The Study Of Qi Zhi Zhou Fei Granule’s Regulation On Alveolar Type Ⅱ Cells’ Nf-κb Signaling Pathway

Chronic obstructive pulmonary disease(COPD) is a kind of irreversible airway disease characterised by airflow limitation, with high incidence, high disabilities and high death rate, etc. Some studies have found that chronic inflammation of airway and lung parenchyma is an important mechanism in the pathogenesis of COPD, but there will be a systemic inflammatory response during the progress in COPD. Some inflammatory mediators which were released from focus not only aggravate the lung inflammation, but also activate the WBC in peripheral blood and affect other organs or tissue. So the regulation of the inflammatory response is an important way to slow the progression of the disease and improve the symptoms of COPD. Studies show that many inflammatory mediators that lead to fibrosis of small airway and the injury of lung structure are related to the incidence of COPD. Controlling the release of inflammatory mediators may affect the effectiveness of the COPD’s treatment and the action point of anti- inflammatory therapy. COPD’s inflammatory reaction involves a variety of cells, among them, the alveolar type Ⅱ epithelial cell(AT-Ⅱ) has attracted much attention in the research of COPD because of its high differentiation potential and involvement in immune defense, regulating lung water transport and secretion of pulmonary surfactant protein and so on. Nuclear factor kappa B(NF-KB) signaling pathway is widely distributed in various tissues, which affecting IL-6 and other inflammatory mediators’ expression and release, plays an important role in the immune response, inflammatory response, cell proliferation and apoptosis. The regulation of NF- KB signaling pathway is one of the important ways to inhibit lung inflammation and the treatment of COPD. In this paper, we observe the Q i Zhi Zhou Fei granule’s effect on regulation of AT-Ⅱ cells’ NF-KB signaling pathway, and explore the NFKB signaling pathway’s function during the Qi Zhi Zhou Fei granule’s function in inflammatory reaction. To provide the experimental basis for studying the mechanism of action of Qi Zhi Zhou Fei Granule on prevention and treatment of COPD. Objective:To observe Qi Zhi Zhou Fei granule’s regulation of NF-KB signaling pathway of AT-Ⅱ cells and affection of the expression of inflammatory mediators. and provide the basis for the study of regulatory effect of Q i Zhi Zhou Fei granule on COPD’s inflammatory reaction. Methods:According to preliminary experiments methods of primary cell culture to culture the AT-II cells, and identify AT-Ⅱ cells with electron microscop y and immunohistochemical staining of SP-B. Selecting 6 bottles of good growth state of AT-cells, dividing them into control group, model group, Q i Zhi Zhou Fei granule of high, medium and low dose group, the model cells were intervented with 20g/ml LPS for 2hours, and then treated with different concentrations of Q i Zhi Zhou Fei Granule(50g/ml, 100g/ml, 200g/ml), culture them for 36 h to 48 h then detect the related index.(1)Observe the cell growth state with inverted microscope and measure cells’ viability with MTT.(2)Detect the gene expression of CD14、TLR-4、NF-?B、IL-6、IL-12 with Q-PCR. Results:(1) MTT results showed that the primary culture of AT-II cells grew well in 10 days, and the most active proliferation time was in 0~72h; A large number of dark granula r AT-II cells were found under the electron microscope. There were no staining cells in the primary culture of AT-II cells by SP-B immunohistochemical staining which indicated the cells had high purity.(2) Q-PCR results showed that compared with the norma l group, the gene expression of model group(CD14、TLR4、NF-?B、IL-6、IL-12) increased significantly(P<0.05); Compared with the model group, the gene transcription level of Q i Zhi Zhou Fei granules group(CD14、TLR4、NF-?B、IL-6、IL-12) decreased in different degree, and showed a certain concentration dependent,the difference was statistically significant(P<0.05). Conclusions:(1) The AT-II cells cultured in this experiment have a high yield and high purity, which can meet the need of experimental research.(2) Through the regulation of NF-?B signaling pathway, CD14, the activation of CD14、TLR-4、NF-?B and the release of inflammatory mediators of IL-6、IL-12, Q i Zhi Zhou Fei granules controled the inflammatory reaction of lung, which may be one of the mechanisms of prevention and treatment of COPD.