Tumor Necrosis Factor-α Induces Fibronectin Synthesis in Coronary Artery Smooth Muscle Cells by a Nitric Oxide–Dependent Posttranscriptional Mechanism

Abstract

Abstract-Postcardiac transplant coronary arteriopathy is associated with tumor necrosis factor-␣ (TNF-␣) induction of fibronectin-dependent smooth muscle cell (SMC) migration into the subendothelium, resulting in occlusive neointimal formation. Because expression of inducible nitric oxide synthase (iNOS) is elevated in neointimal formation after transplantation and upregulated in vascular SMCs by TNF-␣, we investigated whether TNF-␣ induction of fibronectin synthesis in coronary artery (CA) SMCs is mediated by nitric oxide (NO). TNF-␣ caused a dose-dependent increase in reactive oxygen and nitrogen intermediates in CA SMCs (PϽ0.05). This correlated with increased NO production (PϽ0.05) and fibronectin synthesis (PϽ0.05). TNF-␣ induction of fibronectin synthesis was abrogated by the NOS inhibitor N G -monomethyl-L-arginine (L-NMMA) (PϽ0.05) or the flavonoid-containing enzyme inhibitor diphenyleneiodonium (DPI) (PϽ0.05) and reproduced with the NO donor S-nitroso-N-acetyl-penicillamine (SNAP) (PϽ0.05). Northern blotting showed no effect of TNF-␣ on steady-state fibronectin mRNA levels. TNF-␣ increased expression of light chain 3 (LC-3), a protein shown previously to facilitate fibronectin mRNA translation through its interaction with an adenosine-uracil rich element (ARE) in the 3Ј-untranslated region of fibronectin mRNA. RNA gel mobility shift and UV cross-linking assays using CA SMC lysates revealed protein binding complexes with radiolabeled oligonucleotide containing the ARE, similar to those generated with recombinant LC-3. One of these complexes increased after TNF-␣ treatment, an effect inhibited with L-NMMA or DPI. These data demonstrate a novel paradigm whereby cytokines regulate mRNA translation of extracellular matrix proteins through NO-dependent modulation of RNA binding protein interaction with mRNA.

LC3-mediated fibronectin mRNA translation induces fibrosarcoma growth by increasing connective tissue growth factor

“…7). Tumor necrosis factor alpha and TGFβ stimulate production of LC3 (O’Blenes et al, 2001) and FN (Dean et al, 1988), respectively. There was however, no convincing evidence that mRNA expression of LC3 and CTGF tracked with TGFβ or TNFα.…”

“…The mutant LC3 protein was stable and formed a doublet at 16kDa, but it appeared somewhat different from the WT protein in that the upper band was more intense than the lower band and it was also shifted slightly downward. Our previous studies related the lower band of the doublet to the phosphorylated form of LC3 that is associated with the pelleted membranes of the cell and with the polyribosomes necessary for mRNA translation (O’Blenes et al, 2001). The lower band is also associated with a longer sequence (Kabeya et al, 2000).…”

Maternal Hypercholesterolemia and Treatment During Pregnancy Influence the Long-Term Progression of Atherosclerosis in Offspring of Rabbits

“…3,4 This was based on the observation of markedly increased lipid peroxidation in plasma and lesions of human mothers, fetuses, and newborns (now confirmed in the rabbit model), and on the fact that multiple nuclear signaling pathways are oxidation-sensitive. [5][6][7][8][9][10][11][12] Microarray-based determinations in a murine model of fetal atherogenesis indicate that differences in arterial gene expression later in life indeed exist (C. Napoli, F. de Nigris, J. Welch, F. Calara, R. Stuart, C.K. Glass, and W. Palinski, unpublished results, 2001).…”

“…3,4 We also postulated that lipid oxidation plays an important pathogenic role in fetal lesion formation and its postnatal consequences, because oxidation of LDL is prevalent in fetal lesions and because a number of pathways regulating the expression of genes modulating atherogenesis are oxidation-sensitive. [5][6][7][8][9][10][11][12] An important corollary of this hypothesis is that lipid-lowering and antioxidant interventions in mothers during pregnancy should provide long-lasting benefits to their offspring.…”

Developmental expression of LC3α and β: Absence of fibronectin or autophagy phenotype in LC3β knockout mice

“…Increased FN synthesis is associated with enhanced motility of ductus arteriosus versus aortic SMCs (Boudreau et al, 1991), and this feature is necessary in the formation of the intimal cushions required for postnatal closure of the ductus arteriosus (Mason et al, 1999a). In addition, TNF-␣ stimulation of FN synthesis in coronary artery SMC depends on nitric oxide-mediated phosphorylation of LC3 (O'Blenes et al, 2001). LC3 is also a key component of autophagosomes (Kabeya et al, 2000(Kabeya et al, , 2004 assembled under conditions of nutrient starvation (Komatsu et al, 2005).…”