[Effects of aerosolized earthworm fibrinolytic enzyme on bleomycin induced pulmonary fibrosis in rats].

OBJECTIVE: To explore the effects of aerosolized earthworm fibrinolytic enzyme (EFE) on bleomycin-induced pulmonary fibrosis in rats. METHODS: A total of 72 male SD rats were divided randomly into 3 groups of bleomycin (BLM) group with intratracheal BLM (5 mg/kg), control group with the same dose of normal saline, then after both receiving aerosolization of normal saline once daily instead of EFE, EFE group with EFE (2500 U/kg) by aerosolization once daily after BLM instillation. Lung histopathology, immunohistochemistry for transforming growth factor ß(1) (TGF-ß(1)), lung hydroxyproline contents, levels of urokinase PA (uPA), tissue plasminogen activator (tPA) and PA inhibitor 1 (PAI-1) in lung and blood were observed at Days 7, 14 and 28 of experiment, respectively. RESULTS: Compared with BLM group, pulmonary fibrosis improved and the TGF-ß(1) expression of lung tissue decreased (P < 0.01). Hydroxyproline content of lung tissue decreased in EFE group compared with BLM group ((5.8 ± 2.5) vs (9.6 ± 1.3), (6.7 ± 1.4) vs (9.7 ± 1.5), (7.5 ± 1.2) vs (9.7 ± 1.4) mg/L, P < 0.01). Compared with BLM group, the uPA levels of lung were elevated in EFE group at Days 7 and 14 ((1.04 ± 0.36) vs (0.72 ± 0.11), (0.90 ± 0.09) vs (0.75 ± 0.08) µg/L, P < 0.05). Moreover, the plasma levels uPA of increased at Days 14 and 28 ((0.32 ± 0.04) vs (0.25 ± 0.02), (0.36 ± 0.05) vs (0.28 ± 0.04) µg/L, P < 0.05). Consistently, compared with BLM group, the tPA levels of lung increased in EFE group ((4.70 ± 0.87) vs (3.01 ± 0.62), (5.72 ± 0.37) vs (3.00 ± 0.51), (6.73 ± 1.12) vs (3.18 ± 0.38) µg/L, P < 0.01) and the plasma levels of tPA also increased ((3.40 ± 0.36) vs (1.79 ± 0.38), (3.17 ± 0.37) vs (2.18 ± 0.17), (3.85 ± 0.56) vs (2.80 ± 1.06) µg/L, P < 0.01). However, compared with BLM group, the PAI-1 levels of lung decreased in EFE group ((6.04 ± 0.81) vs (8.52 ± 1.01), (6.78 ± 0.81) vs (9.81 ± 1.73), (7.63 + 0.99) vs (11.44 ± 2.54), P < 0.05) and the plasma levels of PAI-1 also decreased in EFE group ((4.82 ± 0.42) vs (6.89 ± 0.84), (5.73 ± 0.40) vs (7.30 ± 1.09), (5.64 ± 0.87) vs (7.98 ± 1.10) µg/L, P < 0.05). CONCLUSIONS: Earthworm fibrinolytic enzyme may decrease bleomycin-induced pulmonary fibrosis and TGF-ß(1) expression while increasing fibrinolytic activation. And fibrinolytic strategies are probably useful for the therapy of fibrotic lung diseases.

[Expression of intercellular cell adhesion molecule-1, interleukin-10 and the activation of activator protein-1 in ventilator-induced lung injury in rabbits].

OBJECTIVE: To study the expression of intercellular cell adhesion molecule-1 (ICAM-1), Interleukin-10 (IL-10) and the activation of transcription factor activator protein-1 (AP-1) in a rabbit model of ventilator-induced lung injury (VILI) and therefore to explore their possible role in VILI. METHODS: The VILI model was established by mechanical ventilation with a large tide volum (V(T)) of 40 ml/kg. Forty healthy male New Zealand rabbits were randomly divided into 5 groups: a control group without mechanical ventilation, a conventional ventilation group, and injurious ventilation with large V(T) for 1 h group, 2 h group and 4 h group. The concentrations of soluble ICAM-1 (sICAM-1) and IL-10 in lung homogenates were measured by enzyme-linked immunosorbent assay (ELISA). The level of mRNA was measured by semiquantitative transcription-polymerase chain reaction (RT-PCR). The DNA-binding activity of AP-1 was measured by electrophoretic mobility shift assay (EMSA). The partial arterial blood pressure of oxygen (PaO2), myeloperoxidase (MPO) in lung homogenate, wet lung weight to dry lung weight ratio (W/D) were observed. RESULTS: (1) The concentrations of sICAM-1 in large V(T) for 2 h group (23 ± 5) ng/L and 4 h group (35 ± 5) ng/L were higher than that in the conventional ventilation group (16 ± 4) ng/L (P all < 0.05), and that in the Large V(T) for 4 h group were higher than that in 1 h group (19 ± 4) ng/L and 2 h group (P all < 0.01). The concentrations of IL-10 in the large V(T) for 2 h group (24 ± 4) ng/L and 4 h group (26 ± 5) ng/L were higher than that in the conventional ventilation group (15 ± 2) ng/L (P all < 0.05), and that in the Large V(T) for 4 h group was higher than that in the 1h group (19 ± 4) ng/L(P < 0.05). The level of ICAM-1 mRNA in the large V(T) for 2 h group (1.18 ± 0.19) and 4 h group (1.29 ± 0.19) were higher than that in the conventional ventilation group (0.84 ± 0.13) (P all < 0.05), and that in Large V(T) for 4 h group was higher than that in 1 h group (0.96 ± 0.24) (P < 0.05). The level of IL-10 mRNA in the large V(T) for 4 h group (1.13 ± 0.17) was higher than that in the conventional ventilation group (0.84 ± 0.20) and Large V(T) for 1 h group (0.86 ± 0.12) (P all < 0.05). (2) The DNA-binding activity of AP-1 in the large V(T) for 2 h group (33.77 ± 8.23) and 4 h group (38 ± 9) were higher than that in the conventional ventilation group (23 ± 9) (P all < 0.01), and that in Large V(T) for 4 h group was higher than that in 1h group (25 ± 9) (P < 0.01). (3) Histopathological findings demonstrated that diffused alveolar damage induced by mechanical ventilation was worse with time, and after mechanical ventilation with large V(T) for 2 h, the level of MPO began to increase, and for 4 h the PaO2 reduced and the W/D increased. CONCLUSIONS: ICAM-1 and IL-10 took part in the inflammatory responses of VILI, and their up-regulation maybe due to the increase of their mRNA. Nuclear transcription factor AP-1 maybe involved in the transcriptional regulation mechanisms of these inflammatory mediators.

[Effects of bone marrow mesenchymal stem cells on bleomycin induced pulmonary fibrosis in rats].

OBJECTIVE: To study the effects of bone marrow mesenchymal stem cells (MSC) on pulmonary fibrosis. METHODS: Bone marrow MSC were harvested from 6 week old male SD rats. Forty-eight female SD rats were randomly divided into six groups. The pulmonary fibrosis models were made by intratracheal instillation of bleomycin (5 mg/kg in 0.3 ml normal saline). The normal controls received intratracheal instillation of NS instead of bleomycin. On the 1st and 7th day after bleomycin administration, the rats received MSC infusion or a same amount of phosphate buffer solution (PBS) as controls via the tail vein, respectively. The rats were sacrificed by the 28 day of experiment, and the pathologic changes and hydroxyproline contents of the lung tissues were investigated. The sry gene of Y chromosome was detected by polymerase chain reaction (PCR). RESULTS: For rats receiving MSC on the 1st and 7th day after bleomycin administration, the lung fibrotic scores were 1.0 +/- 0.2 and 1.6 +/- 0.5, respectively, significantly decreased as compared with rats receiving no MSC (2.5 +/- 0.5 & 2.3 +/- 0.8, respectively). The hydroxyproline contents of lung tissue were (83 +/- 17) microg/mg and (96 +/- 20) microg/mg, also significantly decreased as compared with rats receiving no MSCs [(123 +/- 32) microg/mg & (127 +/- 34) microg/mg, respectively]. Earlier administration of MSCs resulted in more significant improvement of lung injury. The sry gene (322 bp) was detected in lungs of female rats receiving MSC on the first day of bleomycin induced lung injury. CONCLUSIONS: MSC may be involved in the repair of lung injury, especially in the early stage. MSCs are effective in preventing bleomycin induced lung injury and fibrosis.

[Expression of matrix metalloproteinases in lung tissue and levels of some cytokines in bronchoalveolar lavage fluid in the obstructive emphysema rat models].

OBJECTIVE: To observe the changes of matrix metalloproteinase-2 (MMP-2), MMP-9 and interleukin-10 (IL-10), tumor necrosis factor-alpha (TNFalpha) in lung tissue of the obstructive emphysema rat models and to evaluate the relationship between these changes and emphysema formation. METHODS: The rat emphysema models were established by exposure to cigarette smoking. Pulmonary function tests were performed to evaluate the forced expiratory volume in 0.3 second (FEV(0.3)), FEV(0.3)/forced vital capacity (FVC) and functional residual capacity (FRC). The morphological indices of emphysema were measured by computer image analyzer. The protein expression and enzymatic activity of MMPs in lung tissue were observed. The contents of IL-10 and TNFalpha in bronchoalveolar lavage fluid (BALF) were measured. RESULTS: Pulmonary function test showed that in model group the FEV(0.3)/FVC was decreased, whereas the FRC was increased significantly than those in control group (P < 0.01, respectively). There was a significant decrease in the relative content of elastin in lung tissue in model group than that in control group. The expression and enzymatic activity of MMP-2 and MMP-9 in lung tissue, the counts of total leukocytes and neutrophils and the level of TNFalpha in BALF were significantly elevated, but the level of IL-10 in BALF was significantly reduced in model group compared with those in control group (P < 0.01, respectively). There were statistically significant positive correlations between the enzymatic activity of MMP-2, MMP-9 and the total leukocyte counts (P < 0.01, respectively), and significant negative correlations between the enzymatic activity of MMP-2, MMP-9 and the content of elastin (P < 0.05, respectively). CONCLUSIONS: MMP-2, MMP-9, IL-10 and TNFalpha may play an important role in the formation of obstructive emphysema in rat models caused by passive smoking.