Differing Roles of Protein Kinase C on the Antiproliferative Effects of Tumor Necrosis Factor Alpha and Beta on LoVo Cells

Pathobiology ◽  
1990 ◽  
Vol 58 (3) ◽  
pp. 168-171 ◽  
Author(s):  
Nagahide Matsubara ◽  
Sadanori Fuchimoto ◽  
Kunzo Orita
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Tumor Necrosis Factor ◽  
Tumor Necrosis ◽  
Necrosis Factor Alpha ◽  
Antiproliferative Effects ◽  
Kinase C ◽  
Lovo Cells ◽  
Factor Alpha ◽  
Necrosis Factor

Tumor necrosis factor-alpha activates pulmonary artery endothelial protein kinase C

1993 ◽  
Vol 264 (1) ◽  
pp. L7-L14 ◽  
Author(s):  
T. J. Ferro ◽  
D. M. Parker ◽  
L. M. Commins ◽  
P. G. Phillips ◽  
A. Johnson
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Tumor Necrosis Factor ◽  
Pulmonary Artery ◽  
Tumor Necrosis Factor Alpha ◽  
Tumor Necrosis ◽  
Necrosis Factor Alpha ◽  
Kinase C ◽  
Factor Alpha ◽  
Necrosis Factor

We investigated the hypothesis that tumor necrosis factor-alpha (TNF) activates pulmonary endothelial protein kinase C (PKC). Confluent bovine pulmonary artery endothelial monolayers were exposed to recombinant human TNF, and the translocation of PKC, an indicator of enzyme activation, was studied using both slot immunoblotting and immunofluorescence. For slot immunoblot analysis, membrane and cytosol lysate fractions were prepared, and PKC antigen was assessed using MC5 monoclonal anti-PKC antibody. TNF (1,000 U/ml for 15 min) induced translocation of PKC into the membrane. Immunofluorescence analysis with the MC5 antibody was also used. Monolayers treated with culture medium showed diffuse cytoplasmic fluorescence. In contrast, treatment with either TNF (1,000 U/ml for 15 min) or 1,2-dioctanoylglycerol (4 x 10(-5) M for 5 min), a diacylglycerol that activates PKC, resulted in translocation of fluorescence to the cell periphery; fine, punctate PKC-associated fluorescence was localized to the margins of cells. The TNF-induced translocation of PKC was inhibited using either IP-300 polyclonal anti-TNF antibody (indicating that the TNF effect was not due to the vehicle or contaminating endotoxin) or calphostin C (10(-6) M for 15 min), which inhibits PKC activation by interacting with the regulatory diacylglycerol-binding domain. TNF treatment had no effect on either the content of PKC, or of total protein, in the membrane + cytosol, and cycloheximide (40 microM for 5 min) did not alter the translocation of PKC induced by TNF; these results indicate that the effect of TNF on PKC translocation was related to neither de novo membrane synthesis of PKC (as opposed to translocation per se) nor nonspecific augmentation of protein synthesis.(ABSTRACT TRUNCATED AT 250 WORDS)

Tumor necrosis factor-alpha decreases pulmonary artery endothelial nitrovasodilator via protein kinase C

1994 ◽  
Vol 267 (3) ◽  
pp. L318-L325 ◽  
Author(s):  
A. Johnson ◽  
D. T. Phelps ◽  
T. J. Ferro
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Tumor Necrosis Factor ◽  
Pulmonary Artery ◽  
Tumor Necrosis ◽  
Necrosis Factor Alpha ◽  
Calphostin C ◽  
Kinase C ◽  
Factor Alpha ◽  
Necrosis Factor

We postulated that tumor necrosis factor-alpha (TNF) decreases endothelium-derived nitrovasodilator(s) via a protein kinase C (PKC)-dependent pathway. Calf pulmonary artery endothelial monolayers (PAEM) were treated with TNF (10, 100, and 1,000 U/ml) for 15 min or 18 h during an 18-h incubation. At the end of the incubation, the cell lysate and supernatant were harvested. Compared with controls, an 18-h incubation with TNF (100 and 1,000 U/ml) resulted in a decrease in NO2- [the oxidation product of nitric oxide (NO)] in PAEM lysate and supernatant. TNF (100 U/ml) treatment for 15 min did not suppress NO2- levels. The decrease in NO2- and the increase in lipid peroxides in response to TNF were prevented by pretreatment (15 min prior to and throughout the incubation) with either calphostin C (1 microM; a specific PKC inhibitor) or the antioxidants N-acetylcysteine (1 mM), 4,5-dihydroxy-1,3-benzene disulfonic acid (Tiron) (10 mM), and superoxide dismutase (10 U/ml). Treatment with phorbol 12-myristate 13-acetate (PMA, 1 microM for 15 min), an activator of PKC, decreased NO2- similarly to TNF. Pretreatment with calphostin C or N-acetylcysteine prior to TNF (10 U/ml) revealed an increase in NO2- levels above control treatment. Treatment with the NO synthase antagonists NG-monomethyl-L-arginine (1 mM) and N-nitroso-L-arginine (1 mM) induced an L-arginine (1 mM)-dependent decrease in NO2- in control but not in TNF-treated PAEM. The induction of NO2- by calcium ionophore (A23187; 500 nM) was not affected by treatment with TNF.(ABSTRACT TRUNCATED AT 250 WORDS)

Activation of protein kinase C mediates altered pulmonary vasoreactivity induced by tumor necrosis factor-alpha

1994 ◽  
Vol 267 (3) ◽  
pp. L282-L290 ◽  
Author(s):  
G. Serfilippi ◽  
T. J. Ferro ◽  
A. Johnson
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Tumor Necrosis Factor ◽  
Tumor Necrosis ◽  
Necrosis Factor Alpha ◽  
Calphostin C ◽  
Kinase C ◽  
Factor Alpha ◽  
Pkc Activation ◽  
Necrosis Factor

We postulated that tumor necrosis factor-alpha (TNF) "primes" the lung for the development of pulmonary vasoconstriction and edema by activating protein kinase C (PKC). Guinea pigs were injected with TNF (1.6 x 10(5) U/kg i.p.), and the lungs were isolated 4 h later. Compared with controls, TNF pretreatment resulted in greater increases in pulmonary vascular resistance and pressure and lung weight, in response to the thromboxane A2 mimetic, U-46619 (122 pmol/min). Treatment with TNF resulted in 1) pulmonary arterial endothelial PKC activation, 2) increased lung polymorphonuclear neutrophil (PMN) sequestration, 3) increased levels of superoxide radical (O2.) in lung effluent, and 4) decreased nitrite levels (NO2-, oxidation product of nitric oxide) in lung effluent. Intraperitoneal treatment with calphostin C (3 microM, 15 min prior to treatment with TNF) prevented the effects of TNF on 1) PKC activation, 2) the hemodynamic responses to U-46619, and 3) the levels of NO2- and O2(.). PKC activation does not mediate TNF-induced lung sequestration of PMN, since calphostin C had no effect on lung myeloperoxidase activity. The data suggest that PKC activation mediates TNF-induced 1) increases in O2., 2) decreases in NO2-, and 3) increases in vasoreactivity and edema in response to U-46619.

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