%PDF-1.3 % 2 0 obj << /Length 15574 >> stream q 1 i -1 843 597 -844 re 0 842 m W n 0.015 842.01 594.99 -842.01 re W n 0 0 0 0 k /GS2 gs 65 793.85 229.31 -31.84 re 0 842 m f* BT /F6 1 Tf 7 0 0 7 66.0036 787.3962 Tm 0 0 0 1 k 0.0249 Tc 0.1139 Tw (Eur Respir J, 1995, 8, 387391)Tj 0 -1.1429 TD (DOI: 10.1183/09031936.95.08030387)Tj T* (Printed in UK - all rights reserved)Tj ET 0 0 0 0 k 416 794.25 138.22 -32.24 re 179.584 771.396 m f* BT 7 0 0 7 426.0314 787.7977 Tm 0 0 0 1 k 0 Tw (Copyright )Tj /F2 1 Tf 4.92 0 TD 0 Tc ()Tj /F6 1 Tf 0.8125 0 TD 0.0249 Tc 0.1139 Tw (ERS Journals Ltd 1995)Tj -4.3964 -1.1429 TD (European Respiratory Journal)Tj 2.772 -1.1429 TD (ISSN 0903 - 1936)Tj ET 0 0 0 0 k 65 709.93 480.22 -36.35 re 515.612 771.798 m f* BT /F7 1 Tf 15 0 0 15 117.6652 697.4409 Tm 0 0 0 1 k 0 Tc 0 Tw (A)Tj 1 Tr 0 0 0 1 K 0 J 0 j 0.45 w 10 M []0 d 0 0 TD (A)Tj 0 Tr 0.7017 0 TD (c)Tj 1 Tr T* (c)Tj 0 Tr 0.5357 0 TD (t)Tj 1 Tr T* (t)Tj 0 Tr 0.3127 0 TD (i)Tj 1 Tr T* (i)Tj 0 Tr 0.2577 0 TD (v)Tj 1 Tr T* (v)Tj 0 Tr 0.5357 0 TD (a)Tj 1 Tr T* (a)Tj 0 Tr 0.5357 0 TD (t)Tj 1 Tr T* (t)Tj 0 Tr 0.3127 0 TD (i)Tj 1 Tr T* (i)Tj 0 Tr 0.2577 0 TD (o)Tj 1 Tr T* (o)Tj 0 Tr 0.5907 0 TD (n)Tj 1 Tr T* (n)Tj 0 Tr 0.5907 0 TD 0.0936 Tc ( o)Tj 1 Tr 0.3716 0 TD 0 Tc (o)Tj 0 Tr 0.5907 0 TD (f)Tj 1 Tr T* (f)Tj 0 Tr 0.3127 0 TD 0.0936 Tc ( t)Tj 1 Tr 0.3716 0 TD 0 Tc (t)Tj 0 Tr 0.3127 0 TD (h)Tj 1 Tr T* (h)Tj 0 Tr 0.5907 0 TD (e)Tj 1 Tr T* (e)Tj 0 Tr 0.5357 0 TD 0.0936 Tc ( t)Tj 1 Tr 0.3716 0 TD 0 Tc (t)Tj 0 Tr 0.3127 0 TD (r)Tj 1 Tr T* (r)Tj 0 Tr 0.3687 0 TD (a)Tj 1 Tr T* (a)Tj 0 Tr 0.5357 0 TD (n)Tj 1 Tr T* (n)Tj 0 Tr 0.5907 0 TD (s)Tj 1 Tr T* (s)Tj 0 Tr 0.5357 0 TD (c)Tj 1 Tr T* (c)Tj 0 Tr 0.5357 0 TD (r)Tj 1 Tr T* (r)Tj 0 Tr 0.3687 0 TD (i)Tj 1 Tr T* (i)Tj 0 Tr 0.2577 0 TD (p)Tj 1 Tr T* (p)Tj 0 Tr 0.5907 0 TD (t)Tj 1 Tr T* (t)Tj 0 Tr 0.3127 0 TD (i)Tj 1 Tr T* (i)Tj 0 Tr 0.2577 0 TD (o)Tj 1 Tr T* (o)Tj 0 Tr 0.5907 0 TD (n)Tj 1 Tr T* (n)Tj 0 Tr 0.5907 0 TD 0.0936 Tc ( f)Tj 1 Tr 0.3716 0 TD 0 Tc (f)Tj 0 Tr 0.3127 0 TD (a)Tj 1 Tr T* (a)Tj 0 Tr 0.5357 0 TD (c)Tj 1 Tr T* (c)Tj 0 Tr 0.5357 0 TD (t)Tj 1 Tr T* (t)Tj 0 Tr 0.3127 0 TD (o)Tj 1 Tr T* (o)Tj 0 Tr 0.5907 0 TD (r)Tj 1 Tr T* (r)Tj 0 Tr 0.3687 0 TD 0.0936 Tc ( N)Tj 1 Tr 0.3716 0 TD 0 Tc (N)Tj 0 Tr 0.7017 0 TD (F)Tj 1 Tr T* (F)Tj 0 Tr 0.5907 0 TD (-)Tj 1 Tr T* (-)Tj /F2 1 Tf 0 Tr 0.3284 -0.015 TD ()Tj /F7 1 Tf 0.5466 0.015 TD (B)Tj 1 Tr 0 0 TD (B)Tj 0 Tr 0.6991 0 TD 0.091 Tc ( i)Tj 1 Tr 0.369 0 TD 0 Tc (i)Tj 0 Tr 0.2551 0 TD (n)Tj 1 Tr T* (n)Tj 0 Tr 0.5881 0 TD 0.091 Tc ( h)Tj 1 Tr 0.369 0 TD 0 Tc (h)Tj 0 Tr 0.5881 0 TD (u)Tj 1 Tr T* (u)Tj 0 Tr 0.5881 0 TD (m)Tj 1 Tr T* (m)Tj 0 Tr 0.8661 0 TD (a)Tj 1 Tr T* (a)Tj 0 Tr 0.5331 0 TD (n)Tj 1 Tr T* (n)Tj 0 Tr -24.9597 -1.0667 TD (t)Tj 1 Tr 0 0 TD (t)Tj 0 Tr 0.3129 0 TD (r)Tj 1 Tr T* (r)Tj 0 Tr 0.3689 0 TD (a)Tj 1 Tr T* (a)Tj 0 Tr 0.5359 0 TD (c)Tj 1 Tr T* (c)Tj 0 Tr 0.5359 0 TD (h)Tj 1 Tr T* (h)Tj 0 Tr 0.5909 0 TD (e)Tj 1 Tr T* (e)Tj 0 Tr 0.5359 0 TD (o)Tj 1 Tr T* (o)Tj 0 Tr 0.5909 0 TD (b)Tj 1 Tr T* (b)Tj 0 Tr 0.5909 0 TD (r)Tj 1 Tr T* (r)Tj 0 Tr 0.3689 0 TD (o)Tj 1 Tr T* (o)Tj 0 Tr 0.5909 0 TD (n)Tj 1 Tr T* (n)Tj 0 Tr 0.5909 0 TD (c)Tj 1 Tr T* (c)Tj 0 Tr 0.5359 0 TD (h)Tj 1 Tr T* (h)Tj 0 Tr 0.5909 0 TD (i)Tj 1 Tr T* (i)Tj 0 Tr 0.2579 0 TD (a)Tj 1 Tr T* (a)Tj 0 Tr 0.5359 0 TD (l)Tj 1 Tr T* (l)Tj 0 Tr 0.2579 0 TD 0.0938 Tc ( e)Tj 1 Tr 0.3718 0 TD 0 Tc (e)Tj 0 Tr 0.5359 0 TD (p)Tj 1 Tr T* (p)Tj 0 Tr 0.5909 0 TD (i)Tj 1 Tr T* (i)Tj 0 Tr 0.2579 0 TD (t)Tj 1 Tr T* (t)Tj 0 Tr 0.3129 0 TD (h)Tj 1 Tr T* (h)Tj 0 Tr 0.5909 0 TD (e)Tj 1 Tr T* (e)Tj 0 Tr 0.5359 0 TD (l)Tj 1 Tr T* (l)Tj 0 Tr 0.2579 0 TD (i)Tj 1 Tr T* (i)Tj 0 Tr 0.2579 0 TD (a)Tj 1 Tr T* (a)Tj 0 Tr 0.5359 0 TD (l)Tj 1 Tr T* (l)Tj 0 Tr 0.2579 0 TD 0.0938 Tc ( c)Tj 1 Tr 0.3718 0 TD 0 Tc (c)Tj 0 Tr 0.5359 0 TD (e)Tj 1 Tr T* (e)Tj 0 Tr 0.5359 0 TD (l)Tj 1 Tr T* (l)Tj 0 Tr 0.2579 0 TD (l)Tj 1 Tr T* (l)Tj 0 Tr 0.2579 0 TD (s)Tj 1 Tr T* (s)Tj 0 Tr 0.5359 0 TD 0.0938 Tc ( b)Tj 1 Tr 0.3718 0 TD 0 Tc (b)Tj 0 Tr 0.5909 0 TD (y)Tj 1 Tr T* (y)Tj 0 Tr 0.5359 0 TD 0.0938 Tc ( i)Tj 1 Tr 0.3718 0 TD 0 Tc (i)Tj 0 Tr 0.2579 0 TD (n)Tj 1 Tr T* (n)Tj 0 Tr 0.5909 0 TD (f)Tj 1 Tr T* (f)Tj 0 Tr 0.3129 0 TD (l)Tj 1 Tr T* (l)Tj 0 Tr 0.2579 0 TD (a)Tj 1 Tr T* (a)Tj 0 Tr 0.5359 0 TD (m)Tj 1 Tr T* (m)Tj 0 Tr 0.8689 0 TD (m)Tj 1 Tr T* (m)Tj 0 Tr 0.8689 0 TD (a)Tj 1 Tr T* (a)Tj 0 Tr 0.5359 0 TD (t)Tj 1 Tr T* (t)Tj 0 Tr 0.3129 0 TD (o)Tj 1 Tr T* (o)Tj 0 Tr 0.5909 0 TD (r)Tj 1 Tr T* (r)Tj 0 Tr 0.3689 0 TD (y)Tj 1 Tr T* (y)Tj 0 Tr 0.5359 0 TD 0.0938 Tc ( s)Tj 1 Tr 0.3718 0 TD 0 Tc (s)Tj 0 Tr 0.5359 0 TD (t)Tj 1 Tr T* (t)Tj 0 Tr 0.3129 0 TD (i)Tj 1 Tr T* (i)Tj 0 Tr 0.2579 0 TD (m)Tj 1 Tr T* (m)Tj 0 Tr 0.8689 0 TD (u)Tj 1 Tr T* (u)Tj 0 Tr 0.5909 0 TD (l)Tj 1 Tr T* (l)Tj 0 Tr 0.2579 0 TD (i)Tj 1 Tr T* (i)Tj ET 0 0 0 0 k 65 654.3 480.22 -19.05 re 500.701 681.216 m f* BT /F6 1 Tf 11 0 0 11 222.4568 644.7303 Tm 0 Tr 0 0 0 1 k 0.0472 Tc 0.1139 Tw (B. Jany, R. Betz, R. Schreck)Tj /F1 1 Tf 14.506 0 TD 0 Tc 0 Tw (*)Tj ET 0 0 0 0 k 65 619 324 -274.1 re 387.523 644.73 m f* BT /F4 1 Tf 9 0 0 9 66.0036 611.7092 Tm 0 0 0 1 k -0.0026 Tc 0.0951 Tw (Activation of the transcription factor NF-)Tj /F2 1 Tf 16.9507 0 TD 0 Tc 0 Tw ()Tj /F4 1 Tf 0.5466 0 TD -0.0026 Tc 0.0951 Tw (B in human tracheobronchial epithelial cells)Tj -17.4973 -1.1111 TD 0.1025 Tw [(by inflammatory stimuli. B. Jany, R. Betz, R. Schreck. ERS Journals Ltd 1995.)]TJ /F5 1 Tf T* 0.0864 Tw [(ABSTRACT: Recent studies have shown that surface epithelial cells play a major)]TJ T* 0.1863 Tw [(role in the defence and inflammatory reactions of the airways. How extracellu-)]TJ T* 0.2237 Tw (lar stimuli lead to increased gene expression in these epithelial cells is not well)Tj T* 0.0571 Tw [(known. In this study, we asked whether the multiunit transcription factor, nuclear)]TJ T* 0.1959 Tw (factor \(NF\)-)Tj /F2 1 Tf 5.3003 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf 0.5466 0 TD -0.0026 Tc 0.196 Tw (B, which regulates the expression of genes involved in defence and)Tj -5.8469 -1.1111 TD 0.0966 Tw (immune processes, is activated in airway epithelial cells following stimulation with)Tj T* 0.1143 Tw [(inflammatory mediators and hydrogen peroxide. In addition, we studied whether)]TJ T* -0.0082 Tc -0.0169 Tw (this would be followed by upregulation of the NF-)Tj /F2 1 Tf 20.6322 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf 0.541 0 TD -0.0082 Tc -0.0169 Tw (B target gene product granulocyte-)Tj -21.1731 -1.1111 TD -0.0026 Tc 0.1025 Tw (macrophage colony-stimulating factor \(GM-CSF\).)Tj 1 -1.1111 TD 0.167 Tw (Activation of NF-)Tj /F2 1 Tf 7.7328 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf 0.5466 0 TD -0.0026 Tc 0.167 Tw (B in the SV40 transformed human tracheobronchial epithe-)Tj -9.2794 -1.1111 TD 0.1216 Tw [(lial cell line 1HAEo- was measured by electrophoretic mobility shift assays. GM-)]TJ T* 0.051 Tw (CSF concentrations in cell culture supernatants were determined by enzyme-linked)Tj T* 0.1025 Tw (immunosorbent assays.)Tj 1 -1.1111 TD -0.0027 Tc 0 Tw (NF-)Tj /F2 1 Tf 1.658 0 TD 0 Tc ()Tj /F5 1 Tf 0.5466 0 TD -0.0026 Tc 0.046 Tw (B was rapidly activated by exposure of cells to interleukin-1)Tj /F2 1 Tf 25.7279 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf 0.8401 0 TD -0.0026 Tc (\(IL-1)Tj /F2 1 Tf 2.209 0 TD 0 Tc ()Tj /F5 1 Tf 0.5466 0 TD -0.0026 Tc 0.046 Tw (\), phor-)Tj -32.5281 -1.1111 TD 0.1859 Tw (bol myristate acetate \(PMA\), and tumour necrosis factor-)Tj /F2 1 Tf 25.6495 0 TD -0.0024 Tc 0 Tw ()Tj /F5 1 Tf 1.062 0 TD -0.0026 Tc 0.1859 Tw [(\(TNF\). Exposure to)]TJ -26.7115 -1.1111 TD 0 Tc 0 Tw (H)Tj 5.25 0 0 5.25 72.9816 439.4592 Tm (2)Tj 9 0 0 9 75.592 441.7092 Tm (O)Tj 5.25 0 0 5.25 82.57 439.4592 Tm (2)Tj 9 0 0 9 88.2605 441.7092 Tm -0.0026 Tc 0.0947 Tw (platelet activating factor \(PAF\) and lipopolysaccharide \(LPS\) did not lead to)Tj -2.473 -1.1111 TD 0.0126 Tw (increased NF-)Tj /F2 1 Tf 5.9496 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf 0.5466 0 TD -0.0026 Tc 0.0126 Tw [(B activation. Co-stimulation of IL-1)]TJ /F2 1 Tf 15.4672 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf 0.8067 0 TD -0.0026 Tc 0.0125 Tw (with H)Tj 5.25 0 0 5.25 297.1595 429.4592 Tm 0 Tc 0 Tw (2)Tj 9 0 0 9 299.7699 431.7092 Tm (O)Tj 5.25 0 0 5.25 306.7479 429.4592 Tm (2)Tj 9 0 0 9 311.6991 431.7092 Tm -0.0026 Tc 0.0126 Tw (led to augmentation)Tj -27.2995 -1.1111 TD 0.1479 Tw (and prolongation of the effect on NF-)Tj /F2 1 Tf 16.6236 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf 0.5466 0 TD -0.0026 Tc 0.1479 Tw (B activation compared to stimulation with)Tj -17.1702 -1.1111 TD 0 Tw (IL-1)Tj /F2 1 Tf 1.8787 0 TD 0 Tc ()Tj /F5 1 Tf 0.9473 0 TD -0.0026 Tc 0.1533 Tw [(alone. GM-CSF concentrations increased following stimulation with IL-1)]TJ /F2 1 Tf 32.4027 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf -35.2288 -1.1111 TD -0.0026 Tc 0.1121 Tw (and H)Tj 5.25 0 0 5.25 90.6567 399.4592 Tm 0 Tc 0 Tw (2)Tj 9 0 0 9 93.267 401.7092 Tm (O)Tj 5.25 0 0 5.25 100.2451 399.4592 Tm (2)Tj 9 0 0 9 102.8555 401.7092 Tm -0.0026 Tc 0.1121 Tw (, and the effect of IL-1)Tj /F2 1 Tf 9.996 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf 0.5466 0 TD -0.0028 Tc (/H)Tj 5.25 0 0 5.25 207.1949 399.4592 Tm 0 Tc (2)Tj 9 0 0 9 209.8052 401.7092 Tm (O)Tj 5.25 0 0 5.25 216.7833 399.4592 Tm (2)Tj 9 0 0 9 222.6297 401.7092 Tm -0.0026 Tc 0.112 Tw (co-stimulation on GM-CSF concentrations)Tj -17.4029 -1.1111 TD 0.1025 Tw (was additive.)Tj 1 -1.1111 TD -0.003 Tc -0.0221 Tw (These results suggest that NF-)Tj /F2 1 Tf 12.6535 0 TD 0 Tc 0 Tw ()Tj /F5 1 Tf 0.5462 0 TD -0.003 Tc -0.0221 Tw (B may represent an important transcription factor,)Tj -14.1997 -1.1111 TD -0.0026 Tc 0.1025 Tw (controlling the expression of cytokine genes in airway epithelial cells.)Tj /F4 1 Tf T* -0.0025 Tc (Eur Respir J., 1995, 8, 387391.)Tj ET 0 0 0 0 k 407 619 138.22 -230.3 re 187.385 361.709 m f* BT /F1 1 Tf 8 0 0 8 408.0035 612.4082 Tm 0 0 0 1 k 0.0156 Tc 0.2093 Tw (Medizinische Poliklinik der Universitt)Tj 0 -1.125 TD -0.0026 Tc 0.025 Tw [(Wrzburg, Wrzburg, Germany. *Present)]TJ T* -0.0224 Tw (address: Fred Hutchinson Cancer Research)Tj T* -0.0051 Tc -0.02 Tw (Center, 1124 Columbia Street, Seattle, WA)Tj T* -0.0025 Tc 0.1025 Tw (98104, USA.)Tj 0 -2.25 TD -0.0026 Tc (Correspondence: B. Jany)Tj 0 -1.125 TD (Medizinische Poliklinik der Universitt )Tj T* (Klinikstrae 6-8 )Tj T* (97070 Wrzburg )Tj T* 0 Tw (Germany)Tj 0 -2.25 TD 0.1025 Tw (Keywords: Airways )Tj 0 -1.125 TD 0 Tw (asthma)Tj T* 0.1025 Tw (epithelial cells)Tj T* 0 Tw (inflammation )Tj T* 0.1025 Tw (nuclear factor-)Tj /F2 1 Tf 5.921 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1025 Tw (B \(NF-)Tj /F2 1 Tf 2.9479 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0027 Tc (B\))Tj -9.9621 -2.25 TD -0.0026 Tc 0.1025 Tw (Received: April 26 1994)Tj 0 -1.125 TD -0.0121 Tw (Accepted after revision December 11 1994)Tj 0 -2.25 TD 0.1023 Tw (The research was supported by a grant to)Tj 0 -1.125 TD -0.0058 Tc -0.0193 Tw (B.J. from the Deutsche Forschungsgemein-)Tj T* -0.0026 Tc 0.1025 Tw (schaft \(DFG\))Tj ET 0.3 w 398 619 m 398 362.5 l S 0 0 0 0 k 65 339.5 229.31 -254.03 re 451.002 405.408 m f* BT 10 0 0 10 75.0036 331.5103 Tm 0 0 0 1 k 0.114 Tw (Chronic airway diseases, such as asthma and chronic)Tj -0.9 -1.1 TD -0.0166 Tc -0.0085 Tw (bronchitis, are characterized by inflammation of the trache-)Tj T* -0.0026 Tc 0.1045 Tw [(obronchial mucosa. Histologically, the airways show a)]TJ T* 0.0067 Tc 0.2182 Tw (cellular infiltrate with activated eosinophils, lympho-)Tj T* -0.0026 Tc 0.1855 Tw [(cytes and neutrophils [1, 2]. Recent findings indicate)]TJ T* 0.1132 Tw (that in addition to immune cells, airway epithelial cells)Tj T* 0.0219 Tw (are able to act as immune effector cells by secreting pro-)Tj T* 0.1363 Tw (inflammatory mediators. )Tj /F4 1 Tf 10.7317 0 TD (In vivo)Tj /F1 1 Tf 2.8671 0 TD (, an increased number)Tj -13.5988 -1.1 TD 0.0815 Tw (of airway cells in asthmatics express messenger ribonu-)Tj T* 0.1602 Tw (cleic acid \(mRNA\) coding for several interleukins and)Tj T* 0.0035 Tw (granulocyte-macrophage colony-stimulating factor \(GM-)Tj T* 0.0954 Tw [(CSF\) [35]. Cytokines, such as interleukins 5, 6 and 8)]TJ T* 0.0542 Tw (\(IL-5, IL-6 and IL-8\), tumour necrosis factor \(TNF\) and)Tj T* 0.2016 Tw (GM-CSF are released from tracheobronchial epithelial)Tj T* 0.1259 Tw [(cells in culture [6, 7]. An important role for GM-CSF)]TJ T* 0.0115 Tc 0.2134 Tw (in inflammatory airway diseases has been suggested)Tj T* -0.0026 Tc 0.0418 Tw (by studies indicating that this cytokine promotes eosino-)Tj T* -0.0098 Tc -0.0153 Tw (phil [5, 8], neutrophil [9], and macrophage [10] accumula-)Tj T* -0.0026 Tc 0.0299 Tw (tion in the airways, probably by enhancing their survival)Tj T* -0.0025 Tc 0.1025 Tw ([11, 12].)Tj 0.9 -1.1 TD -0.0026 Tc 0.0692 Tw (The intracellular mechanisms leading to the increased)Tj -0.9 -1.1 TD -0.0019 Tc 0.2268 Tw (biosynthesis of proinflammatory cytokines by airway-)Tj T* -0.0099 Tc -0.0152 Tw [(epithelial cells have not been well studied. Some cytokine)]TJ ET 0 0 0 0 k 315.91 339.5 229.31 -253.83 re 293.214 89.51 m f* BT 10 0 0 10 316.9134 331.5103 Tm 0 0 0 1 k -0.0026 Tc 0.1072 Tw (genes, such as GM-CSF, contain recognition sequences)Tj T* -0.0079 Tc -0.0172 Tw (for the multi-unit transcription factor, nuclear factor \(NF\)-)Tj /F2 1 Tf T* 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1444 Tw [(B [13, 14]. Cleavage of an inhibitory \(I\)-)]TJ /F2 1 Tf 17.6519 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0025 Tc 0.1443 Tw (B subunit)Tj -18.745 -1.1 TD -0.0026 Tc 0.0304 Tw (from a cytoplasmic, inactive form leads to nuclear trans-)Tj T* -0.0056 Tc -0.0195 Tw (location, binding of the p50 and p65 subunit to the corres-)Tj T* -0.0183 Tc -0.0068 Tw (ponding cis-acting deoxyribonucleic acid \(DNA\) sequence,)Tj T* -0.0026 Tc 0.0548 Tw (and transcriptional activation of the respective gene [13,)Tj T* 0.0723 Tw [(15]. NF-)]TJ /F2 1 Tf 3.8155 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0723 Tw (B may act as an important second messenger)Tj -4.362 -1.1 TD 0.1458 Tw (system in defence processes by activating transcription)Tj T* 0.0471 Tw (of genes of the acute phase response, immune receptors,)Tj T* 0.1048 Tw (or by inducing the interferon-)Tj /F2 1 Tf 12.1726 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.7607 0 TD -0.0027 Tc (\(IFN-)Tj /F2 1 Tf 2.2637 0 TD 0 Tc ()Tj /F1 1 Tf 0.4084 0 TD -0.0026 Tc 0.1048 Tw [(\) gene [16]. NF-)]TJ /F2 1 Tf -15.6053 -1.1 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1624 Tw (B-inducing factors include TNF-)Tj /F2 1 Tf 13.5439 0 TD -0.0024 Tc 0 Tw ()Tj /F1 1 Tf 1.286 0 TD -0.0027 Tc (IL-1)Tj /F2 1 Tf 1.7664 0 TD 0 Tc ()Tj /F1 1 Tf 0.9565 0 TD -0.0026 Tc 0.1624 Tw ([17], radia-)Tj -18.0994 -1.1 TD 0.0078 Tw (tion [18], lipopolysaccharide \(LPS\), bacteria and exotox-)Tj T* 0.1366 Tw (ins [19], double-stranded ribonucleic acid \(RNA\), viral)Tj T* 0.1021 Tw (proteins, ultraviolet \(UV\) light and oxidative stress \(for)Tj T* 0.0822 Tw [(review see [20, 21]\). The activation of the cytoplasmic)]TJ T* 0.1712 Tw (form of NF-)Tj /F2 1 Tf 5.2018 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1713 Tw (B is probably mediated by reactive oxy-)Tj -5.7484 -1.1 TD 0.1238 Tw [(gen intermediates [22]. The role of NF-)]TJ /F2 1 Tf 16.8147 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1238 Tw (B as a major)Tj -17.3613 -1.1 TD 0.1777 Tw (intracellular transducer of a variety of external signals)Tj T* 0.0536 Tw (has been investigated mainly in immune cells and fibro-)Tj T* 0.0404 Tw [(blasts [1621]. Less is known about the function of this)]TJ T* 0.1025 Tw (transcription factor in normal epithelial cells.)Tj 0.9 -1.1 TD 0.0215 Tw (The tracheobronchial epithelium is constantly exposed)Tj ET Q endstream endobj 3 0 obj << /ProcSet [/PDF /Text ] /Font << /F1 4 0 R /F2 5 0 R /F4 6 0 R /F5 7 0 R /F6 8 0 R /F7 9 0 R >> /ExtGState << /GS2 10 0 R >> >> endobj 13 0 obj << /Length 13331 >> stream q 1 i -1 843 597 -844 re 0 842 m W n 0.015 842.01 594.99 -842.01 re W n 0 0 0 0 k /GS2 gs 50.33 762.01 229.31 -682.74 re 0 842 m f* BT /F1 1 Tf 10 0 0 10 51.328 754.0183 Tm 0 0 0 1 k -0.0026 Tc 0.033 Tw (to a wide spectrum of inhaled potentially noxious agents)Tj 0 -1.1 TD -0.0066 Tc -0.0185 Tw [(that may play a role in the pathogenesis of airway disease.)]TJ T* -0.0039 Tc -0.0212 Tw (We hypothesized that, in tracheobronchial surface epithe-)Tj T* -0.0026 Tc 0.0161 Tw (lial cells, the transcription activator NF-)Tj /F2 1 Tf 15.9662 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0161 Tw (B could rapidly)Tj -16.5128 -1.1 TD 0.0383 Tw (mediate the synthesis of defence and signalling proteins.)Tj T* 0.0773 Tw (Therefore, we investigated whether factors known to be)Tj T* -0.0114 Tc -0.0137 Tw (active in inflammatory airway disease and oxygen radicals)Tj T* 0.0041 Tc 0.2208 Tw [(would lead to activation of NF-)]TJ /F2 1 Tf 13.8121 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5532 0 TD 0.0041 Tc 0.2208 Tw (B in human airway)Tj -14.3653 -1.1 TD -0.0026 Tc 0.0705 Tw [(epithelial cells. In addition, we studied whether this re-)]TJ T* 0.0765 Tw (sults in the release of the cytokine GM-CSF in cell cul-)Tj T* 0.1025 Tw (ture supernatants.)Tj /F5 1 Tf 9.5011 -3.3 TD 0 Tw (Methods)Tj /F4 1 Tf -9.5011 -3.3 TD 0.1025 Tw (Cells and cell culture)Tj /F1 1 Tf 0.9 -2.2 TD -0.0351 Tc 0.0741 Tw (Normal human tracheobronchial epithelial cells \(1HAEo-\))Tj -0.9 -1.1 TD -0.0026 Tc 0.0911 Tw (were transformed with an origin of replication-deficient)Tj T* 0.0638 Tw [(SV 40 virus [23]. The criteria for characteristics of pri-)]TJ T* 0.1257 Tw (mary airway epithelial cells are: 1\) the cells form tight)Tj T* -0.0043 Tw (junctions and respond to )Tj /F2 1 Tf 9.9736 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc -0.0044 Tw (-adrenergic agonists by elevat-)Tj -10.5201 -1.1 TD 0.1625 Tw (ed cyclic adenosine monophosphate \(cAMP\) levels; 2\))Tj T* -0.0138 Tc -0.0113 Tw (phase contrast microscopy shows the typical "cobblestone-)Tj T* -0.0026 Tc 0.0045 Tw (like" appearance of epithelial cells in culture; 3\) cytospin)Tj T* 0.0092 Tc 0.2157 Tw (preparations of these cells shows immunofluorescent)Tj T* -0.0026 Tc 0.1359 Tw (staining with a monoclonal antibody to cytokeratin 19;)Tj T* 0.1813 Tw (and 4\) using the polymerase chain reaction, we found)Tj T* 0.145 Tw (expression of the MUC 2 mucin gene in this cell line,)Tj T* 0.0492 Tw (consistant with its origin from human airway epithelium)Tj T* 0.0791 Tw [(\(data not shown [24]\). Cells were cultured at 37C and)]TJ T* -0.0017 Tc 0.2266 Tw (5% CO)Tj 5.83 0 0 5.83 83.2299 399.5183 Tm -0.0016 Tc 0 Tw (2 )Tj 10 0 0 10 88.9046 402.0183 Tm -0.0017 Tc 0.2266 Tw (in F12/Dulbecco's modified Eagles's medium)Tj -3.7577 -1.1 TD -0.0026 Tc 0.0716 Tw (\(DMEM\) supplemented with 10% foetal calf serum and)Tj T* -0.0025 Tc 0.1436 Tw (100 gml)Tj 5.83 0 0 5.83 93.0965 382.5183 Tm -0.0028 Tc 0 Tw (-1)Tj 10 0 0 10 101.8339 380.0183 Tm -0.0026 Tc 0.1436 Tw (penicillin/streptomycin \(all purchased from)Tj -5.0506 -1.1 TD -0.0009 Tw (Gibco Laboratories\). Tissue culture flasks \(Falcon\) were)Tj T* 0.0773 Tw (coated with a mixture of fibronectin \(Becton Dikinson\),)Tj T* 0.0032 Tc 0.2217 Tw (bovine serum albumin \(Sigma\) and collagen type IV)Tj T* 0.0175 Tc 0.2074 Tw [(\(Serva\) as described previously [25]. Medium was)]TJ T* 0.0066 Tc 0.2183 Tw [(changed every 34 days. Exposure to inflammatory)]TJ T* -0.0026 Tc 0.048 Tw (mediators for up to 48 h did not change the morphology)Tj T* 0.1795 Tw (or growth characteristics of the cells at the concentra-)Tj T* 0.0899 Tw (tions used \(see below\) for electrophoretic mobility shift)Tj T* 0.1551 Tw (assay \(EMSA\) or enzyme-linked immunosorbent assay)Tj T* 0.1909 Tw [(\(ELISA\) experiments. Stimulation of cells with H)]TJ 5.83 0 0 5.83 265.628 267.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 268.5284 270.0184 Tm (O)Tj 5.83 0 0 5.83 275.7227 267.5183 Tm (2)Tj 10 0 0 10 51.328 259.0183 Tm -0.0026 Tc 0.0707 Tw [(up to 100 M did not lead to increased lactate dehydro-)]TJ T* -0.0028 Tc -0.0223 Tw (genase \(LDH\) concentrations in cell culture supernatants,)Tj T* -0.0123 Tc -0.0128 Tw (excluding toxic effects of H)Tj 5.83 0 0 5.83 159.0196 234.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 161.8636 237.0184 Tm (O)Tj 5.83 0 0 5.83 168.9611 234.5183 Tm (2)Tj 10 0 0 10 174.0551 237.0184 Tm -0.0123 Tc -0.0128 Tw (on cell membrane integrity)Tj -12.2727 -1.1 TD -0.0026 Tc 0.1025 Tw (at the concentrations used.)Tj /F4 1 Tf 0 -3.3 TD (Electrophoretic mobility shift assays \(EMSA\))Tj /F1 1 Tf 0.9 -2.2 TD 0.1962 Tw (1HAEo- cells were grown to a confluence of about)Tj -0.9 -1.1 TD 0.0663 Tw (70% in uncoated Petri dishes [24] and incubated for 30,)Tj T* 0.0196 Tw (90 or 180 min with phorbol myristate acetate \(PMA\) \(50)Tj T* 0 Tw (ngml)Tj 5.83 0 0 5.83 74.2599 140.5183 Tm -0.0027 Tc (-1)Tj 10 0 0 10 79.0858 138.0183 Tm -0.0026 Tc 0.092 Tw (; Sigma\), human TNF-)Tj /F2 1 Tf 9.335 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.9681 0 TD -0.0026 Tc 0.092 Tw (\(30 ngml)Tj 5.83 0 0 5.83 221.6959 140.5183 Tm -0.0028 Tc 0 Tw (-1)Tj 10 0 0 10 226.5218 138.0183 Tm -0.0026 Tc 0.092 Tw (; Saxon Bio-)Tj -17.5194 -1.1 TD 0.101 Tw (chemicals\), human IL-1)Tj /F2 1 Tf 9.7257 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.8951 0 TD -0.0026 Tc 0.101 Tw (\(30 ngml)Tj 5.83 0 0 5.83 197.2058 129.5183 Tm -0.0028 Tc 0 Tw (-1)Tj 10 0 0 10 202.0316 127.0183 Tm -0.0026 Tc 0.101 Tw (; Saxon Biochemi-)Tj -15.0704 -1.1 TD 0.0721 Tw (cals\), LPS \(10 gml)Tj 5.83 0 0 5.83 135.4741 118.5183 Tm -0.0028 Tc 0 Tw (-1)Tj 10 0 0 10 140.3 116.0183 Tm -0.0026 Tc 0.0721 Tw (; Sigma\), platelet activating factor)Tj -8.8972 -1.1 TD 0.1282 Tw (\(PAF\) \(1 M; Sigma\) or H)Tj 5.83 0 0 5.83 164.0573 102.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 166.9578 105.0183 Tm (O)Tj 5.83 0 0 5.83 174.152 102.5183 Tm -0.0025 Tc (2 )Tj 10 0 0 10 179.243 105.0183 Tm -0.0026 Tc 0.1282 Tw (\(100 M; Merck\) alone,)Tj -12.7915 -1.1 TD 0.0014 Tc 0.2235 Tw (or co-incubated with H)Tj 5.83 0 0 5.83 150.8261 91.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 153.7499 94.0183 Tm (O)Tj 5.83 0 0 5.83 160.9843 91.5183 Tm (2)Tj 10 0 0 10 168.6581 94.0183 Tm 0.0014 Tc 0.2235 Tw (and either human TNF-)Tj /F2 1 Tf 10.1153 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.6326 0 TD (,)Tj -22.4809 -1.1 TD -0.0026 Tc 0.0331 Tw (human IL-1)Tj /F2 1 Tf 4.7562 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0331 Tw [(, LPS or PAF \(same concentrations\). After)]TJ ET 0 0 0 0 k 301.24 762.01 229.31 -680.01 re 278.614 83.018 m f* BT 10 0 0 10 302.2378 754.0183 Tm 0 0 0 1 k 0.0031 Tc 0.2218 Tw (washing twice with phosphate buffered saline \(PBS\),)Tj 0 -1.1 TD -0.0038 Tc -0.0213 Tw (whole cell extracts were isolated as described by S)Tj 7 0 0 7 501.341 743.0183 Tm -0.0053 Tc 0 Tw (CHRECK)Tj 10 0 0 10 302.2378 732.0183 Tm -0.0025 Tc 0.1074 Tw (and B)Tj 7 0 0 7 326.7961 732.0183 Tm -0.0037 Tc 0 Tw (UERLE)Tj 10 0 0 10 357.7995 732.0183 Tm -0.0026 Tc 0.1075 Tw [([26]. Protein concentrations of the whole)]TJ -5.5562 -1.1 TD -0.0035 Tc -0.0216 Tw [(cell extracts were determined by the Bradford microassay)]TJ T* -0.0026 Tc 0.2029 Tw [(\(Biorad\). The extracts were frozen in liquid nitrogen)]TJ T* -0.0051 Tw [(and stored at -80C. Equal amounts of protein were used)]TJ T* -0.0067 Tc -0.0184 Tw [(for EMSAs. Binding reactions were performed according)]TJ T* -0.0026 Tc 0.1719 Tw (to Z)Tj 7 0 0 7 320.2437 677.0183 Tm -0.0037 Tc 0 Tw (ABEL)Tj /F4 1 Tf 10 0 0 10 342.612 677.0183 Tm -0.0026 Tc 0.1719 Tw (et al.)Tj /F1 1 Tf 2.5759 0 TD 0.1718 Tw ([27] for 30 min on ice with 1015 g)Tj -6.6133 -1.1 TD 0.1863 Tw [(protein in 16 l of 8 mM Tris-HCl \(pH 7.5\), 40 mM)]TJ T* -0.009 Tc -0.0161 Tw (NaCl, 0.4 mM ethylene diamine tetra-acetic acid \(EDTA\),)Tj T* -0.0026 Tc 0.1971 Tw [(3% glycerol, 15 g bovine serum albumin, 2 g poly)]TJ T* 0.1717 Tw (\(dI-dC\) \(Boehringer Mannheim\), 1 mM phenylmethyl-)Tj T* (sulphonylfluoride \(Sigma\) and about 8,500 c.p.m. NF-)Tj /F2 1 Tf T* 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1411 Tw (B oligonucleotide \(Promega\), labelled with T4 kinase)Tj -0.5466 -1.1 TD 0.0039 Tc 0.221 Tw (\(Gibco/BRL\) and \()Tj /F2 1 Tf 8.1179 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.4148 0 TD (-)Tj 5.83 0 0 5.83 390.9329 602.5183 Tm 0.0039 Tc (32)Tj 10 0 0 10 396.809 600.0183 Tm 0.221 Tw (P\)-adenosine triphoshate \(ATP\))Tj -9.4571 -1.1 TD -0.0026 Tc 0.0164 Tw (\(3,000 Cimmol)Tj 5.83 0 0 5.83 365.6624 591.5183 Tm -0.0028 Tc 0 Tw (-1)Tj 10 0 0 10 370.4883 589.0183 Tm -0.0026 Tc 0.0163 Tw [(; Amersham\). The sequence of the NF-)]TJ /F2 1 Tf -6.8251 -1.1 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1025 Tw (B oligonucleotide is as follows: )Tj 2.1299 -1.9 TD (5 -AGTTGAGGGGACTTTCC)Tj ET 0 0 0 1 K 0 J 0 j 0.49 w 10 M []0 d 388.919 557.928 m 455.907 557.928 l S BT 10 0 0 10 455.8817 559.0183 Tm (CAGGC- 3)Tj -12.661 -1.1 TD (3 -TCAACTCCCCTGAAAGGGTCCG- 5)Tj -1.8034 -1.9 TD -0.0133 Tw (DNA-protein complexes were separated from unbound)Tj -0.9 -1.1 TD 0.204 Tw (DNA on native 4.0% polyacrylamide gels in 6.8 mM)Tj T* 0.068 Tw (Tris-HCl \(pH 7.5\), 3.4 mM sodium acetate and 1.0 mM)Tj T* 0.0928 Tw [(EDTA \(pH 8.0\). Gels were vacuum dried and exposed)]TJ T* 0.1025 Tw [(to Kodak \(X-OMAT\) films at -80C for 1024 h.)]TJ /F4 1 Tf 0 -2.6 TD (GM-CSF ELISA)Tj /F1 1 Tf 0.9 -1.9 TD -0.0101 Tc -0.015 Tw (1HAEo- cells were grown in coated tissue culture flasks)Tj -0.9 -1.1 TD -0.027 Tc 0.0019 Tw (to a confluence of about 70%. About 2 h before stimulation,)Tj T* -0.0026 Tc 0.0776 Tw [(culture medium was changed. Cells were incubated for)]TJ T* -0.0025 Tc 0.1112 Tw (30 min, 90 min, 3, 6, 12, 24 or 48 h with either PMA,)Tj T* -0.0026 Tc 0.181 Tw (human IL-1)Tj /F2 1 Tf 4.904 0 TD -0.0024 Tc 0 Tw ()Tj /F1 1 Tf 0.975 0 TD -0.0026 Tc 0.181 Tw (or H)Tj 5.83 0 0 5.83 380.7848 393.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 383.6852 396.0183 Tm (O)Tj 5.83 0 0 5.83 390.8795 393.5183 Tm (2)Tj 10 0 0 10 393.7799 396.0183 Tm -0.0026 Tc 0.181 Tw (, or co-incubated with H)Tj 5.83 0 0 5.83 497.8795 393.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 500.7799 396.0183 Tm (O)Tj 5.83 0 0 5.83 507.9742 393.5183 Tm (2)Tj 10 0 0 10 515.1591 396.0183 Tm -0.0026 Tc (and)Tj -21.2921 -1.1 TD -0.0045 Tc -0.0206 Tw (human IL-1)Tj /F2 1 Tf 4.6832 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.7696 0 TD -0.0045 Tc -0.0206 Tw [(as indicated. After incubation, supernatants)]TJ -5.4528 -1.1 TD -0.0026 Tc 0.0115 Tw (were collected, divided into aliquots and stored at -80C.)Tj T* 0.1101 Tw (The amount of immunoreactive human GM-CSF in the)Tj T* -0.0153 Tc -0.0098 Tw (supernatants was determined by ELISA \(GM-CSF ELISA;)Tj T* -0.0026 Tc 0.1163 Tw [(Biomar, Germany\). All measurements were performed)]TJ T* 0.0017 Tc 0.2232 Tw (in duplicate, according to the manufacturer, using an)Tj T* -0.0026 Tc 0.1025 Tw (ELISA reader \(Labtek\).)Tj /F5 1 Tf 9.8067 -2.6 TD 0 Tw (Results)Tj /F4 1 Tf -9.8067 -2.6 TD 0.1025 Tw (Electrophoretic mobility shift assays)Tj /F1 1 Tf 0.9 -1.9 TD 0.0469 Tw (The strongest activators of NF-)Tj /F2 1 Tf 12.6052 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.047 Tw (B DNA binding were)Tj -14.0518 -1.1 TD 0.0997 Tw (PMA, TNF-)Tj /F2 1 Tf 4.9654 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.9758 0 TD -0.0026 Tc 0.0997 Tw (and IL-1)Tj /F2 1 Tf 3.5499 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0997 Tw [(. Incubation of a radiolabelled)]TJ -10.0377 -1.1 TD -0.0027 Tc 0 Tw (NF-)Tj /F2 1 Tf 1.6031 0 TD 0 Tc ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.22 Tw (B oligonucleotide with cell extracts from treated)Tj -2.1497 -1.1 TD -0.0162 Tc -0.0089 Tw (cells resulted in the retardation of a single specific complex)Tj T* -0.0026 Tc 0.1267 Tw [(\(fig. 1\). This effect occurred rapidly within 30 min of)]TJ T* -0.0005 Tw (stimulation \(fig. 1, lanes 24\). The signal was detectable)Tj T* 0.0868 Tw (after stimulation with PMA and TNF at about the same)Tj T* 0.1652 Tw (intensity after 90 and 180 min \(fig. 1, lanes 2 and 3\).)Tj T* -0.0027 Tc 0 Tw (NF-)Tj /F2 1 Tf 1.6031 0 TD 0 Tc ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.2271 Tw (B activation induced by IL-1)Tj /F2 1 Tf 12.3903 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 1.0212 0 TD -0.0026 Tc 0.2271 Tw (was a strong but)Tj -15.5612 -1.1 TD 0.1659 Tw (transient effect that was lost after 90 min \(fig. 1, lane)Tj T* 0 Tw (4\).)Tj 0.9 -1.1 TD -0.0156 Tc -0.0095 Tw (To ensure the specificity of the DNA-protein interaction,)Tj -0.9 -1.1 TD -0.0026 Tc 0.191 Tw (we performed competition studies with unlabelled oli-)Tj T* -0.0032 Tc -0.0219 Tw [(gonucleotides. Addition of a 50 fold excess of unlabelled)]TJ T* -0.0034 Tc -0.0217 Tw (oligonucleotide resulted in complete disappearance of the)Tj ET 0 0 0 0 k 50.33 789.01 480.22 -21.01 re 529.51 94.018 m f* BT 7.7 0 0 7.7 218.9132 780.3193 Tm 0 0 0 1 k 0 Tc 0 Tw (B)Tj 11 0 0 11 224.8464 780.3193 Tm 0.0725 Tc (. )Tj 7.7 0 0 7.7 233.0689 780.3192 Tm 0.1035 Tc [(J)59.8(ANY)]TJ 11 0 0 11 254.4766 780.3192 Tm 0.0725 Tc (, )Tj 7.7 0 0 7.7 262.699 780.3192 Tm 0 Tc (R)Tj 11 0 0 11 268.6322 780.3191 Tm 0.0725 Tc (. )Tj 7.7 0 0 7.7 276.8547 780.3191 Tm 0.1034 Tc (BETZ)Tj 11 0 0 11 299.2906 780.3191 Tm 0.0725 Tc (, )Tj 7.7 0 0 7.7 307.513 780.3191 Tm 0 Tc (R)Tj 11 0 0 11 313.4463 780.3191 Tm 0.0725 Tc (. )Tj 7.7 0 0 7.7 321.6687 780.3191 Tm 0.1035 Tc (SCHRECK)Tj 9 0 0 9 51.328 779.7092 Tm 0.0225 Tc (388)Tj ET Q endstream endobj 14 0 obj << /ProcSet [/PDF /Text ] /Font << /F1 4 0 R /F2 5 0 R /F4 6 0 R /F5 7 0 R >> /ExtGState << /GS2 10 0 R >> >> endobj 16 0 obj << /Length 22023 >> stream q 1 i -1 843 597 -844 re 0 842 m W n 0.015 842.01 594.99 -842.01 re W n 0 0 0 0 k /GS2 gs 315.91 762.01 229.31 -294.7 re 0 842 m f* BT /F1 1 Tf 10 0 0 10 316.9134 754.0183 Tm 0 0 0 1 k 0.0019 Tc 0.223 Tw (compared to IL-1)Tj /F2 1 Tf 7.4751 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 1.0261 0 TD 0.002 Tc 0.2229 Tw (incubation alone \(compare fig. 1,)Tj -8.5012 -1.1 TD -0.0026 Tc 0.0396 Tw [(lanes 4 and 8\). The activation of NF-)]TJ /F2 1 Tf 15.2441 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0397 Tw (B induced by IL-)Tj -15.7907 -1.1 TD 0 Tc 0 Tw (1)Tj /F2 1 Tf 0.4786 0 TD ()Tj /F1 1 Tf 0.7527 0 TD -0.0215 Tc -0.0036 Tw (together with H)Tj 5.83 0 0 5.83 388.7017 729.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 391.492 732.0183 Tm (O)Tj 5.83 0 0 5.83 398.4973 729.5183 Tm (2)Tj 10 0 0 10 403.5376 732.0183 Tm -0.0215 Tc -0.0036 Tw (was the strongest of all combinations)Tj -8.6624 -1.1 TD -0.0026 Tc 0.1547 Tw [(tested. Taken together, these experiments demonstrate)]TJ T* -0.0139 Tc -0.0112 Tw (that the transcription factor NF-)Tj /F2 1 Tf 12.2154 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5352 0 TD -0.0139 Tc -0.0112 Tw (B is activated by different)Tj -12.7507 -1.1 TD -0.0026 Tc 0.0207 Tw (inflammatory stimuli in tracheobronchial epithelial cells. )Tj 0.9 -1.1 TD -0.0103 Tc -0.0148 Tw (In all experiments, NF-)Tj /F2 1 Tf 9.0514 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5389 0 TD -0.0103 Tc -0.0148 Tw (B activation was clearly depen-)Tj -10.4903 -1.1 TD -0.0026 Tc 0.071 Tw [(dent on the cell confluency. Confluent cultures showed)]TJ T* 0.0727 Tw (lower activation of NF-)Tj /F2 1 Tf 9.5734 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0728 Tw (B DNA-binding compared to a)Tj -10.12 -1.1 TD 0.2192 Tw [(confluency of about 70%. This was determined in a)]TJ T* 0.1025 Tw (series of preliminary experiments \(data not shown\).)Tj /F4 1 Tf 0 -2.6 TD (GM-CSF ELISA)Tj /F1 1 Tf 0.9 -1.8 TD -0.0036 Tc -0.0215 Tw (1HAEo- cells were incubated with IL-1)Tj /F2 1 Tf 15.617 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.7706 0 TD -0.0036 Tc -0.0215 Tw (or H)Tj 5.83 0 0 5.83 507.4818 597.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 510.3764 600.0183 Tm (O)Tj 5.83 0 0 5.83 517.5608 597.5183 Tm (2)Tj 10 0 0 10 522.7055 600.0183 Tm -0.0036 Tc (alone)Tj -20.5792 -1.1 TD -0.0026 Tc -0.0169 Tw (and co-incubated with IL-1)Tj /F2 1 Tf 10.7961 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0027 Tc (/H)Tj 5.83 0 0 5.83 440.2874 586.5183 Tm 0 Tc (2)Tj 10 0 0 10 443.1878 589.0183 Tm (O)Tj 5.83 0 0 5.83 450.3821 586.5183 Tm (2)Tj 10 0 0 10 455.5886 589.0183 Tm -0.0026 Tc -0.0169 Tw (at the same concentra-)Tj -13.8675 -1.1 TD 0.0553 Tw [(tions used in the EMSAs. Immunoreactive human GM-)]TJ T* 0.0751 Tw (CSF was measured in cell culture supernatants up to 48)Tj T* 0.1034 Tw [(h after stimulation by ELISA. Stimulation with IL-1)]TJ /F2 1 Tf 22.1819 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf -22.1819 -1.1 TD -0.0026 Tc 0.0568 Tw (led to GM-CSF concentrations above the detection limit)Tj T* 0.0932 Tw [(as early as 90 min following incubation \(fig. 2\). There)]TJ T* 0.1431 Tw (was a linear increase of GM-CSF concentrations up to)Tj T* 0.0443 Tw [(48 h following stimulation. Incubation with H)]TJ 5.83 0 0 5.83 505.498 509.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 508.3984 512.0183 Tm (O)Tj 5.83 0 0 5.83 515.5928 509.5183 Tm (2)Tj 10 0 0 10 521.4105 512.0183 Tm -0.0026 Tc 0.0442 Tw (led to)Tj -20.4497 -1.1 TD 0.1121 Tw [(increased GM-CSF concentrations after only 6 h. This)]TJ T* -0.0282 Tc 0.0031 Tw (delay compared to IL-1)Tj /F2 1 Tf 8.7963 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.7456 0 TD -0.0283 Tc 0.0033 Tw (lasted up to 12 h. H)Tj 5.83 0 0 5.83 487.3862 487.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 490.1353 490.0183 Tm (O)Tj 5.83 0 0 5.83 497.07 487.5183 Tm (2)Tj 10 0 0 10 502.0693 490.0183 Tm -0.0286 Tc (stimulation)Tj -18.5156 -1.1 TD -0.0026 Tc 0.12 Tw (reached the IL-1)Tj /F2 1 Tf 6.8063 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.9141 0 TD -0.0025 Tc 0.12 Tw [(induced values at 24 and 48 h. Co-)]TJ ET 0 0 0 0 k 65 789 480.22 -21.49 re 544.2 479.018 m f* BT 7.7 0 0 7.7 160.8781 780.3113 Tm 0 0 0 1 k 0.1034 Tc 0 Tw [(A)39.7(CTIV)134.6(A)110.7(TION)-607.3(OF)-607.3(NK)]TJ 11 0 0 11 247.6128 780.3112 Tm 0 Tc (-)Tj /F2 1 Tf 0.4053 0 TD ()Tj /F1 1 Tf 7.7 0 0 7.7 258.358 780.3113 Tm 0.1034 Tc [(B)-607.3(I)0.1(N)-607.3(AIR)54.8(W)119.9(A)104.9(Y)-607.2(EPITHELIUM)-607.3(INFLAMMA)110.7(TION)]TJ 9 0 0 9 530.3182 779.7092 Tm 0.0225 Tc (389)Tj ET 0 0 0 0 k 65 406.4 229.31 -136.39 re 544.426 779.709 m f* BT 8 0 0 8 66.0036 399.8082 Tm 0 0 0 1 k -0.0025 Tc 0.0083 Tw [(Fig. 1. NF-)]TJ /F2 1 Tf 5.6987 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0083 Tw (B activating effects of inflammatory stimuli in 1HAEo-)Tj -6.2453 -1.125 TD 0.1645 Tw [(cells. Cells at 70% confluency were stimulated for 30, 90 and 180)]TJ T* -0.0103 Tc -0.0148 Tw [(min. The figure represents a composite of the three gels after stimulation)]TJ T* -0.0026 Tc 0.1926 Tw [(for the times indicated. Lane 1: unstimulated control cells. Lanes)]TJ T* 0.1024 Tw [(27: incubation with single stimuli. Lanes 811: co-stimulation with)]TJ T* 0 Tc 0 Tw (H)Tj 4.66 0 0 4.66 71.759 352.8082 Tm (2)Tj 8 0 0 8 74.0793 354.8082 Tm (O)Tj 4.66 0 0 4.66 79.8348 352.8082 Tm (2)Tj 8 0 0 8 82.1551 354.8082 Tm -0.0026 Tc 0.1317 Tw [(. Lanes 12 and 13: DNA-protein interaction after addition of a)]TJ -2.0189 -1.125 TD 0.1925 Tw (50 fold excess of unlabelled oligonucleotide to control cell extracts)Tj T* 0.1358 Tw (and IL-1/H)Tj 4.66 0 0 4.66 102.6505 334.8082 Tm 0 Tc 0 Tw (2)Tj 8 0 0 8 104.9708 336.8082 Tm (O)Tj 4.66 0 0 4.66 110.7263 334.8082 Tm (2)Tj 8 0 0 8 113.0466 336.8082 Tm -0.0026 Tc 0.1358 Tw [(-stimulated cell extracts. This competition experiment)]TJ -5.8804 -1.125 TD 0.0187 Tw [(is shown only for 90 min stimulation. Filled arrowheads: retarded NF-)]TJ /F2 1 Tf T* 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1614 Tw (B/DNA complex: open arrowheads: unspecific DNA-binding; open)Tj -0.5466 -1.125 TD 0.2038 Tw (circle: position of unbound DNA \(shown only for one gel, 90 min)Tj T* 0.1509 Tw [(simulation\). NF-)]TJ /F2 1 Tf 7.1747 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1509 Tw (B: nuclear factor-)Tj /F2 1 Tf 7.3077 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1509 Tw (B; IL-1)Tj /F2 1 Tf 3.1046 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1509 Tw (: interleukin-1)Tj /F2 1 Tf 5.8059 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1509 Tw (; DNA)Tj -25.5792 -1.125 TD 0.1955 Tw (deoxyribonucleic acid; PMA: phorbol myristate acetate; TNF-)Tj /F2 1 Tf 25.9611 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.6286 0 TD -0.0026 Tc 0.1955 Tw (: tu-)Tj -26.5897 -1.125 TD 0.0314 Tw (mour necrosis factor-)Tj /F2 1 Tf 8.561 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.6286 0 TD -0.0026 Tc 0.0314 Tw (; LPS: lipopolysaccharide; PAF: platelet-activa-)Tj -9.1896 -1.125 TD 0.1025 Tw (ting factor.)Tj ET 0 0 0 0 k 315.91 465.21 229.31 -280.1 re 101.676 273.808 m f* Q BT /F10 1 Tf 9.322 0 0 9.322 370.2555 211.5171 Tm 0 0 0 1 k /GS3 gs 0 Tc 0 Tw (0)Tj 1.3584 0.0216 TD (30 min)Tj 2.6731 -0.0432 TD (90 min)Tj 3.238 0.0432 TD [(3 h)-1262(6)0( h)-1251.2(12 h)-825.9(24 h)]TJ 9.5928 -0.0072 TD (48 h)Tj -10.6211 -1.6735 TD (Stimulation time)Tj 9.3266 0 0 9.3266 340.3611 430.2143 Tm (350)Tj -0.0166 2.7182 TD (400)Tj 0.0438 -5.4432 TD (300)Tj -0.0602 -2.8796 TD (250)Tj 0.0102 -2.8204 TD (200)Tj 0.0182 -2.7488 TD (150)Tj -0.025 -2.7012 TD (100)Tj 0.4255 -2.9546 TD (50)Tj 0.5017 -2.8238 TD (0)Tj 0 9.3264 -9.3268 0 325.6422 320.3688 Tm (GM-CSF pgml)Tj 0 7.4611 -7.4615 0 322.9561 373.4832 Tm (-1)Tj ET 0 0 0 1 K 0 J 0 j 0.198 w 3.864 M []0 d /GS4 gs 1 i 440.67 256.376 m 443.799 256.376 l 440.67 256.808 m 443.799 256.808 l S BT /F8 1 Tf 3.3425 0 0 5.651 417.3745 252.2367 Tm /GS3 gs ()Tj -7.0059 -0.2245 TD ()Tj ET 0.422 w /GS4 gs 464.933 314.821 m 443.414 268.468 l S 0.375 w 429.253 253.983 m 418.821 253.621 l 428.281 255.69 m 419.257 255.221 l 419.257 255.559 m 430.225 260.957 l 500.058 266.754 m 488.813 262.999 l 511.255 271.901 m 501.863 267.764 l 499.919 357.193 m 489.09 341.702 l 511.859 375.502 m 501.169 359.306 l S BT 3.331 0 0 5.6317 440.6978 253.4323 Tm /GS3 gs ()Tj ET /GS4 gs 362.577 226.037 m 362.577 460.94 l 362.577 460.94 m 360.113 460.94 l 362.577 434.47 m 360.113 434.47 l 362.577 408.181 m 360.113 408.181 l 362.577 381.892 m 360.113 381.892 l 362.577 355.603 m 360.113 355.603 l 362.577 329.784 m 360.113 329.784 l 362.577 303.965 m 360.113 303.965 l 362.577 277.206 m 360.113 277.206 l 362.577 250.448 m 360.113 250.448 l 371.74 226.037 m 371.74 221.994 l 395.342 226.037 m 395.342 221.994 l 418.666 226.037 m 418.666 221.994 l 442.268 226.037 m 442.268 221.994 l 465.593 226.037 m 465.593 221.994 l 488.917 226.037 m 488.917 221.994 l 511.963 226.037 m 511.963 221.994 l 535.565 226.037 m 535.565 221.994 l S BT 3.331 0 0 5.6317 533.9573 343.8523 Tm /GS3 gs ()Tj -7.027 -4.7557 TD ()Tj -6.9908 -7.2521 TD ()Tj -6.9302 -2.5311 TD ()Tj ET /GS4 gs 535.527 442.215 m 535.527 422.681 l S 0.198 w 534.038 442.418 m 536.988 442.418 l 534.002 422.478 m 536.952 422.478 l S 0.375 w 535.426 354.931 m 535.426 330.702 l S 0.198 w 533.901 354.931 m 536.851 354.931 l 533.935 349.448 m 536.886 349.448 l 533.938 330.702 m 536.889 330.702 l 533.917 333.486 m 536.867 333.486 l S 0.375 w 535.426 289.177 m 535.426 275.745 l S 0.198 w 534.021 289.38 m 536.971 289.38 l 533.901 275.745 m 536.851 275.745 l S 0.375 w 512.118 387.76 m 512.118 362.357 l S 0.198 w 510.592 387.963 m 513.542 387.963 l 510.731 362.357 m 513.681 362.357 l S 0.375 w 512.118 330.723 m 512.118 305.554 l S 0.198 w 510.611 330.926 m 513.561 330.926 l 510.712 313.236 m 513.662 313.236 l 510.63 307.305 m 513.58 307.305 l 510.611 305.554 m 513.561 305.554 l S 0.375 w 511.998 276.502 m 511.998 267.53 l S 0.198 w 510.472 267.295 m 513.423 267.295 l 510.472 276.737 m 513.423 276.737 l S 0.375 w 488.794 349.501 m 488.794 333.721 l S 0.198 w 487.388 333.753 m 490.338 333.753 l 487.287 349.703 m 490.237 349.703 l S 0.375 w 488.674 296.219 m 488.674 289.594 l S 0.198 w 487.148 289.594 m 490.098 289.594 l 487.249 296.39 m 490.199 296.39 l S 0.375 w 488.674 282.84 m 488.674 273.163 l S 0.198 w 487.249 273.163 m 490.199 273.163 l 487.129 283.011 m 490.08 283.011 l 486.921 264.787 m 491.041 264.787 l S 0.375 w 465.766 322.774 m 465.766 306.995 l S 0.198 w 464.222 306.76 m 467.172 306.76 l 464.222 322.977 m 467.172 322.977 l S 0.375 w 465.766 287.534 m 465.766 280.439 l S 0.198 w 464.222 280.439 m 467.172 280.439 l 464.222 287.737 m 467.172 287.737 l 464.083 255.723 m 468.171 255.723 l 464.064 259.811 m 468.171 259.811 l 463.941 261.374 m 467.211 261.374 l 464.241 265.855 m 467.191 265.855 l 440.56 253.348 m 443.689 253.348 l 440.639 274.412 m 443.589 274.412 l S 0.375 w 442.025 274.155 m 442.025 262.835 l S 0.198 w 440.5 262.601 m 443.45 262.601 l S 0.375 w 418.821 252.945 m 418.821 251.569 l S 0.198 w 417.371 251.2 m 420.322 251.2 l 393.772 251.003 m 397.279 251.003 l 393.736 250.025 m 397.224 250.025 l 393.764 256.824 m 396.98 256.824 l S 0.375 w 395.446 252.735 m 395.446 250.16 l S 0.198 w 370.609 252.241 m 373.56 252.241 l 370.628 256.461 m 373.579 256.461 l S 0.375 w 373.111 254.297 m 394.388 254.524 l 396.347 252.436 m 418.146 260.253 l 396.036 254.673 m 418.821 254.735 l 442.685 272.042 m 465.072 283.725 l 443.084 256.879 m 465.488 257.85 l 442.025 255.089 m 465.072 262.835 l 466.148 263.827 m 488.674 277.153 l 466.738 257.544 m 488.985 262.409 l 466.148 284.248 m 488.257 292.88 l 488.361 341.52 m 466.044 315.413 l 513.094 272.104 m 534.906 282.285 l 535.287 432.357 m 512.553 375.502 l 513.126 311.804 m 534.906 337.946 l 512.414 319.168 m 534.767 344.284 l S 0.198 w 440.55 258.569 m 443.678 258.569 l 440.448 254.855 m 443.577 254.855 l 440.531 269.347 m 443.659 269.347 l 417.188 259.152 m 421.444 259.152 l 417.207 262.608 m 421.211 262.608 l 417.188 257.403 m 420.316 257.403 l S 0 w 416.91 253.681 m 420.038 253.681 l 417.087 254.126 m 420.215 254.126 l S 0.198 w 440.411 271.989 m 443.362 271.989 l S 0.375 w 395.932 253.212 m 418.821 252.945 l 396.452 253.2 m 418.285 254.385 l 417.763 253.553 m 419.88 253.553 l S 0 0 0 0 k 0.353 w 440.776 268.536 m 443.414 268.536 l 442.095 264.645 l 440.776 268.536 l b* 0.375 w 362.577 226.037 m 428.975 226.037 l 431.752 226.219 m 498.948 226.219 l S 0.353 w 464.319 316.419 m 466.957 316.419 l 465.638 312.529 l 464.319 316.419 l b* 487.483 343.647 m 490.121 343.647 l 488.802 339.756 l 487.483 343.647 l b* 510.748 376.977 m 513.386 376.977 l 512.067 373.087 l 510.748 376.977 l b* 534.038 433.129 m 536.676 433.129 l 535.357 429.238 l 534.038 433.129 l b* 0.375 w 499.295 362.24 m 503.043 355.902 l 498.153 360.362 m 501.901 354.025 l 499.518 305.437 m 503.267 299.1 l 498.377 303.559 m 502.125 297.222 l 500.074 270.933 m 503.822 264.596 l 498.932 269.055 m 502.681 262.718 l 511.026 317.291 m 502.002 300.625 l 501.308 298.982 m 489.772 277.857 l 511.443 309.545 m 501.308 302.268 l 499.642 301.262 m 489.368 292.812 l 499.015 230.06 m 502.764 223.722 l 497.873 228.182 m 501.622 221.844 l 501.308 226.219 m 535.565 226.219 l 429.252 229.861 m 433 223.523 l 428.11 227.983 m 431.858 221.645 l 428.713 267.647 m 432.461 261.31 l 427.571 265.769 m 431.32 259.432 l 428.671 258.767 m 432.419 252.43 l 427.529 256.889 m 431.277 250.552 l 441.054 271.05 m 429.947 266.121 l 428.003 264.948 m 419.673 260.957 l 432.029 261.896 m 441.054 267.06 l 440.967 256.879 m 430.502 255.794 l 440.967 255.089 m 431.335 253.916 l S 0.198 w 393.672 252.138 m 397.16 252.138 l 393.686 252.559 m 397.174 252.559 l 417.339 251.636 m 420.289 251.636 l 440.67 256.073 m 443.799 256.073 l 440.707 256.376 m 443.836 256.376 l S 0.466 w 370.164 254.451 m 370.164 255.513 371.025 256.373 372.087 256.373 c 373.149 256.373 374.01 255.513 374.01 254.451 c 374.01 253.389 373.149 252.528 372.087 252.528 c 371.025 252.528 370.164 253.389 370.164 254.451 c b 393.528 254.742 m 393.528 255.804 394.389 256.665 395.45 256.665 c 396.512 256.665 397.373 255.804 397.373 254.742 c 397.373 253.68 396.512 252.819 395.45 252.819 c 394.389 252.819 393.528 253.68 393.528 254.742 c b 416.875 255.265 m 416.875 256.327 417.736 257.187 418.798 257.187 c 419.86 257.187 420.72 256.327 420.72 255.265 c 420.72 254.203 419.86 253.342 418.798 253.342 c 417.736 253.342 416.875 254.203 416.875 255.265 c b 0 0 0 1 k 417.764 262.432 3.048 -3.048 re B 440.609 273.18 3.048 -3.048 re B 440.147 256.548 m 440.147 257.61 441.007 258.471 442.069 258.471 c 443.131 258.471 443.992 257.61 443.992 256.548 c 443.992 255.486 443.131 254.626 442.069 254.626 c 441.007 254.626 440.147 255.486 440.147 256.548 c s 0 0 0 0 k 464.16 257.655 m 464.16 258.717 465.02 259.578 466.082 259.578 c 467.144 259.578 468.005 258.717 468.005 257.655 c 468.005 256.593 467.144 255.733 466.082 255.733 c 465.02 255.733 464.16 256.593 464.16 257.655 c b 486.995 262.476 m 486.995 263.538 487.856 264.399 488.918 264.399 c 489.979 264.399 490.84 263.538 490.84 262.476 c 490.84 261.414 489.979 260.554 488.918 260.554 c 487.856 260.554 486.995 261.414 486.995 262.476 c b 0 0 0 1 k 464.083 285.514 3.047 -3.048 re B 0 0 0 0 k 510.12 271.871 m 510.12 272.933 510.981 273.794 512.042 273.794 c 513.104 273.794 513.965 272.933 513.965 271.871 c 513.965 270.809 513.104 269.949 512.042 269.949 c 510.981 269.949 510.12 270.809 510.12 271.871 c b 0 0 0 1 k 487.156 294.37 3.047 -3.048 re B 0 0 0 0 k 533.591 282.475 m 533.591 283.537 534.452 284.398 535.513 284.398 c 536.575 284.398 537.436 283.537 537.436 282.475 c 537.436 281.413 536.575 280.552 535.513 280.552 c 534.452 280.552 533.591 281.413 533.591 282.475 c b 0 0 0 1 k 510.519 311.965 3.047 -3.048 re B 534.057 339.42 3.048 -3.047 re B 0.198 w 486.79 260.567 m 490.911 260.567 l S q -1 843 597 -844 re 0 842 m W n 0.015 842.01 594.99 -842.01 re W n 0 0 0 0 k /GS1 gs 315.91 187 229.31 -109.13 re f* BT /F1 1 Tf 8 0 0 8 316.9134 180.4082 Tm 0 0 0 1 k /GS2 gs 0.0224 Tc 0.1192 Tw [(Fig. 2. Effect of IL-1)]TJ /F2 1 Tf 11.0542 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5716 0 TD 0.0225 Tc 0.1192 Tw (, H)Tj 4.66 0 0 4.66 421.1886 178.4082 Tm 0 Tc 0 Tw (2)Tj 8 0 0 8 423.6255 180.4082 Tm (O)Tj 4.66 0 0 4.66 429.5809 178.4082 Tm (2)Tj 8 0 0 8 435.1515 180.4082 Tm 0.0224 Tc 0.1192 Tw (and combination of IL-1)Tj /F2 1 Tf 10.6771 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5716 0 TD 0.0223 Tc (/H)Tj 4.66 0 0 4.66 533.499 178.4082 Tm 0 Tc (2)Tj 8 0 0 8 535.9359 180.4082 Tm (O)Tj 4.66 0 0 4.66 541.8914 178.4082 Tm (2)Tj 8 0 0 8 316.9134 171.4082 Tm 0.0224 Tc 0.1179 Tw [(on the secretion of GM-CSF by 1HAEo-cells. Cells were grown)]TJ 0 -1.125 TD 0.1036 Tw (to a confluence of 70% and incubated with 30 ngml)Tj 4.66 0 0 4.66 500.8683 164.4082 Tm 0.023 Tc 0 Tw (-1)Tj 8 0 0 8 507.9705 162.4082 Tm 0.0224 Tc 0.1035 Tw (human IL-)Tj -23.8821 -1.125 TD 0 Tc 0 Tw (1)Tj /F2 1 Tf 0.5225 0 TD ()Tj /F1 1 Tf 0.5716 0 TD 0.0225 Tc 0.1828 Tw [(, 100 M H)]TJ 4.66 0 0 4.66 368.7404 151.4082 Tm 0 Tc 0 Tw (2)Tj 8 0 0 8 371.1773 153.4082 Tm (O)Tj 4.66 0 0 4.66 377.1327 151.4082 Tm (2)Tj 8 0 0 8 383.2122 153.4082 Tm 0.0224 Tc 0.1828 Tw (or combined IL-1)Tj /F2 1 Tf 7.7781 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5716 0 TD 0.0223 Tc (/H)Tj 4.66 0 0 4.66 458.3679 151.4082 Tm 0 Tc (2)Tj 8 0 0 8 460.8048 153.4082 Tm (O)Tj 4.66 0 0 4.66 466.7602 151.4082 Tm (2)Tj 8 0 0 8 472.8397 153.4082 Tm 0.0224 Tc 0.1828 Tw (using the same con-)Tj -19.4908 -1.125 TD 0.0193 Tc -0.0194 Tw [(centrations. At the different time-points indicated, cell supernatants)]TJ T* 0.0224 Tc 0.0601 Tw [(were tested for human GM-CSF by ELISA. Data points represent)]TJ T* 0.0364 Tc 0.2135 Tw (the median)Tj 5.6 0 0 5.6 361.6659 126.4082 Tm 0.052 Tc 0 Tw (SD)Tj 8 0 0 8 373.4059 126.4082 Tm 0.0364 Tc 0.2135 Tw [(of three independent sets of experiments. All)]TJ -7.0616 -1.125 TD 0.0224 Tc 0.0158 Tw [(measurements were performed in duplicate. IL-1)]TJ /F2 1 Tf 20.9471 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5716 0 TD 0.0224 Tc 0.0158 Tw (: interleukin-1)Tj /F2 1 Tf 6.0457 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5716 0 TD (;)Tj -28.1359 -1.125 TD 0.0033 Tc -0.0034 Tw (GM-CSF: granulocyte-macrophage colony-stimulating factor; ELISA:)Tj T* 0.037 Tc 0.2129 Tw [(enzyme-linked immunosorbent assay. )]TJ /F2 1 Tf 7 0 0 7 475.472 99.4082 Tm 0 Tc 0 Tw ()Tj /F1 1 Tf 8 0 0 8 495.7216 99.4082 Tm 0.0369 Tc 0.2129 Tw (: IL-1)Tj /F2 1 Tf 2.7395 0 TD 0.0372 Tc 0 Tw ( )Tj /F1 1 Tf 0.9013 0 TD 0 Tc (H)Tj 4.66 0 0 4.66 530.9194 97.4082 Tm (2)Tj 8 0 0 8 533.4241 99.4082 Tm (O)Tj 4.66 0 0 4.66 539.496 97.4082 Tm (2)Tj 8 0 0 8 542.0009 99.4082 Tm 0.0369 Tc (; )Tj /F8 1 Tf 5 0 0 5 331.9134 90.4082 Tm 0 Tc ()Tj /F1 1 Tf 8 0 0 8 350.8557 90.4082 Tm 0.0224 Tc 0.1025 Tw (: H)Tj 4.66 0 0 4.66 362.2137 88.4082 Tm 0 Tc 0 Tw (2)Tj 8 0 0 8 364.6506 90.4082 Tm (O)Tj 4.66 0 0 4.66 370.606 88.4082 Tm (2)Tj 8 0 0 8 373.0429 90.4082 Tm 0.125 Tc [(;)102.5( )]TJ /F8 1 Tf 5 0 0 5 390.4456 90.4082 Tm 0 Tc ()Tj /F1 1 Tf 8 0 0 8 409.4554 90.4082 Tm 0.0223 Tc 0.1025 Tw (: IL-1)Tj /F2 1 Tf 2.5417 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5716 0 TD 0.125 Tc [(;)102.5( )]TJ /F8 1 Tf 5 0 0 5 454.7642 90.4082 Tm 0 Tc ()Tj /F1 1 Tf 8 0 0 8 474.3332 90.4082 Tm 0.0224 Tc 0.1025 Tw (: control.)Tj ET 0 0 0 0 k 65 263.6 229.31 -184.89 re 505.833 90.408 m f* BT 10 0 0 10 66.0036 255.6103 Tm 0 0 0 1 k -0.0026 Tc 0.1567 Tw (retarded DNA-protein complex in unstimulated control)Tj 0 -1.1 TD 0.1655 Tw [(cells \(fig. 1, 90 min, lane 12\). A faint band was still)]TJ T* 0.0401 Tw (visible when a 50 fold excess of unlabelled oligonucleo-)Tj T* 0.2199 Tw (tide was used in cell extracts of IL-1/H)Tj 5.83 0 0 5.83 236.16 220.1102 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 239.0604 222.6103 Tm (O)Tj 5.83 0 0 5.83 246.2547 220.1102 Tm -0.0025 Tc (2 )Tj 10 0 0 10 251.8803 222.6103 Tm -0.0026 Tc (stimulated)Tj -18.5877 -1.1 TD 0.0925 Tw [(cells \(fig. 1, 90 min, lane 13\). Independent of the time)]TJ T* 0.1935 Tw (of incubation, LPS \(10 gml)Tj 5.83 0 0 5.83 189.6943 203.1103 Tm -0.0028 Tc 0 Tw (-1)Tj 10 0 0 10 194.5201 200.6102 Tm -0.0026 Tc 0.1935 Tw (\) alone did not activate)Tj -12.8516 -1.1 TD -0.0027 Tc 0 Tw (NF-)Tj /F2 1 Tf 1.6031 0 TD 0 Tc ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0569 Tw [(B \(fig 1, lane 5\). Co-stimulation of LPS and H)]TJ 5.83 0 0 5.83 280.3037 187.1102 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 283.2042 189.6103 Tm (O)Tj 5.83 0 0 5.83 290.3984 187.1102 Tm (2)Tj 10 0 0 10 66.0036 178.6102 Tm -0.0026 Tc -0.0059 Tw (resulted in a transient NF-)Tj /F2 1 Tf 10.349 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc -0.0059 Tw (B activation after 90 min \(fig.)Tj -10.8956 -1.1 TD 0.192 Tw [(1, lane 9\). The lipid mediator PAF had a minor and)]TJ T* 0.0879 Tw [(transient effect \(fig. 1, 30 min, lane 7\). When the cells)]TJ T* 0.0209 Tw (were co-stimulated with PAF and hydrogen peroxide, no)Tj T* -0.008 Tw (activation of DNA-binding could be detected \(fig. 1, lane)Tj T* 0.1573 Tw [(11\). Incubation with H)]TJ 5.83 0 0 5.83 165.3056 121.1102 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 168.206 123.6103 Tm (O)Tj 5.83 0 0 5.83 175.4003 121.1102 Tm (2)Tj 10 0 0 10 182.3487 123.6103 Tm -0.0026 Tc 0.1573 Tw (alone at the concentrations)Tj -11.6345 -1.1 TD 0.1651 Tw [(tested \(up to 100 M\) had no effect. Interestingly, in)]TJ T* 0.1237 Tw (the case of IL-1)Tj /F2 1 Tf 6.6325 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1237 Tw (, co-stimulation with H)Tj 5.83 0 0 5.83 233.69 99.1102 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 236.5905 101.6102 Tm (O)Tj 5.83 0 0 5.83 243.7848 99.1102 Tm (2)Tj 10 0 0 10 250.3973 101.6102 Tm -0.0026 Tc 0.1237 Tw (resulted in)Tj -18.4394 -1.1 TD 0.0026 Tc 0.2223 Tw (a markedly prolonged and stronger NF-)Tj /F2 1 Tf 17.0672 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5518 0 TD 0.0026 Tc 0.2223 Tw (B activation)Tj ET 0.3 w 10 M 325.63 92.51 m 343.37 92.51 l 384.13 92.51 m 401.87 92.51 l 448.5 92.51 m 466.25 92.51 l 476 103 m 467.5 103 l 487.75 102.05 m 479.25 102.05 l S 0 0 0 0 k 65 762.01 229.31 -349.01 re 293.34 90.61 m f* Q q 65 762.01 229.31 -349.01 re W* n /GS1 gs q 229.3171 0 0 349.0115 65 412.9985 cm /Im1 Do Q Q endstream endobj 17 0 obj << /ProcSet [/PDF /Text /ImageC ] /Font << /F1 4 0 R /F2 5 0 R /F4 6 0 R /F8 18 0 R /F10 19 0 R >> /XObject << /Im1 20 0 R >> /ExtGState << /GS1 21 0 R /GS2 10 0 R /GS3 22 0 R /GS4 23 0 R >> >> endobj 25 0 obj << /Length 15131 >> stream q 1 i -1 843 597 -844 re 0 842 m W n 0.015 842.01 594.99 -842.01 re W n 0 0 0 0 k /GS2 gs 50.33 762.01 229.31 -679.54 re 0 842 m f* BT /F1 1 Tf 10 0 0 10 51.328 754.0183 Tm 0 0 0 1 k -0.0026 Tc 0.1594 Tw (stimulation with IL-1)Tj /F2 1 Tf 8.8202 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0027 Tc (/H)Tj 5.83 0 0 5.83 154.9431 751.5183 Tm 0 Tc (2)Tj 10 0 0 10 157.8435 754.0183 Tm (O)Tj 5.83 0 0 5.83 165.0378 751.5183 Tm (2)Tj 10 0 0 10 172.0075 754.0183 Tm -0.0026 Tc 0.1594 Tw (showed an additive effect)Tj -12.0679 -1.1 TD 0.1025 Tw [(after 6 h, that continued up to 48 h of cell culture. )]TJ /F5 1 Tf 9.7405 -3.3 TD 0 Tw (Discussion)Tj /F1 1 Tf -8.8405 -2.2 TD 0.0426 Tw (In this report we show that exposure of human airway)Tj -0.9 -1.1 TD 0.168 Tw (epithelial cells )Tj /F4 1 Tf 6.3466 0 TD (in vitro)Tj /F1 1 Tf 3.4799 0 TD (to inflammatory stimuli known)Tj -9.8265 -1.1 TD -0.0148 Tw (to be active in inflammatory chronic airway disease leads)Tj T* 0.1521 Tw (to the activation and DNA-binding of the transcription)Tj T* -0.0126 Tw (factor NF-)Tj /F2 1 Tf 4.1541 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc -0.0126 Tw [(B. This is followed by an increased secretion)]TJ -4.7007 -1.1 TD 0.1025 Tw (of the NF-)Tj /F2 1 Tf 4.3451 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1025 Tw (B responsive gene product GM-CSF.)Tj -3.9917 -1.1 TD 0.1148 Tw (Firstly, we focused on the possible NF-)Tj /F2 1 Tf 16.3372 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1148 Tw (B activating)Tj -17.7838 -1.1 TD 0.1771 Tw (role of the inflammatory mediators IL-1)Tj /F2 1 Tf 16.8087 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.9712 0 TD -0.0026 Tc 0.1771 Tw (and TNF-)Tj /F2 1 Tf 4.0724 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.6286 0 TD (,)Tj -22.481 -1.1 TD -0.0026 Tc 0.1043 Tw (since IL-1 receptors have been demonstrated on human)Tj T* 0.1676 Tw [(airway epithelial cells [6]. IL-1 and TNF may play a)]TJ T* -0.0035 Tc -0.0216 Tw (key role in allergic airway disease; a selective IL-1 recep-)Tj T* -0.0026 Tc 0.1452 Tw (tor antagonist is able to prevent airway hyperreactivity)Tj T* 0.0295 Tw (and eosinophil accumulation in a guinea-pig model [28].)Tj T* 0.0849 Tw (In our model, both cytokines lead to a strong activation)Tj T* -0.0032 Tc -0.0219 Tw (of NF-)Tj /F2 1 Tf 2.6532 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.546 0 TD -0.0031 Tc -0.022 Tw [(B in human airway epithelial cells. This supports)]TJ -3.1993 -1.1 TD -0.0037 Tc -0.0214 Tw (the role of these factors in the regulation of the inflamma-)Tj T* -0.0027 Tc -0.0032 Tw (tory response of epithelial cells. In addition, NF-)Tj /F2 1 Tf 19.559 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc -0.0033 Tw (B may)Tj -20.1056 -1.1 TD 0.0946 Tw (represent an important trans-acting factor in these cells,)Tj T* 0.0515 Tw [(leading to upregulation of responsive genes. In fact, we)]TJ T* 0.0973 Tw (were able to show that one NF-)Tj /F2 1 Tf 13.0596 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0973 Tw (B target gene product,)Tj -13.6062 -1.1 TD 0.2147 Tw (GM-GSF, is released by these cells upon stimulation,)Tj T* -0.0098 Tc -0.0153 Tw (following the activation of NF-)Tj /F2 1 Tf 12.1347 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5394 0 TD -0.0097 Tc -0.0154 Tw [(B. This strongly suggests)]TJ -12.6741 -1.1 TD -0.0026 Tc 0.0821 Tw (that NF-)Tj /F2 1 Tf 3.4223 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0821 Tw (B activation is one of the cytoplasmic-nuclear)Tj -3.9689 -1.1 TD 0.0004 Tc 0.2245 Tw (signalling pathways used in airway epithelial cells in)Tj T* -0.0026 Tc 0.1025 Tw (response to inflammatory stimuli.)Tj 0.9 -1.1 TD 0.0086 Tc 0.2163 Tw (In contrast to Jurkat T-cells, mouse fibroblast and)Tj -0.9 -1.1 TD -0.0026 Tc 0.2082 Tw (mouse pre-B-cells [21], LPS and H)Tj 5.83 0 0 5.83 201.9757 399.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 204.8762 402.0183 Tm (O)Tj 5.83 0 0 5.83 212.0704 399.5183 Tm -0.0025 Tc (2 )Tj 10 0 0 10 217.6278 402.0183 Tm -0.0026 Tc 0.2082 Tw (alone failed to)Tj -16.63 -1.1 TD 0.1684 Tw (activate NF-)Tj /F2 1 Tf 5.108 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1685 Tw [(B in human airway epithelial cells. The)]TJ -5.6546 -1.1 TD 0.0002 Tw (reason for this is unclear. It is conceivable that in airway)Tj T* 0.1531 Tw (epithelial cells higher levels of enzymes regulating the)Tj T* 0.2143 Tw (intracellular concentrations of reactive oxygen interm-)Tj T* -0.0154 Tc -0.0097 Tw [(ediates, such as superoxide dismutase [29], the glutathione)]TJ T* -0.0026 Tc 0.2061 Tw (\(reduced form\) GSH peroxidase/GSH system, catalase)Tj T* -0.0147 Tw [(or peroxidases, prevent H)]TJ 5.83 0 0 5.83 155.3194 322.5184 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 158.2198 325.0183 Tm (O)Tj 5.83 0 0 5.83 165.4141 322.5184 Tm (2)Tj 10 0 0 10 170.6423 325.0183 Tm -0.0026 Tc -0.0147 Tw (and other resulting reactive)Tj -11.9314 -1.1 TD 0.0077 Tw (oxygen intermediates from activating the latent cytoplas-)Tj T* 0.1794 Tw (mic form of NF-)Tj /F2 1 Tf 7.1369 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1794 Tw (B compared to cells of the immune)Tj -7.6835 -1.1 TD 0.1208 Tw [(system. Alternatively, longer exposure times or higher)]TJ T* 0.0846 Tw (concentrations of H)Tj 5.83 0 0 5.83 131.3992 278.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 134.2996 281.0183 Tm (O)Tj 5.83 0 0 5.83 141.4939 278.5183 Tm (2)Tj 10 0 0 10 147.7157 281.0183 Tm -0.0026 Tc 0.0846 Tw (may be required to activate NF-)Tj /F2 1 Tf -9.6388 -1.1 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0025 Tc (B.)Tj 0.3534 -1.1 TD -0.0082 Tc -0.0169 Tw (Although PAF has a wide spectrum of proinflammatory)Tj -0.9 -1.1 TD -0.0026 Tc 0.1321 Tw (effects and may play a role in mediating the effects of)Tj T* -0.0092 Tw (ozone in the airways [30], this mediator had a very minor)Tj T* -0.0279 Tc 0.0028 Tw (effect on NF-)Tj /F2 1 Tf 5.0303 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5213 0 TD -0.0277 Tc 0.0026 Tw (B activation. A combination of inflammatory)Tj -5.5516 -1.1 TD -0.0026 Tc 0.0538 Tw (mediators different to those used in our experiment may)Tj T* 0.0068 Tc 0.2181 Tw (lead to activation of NF-)Tj /F2 1 Tf 10.8749 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.556 0 TD 0.0068 Tc 0.2181 Tw (B, since cytokine-cytokine)Tj -11.4309 -1.1 TD -0.0026 Tc 0.2021 Tw (interaction, like the synergy caused by IL-1 and TNF)Tj T* -0.0052 Tc -0.0199 Tw ([31], plays an important role in several biological systems)Tj T* -0.0026 Tc 0 Tw ([32].)Tj 0.9 -1.1 TD 0.0902 Tw (LPS co-stimulation with H)Tj 5.83 0 0 5.83 169.8946 157.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 172.795 160.0183 Tm (O)Tj 5.83 0 0 5.83 179.9893 157.5183 Tm (2)Tj 10 0 0 10 186.2664 160.0183 Tm -0.0026 Tc 0.0902 Tw (led to transient NF-)Tj /F2 1 Tf 8.0236 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD (B)Tj -22.064 -1.1 TD -0.0026 Tc 0.1153 Tw [(DNA-binding after 90 min. In pre-B-cells, PMA leads)]TJ T* 0.1035 Tw (to a much faster NF-)Tj /F2 1 Tf 8.638 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1035 Tw (B activation than LPS, similar to)Tj -9.1845 -1.1 TD 0.1308 Tw [(the kinetics in our experiments [17]. Interestingly, co-)]TJ T* 0.0974 Tw (stimulation with H)Tj 5.83 0 0 5.83 127.8185 113.5183 Tm 0 Tc 0 Tw (2)Tj 10 0 0 10 130.7189 116.0183 Tm (O)Tj 5.83 0 0 5.83 137.9132 113.5183 Tm (2)Tj 10 0 0 10 144.2621 116.0183 Tm -0.0026 Tc 0.0973 Tw (and IL-1)Tj /F2 1 Tf 3.5476 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.8914 0 TD -0.0026 Tc 0.0974 Tw (led to a longer lasting)Tj -13.7324 -1.1 TD 0.1346 Tw (effect compared to IL-1)Tj /F2 1 Tf 9.8678 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.9287 0 TD -0.0026 Tc 0.1347 Tw [(alone. This may be relevant)]TJ -10.7965 -1.1 TD 0.113 Tw (to the )Tj /F4 1 Tf 2.708 0 TD (in vivo)Tj /F1 1 Tf 3.1494 0 TD (situation, since in the presence of airway)Tj ET 0 0 0 0 k 301.24 762.01 229.31 -671.54 re 278.614 94.018 m f* BT 10 0 0 10 302.2378 754.0183 Tm 0 0 0 1 k 0.058 Tw (inflammation, the additional appearance of oxygen radi-)Tj 0 -1.1 TD 0.1488 Tw (cals from any source may lead to prolonged and addi-)Tj T* 0.1329 Tw (tive inflammatory effects, mediated in part through the)Tj T* 0.0945 Tw (transcription factor NF-)Tj /F2 1 Tf 9.6243 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0946 Tw [(B. How this synergistic effect)]TJ -10.1709 -1.1 TD 0.1985 Tw [(is mediated, remains unknown. The variations of the)]TJ T* 0.1308 Tw (effect of the different agents on NF-)Tj /F2 1 Tf 15.1286 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1308 Tw (B activation may)Tj -15.6752 -1.1 TD 0.0435 Tw (reflect qualitative or quantitative differences in the com-)Tj T* 0.1025 Tw (ponents constituting the signalling pathway.)Tj 0.9 -1.1 TD 0.0612 Tw (GM-CSF may play an important role in inflammatory)Tj -0.9 -1.1 TD -0.0075 Tc -0.0176 Tw [(airway diseases [5]. In addition, GM-CSF may be involv-)]TJ T* -0.005 Tc -0.0201 Tw (ed in the inflammatory changes in human airways follow-)Tj T* -0.0026 Tc 0.0508 Tw [(ing ozone exposure [33]. A role for GM-CSF in airway)]TJ T* 0.1061 Tw (disease has been further emphasized by a number of )Tj /F4 1 Tf 21.9557 0 TD -0.0027 Tc 0 Tw (in)Tj -21.9557 -1.1 TD -0.0026 Tc (vitro)Tj /F1 1 Tf 2.135 0 TD 0.0116 Tw [(studies [7, 9, 10, 33]. We found that human airway)]TJ -2.135 -1.1 TD 0.2169 Tw (epithelial cells release the NF-)Tj /F2 1 Tf 12.9482 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.217 Tw (B target gene product)Tj -13.4947 -1.1 TD 0.0397 Tw (GM-CSF into the culture medium upon stimulation with)Tj T* -0.0027 Tc 0 Tw (IL-1)Tj /F2 1 Tf 1.7664 0 TD 0 Tc ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0526 Tw [(. This effect was additive when the cells were co-)]TJ -2.313 -1.1 TD -0.011 Tc -0.0141 Tw (stimulated with IL-1)Tj /F2 1 Tf 7.9746 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.7632 0 TD -0.0109 Tc -0.0142 Tw (and hydrogen peroxide, comparable)Tj -8.7378 -1.1 TD -0.0026 Tc 0.1718 Tw (to results in human primary airway epithelial cell cul-)Tj T* 0.1309 Tw [(tures [8]. The effects of IL-1)]TJ /F2 1 Tf 12.3944 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0027 Tc (/H)Tj 5.83 0 0 5.83 441.5955 542.5183 Tm 0 Tc (2)Tj 10 0 0 10 444.496 545.0183 Tm (O)Tj 5.83 0 0 5.83 451.6903 542.5183 Tm (2)Tj 10 0 0 10 458.3745 545.0183 Tm -0.0026 Tc 0.1309 Tw (co-stimulation on)Tj -15.6137 -1.1 TD 0.168 Tw (GM-CSF secretion are strikingly similar to the pattern)Tj T* 0.2032 Tw (of NF-)Tj /F2 1 Tf 2.8816 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.2032 Tw [(B activation. Treatment of cells with NF-)]TJ /F2 1 Tf 18.0892 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD (B)Tj -22.064 -1.1 TD -0.0026 Tc 0.1123 Tw (anti-sense oligonucleotides inhibited GM-CSF gene ex-)Tj T* 0.0098 Tw (pression [34], supporting the role of NF-)Tj /F2 1 Tf 16.201 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0098 Tw (B in regulating)Tj -16.7476 -1.1 TD 0.1715 Tw [(the biosynthesis of this cytokine. Taken together, our)]TJ T* 0.0115 Tc 0.2134 Tw (findings strongly suggest that GM-CSF secretion by)Tj T* -0.0026 Tc 0.1341 Tw (tracheobronchial epithelial cells is mediated, at least in)Tj T* 0.0486 Tw (part, by the activation of the transcription factor NF-)Tj /F2 1 Tf 21.2699 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0025 Tc (B.)Tj -20.9165 -1.1 TD -0.0026 Tc 0.1931 Tw (In conclusion, we have shown that the transcription)Tj -0.9 -1.1 TD 0.0005 Tc 0.2244 Tw (factor NF-)Tj /F2 1 Tf 4.4229 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5498 0 TD 0.0006 Tc 0.2243 Tw (B, known to regulate the expression of a)Tj -4.9727 -1.1 TD -0.0026 Tc 0.1089 Tw (variety of genes involved in defence and immune reac-)Tj T* 0.0303 Tw (tions, is activated in human airway epithelial cells by in-)Tj T* -0.0031 Tc -0.022 Tw [(flammatory stimuli. The activation of NF-)]TJ /F2 1 Tf 16.8767 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5461 0 TD -0.0031 Tc -0.022 Tw (B is followed)Tj -17.4228 -1.1 TD -0.0026 Tc 0.0225 Tw (by secretion of the NF-)Tj /F2 1 Tf 9.3066 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.0225 Tw (B target gene product GM-CSF.)Tj -9.8531 -1.1 TD 0.1153 Tw (Hydrogen peroxide has a modulating effect on the NF-)Tj /F2 1 Tf T* 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1999 Tw (B activation induced by IL-1)Tj /F2 1 Tf 12.2814 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1999 Tw [(. Hence, NF-)]TJ /F2 1 Tf 5.9813 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1999 Tw (B may)Tj -19.9024 -1.1 TD 0.1171 Tw (play a major role in tracheobronchial epithelial cells as)Tj T* 0.0567 Tw (a second messenger system following stimulation by in-)Tj T* 0.0202 Tw [(flammatory mediators. Investigation of compounds cap-)]TJ T* 0.1374 Tw (able of inhibiting the activation of NF-)Tj /F2 1 Tf 16.2201 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1374 Tw (B may lead to)Tj -16.7666 -1.1 TD 0.0736 Tw (the development of new treatment strategies for chronic)Tj T* 0.1039 Tw (inflammatory airway diseases, such as chronic bronchi-)Tj T* 0.1025 Tw (tis and asthma.)Tj /F11 1 Tf 8 0 0 8 346.2378 259.0183 Tm 0 Tw (Acknowledgements)Tj /F1 1 Tf 7.9037 0 TD 0.1509 Tw [(: The authors thank D.C. Gruenert,)]TJ -10.0287 -1.125 TD -0.0045 Tc -0.0206 Tw (Cardiovascular Research Institute, University of San Francisco,)Tj T* -0.0026 Tc 0.0167 Tw (for the human airway cell line used in this work, and A. Borde)Tj T* 0.1025 Tw (for her excellent technical assistance. )Tj /F5 1 Tf 9 0 0 9 394.9107 198.0184 Tm 0 Tw (References)Tj /F1 1 Tf -10.297 -2.5556 TD 0.1357 Tw [(1. )-1375.9(Jeffery PK. 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Haematopoietic colony-stimulating factors promote)]TJ T* -0.001 Tc 0.2259 Tw (cell survival by suppressing apoptosis. )Tj /F4 1 Tf 17.2314 0 TD 0 Tw (Nature)Tj /F1 1 Tf 3.2471 0 TD (1990;)Tj -20.4785 -1.1111 TD -0.0025 Tc 0.1025 Tw (343: 7679.)Tj -2.504 -1.1111 TD -0.0026 Tc 0.0884 Tw [(13. )-925.7(Baeuerle PA. The inducible transcription activator NF-)]TJ /F2 1 Tf 2.504 -1.1111 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5496 0 TD 0.0004 Tc 0.2245 Tw (B: regulation by distinct protein subunits. )Tj /F4 1 Tf 18.702 0 TD 0 Tw (Biochem)Tj -19.2515 -1.1111 TD -0.0026 Tc 0.1025 Tw (Biophys Acta )Tj /F1 1 Tf 5.7265 0 TD -0.0025 Tc (1991; 1072: 6380.)Tj -8.2305 -1.1111 TD -0.0026 Tc 0.057 Tw [(14. )-957.1(Schreck R, Baeuerle PA. NF-)]TJ /F2 1 Tf 14.7976 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.057 Tw (B as inducible transcrip-)Tj -12.8401 -1.1111 TD 0.1624 Tw (tional activator of the granulocyte-macrophage colony-)Tj T* -0.0299 Tc 0.0048 Tw (stimulating factor gene. )Tj /F4 1 Tf 9.2129 0 TD (Mol Cell Biol )Tj /F1 1 Tf 5.2908 0 TD (1990; 10: 12811286.)Tj -17.0077 -1.1111 TD -0.0026 Tc 0.0672 Tw [(15. )-946.9(Baeuerle PA, Baltimore D. Activation of DNA-binding)]TJ 2.504 -1.1111 TD 0.219 Tw (activity in an apparently cytoplasmic precursor of the)Tj T* -0.0027 Tc 0 Tw (NF-)Tj /F2 1 Tf 1.6031 0 TD 0 Tc ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1025 Tw (B transcription factor. )Tj /F4 1 Tf 9.6492 0 TD -0.0027 Tc 0 Tw (Cell)Tj /F1 1 Tf 2.0065 0 TD -0.0025 Tc 0.1025 Tw (1988; 53: 211217.)Tj -16.3094 -1.1111 TD -0.0026 Tc 0.1566 Tw [(16. )-857.5(Lenardo MJ, Fan C-H, Maniatis T, Baltimore D. The)]TJ 2.504 -1.1111 TD 0.1964 Tw (involvement of NF-)Tj /F2 1 Tf 8.2903 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0025 Tc 0.1964 Tw (B in )Tj /F2 1 Tf 2.3252 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1964 Tw (-interferon gene regulation)Tj -11.7086 -1.1111 TD 0.1884 Tw (reveals its role as widely inducible mediator of signal)Tj T* 0.1025 Tw (transduction. )Tj /F4 1 Tf 5.8605 0 TD -0.0027 Tc 0 Tw (Cell)Tj /F1 1 Tf 2.0065 0 TD -0.0025 Tc 0.1025 Tw (1989; 57: 287294.)Tj -10.371 -1.1111 TD -0.0026 Tc 0.0997 Tw [(17. )-914.4(Sen R, Baltimore D. Inducibility of )]TJ /F2 1 Tf 17.7453 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc (-immunoglobulin)Tj -15.7878 -1.1111 TD 0.1848 Tw (enhancer-binding protein NF-)Tj /F2 1 Tf 12.2392 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1848 Tw (B by a posttranslational)Tj -12.7857 -1.1111 TD 0 Tw (mechanism. )Tj /F4 1 Tf 5.129 0 TD -0.0027 Tc (Cell)Tj /F1 1 Tf 2.0065 0 TD -0.0025 Tc 0.1025 Tw (1986; 47: 921928.)Tj -9.6396 -1.1111 TD -0.0026 Tc -0.0009 Tw [(18. )-1015(Brach M, Hass R, Sherman ML, Gunji H, Weichselbaum)]TJ 2.504 -1.1111 TD 0.1668 Tw [(R, Kufe D. Ionizing radiation induces expression and)]TJ T* 0.1704 Tw (binding of the nuclear factor )Tj /F2 1 Tf 12.4105 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0025 Tc 0.1704 Tw (B. )Tj /F4 1 Tf 1.7478 0 TD -0.0026 Tc (J Clin Invest )Tj /F1 1 Tf 5.78 0 TD -0.0025 Tc 0 Tw (1991;)Tj -20.4848 -1.1111 TD 0.1025 Tw (88: 691695.)Tj -2.504 -1.1111 TD -0.0026 Tc 0.0899 Tw [(19. )-924.2(Busam K, Gieringer C, Freudenberg M, Hohmann. H-P)]TJ /F4 1 Tf 2.504 -1.1111 TD 0.0099 Tc 0.215 Tw (Staphylococcus aureus)Tj /F1 1 Tf 10.0916 0 TD (and derived exotoxins induce)Tj -10.0916 -1.1111 TD -0.0026 Tc 0.1641 Tw (nuclear factor )Tj /F2 1 Tf 6.064 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1641 Tw (B-like activity in murine bone marrow)Tj ET 0 0 0 0 k 315.91 762.01 229.31 -671.54 re 293.292 124.717 m f* BT 9 0 0 9 339.4494 754.7172 Tm 0 0 0 1 k 0.1025 Tw (macrophages. )Tj /F4 1 Tf 6.195 0 TD (Infect Immunol )Tj /F1 1 Tf 6.4982 0 TD -0.0025 Tc (1992; 60: 20082015.)Tj -15.1971 -1.1111 TD -0.0026 Tc 0.0871 Tw [(20. )-927(Schreck R, Albermann K, Baeuerle PA. Nuclear factor)]TJ /F2 1 Tf 2.504 -1.1111 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc 0.1602 Tw (B: an oxidative stress-response transcription factor of)Tj -0.5466 -1.1111 TD 0.0744 Tw (eukaryotic cells \(a review\). )Tj /F4 1 Tf 11.6792 0 TD 0.0745 Tw (Free Rad Res Comm )Tj /F1 1 Tf 8.8056 0 TD -0.0025 Tc 0 Tw (1992;)Tj -20.4848 -1.1111 TD 0.1025 Tw (17: 221237.)Tj -2.504 -1.1111 TD -0.0026 Tc 0.1561 Tw [(21. )-858(Schreck, R., Rieber P, Baeuerle PA. Reactive oxygen)]TJ 2.504 -1.1111 TD 0.1671 Tw (intermediates as apparently widely used messengers in)Tj T* -0.0114 Tw (the activation of the NF-)Tj /F2 1 Tf 9.7217 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0026 Tc -0.0114 Tw (B transcription factor and HIV-)Tj -10.2683 -1.1111 TD -0.0025 Tc 0.1025 Tw (1. )Tj /F4 1 Tf 1.445 0 TD -0.0026 Tc (EMBO J )Tj /F1 1 Tf 3.9078 0 TD -0.0025 Tc (1991; 10: 22472258.)Tj -7.8568 -1.1111 TD -0.0026 Tc 0.084 Tw [(22. )-930.1(Schreck R, Baeuerle PA. A role for oxygen radicals as)]TJ 2.504 -1.1111 TD 0.1418 Tw (second messengers. )Tj /F4 1 Tf 8.761 0 TD (Trends Cell Biology)Tj /F1 1 Tf 8.6799 0 TD -0.0025 Tc (1991; 1: 39)Tj -17.4409 -1.1111 TD 0 Tw (42.)Tj -2.504 -1.1111 TD -0.0026 Tc 0.1036 Tw [(23. )-910.5(Cozens AL, Yezzi MJ, Chin L, Simon EM, Friend DS,)]TJ 2.504 -1.1111 TD 0.0068 Tc 0.2181 Tw [(Gruenert DC. Chloride ion transport in transformed)]TJ T* -0.0026 Tc 0.0543 Tw (normal and cystic fibrosis epithelial cells. )Tj /F4 1 Tf 17.4254 0 TD -0.0027 Tc 0 Tw (In)Tj /F1 1 Tf 0.8278 0 TD -0.0026 Tc 0.0543 Tw (: Tsui L-C,)Tj /F4 1 Tf -18.2531 -1.1111 TD -0.0043 Tc -0.0208 Tw (et al.)Tj /F1 1 Tf 1.9533 0 TD [(, eds. The Identification of the Cystic Fibrosis Gene.)]TJ -1.9533 -1.1111 TD -0.0026 Tc 0.1025 Tw (New York, Plenum Press, 1991.)Tj -2.504 -1.1111 TD -0.0025 Tc 0.0921 Tw [(24. )-921.9(Jany B, Gallup M, Yan P, Gum J, Kim Y, Basbaum C.)]TJ 2.504 -1.1111 TD -0.0026 Tc 0.1304 Tw (Human bronchus and intestine express the same mucin)Tj T* -0.0025 Tc 0.1025 Tw (gene. )Tj /F4 1 Tf 2.8252 0 TD -0.0026 Tc (J Clin Invest )Tj /F1 1 Tf 5.5763 0 TD -0.0025 Tc (1991; 87: 7782.)Tj -10.9055 -1.1111 TD -0.003 Tc -0.0221 Tw [(25. )-1036.6(Gruenert DC, Basbaum CB, Widdicombe JH. Long-term)]TJ 2.504 -1.1111 TD -0.0074 Tc -0.0177 Tw (culture of normal and cystic fibrosis epithelial cells grown)Tj T* -0.005 Tc -0.0201 Tw (under serum-free conditions. )Tj /F4 1 Tf 11.7943 0 TD (In vitro Cell Dev Biol )Tj /F1 1 Tf 8.7003 0 TD 0 Tw (1990;)Tj -20.4946 -1.1111 TD -0.0025 Tc 0.1025 Tw (26: 411418.)Tj -2.504 -1.1111 TD -0.0026 Tc 0.1874 Tw [(26. )-826.7(Schreck R, Baeuerle P. Assessing oxygen radicals as)]TJ 2.504 -1.1111 TD 0.0211 Tw (mediators in activation of inducible eukaryotic transcrip-)Tj T* -0.0154 Tc -0.0097 Tw (tion factor NF-)Tj /F2 1 Tf 5.7486 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5338 0 TD -0.0153 Tc -0.0097 Tw (B. )Tj /F4 1 Tf 1.3364 0 TD (Methods Enzymol)Tj /F1 1 Tf 7.1227 0 TD (1994; 234: 151163. )Tj -17.2455 -1.1111 TD 0.0021 Tc 0.2228 Tw [(27. )-786.6(Zabel U, Schreck R, Baeuerle PA. DNA-binding of)]TJ 2.504 -1.1111 TD -0.0026 Tc 0.0675 Tw (purified transcription factor NF-)Tj /F2 1 Tf 13.0304 0 TD 0 Tc 0 Tw ()Tj /F1 1 Tf 0.5466 0 TD -0.0025 Tc 0.0675 Tw (B. )Tj /F4 1 Tf 1.5421 0 TD -0.0026 Tc (J Biol Chem)Tj /F1 1 Tf 5.3658 0 TD -0.0025 Tc 0 Tw (1991;)Tj -20.4848 -1.1111 TD 0.1025 Tw (266: 252260.)Tj -2.504 -1.1111 TD 0.0217 Tc 0.2032 Tw [(2)24.2(8)24.2(.)-1237.3(Watson ML, Smith D, Bourne AD, Thompson RC,)]TJ 2.504 -1.1111 TD -0.0026 Tc 0.0246 Tw [(Westwick J. Cytokines contribute to airway dysfunction)]TJ T* 0.1735 Tw (in antigen-challenged guinea-pigs: inhibition of airway)Tj T* 0.0015 Tw (hyperreactivity, pulmonary eosinophil accumulation, and)Tj T* 0.0215 Tw (tumor necrosis factor generation by pretreatment with an)Tj T* 0.0533 Tw (interleukin-1 receptor antagonist. )Tj /F4 1 Tf 13.9229 0 TD (Am J Respir Cell Mol)Tj -13.9229 -1.1111 TD 0 Tw (Biol )Tj /F1 1 Tf 2.0065 0 TD -0.0025 Tc 0.1025 Tw (1993; 8: 365369.)Tj -4.5105 -1.1111 TD 0 Tw (29. )Tj 2.504 0 TD -0.0191 Tc -0.006 Tw [(Warner B, Papes R, Heile M, Spitz D, Wispe J. Expression)]TJ 0 -1.1111 TD -0.0099 Tc -0.0152 Tw (of human Mn SOD in Chinese hamster ovary cells confers)Tj T* -0.0026 Tc 0.0193 Tw (protection from oxidant injury. )Tj /F4 1 Tf 12.8991 0 TD (Am J Physiol )Tj /F1 1 Tf 5.5511 0 TD -0.0025 Tc (1993; 264:)Tj -18.4502 -1.1111 TD 0 Tw (L598L605.)Tj -2.504 -1.1111 TD -0.0026 Tc 0.042 Tw [(30.)-1261.6(Samet JM, Noah TL, Devlin RB, )]TJ /F4 1 Tf 16.4401 0 TD (et al.)Tj /F1 1 Tf 2.6053 0 TD (Effect of ozone)Tj -16.5414 -1.1111 TD 0.2187 Tw (on platelet-activating factor production in phorbol-dif-)Tj T* 0.0496 Tw (ferentiated HL60 cells, a human bronchial epithelial cell)Tj T* 0.174 Tw (line \(BEAS S6\), and primary human bronchial epithe-)Tj T* 0.2167 Tw (lial cells. )Tj /F4 1 Tf 4.7274 0 TD (Am J Respir Cell Mol Biol)Tj /F1 1 Tf 12.0661 0 TD -0.0025 Tc (1992; 7: 514)Tj -16.7935 -1.1111 TD 0 Tw (522.)Tj -2.504 -1.1111 TD -0.0103 Tc -0.0148 Tw [(31. )-1036.6(Tanner WG, Welborn MB, Shepherd VL. 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