Curcuma Oil Intervention In Pulmonary Fibrosis Epithelial Cell Transdifferentiation Of Experimental Research

Pulmonary fibrosis(PF) is a kind of diffuse interstitial lung disease which caused by many causes and has similar pathogenesis, Its pathogenic characterize is which alveolar structure destruction and diffuse alveolitise in the early, then cause over aggradation of extracellular matrix(ECM) and gaudily leading to fibrosis, consequently cause respiratory function destruction. Most of the patients died from respiratory failure from progressive pulmonary fibrosis. In clinic idiopathic pulmonary fibrosis(IPF) is the most common. IPF is a kind of pulmonary fibrosis, however, the mechanisma are still unknown. An emerging concept is that the alveolar typeⅡepithelial cells transition into mesenchymal cells via a process termed epithelial-mesenchymal transition(EMT). As pollution, environmental degradation, and so the number of seizures have increased trend every year. The laboratory studies of chinese medicine in the control organs fibrosis found that Zedoary Oil has the effect of remedying pulmonary fibrosis pharmacology. Zedoary Oil is an effective part in Zedoary. Studies show that Zedoary Oil can inhibit the donor cell proliferation, promote the donor cell apoptosis. But the control mechanisms of cellular and molecular about Zedoary Oil are not clear yet. To evaluate the effect of the pathogenesis and role of pulmonary fibrosis induced by Zedoary Oil. Mice model induced by Bleomycin(BLM) and alveolar epithelial cellⅡ(A549)as a model, and describes the effect of Zedoary Oil proliferation, and to discuss the potential mechanism of the inhibiting effect of Zedoary Oil pulmonary fibrosis.Methods & Results1 The chemical constituents and yield of the Zedoary Oil1.1 Method The chemical componentswere of Zedoary Oil are separated and identified by GC/MS. Taking Nist98 standard mass spectrometer database and contrast as control, the structure of themass spectrogram was identified, and relatively reserved time and peak area percentage occupy the peak total area percentage.1.2 Results 23 compositions were identified from Zedoary Oil. Three single peaked area are larger than ten percentage of the peak total area. Three single peaked area are one out of five or ten of the peak total area,β-Elemene and Curcumol can inhibit the tumor cells growth in the 23 compositions.2 To study medicine of the Zedoary Oil2.1 Methods①96 IR mice was divided into four groups:modle group (BLM), medium dose group(Oil M), the high dose group(Oil H) and sham operation group (Sham). A single dose of 0.05 U bleomycin in 100μL of saline'solution was administered by transtracheal puncture. Oil M and Oil H reeeived the same amount of bleomycin as BLM, followed by different amouts Zedoary oil. Sham received normal saline'solution by tratracheally. These mice were killed on 7,14,28 day post bleomycin.The lung tissues were obtained.②Apply the HE staining to investigate the lung inflammation and lung fibrosis under the routine mieroseope.③Use the immunohistochemistry staining and western blotting to study the expression of collagenⅠ,collagenⅢ,TGF-β1,Smad3,Cat B,Cat D and Cat K. Use image analysis software to measure the density.④Use the real time PCR to measure the expression of Cat B. Cat D,Cat K and Cat S.⑤The lung hydroxyproline content was analyzed using a enzymes response method in minced lung.2.2 Results HE staining display that the alveolar structure of sham is complete and clear. There were no inflammatory cells in alveolus. pulmonary fibrosis were obviously in sham. Consolidation area appeared around bronchus:abnormal thickening of alveolar walls, architectural distortion of alveolar, exudation in alveolus. As time went on, pulmonary fibrosis become serious. thickening and Exudation were seen in sham group that didn't exist in rest group.Mallory staining results:the collagen layer of tissue section of lung of mice were thin around bronchus. There were the change visible in alveolus.Collagen fibers and deposition of BLM group increased considerably in interstitial lung, alveolar, bronchus, vessels.Blue fabric of BLM group increased obviously in alveolus interval.The changes of two treatment groups is lower than BLM group.Immunohistochemistry(IHC) staining results:normal structure of alveolar was destroyed by the deposition of I type collagen as fibrosis went on. The changes of two treatment groups is lower than BLM group.Ⅲtype collagen primarily on the vessels and bronchi. The collagen increased day by day and reached the highest at 7th day. deposition ofⅢtype collagen on the lung break down along with times.The expression ofⅢtype collagen approximate the normal level at 28th day.Compare to sham grounp, the content ofⅢtype collagen increassd day by day and reached the highest at 7th day, second at 14th day.The content get less at 28th day. two treatment groups can reduce the deposition of III type collagen on the lung.EMT of signal elements and Cathepisins of IHC staining results:EMT were urging TGF-β1 activated. The expression ofα-SMA in alveolar increased, and so the expression of Smad3. activated Cat K,Cat B and Cat S can promote degradation of collagen. Activated Cat D is correlated with pulmonary fibrosis. Compare to sham grounp, the expression of singnal in EMT decreased. the expression of Cat K,Cat B and Cat S increased. Cat D was inhibited.3 The pharmacologic mechanism of proliferation of A549 cell by the Zedoary Oil3.1 Methods The experiment measures that the effect on the regeneration of cell, the regulation of cell, the apoptosis of cell, the deposition of collagen and up cathepsins expression when the Zedoary Oil act on open educational A549 cell.3.2 Results when the concentration of the Zedoary Oil is 80 mg·mL-1, cell cycle was arrested at S stage. At the same time, some cell was apoptosis or necrosis. the expression of Cat D and Cat K increased.Conclusion1 The chemical componentswere separated and identified by GC/MS.23 compositions were identified from Zedoary Oil,β-Elemene and Curcumol which can inhibit the tumor cells growth.2 The Zedoary Oil can promote the gas eschange and the survival rate through maintaining the normal structure of alveolar. The exhibition of basis of cell give priority to the deposition of I type collagen.at the same time, the deposition of III type collagen increased.3 The Zedoary Oil can prevent the formation of Pulmonary fibrosis through inhibiting EMT in the modle of Pulmonary fibrosis which induced by BLM.4 The Zedoary Oil can prevent the formation of Pulmonary fibrosis throug hinhibiting some immunocompetence of TGF-β1, increasing the expression of Cat K,Cat S and Cat B mRNA and reducing the deposition of EMT. Cat D different from Cat K,Cat S and Cat B,5 The Zedoary Oil can inhibit the regeneration of cell through arresetting cell cycle at S stage and reducing the rate of cell division.80 mg·mL-1 can increase the expression of Cat D though accelerating cell senescence.