient cells or in naive cells treated with the PKC inhibitor H-7. However, these cells proliferate in response to serum, epidermal growth factor, and dibutyryl cyclic AMP. Similar results are obtained in normal FBAE AG 7680 cells. These data indicate that activation of PKC is responsible for the mitogenic activity of bFGF in FBAE cells. On the contrary, the PA-inducing activity of bFGF is unaffected by down-regulation of PKC or by treatment with the PKC inhibitor H-7 in both transformed GM 7373 and normal AG 7680 cells. bFGF induces a rapid 45Ca influx in naive and in PKC-deprived GM 7373 cells. In these cells, addition of EGTA to the incubation medium prevents both the 45Ca influx and the increase in PA activity induced by bFGF, without affecting its mitogenic activity. Even though the involvement of PKC in the increase of cell-associated PA activity induced by bFGF cannot be completely dismissed, the present results suggest a role of calcium entry in the modulation of the PA-inducing activity of bFGF
Endogenous interleukin 1 alpha must be transported to the nucleus to exert its activity in human endothelial cells
No full textWe have previously shown that the signal peptideless cytokine interleukin 1 alpha (IL-1 alpha) may play a role as an intracellular regulator of human endothelial cell senescence (J. A. M. Maier, P. Voulalas, D. Roeder, and T. Maciag, Science 249:1570-1574, 1990). To investigate the potential intracellular function of IL-1 alpha, transformed endothelial cells were transfected with the human cDNAs that code for the two forms of IL-1 alpha, the precursor molecule IL-1(1-271) and the mature protein IL-1(113-271). The subcellular localization of the two different polypeptides was investigated directly or by using chimeric genes constructed by fusion of different fragments of the IL-1 alpha gene and the beta-galactosidase open reading frames. The IL-1(113-271) protein was cytoplasmic, while IL-1(1-271) was nuclear. The basic cluster at the NH2 terminus of IL-1, KVLKKRR, has been shown to mediate IL-1 alpha nuclear targeting. Moreover, nuclear localization of IL-1 alpha correlates with impaired cell growth and expression of some IL-1 alpha-inducible genes. These results suggest that transport of endogenous IL-1(1-271) into the nucleus is required for it to modulate endothelial cell function
A method for oriented Epon embedding of cell monolayers to obtain orizontal ultrathin sections for T.E.M. studies
No full textInduction of plasminogen activator activity by phorbol ester in transformed fetal bovine aortic endothelial cells. Possible independence from protein kinase C
No full text12-O-tetradecanoyl phorbol 13-acetate (TPA) and 1,2-dioctanoyl-sn-glycerol (diC8) activate protein kinase C (PKC) in transformed fetal bovine aortic endothelial GM 7373 cells. Both molecules cause a similar increase in membrane-associated PKC activity and in the phosphorylation of a PKC-specific endogenous 87-kDa substrate in intact cells. Even though both TPA and diC8 exert a mitogenic activity in GM 7373 cells, only TPA induces also an increase in cell-associated plasminogen activator (PA) activity. Down-regulation of PKC which follows TPA-pretreatment completely abolishes the mitogenic activity of diC8 and the mitogenic and PA-inducing activity of TPA. However, both the PKC inhibitor H-7 and the down-regulation of PKC which follows a prolonged stimulation with diC8 do not abolish the PA-inducing activity of TPA. The PA-inducing activity of TPA is instead inhibited in cultures incubated in the presence of 1 mM EGTA or in a calcium-free medium. The data indicate that TPA and diC8 induce a different pattern of cellular activation in GM 7373 cells and that the PA-inducing activity of TPA might not be mediated by PKC
Basic fibroblast growth factor in ovulatory cycle and postmenopausal human endometrium.
No full textBiopsies of human endometrium were studied for the presence of basic fibroblast growth factor (bFGF). An immunoreactive Mr 18,000 bFGF-like molecule was detected at high levels both in ovulatory cycle and postmenopausal endometrium. This molecule was identified as bFGF on the basis of its molecular weight, its affinity for heparin, its capacity to induce plasminogen activator production and cell proliferation in endothelial GM 7373 cells, and its cross-reactivity with various anti-bFGF antibodies. The levels of endometrial bFGF do not change during the menstrual cycle but they increase significantly after menopause, as evaluated both by biological and immunological assays. Lower levels of an acidic FGF-like activity were also evident in ovulatory cycle endometrium but, at variance with bFGF, no significant increase of this activity was observed in postmenopausal endometrium. These data represent the first characterization of a polypeptide growth factor present in human endometrium
Influence on cell-cell communication (dye-transfer) of the oncogenic beta-blocker DL-ZAMI 1305: possible relation to tumor promotion.
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