Bleomycin-induced Pulmonary Fibrosis In Hamsters Research Articles

Effects of vitamin E deficiency on bleomycin-induced pulmonary fibrosis in hamsters

Effects of vitamin E deficiency on bleomycin-induced pulmonary fibrosis in hamsters

The anti-fibrotic agent SMP-534 attenuates bleomycin-induced pulmonary fibrosis in hamsters

Pulmonary fibrosis is a progressive and lethal lung disease characterized by accumulation of ECM and loss of pulmonary function. However, no cure exists for this disease, and current treatments often fail to slow its progression or relieve its symptoms. We have previously reported that the anti-fibrotic agent SMP-534 has beneficial effects on renal fibrosis in animal model of nephropathy. In this study, we examined whether SMP-534 has beneficial effects on pulmonary fibrosis in bleomycin-treated hamsters. Treatment with SMP-534 [low dose (70 mg/kg) or high dose (110 mg/kg)] counteracted inhibition of body weight increase induced by bleomycin. In addition, SMP-534 significantly inhibited bleomycin-induced increase in lung hydroxyproline level, an index of collagen formation. Moreover, SMP-534 significantly ameliorated histological pulmonary fibrotic changes induced by bleomycin. The results of this study indicate that the anti-fibrotic agent SMP-534 may offer a new therapeutic option for the treatment of pulmonary fibrosis.

A specific chymase inhibitor, NK3201, suppresses bleomycin-induced pulmonary fibrosis in hamsters

We evaluated whether a chymase inhibitor, 2-(5-formylamino-6-oxo-2-phenyl-1,6-dihydropyrimidine-1-yl)- N-[{3,4-dioxo-1-phenyl-7-(2-pyridyloxy)}-2-heptyl]acetamide (NK3201), suppressed bleomycin-induced pulmonary fibrosis. Hamsters were orally administered NK3201 (30 mg/kg per day) or placebo, beginning 5 days before intratracheal instillation of bleomycin (10 mg/kg). Four weeks after the instillation of bleomycin, pulmonary chymase activity in placebo-treated hamsters was significantly higher than in control hamsters, whereas the activity in NK3201-treated hamsters was significantly lower than in placebo-treated hamsters. The ratio of fibrotic area to total area in NK3201-treated hamsters was significantly decreased to 54.0% of the ratio in placebo-treated hamsters. Therefore, NK3201 may be useful in the prevention of pulmonary fibrosis.

Administration of truncated secretory leukoprotease inhibitor ameliorates bleomycin-induced pulmonary fibrosis in hamsters.

The purposes of this study were to investigate the effect of our recently developed truncated secretory leukoprotease inhibitor (SLPI) on bleomycin (BLM)-induced lung injury and fibrosis in hamsters, which is widely used as a model of interstitial pulmonary fibrosis, and to assess the possible involvement of neutrophil elastase in the pathogenesis of pulmonary fibrosis. The fibrosis model was prepared by administration of BLM (10 mg/kg) intratracheally to hamsters. The truncated SLPI was administered at a dose 5 or 15 mg/kg intraperitoneally twice a day only for 10 d starting from the time of BLM administration. BLM administration resulted in decreases in body weight and survival rate. Elastase activity in the supernatant of bronchoalveolar lavage (BAL) fluids was increased at 3 and 7 d after BLM administration in parallel with the increase in the number of neutrophils in the fluids. Histopathologically, at 14 and 28 d after BLM administration, diffuse thickening and fibrosis of alveolar walls with marked cell infiltration and severe distortion of alveolar structure, including honeycomb lung, were observed to have occurred. In this model, the decreases in body weight and survival rate and the increase in elastase activity were inhibited by the truncated SLPI dose-dependently, and pulmonary fibrosis was significantly ameliorated by the administration of this shortened form of SLPI. These results suggest that neutrophil elastase may be implicated in the pathogenesis of BLM-induced pulmonary fibrosis and that the truncated SLPI might be a promising therapeutic in the treatment of interstitial pulmonary fibrosis.

Amelioration of bleomycin-induced pulmonary fibrosis in hamsters by combined treatment with taurine and niacin

Interstitial pulmonary fibrosis induced by bleomycin (BL) involves the production of reactive oxygen species and the impairment of repair of damaged epithelial cells. We have shown previously that taurine or niacin treatment partially attenuates BL-induced lung fibrosis in hamsters and that the two agents probably act through different mechanisms. In the present investigation, we have demonstrated that taurine and niacin in combination provide nearly complete protection against BL-induced pulmonary fibrosis. Based on the findings of this investigation, it is suggested that combined treatment with taurine and niacin offers the potential for a novel pharmacological approach in the prevention of lung fibrosis in humans.

Changes of lung surfactant and pressure-volume curve in bleomycin-induced pulmonary fibrosis

We investigated whether alveolar surface force increased and participated in the lung pressure-volume relationship in bleomycin-induced pulmonary fibrosis in hamsters and, if so, whether lung surfactant was hampered in the lungs. On the air-filled pressure-volume curve, decreases of lung volume from control level were significantly higher at 3-8 cmH2O pressure on day 10 than on day 30. Because the change of lung tissue elasticity evaluated from the saline-filled pressure-volume curve was equal for the 2 days, the higher decrease of air volume on day 10 was due primarily to contribution of alveolar surface force. Pressure differences between deflation limbs of air-filled and saline-filled pressure-volume curves, which represented net alveolar surface force, were significantly higher at any lung volume between 50 and 90% total lung capacity on day 10, but almost no significance was observed on day 30. Phospholipid concentration in bronchoalveolar lavage fluid significantly decreased on day 10 but had improved by day 30. Analysis of phospholipid species in purified lung surfactant showed decreased fractions of disaturated phosphatidylcholine and phosphatidylglycerol on day 10. Surface-active properties of the surfactant, measured by a modified Wilhelmy balance, were remarkably hampered on day 10, but most of them had improved by day 30. We consider that the quantitative and functional abnormalities of lung surfactant have a part in the aggravation of lung mechanics in the acute phase of pulmonary fibrosis.

The effect of diclofenac acid (Voltaren ®) on bleomycin-induced pulmonary fibrosis in hamsters

Bleomycin is an anti-neoplastic compound which produces a time- and dose-dependent pulmonary fibrosis. The mechanisms which cause this fibrosis are not known. However, the ability of bleomycin to modulate prostaglandin synthesis, degradation and circulating levels appears to be central to the fibrosis. Previous studies, which have attempt to modulate bleomycin-induced fibrosis by prevention of prostaglandin synthesis have conflicting results. Therefore, the present study was designed to determine the effects of diclofenac acid, a nonsteroidal anti-inflammatory compound, on the development of bleomycin-induced pulmonary fibrosis. Diclofenac acid pretreatment and daily injections prevented lung collagen accumulation after intratracheal bleomycin. In addition diclofenac acid treatment resulted in significantly lower lung collagen level after intratracheal bleomycin at 14 and 21 days when compared with bleomycin alone. These data indicate that diclofenac acid treatment inhibits bleomycin-induced lung fibrosis possible through the prevention of prostaglandins synthesis.

The Effect of Cigarette Smoke on Bleomycin-induced Pulmonary Fibrosis in Hamsters

To investigate the effect of cigarette smoke on the development of bleomycin (BLM)-induced pulmonary fibrosis in hamsters, four experimental groups were studied: a control group (C), a cigarette smoke-inhaled group (T), a BLM-administered group (B), and a cigarette smoke-inhaled plus BLM-administered group (TB). Groups T and TB were exposed to sidestream smoke of cigarettes for 30 min/day, 5 days/wk. Groups B and TB were administered 0.5 mg BLM hydrochloride per 100 g body weight endotracheally once on day 30 (Day 0) after housing start. Quantitative morphometry of the lungs revealed that Group TB showed less lung fibrotic change compared with Group B, but based on qualitative observation, the fibrotic lesions of Group TB were intermingled with slight emphysematous changes. Neutrophils in bronchoalveolar lavage fluid were remarkably increased in both the groups, with a peak on Day 1, but the increase in Group TB lasted longer. Alveolar macrophages were increased in both smoking groups (T and TB) compared to the non-smoking groups (C and B). These results suggest that cigarette smoke reduces BLM-induced lung fibrotic changes; however, it simultaneously causes derangement of alveolar architecture. The persistence of increased neutrophils in the early phase after BLM accompanied by exposure to cigarette smoke may play an important role in the mechanism by which smoke ameliorates the effect of BLM.

Increased synthesis of elastin in amiodarone-induced pulmonary fibrosis.

Both synthesis and total content of lung elastin were measured after induction of interstitial pulmonary fibrosis in hamsters by a single intratracheal insufflation of amiodarone. Elastin synthesis, as measured by 14C-lysine incorporation into desmosine and isodesmosine, was significantly elevated (P less than 0.05) above control values for a 3-week interval after induction of lung injury. Total lung elastin content in the amiodarone-treated animals was 32% greater than in controls (P less than 0.05) 2 weeks after insufflation of the agent. Furthermore, the time course of elastin synthesis in this experimental model was similar to that observed in bleomycin-induced pulmonary fibrosis in hamsters. Increases in elastin may therefore be a common feature of interstitial pulmonary fibrosis and may contribute to the altered lung mechanics seen in this disease.

Analysis of Bronchoalveolar Lavage Fluid from Bleomycin-Induced Pulmonary Fibrosis in Hamsters

The purpose of this study was to analyze the cellular and noncellular components of bronchoalveolar lavage fluid (BALF) at varying times during the development of pulmonary fibrosis induced by bleomycin. Hamsters were killed and lavaged in situ following the administration of a single intratracheal injection of 1 unit of bleomycin or an equivalent volume of sterile isotonic saline. The results show that the total cell counts in the BALF of bleomycin-treated hamsters, as compared with controls, were increased 7.7, 4.4, 2.4, 1.6, and 1.9-fold at 2, 4, 7, 14, and 21 days after treatment, respectively. The predominant cell types in the BALF of control animals were macrophages which constituted 84% of the total cells, followed by lymphocytes, 11%. The predominant cell types in the BALF of bleomycin-treated animals were polymorphonuclear leukocytes (PMN) which constituted 65% at two days and approximately 50% of the total at 4, 7, and 14 days; at 21 days macrophages were the predominant cell type constituting 50%, followed by lymphocytes at 30%. However, the total number of lymphocytes was not increased at 21 days compared to previous times. The noncellular protein content of BALF from bleomycin-treated hamsters, an index of pulmonary vascular permeability, was increased to 224, 559, 637, and 270% of control (2.7 mg/lung) at 2, 4, 7, and 14 days after treatment, respectively, and returned to control levels at 21 days. The acid phosphatase activity in the supernatant of BALF of bleomycin-treated animals was significantly increased to 181, 181, 199, 176, and 125% of control (258 units/lung) at 2, 4, 7, 14, and 21 days, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Bleomycin-induced pulmonary fibrosis in hamsters. An alveolar macrophage product increases fibroblast prostaglandin E2 and cyclic adenosine monophosphate and suppresses fibroblast proliferation and collagen production.

Bleomycin-induced pulmonary fibrosis in hamsters is associated with collagen accumulation that results from increased lung collagen synthesis rates. However, 1-2 wk after intratracheal instillation of bleomycin, lung collagen synthesis rates decline toward control values. To evaluate the potential role of the bronchoalveolar macrophage in regulating lung collagen production, we studied the effects of macrophages from normal and bleomycin-treated hamsters upon fibroblasts in vitro. We observed: (a) Medium from macrophage cultures decreased fibroblast [3H]thymidine incorporation and nondialyzable [3H]hydroxyproline production in a dose-dependent manner. Fibroblast cell counts were decreased in exposed cultures, and fibroblast viability was unchanged. Procollagen prolyl hydroxylation and prolyl-transfer RNA-specific activity were not altered by macrophage medium; this indicates that [3H]hydroxyproline reflects collagen production rate under the experimental conditions. (b) The suppressive effect of macrophage medium was selective for collagen since collagen production decreased more than noncollagen protein production. (c) Medium from bleomycin-treated hamster macrophages suppressed fibroblast proliferation and collagen production to a greater degree than medium from normal hamster macrophages. (d) Fibroblast suppression by macrophage medium was associated with increased fibroblast endogenous prostaglandin E2 production and intracellular cyclic AMP (cAMP). (e) Incubation of fibroblasts with indomethacin before exposure completely inhibited prostaglandin E2 production and increases in cAMP, and eliminated suppression of fibroblast proliferation and collagen production. The macrophage-derived suppressive factor has an apparent molecular weight of 20,000-30,000 and is heat stable. It does not appear to be species restricted since both hamster and human lung fibroblasts are similarly suppressed. It is at least in part preformed in macrophages obtained by lavage, but its production can also be stimulated in vitro. We concluded that alveolar macrophages release a product that stimulates endogenous fibroblast prostaglandin E2 production and cAMP formation with resultant suppression of fibroblast proliferation and collagen production. Enhanced release of suppressive factor by macrophages during a time when lung collagen production is declining in bleomycin-induced pulmonary fibrosis suggests that macrophages may limit collagen accumulation in pulmonary fibrosis.

The [formula omitted] synthesis and degradation of prostaglandins during the development of bleomycin-induced pulmonary fibrosis in hamsters 1−3

Subsequent to optimization of conditions for enzyme assay, we examined the in vitro synthesis and degradation of prostaglandins by the lung during the devel opment of bleomycin-induced pulmonary fibrosis in hamsters. It was found that the microsomal protein content on a per lung basis was significantly increased to 144, 129, 134, and 121% of control (2.3 mg protein/lung) at 4, 7, 14 and 21 days post-treatment, respectively. The synthesis of PGD 2 was significantly elevated to 10.2, 10.8, and 12.5 moles/lung at 7, 21 and 28 days, respectively, as compared to the control value of 5.6 moles/lung. Significant increases in PGF synthesis from the control value of 3.3 moles/lung to 5.2, 8.2 and 5.5 moles/lung were found at 4, 7 and 21 days post-treatment, respectively. The synthesis of PGE 2 also showed significant increases above above the control value of 6.1 moles/lung to 10.5, 12.2 and 11.0 nmoles/lung at 7, 21 and 28 days post-treatment, respectively. Similarly, the synthesis of 6-keto-PGF was significantly increased to 7.4, 7.5 and 8.6 moles/lung at 7, 21 and 28 days post-treatment, respectively, as compared to the control value of 4.4 moles/lung. The synthesis of TxB was also significantly increased from the control value of 3.9 nmoles/lung to 7.5 and 6.4 moles/lung at 7 and 21 days post-treatment, respectively. Accompanying the increased synthesis of prostaglandins in general, the in vitro degradation of PGF was significantly increased from the control v of 71.1 nmoles/lung to 173.5, 131.7 and 143.3 nmoles/lung at at 2, 4 and 7 days after bleomycin treatment, respectively. We conclude that bleomycin-induced pulmonary fibrosis leads to changes in prostaglandin synthesis and degradation possibly as a result of an accompanying inflammatorv response and resident cellular oroliferation.

Bleomycin-induced pulmonary fibrosis in hamsters: effect of neutrophil depletion on lung collagen synthesis.

The development of pulmonary fibrosis in hamsters after the intratracheal instillation of bleomycin is attended by exudation of polymorphonuclear leukocytes (PMNs), but the function of these cells is unknown. In this study, we examined the effect of PMN depletion on lung collagen synthetic rate in bleomycin-treated hamsters. Antiserum to hamster PMNs was produced in rabbits and injected intraperitoneally into hamsters on the fourth through seventh days after intratracheal instillation of bleomycin or saline. Control animals received either normal rabbit serum (NRS) or no serum. Anti-PMN serum (APS) significantly reduced PMN exudation after bleomycin treatment through the eighth day (p less than 0.01). Collagen synthetic rate measured in lung explant cultures was increased in all bleomycin-treated groups compared with that in the saline injected control group (p less than 0.005). Collagen synthesis was greater in neutrophil-depleted bleomycin-treated hamsters than in bleomycin-treated animals receiving NRS or no serum on Days 8 (p less than 0.01) and 12 (p less than 0.05). In APS-treated hamsters, the proportion of protein synthesis directed toward collagen synthesis was increased above other bleomycin-treated groups, suggesting that collagen synthesis is selectively increased in neutrophil-depleted hamsters with bleomycin-induced pulmonary fibrosis.

Modulation of collagen production following bleomycin-induced pulmonary fibrosis in hamsters. Presence of a factor in lung that increases fibroblast prostaglandin E2 and cAMP and suppresses fibroblast proliferation and collagen production.

To elucidate mechanisms involved in the regulation of lung collagen content we studied hamsters with bleomycin-induced pulmonary fibrosis. Lung collagen in this model is increased as the result of greatly increased lung collagen synthesis rates. However, collagen synthesis rates are subsequently restored to normal. Hamster lung explants from both normal and bleomycin-exposed hamsters were cultured, and the effects of explant conditioned medium (CM) on lung fibroblast (IMR-90) proliferation and collagen production in vitro were determined. Lung explant CM increased fibroblast prostaglandin (PG)E2 production and intracellular cAMP, and decreased both fibroblast proliferation and collagen production in a dose-dependent manner. Greater activity was observed with lung explant CM from bleomycin-exposed lungs. Incubation of fibroblasts with indomethacin prior to addition of CM blocked CM-mediated changes in PGE2 and cAMP and inhibited changes in fibroblast proliferation and collagen production. Exogenous PGE2 or dibutyryl cAMP also suppressed fibroblast proliferation and collagen production. The suppressive activity in lung-conditioned medium is nondialyzable, has an apparent molecular weight of 15,000-20,000 by gel filtration, and is heat-stable. It is not species-restricted since CM from hamster lung affected human and hamster lung fibroblasts similarly. Activity is present preformed in lung and bronchoalveolar lavage fluid, although bronchoalveolar macrophages produce a nondialyzable factor in culture which suppresses fibroblast proliferation. The suppressive activity identified in fibrotic lung may represent a means for limiting collagen accumulation following tumor injury.

Elevation of glucosamine 6-phosphate synthetase activity in bleomycin-induced pulmonary fibrosis in hamsters.

The single intratracheal instillation of bleomycin sulfate (0.5 mg (potency) per 100 g body weight) in hamsters rapidly increased the activity of glucosamine 6-phosphate synthetase (EC 2.6.1.16) of the lung, a major regulatory enzyme for the synthesis of acidic glycosaminoglycan (AGAG). The activity increased as early as day 2, reached maximum level at day 10, then decreased and returned to the control level at day 45. The content of AGAG was also increased by bleomycin treatment, but the increase of AGAG followed the elevation of the enzyme activity. These results suggest that the early elevation of glucosamine 6-phosphate synthetase activity is closely related to the accumulation of AGAG in the fibrosing lung caused by bleomycin.