Tumor necrosis factor-α enhances neutrophil adhesiveness:: Induction of vascular cell adhesion molecule-1 via activation of Akt and CaM kinase II and modifications of histone acetyltransferase and histone deacetylase 4 in human tracheal smooth muscle cells

Up-regulation of vascular cell adhesion molecule-1 (VCAM-1) involves adhesions between both circulating and resident leukocytes and the human tracheal smooth muscle cells (HTSMCs) during airway inflammatory reaction. We have demonstrated previously that tumor necrosis factor (TNF)-alpha-induced VCAM-1 expression is regulated by mitogen-activated protein kinases, nuclear factor-kappa B, and p300 activation in HTSMCs. In addition to this pathway, phosphorylation of Akt and CaM kinase II has been implicated in histone acetyltransferase and histone deacetylase 4 (HDAC4) activation. Here, we investigated whether these different mechanisms participated in TNF-alpha-induced VCAM-1 expression and enhanced neutrophil adhesion. TNF-alpha significantly increased HTSMC-neutrophil adhesions, and this effect was associated with increased expression of VCAM-1 on the HTSMCs and was blocked by the selective inhibitors of Src [4-amino-5-(4-methyl-phenyl)7-(t-butyl)pyrazolo[3,4-d]-pyrimidine (PP1)], epidermal growth factor receptor [EGFR; 4-(3' -chloroanilino)-6,7-dimethoxy-quinazoline, (AG1478)], phosphatidylinositol 3-kinase (PI3K) [2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride(LY294002) andwortmannin],calcium[1,2-bis(2- aminophenoxy) ethane-N,N,N', N' -tetraacetic acid- acetoxymethyl ester; BAPTA-AM], phosphatidylinositol-phospholipase C ( PLC) [1-[6-[[17 beta-methoxyestra- 1,3,5( 10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-dione(U73122)], protein kinase C (PKC) [12-(2-cyanoethyl)-6,7,12, 13-tetrahydro-13-methyl-5-oxo-5H-indolo(2,3-a)pyrrolo(3,4-c)carbazole (Go6976), rottlerin, and 3-1-[3-(amidinothio)propyl-1H-indol-3-yl]- 3-(1-methyl-1H-indol-3-yl) maleimide (bisindolylmaleimide IX) (Ro 31-8220)], CaM ( calmidazolium chloride), CaM kinase II [(8R*, 9S*, 11S*)-( -)- 9-hydroxy-9-methoxycarbonyl-8methyl-14-n-propoxy-2,3,9,10-tetrahydro-8,11-epoxy,1H,8H,11H- 2,7b, 11a-triazadibenzo[a,g]cycloocta[cde]trinden-1-one(KT5926) and 1-[N,O-bis( 5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]4- phenylpiperazine (KN62)], p300 (curcumin), and HDAC ( trichostatin A) or transfection with short interfering RNAs for Src, Akt, PKC alpha, PKC mu, and HDAC4. At gene regulation level, reverse-transcriptase polymerase chain reaction and promoter assays revealed that expression of VCAM-1 was also attenuated by these signaling molecule inhibitors. Moreover, TNF-alpha induced Akt and CaM kinase II phosphorylation via cascades through Src/EGFR/PI3K and PLC/calcium/CaM, respectively. Finally, activation of Akt and CaM kinase II may eventually lead to the acetylation of histone residues and phosphorylation of histone deacetylase. These findings revealed that TNF-alpha induced VCAM-1 expression via multiple signaling pathways. Blockade of these pathways may be selectively targeted to reduce neutrophil adhesion via VCAM-1 suppression and attenuation of the inflammatory responses in airway diseases.