Background: The role of lymphocytes and their subpopulations in lung fibrosis is as yet unclear. Objective: To define the role of immunomodulation in bleomycin-induced inflammatory fibrotic lung injury, by testing the effect of two known Th1 inhibitors: linomide and pentoxifylline. Methods: C57BL/6 mice were treated by a single intratracheal instillation of 0.06 mg bleomycin in 0.01 ml saline or saline alone. Treatment groups included: (1) intratracheal bleomycin and daily treatment with linomide or pentoxifylline; (2) intratracheal bleomycin and daily water; (3) intratracheal saline and daily linomide or pentoxifylline; (4) intratracheal saline and daily water. Linomide and pentoxifylline were available per os in the drinking water from 1 day prior to intratracheal instillation. Animals were studied 14 days after intratracheal instillation. Lung injury was evaluated by total and differential cell count in bronchoalveolar lavage fluid, by a semiquantitative morphological index of lung injury and a quantitative image analysis of cellularity, fibrosis fraction and alveolar wall area fraction, and by biochemical analysis of lung hydroxyproline content. Results: Linomide or pentoxifylline did not cause any lung injury in saline-treated control mice. Overt signs of lung injury were apparent in bleomycin-treated mice. These changes were not affected by daily treatment with linomide or pentoxifylline, which were given in the highest tolerable dose. Conclusion: This study does not support the use of linomide or pentoxifylline to prevent or ameliorate lung fibrosis and may suggest that drug-induced differentiation of T lymphocytes into Th1/th2 subpopulations does not affect the evolution of bleomycin-induced lung injury.

Keywords:
Bleomycin, Linomide, Pentoxifylline, Lung, Mice
1.
Sulavik SB: A clinician’s view; in Phan SH, Thrall RS (eds): Pulmonary Fibrosis. Lung Biology in Health and Disease. New York, Dekker, 1995, vol 80, pp 1–58.
2.
Turner-Warwick M, Burrows B, Johnson A: Cryptogenic fibrosing alveolitis: Response to corticosteroid treatment and its effect on survival. Thorax 1980;35:593–599.
3.
Stack BHR, Choo-Kang YFJ, Heard BE: The prognosis of cryptogenic fibrosing alveolitis. Thorax 1972;17:535–542.
4.
Turner-Warwick M, Burrows B, Johnson A: Cryptogenic fibrosing alveolitis: Clinical features and their influence on survival. Thorax 1980;35:171–180.
5.
Snider GL, Celli RR, Goldstein RH, O’Brien JJ, Lucei EC: Chronic interstitial pulmonary fibrosis produced in hamsters by endotracheal bleomycin. Am Rev Respir Dis 1978; 117:289–297.
6.
Schrier DJ, Phan SH, McGarry BM: The effect of the nude (nu\nu) mutation on bleomycin-induced pulmonary fibrosis. Am Rev Respir Dis 1983;127:614–617.
7.
Sharma SK, MacLean JA, Pinto C, Kradin RL: The effect of an anti CD3 monoclonal antibody on bleomycin induced lymphokine production and lung injury. Am J Respir Crit Care Med 1996;154:193–200.
8.
Thrall RS, McCormick JR, Jack RM, Phan SH, Ward PA: The effect of antilymphocytic globulin on the development of bleomycin induced pulmonary fibrosis in the rat. Am Rev Respir Dis 1979;119(s):83.
9.
Thrall RS, Lovely EJ III, Baron RW, McCormick JR, Phan SH, Ward PA: The effect of T cell depletion on the development of bleomycin induced pulmonary fibrosis in the rat. Am Rev Respir Dis 1980;121(s):99.
10.
Lossos IS, Or R, Goldstein RH, Conner MW, Breuer R: Amelioration of bleomycin-induced pulmonary injury by cyclosporin A. Exp Lung Res 1996;22:337–349.
11.
Sendelbach LE, Lindenschmidt RC, Witschi HP: The effect of cyclosporin A on pulmonary fibrosis induced by butylated hydroxytoluene, bleomycin and beryllium sulfate. Toxicol Lett 1985;26:169–173.
12.
Szapiel SV, Elson NA, Fulmer JD, Hunninghake GW, Crystal RG: Bleomycin induced interstitial pulmonary disease in the nude, athymic mice. Am Rev Respir Dis 1979;120:893–899.
13.
Buckner JO, Ranges GE, Hacker MP: Effect of cytotoxic monoclonal antibody depletion of T lymphocytes subpopulations on bleomycin-induced lung damage in C57BL/6 mice. Toxicol Appl Pharmacol 1989;100:474–484.
14.
Karussis DM, Lehmann D, Brenner T, Wirguin I, Mizrachi R, Sicsic C, Abramsky O: Immunomodulation of experimental autoimmune myasthenia gravis with linomide. J Neuroimmunol 1994;55:187–193.
15.
Karussis DM, Lehmann D, Slavin S, Karussis UV, Mizrachi R, Ovadia H, Kalland T, Abramsky O: Treatment of chronic-relapsing experimental autoimmune encephalomyelitis with synthetic immunomodulator linomide (quinoline-3-carboxamide). Proc Natl Acad Sci USA 1993;90:6400–6404.
16.
Gross DJ, Sidi H, Weiss L, Kalland T, Rosenmann E, Slavin S: Prevention of diabetes mellitus in non-obese diabetic mice by linomide, a novel immunomodulating drug. Diabetologia 1994;37:1195–1201.
17.
Rapoport MJ, Weiss L, Mor A, Bistritzer T, Ramot Y, Slavin S: Prevention of autoimmune diabetes by linomide in nonobese diabetic (NOD) mice is associated with up-regulation of the TCR-mediated activation of p21 ras1. J Immunol 1996;157:4721–4725.
18.
Constantinescu CS, Hillard B, Lavi E, Ventura E, Venkatech V, Rostami A: Suppression of experimental autoimmune neuritis by PDE inhibitor pentoxifylline. J Neurol 1996;143:14–18.
19.
Rott O, Cash E, Fleischer B: Phosphodiesterase inhibitor pentoxifylline, a selective suppressor of T helper type 1 but not type 2 associated lymphokine production, prevents induction of experimental autoimmune encephalomyelitis in Lewis rats. J Immunol 1993;23:1745–1751.
20.
Snedecor GW, Cochran WG: Statistical Methods, ed 6. Ames, Iowa State University Press, 1967; pp 258–298.
21.
Karussis DM, Meiner Z, Lehmann D, Gomori JM, Schwarz A, Linde A, Abramsky O: Treatment of secondary progressive multiple sclerosis with the immunomodulator linomide. Neurology 1996;47:341–346.
22.
Arad G, Katzenellenbogen M, Levy R, Slavin S, Kaempfer R: Linomide, an immunomodulator that inhibits Thl cytokine gene expression. Int Immunol 1996;8:1603–1607.
23.
Rieneck K, Diamant M, Haahr PM, Schonharting M, Bendtzen K: In vitro immunomodulatory effects of pentoxifylline. Immunol Lett 1993;37:131–138.
24.
Thanhauser A, Reiling N, Bohle A, Toellner KM, Duchrow M, Scheel D, Schluter C, Ernst M, Flad HD, Ulmer AJ: Pentoxifylline: A potent inhibitor of IL-2 and IFNγ biosynthesis and BCG-induced cytotoxicity. Immunology 1993;80:151–156.
25.
Hecht M, Muller M, Lohmann-Matthes ML, Emmendorffer A: In vitro and in vivo effects of pentoxifylline on macrophages and lymphocytes derived from autoimmune MRL-lpr/lpr mice. J Leuk Biol 1995;57:242–249.
26.
Zabel P, Entzian P, Dalhoff K, Schlaak M: Pentoxifylline in treatment of sarcoidosis. J Respir Crit Care Med 1997;155:1665–1669.
27.
Bursten SL, Federighi DA, Parsons P, Harris WE, Abraham E, Moore EE Jr, Moore FA, Bianco JA, Singer JW, Repine JE: An increase in serum C18 unsaturated free fatty acids as a predictor of the development of acute respiratory distress syndrome. Crit Care Med 1996;24:1129–1136.
28.
Entzian P, Gerlach C, Gerdes J, Schlaak M, Zabel P: Pentoxifylline inhibits experimental bleomycin induced fibrosing alveolitis. Pulmonologie 1997;51:375–380.
29.
Thrall RS, Barton RW, D’Amato DA, Sulavik SB: Differential cellular analysis of bronchoalveolar lavage fluid obtained at various stages during the development of bleomycin-induced pulmonary fibrosis in the rat. Am Rev Respir Dis 1982;126:488–492.
30.
Thrall RS, Barton RW: A comparison of lymphocyte populations in lung tissue and in bronchoalveolar lavage fluid of rats at various times during the development of bleomycin-induced pulmonary fibrosis. Am Rev Respir Dis 1984;129:279–283.
31.
Lossos IS, Breuer R, Shriki M, Or R: Peribronchial lymphocyte activation in bleomycin-induced lung injury. Life Sci 1998;63:1883–1893.
32.
Sigiura Y, Kikuchi T: Formation of superoxide and hydroxy radicals in iron (II)-bleomycin-oxygen system: Electron spin resonance detection by spin trapping. J Antibiot 1978;31:1310–1312.
33.
Thrush MA, Mimnaugh ES, Ginsburg E, Gram TE: Studies on the interaction of bleomycin A2 with rat lung microsomes. II: Involvement of adventitious iron and reactive oxygen in bleomycin-mediated chain breakage. J Pharmacol Exp Ther 1982;221:159–165.
34.
Tom WM, Montgomery MR: Bleomycin toxicity: Alterations in oxidative metabolism in lung and liver microsomal fractions. Biochem Pharmacol 1980;29:3239–3244.
35.
Wahl SM, Wahl LM, McCarthy LB: Lymphocyte-mediated activation of fibroblast proliferation and collagen production. J Immunol 1978;121:942–946.
36.
Johnson RL, Ziff M: Lymphocyte stimulation of collagen accumulation. J Clin Invest 1979;58:240–248.
37.
Piguet PF: Cytokines involved in pulmonary fibrosis. Int Rev Exp Pathol 1993;34:173–181.
38.
Schrier DJ, Phan SH: Modulation of bleomycin-induced pulmonary fibrosis in the BALB/c mouse by cyclophosphamide-sensitive T cells. Am J Pathol 1984;116:270–278.
39.
Gillery P, Fertin C, Nicolas JF, Chastang F, Kalis B, Banchereau J, Maquart FX: Interleukin-4 stimulates collagen gene expression in human fibroblast monolayer cultures. Potential role in fibrosis. FEBS Lett 1992;302:231–234.
40.
Fertin C, Nicolas JF, Gillery P, Kalis B, Banchereau J, Maquart FX: Interleukin-4 stimulates collagen synthesis by normal and scleroderma fibroblasts in dermal equivalents. Cell Mol Biol 1991;37:823–829.
41.
Janick Buckner D, Ranges GE, Hacker MP: Alteration of bronchoalveolar lavage cell populations following bleomycin treatment. Toxicol Appl Pharmacol 1985;100:4651–473.
42.
Gharaee-Kermain M, Phan SH: Lung interleukin-5 expression in murine bleomycin-induced pulmonary fibrosis. Am J Respir Cell Mol Biol 1997;16:438–447.
43.
Or R, Lossos IS, Hirschfeld E, Breuer R: Effect of interleukin-2 and interleukin-4 on lymphocytes from peribronchial lymphatic tissue. Exp Lung Res 1996;22:245–253.
1999
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