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Journal of Chinese Integrative Medicine ›› 2006, Vol. 4 ›› Issue (4): 402-407.doi: 10.3736/jcim20060417

• Original Experimental Research • Previous Articles     Next Articles

Feature changes of MMP-2/9 activities and TIMP-1/2 protein expressions during the progression of pulmonary fibrosis in rats

Shan-zhong Tan1, Cheng-hai Liu1, Wei Zhang2, Xiong Lu3, Wei-cheng Ye4, Zhen-zhen Cai5, Ping Liu1   

  1. 1. E-institute of Internal Medicine of Traditional Chinese Medicine, Shanghai Municipal Education Commission, Institute of Liver Diseases, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
    2. Department of Respiratory Diseases, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
    3. Central Laboratory, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
  • Online:2006-07-31 Published:2006-07-15

Objective

To investigate the dynamic trends of activities of matrix metalloproteinase (MMP)-2/9 and protein expressions of their inhibitors-tissue inhibitor of matrix metalloproteinase (TIMP)-1/2 during the progression of pulmonary fibrosis in rats so as to get insight of the roles played by MMP-2 /9 in lung injury and fibrogenesis.

Methods

Forty-eight SD rats were randomly divided into normal control group (n=18) and bleomycin (BLM)-treated group (n=30). The pulmonary fibrosis was induced by intratracheal injection of BLM once. At the consecutive time of 1 day, 3 days, 1 week, 2, 3, and 4 weeks after intoxication, the lung-to-body weight ratio was calculated and the inflammation and collagen deposition in lung tissue were checked by HE and Masson stainings respectively. Meanwhile, the content of hypdroxyproline (Hyp) in lung tissue was assayed with Jamall’s method, the protein expressions of MMP-2/9, TIMP-1/2 were examined by Western blotting, and the activities of MMP-2/9 were detected by gelatin zymography.

Results

The histopathological changes in lung tissue in the BLM-treated group from 1 day to 2 weeks after intoxication presented local lesions, broadened alveolar wall and septum, infiltration with lots of inflammatory cells and few of fibroblasts inside alveolar space and septum. At this early stage in the BLM-treated group, the lung-to-body weight ratio was increased significantly, the protein expressions and activities of MMP-2/9 were obviously increased especially for activity of active MMP-2, and the protein expressions of TIMP-1/2 were also increased gradually, as compared with those in the normal control group. From 3 to 4 weeks after intoxication in the BLM-treated group, the alveolar structure was damaged, parts of the alveolar space collapsed and replaced by collagens and fibroblasts, and the alveolar wall and septum obviously widened with remarkable fibrotic characteristics, as compared with those in the normal control group. Meanwhile, the lung-to-body weight ratio and the activities of MMP-2/9 were decreased in the BLM-treated group as compared with those in the same group at 2 weeks after intoxication, but the content of Hyp and the protein expressions of TIMP-1/2 were both increased dramatically, especially at 4 weeks after intoxication.

Conclusions

During the lung fibrogenesis induced by BLM in rats, the alveolar inflammation is the most important alteration with enhanced MMP-2/9 activities in the early stage. While in the late stage, the main change is displayed as pulmonary fibrosis, characterized by increased TIMP-1/2 and declined MMP-2/9 activities.

Key words: Bleomycin, Alveolitis, Pulmonary fibrosis, MMP-2, MMP-9, TIMP-1, TIMP-2

CLC Number: 

  • R563.9

Table 1

Dynamic trend of body weight, lung-to-body weight ratio and Hyp contentin lung tissue of rats during pulmonary fibrogenesis"

Group n Body weight (g) Lung / body weight ratio (mg/g) Hyp (μg/mg)
Normal control
1 day after intoxication 3 206.67±5.77 1.45±0.04 1.81±0.23
3 days after intoxication 3 203.33±20.82 1.45±0.02 1.78±0.18
1 week after intoxication 3 236.67±15.28 1.44±0.03 1.70±0.10
2 weeks after intoxication 3 254.67±3.06 1.46±0.17 1.65±0.03
3 weeks after intoxication 3 287.33±23.01 1.33±0.03 1.80±0.38
4 weeks after intoxication 3 322.67±15.01 1.30±0.10 1.88±0.06
Bleomycin-treated
1 day after intoxication 5 184.00±6.52* 1.63±0.06 1.79±0.18
3 days after intoxication 5 180.00±15.81 1.88±0.38 1.71±0.39
1 week after intoxication 5 165.00±35.36* 4.35±2.21* 1.87±0.80
2 weeks after intoxication 5 221.60±61.50 1.56±0.09 1.74±0.50
3 weeks after intoxication 3 208.67±94.75 1.56±0.10 2.04±0.71
4 weeks after intoxication 5 263.60±24.02* 1.63±0.30 2.30±0.33*

Figure 1

Histopathological changes of lung tissue in rats during pulmonary fibrogenesis (HE staining, ×200) A: Normal control group; B: Three days after intoxication in bleomycin-treated group; C: Two weeks after intoxication in bleomycin-treated group; D: Four weeks after intoxication in bleomycin-treated group."

Figure 2

Collagen deposition in lung tissue of rats during pulmonary fibrogenesis (Masson staining, ×200) A: Normal control group; B: Three days after intoxication in bleomycin-treated group; C: Two weeks after intoxication in bleomycin-treated group; D: Four weeks after intoxication in bleomycin-treated group."

Figure 3

Dynamic trends of MMP-2/9 protein expressions in lung tissue of rats duringpulmonary fibrogenesis (Western blotting method) A: Expressions of MMP-2/9 and β-actin, which were consecutively blotted by incubation with different antibodies on the same membrane;B: Semi-quantification results of MMP-2 and MMP-9 with β-actin as the internal control (n=3); N: Normal control."

Figure 4

Dynamic trends of MMP-2/9 activitis in lung tissue of rats during pulmonary fibrogenesis A: Gelatin zymography Zymography of MMP-2 and MMP-9; B: Semi-quantification results of MMP-2 and MMP-9 with β-actin as the internal control (n=3); N:Normal control."

Figure 5

Dynamic trends of TIMP-1/2 protein expressions in lung tissue of rats duringpulmonary fibrogenesis (Western blotting method) A: Expressions of TIMP-1/2 and β-actin, which were consecutively blotted by incubation with different antibodies on the same membrane; B: Semi-quantification results of TIMP-1/2 with β-actin as the internal control (n=3); N: Normal control."

[1] Cook DN, Brass DM, Schwartz DA . A matrix for new ideas in pulmonary fibrosis[J]. Am J Respir Cell Mol Boil, 2002,27(2):122-124
[2] Taooka Y, Maeda A, Hiyama K , et al. Effects of neutrophil elastase inhibitor on bleomycin-induced pulmonary fibrosis in mice[J]. Am J Respir Crit Care Med, 1997,156(1):260-265
doi: 10.1164/ajrccm.156.1.9612077 pmid: 9230758
[3] Jamall IS, Finelli VN, Que Hee SS . A simple method to determine nanogram levels of 4-hydroxyproline in biological tissues[J]. Anal Biochem, 1981,112(1):70-75
doi: 10.1016/0003-2697(81)90261-X pmid: 7258630
[4] K1einer DE , Stetler-Stevenson WG .. Quantitative zymography: detection of picogran quantities of gelatinases[J].Anal Biochem, 1994, 325-329
doi: 10.1006/abio.1994.1186 pmid: 8074288
[5] 侯显明, 于润江 . 间质性肺病学(第1版)[M]. 北京: 人民卫生出版社, 2003,( 第1版):110-124
[6] 章巍, 刘新民, 李海朝 , 等. 博莱霉素致大鼠肺纤维化模型肺组织MMP-2、MMP-9、MT1-MMP及TIMP-1、TIMP-2的表达[J]. 北京大学学报(医学版), 2002,34(6):716-721
[7] Blackwell TS, Christman JW . The role of nuclear factor-kappa B in cytokine gene regulation[J]. Am J Respir Cell Mol Biol, 1977,17(1):3-9
doi: 10.1165/ajrcmb.17.1.f132 pmid: 9224203
[8] Selman M, Ruiz V, Cabrera S , et al. TIMP-1, -2, -3, and -4 in idiopathic pulmonary fibrosis. A prevailing nondegradative lung microenviroment[J]. Am J Physiol Lung Cell Mol Physiol, 2000,279(3):L562-L574
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