Changes In Proteomic Expression Of Bronches And Lung Of Rats Exposed To Cigarette Smoke And Radon

Objective: To study the early biological effect induced by cigarette smoke and radon in terms of changed in proteomic expressions.Methods: (1) Two dimensional electrophoresis was used to screen for differential protein points. (2) Wistar rats were exposed to cigarette smoke with a gas inhalation equipment with the smoke concentration controlled at the range of (20±5)％and oxygen concentration at 25％. The rats were exposed twice a day,30min each at low,middle and high dose for 1,2 and 4 months. The total protein of tracheal epithelium cells and lung tissue were extracted for 2-DE and silver staining or coomassie brilliant blue staining, and MALDI-TOF or MALDI-TOF-TOF were conducted for protein identification. (3) Wistar rats were exposed to radon and its progeny with a multifunction ecological radon room at a constant level of 100000Bq/KL. The low,middle and high dose group were exposed up to a cumulative dose of 100,200 and 400WLM respectively. The total protein of rat lung were extracted for 2-DE and MALDI-TOF or MALDI-TOF-TOF were conducted for protein identification. (4) The proteins of RAGE and TRX were analyzed by Western blot to confirm the 2-DE results.Results: The two dimensional electrophoresis technique with silver staining resulted in 450 proteins at condition of pH3～10,11cm IPG strip,625 proteins at pH3～10,24cm IPG strip and 1248 proteins at pH4～7,24cm IPG strip which was chosen for later experiment. (2) In rat exposed to smoke, pulmonary emphysema, bronchitis response and fibroglia fibrils were observed especially in the high dose group. 23 proteins were found in the tracheal epithelium cells, in which 11 spots showed an evident dose-effect relationship. 18 different expression proteins were identified in the lung through 2-DE and mass spectrometry, including 10 upwards expression (beta actin, typeⅡkeratin, receptor for advanced glycosylation endproducts, pyruvate dehydrogenase, thioredoxin, ATP synthase, peroxiredoxin6, dnak type molecular chaperones, moesin and AnnexinⅢ),and 8 downwards expression (tropomyosin4, myosin light chain4, serine protease inhibitor B11, dimethyl arginine dimethyl aminohydrolasel, et al). (3) In rat exposed to radon, hyperaemia, hydropsia of the pneumono interstitium, thickening of the lung interval, inflammatory cell infiltration, collagen fibrils and pulmonary emphysema were observed. 15 different expression proteins were identified in the lung through 2-DE and mass spectrometry, including 9 upwards expression (Zinc finger4, alpha enolase, tubulin beta, HSP27, beta actin, thioredoxin, receptor for advanced glycosylation endproducts, S100A6 and Annexin A2), and 6 downwards expression (cytochrome C oxidase, serine protease inhibitor, gamma actin, aldehyde dehydrogenase, myosin light chain4, LOC500450, et al). (4) The proteins of RAGE and TRX were confirmed to show a dose-response relationship with exposures to cigarette smoke and radon, which was coincident with the 2-DE results.Conclusion: The two dimensional electrophoresis technique was established for measurement of proteomic expression of lung tissue in rat exposed to cigarette smoke and radon. Some proteins are commonly expressed such as RAGE, TRX, Serpin, Annexin. Smoke specific expression proteins include typeⅡkeratin, peroxiredoxin6, HSP70, dimethyl arginine dimethyl aminohydrolasel, and radon specific expression proteins include zinc finger4, HSP27, S100A6, cytochrome C oxidase.