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Effects of N-acetylcysteine on Clara cells in rats with cigarette smoke exposure

Chinese Medical Journal 2010年第4期123卷 412-417页

作者： LIAO Ji-ping CHI Chun-hua LI Hai-chao TANG Xiu-ying Department of Respiratory Medicine Peking University First Hospital Beijing 100034 China Department of Pathology Peking University First Hospital Beijing 100034 China

Background The number of Clara cells and the Clara cell 16-kDa protein （CC16） levels of the lung decrease in patients with chronic obstructive pulmonary disease （COPD）. N-acetylcysteine （NAC） is a powerful antioxidant and can reduce the frequency of acute exacerbations of COPD. But the exact mechanism is unclear. The present study was designed to investigate the effects of NAC on Clara cells in rats with cigarette smoke exposure. Methods Eighteen adult male Wistar rats were randomly divided into 3 groups, 12 exposed to cigarette smoke （CS） thrice a day, 10 cigarettes for 30 minutes each time for 1 week, without （CS group） or with （CS＋NAC group） oral intake of NAC 80 mg·kg^-1·d^-1, and another 6 rats exposed to fresh air （control group）. Clara cells were observed by an electron microscope. The mRNA expression of CC16 and CC16 protein in lungs were determined by reverse transcription polymerase chain reaction （RT-PCR） and immunohistochemistry respectively. The glutathion （GSH） level in plasma and lung tissue were tested by fluorimetry assay. Results Compared with the controls, the pathologic score of small airways significantly increased in the CS exposed rats （20.3±14.7 vs. 53.7±11.5, P 〈0.05）. The Clara cell particles in cytoplasm decreased in the CS group （P 〈0.05）. The percentage of CC16-positive cells in bronchioles in the CS group （27.8±4.3 and 29.5±2.4 in terminal bronchioles and respiratory bronchioles, respectively） significantly decreased as compared with the control group （37.1±3.8 and 43.8±5.8 in terminal bronchioles and respiratory bronchioles, respectively）（P 〈0.05）. No significant difference was observed in GSH level （（181±26） nmol/L in the control group vs. （170±18） nmol/L in the CS group） between the two groups. After treatment with NAC, the pathologic score of small airways （24.1±17.5） decreased （P 〈0.05）. Clara cell particles in cytoplasm of Clara cells increased and GSH level in plasma （（213±40） nmol/L vs. （170±18） nmol/L in the CS group） increase

关键词： tobacco smoke pollution Clara cells N-acetylcysteine glutathion

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Effect of cigarette smoke extract on proliferation of rat pulmonary artery smooth muscle cells and the relevant roles of protein kinase C

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Chinese Medical Journal 2007年第17期120卷 1523-1528页

作者： HU Jing XU Yong-jian ZHANG Zhen-xiang TIAN Feng Department of Respiratory Medicine Tongji Hospital Affiliated toTongji Medical College Huazhong University of Science andTechnology Wuhan 430030 China

Background Increased proliferation of pulmonary vascular cells and muscularisation of pulmonary vessels are frequently observed in human smokers and in animals exposed to cigarette smoke. To elucidate the molecular mechanisms leading to these changes, we studied the in vitro effect of cigarette smoke extract （CSE） on proliferation of pulmonary artery smooth muscle cells （PASMCs） and activation of protein kinase C （PKC）, an important kinase implicated in cell proliferation. Methods PASMCs cultured from 12 normal Wistar rats were studied in the following conditions：（1） PASMCs were exposed to different concentrations of CSE for 24 hours, then MTT colorimetric assay was used for detection of cell proliferation. Cell viability was assessed by trypan blue exclusion. （2） PASMCs were pre-incubated with phorbol 12-myristate 13-acetate （PMA） for 24 hours or Ro31-8220 for 30 minutes before exposure to 5% CSE for 24 hours. Cell proliferation was examined by MTT colorimetric assay, cell cycle analysis and proliferating cell nuclear antigen （PCNA） immunocytochemical staining. （3） PASMCs were exposed to 5% CSE for 24 hours. Then PKC-a mRNA expression was detected by reverse transcription-polymerase chain reaction （RT- PCR） and protein expression by Western blotting, while PKC-α translocation was observed by immunofluorescence staining and confocal microscopy. （4） PASMCs were transfected with specific antisense oligodeoxynucleotides against PKC-a 6 hours before exposure to 5% CSE for 24 hours. PKC-α protein expression and cell proliferation were detected by methods described previously. Results （1） Low concentration of CSE （5%） increased proliferation of PASMCs, whereas high concentrations （20%, 30%） were inhibitory as a result of cytotoxicity. （2） The value of absorbance （Value A）, proliferation index （PI）, S-phase cell fraction （SPF） and average optical density of PCNA staining in PASMCs from 5% CSE exposure group （0.306 ± 0.033, 0.339 ± 0.033, 0.175 ± 0.021, 0.315 ± 0.038,

关键词： tobacco smoke pollution protein kinase C pulmonary artery muscle, smooth cell proliferation

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