P53 And Bcl-2 Expressions In The Eosinophils Of Asthmatic Rats Model And Effect Of Glucocorticoid

PrefaceEosinophil (EOS) infiltration in the airway is the characteristic of asthma. Recently, it has been found a failure in EOS apoptosis contributed to the prolonged EOS living, which was related to asthma inflammation, but the exact mechanism of apoptosis is not clear. B - cell lymphoma/ leukima - 2 ( Bcl - 2) is the first - studied gene related to apoptosis. It can inhibit cell apoptosis by changing cell cycles and activing anti -oxide mechanism. p53 is a oncogene Recently is discovered to be closely correlated with apoptossis. But further study should be done to find if p53 participates in EOS apoptosis in asthma. Glucocorticoids (GCs) are potent anti - inflammatory drugs in the asthma therapy, but the mechanism is still unknown .In our study, we remade asthma rat model and treated it using dexametha-sone( DEX). Then the expressions of apoptosis gene Bcl -2 and p53 in EOS of BALF were measured to study the effect of Bcl - 2 and p53 in EOS apoptosis and the role of GCs in changing Bcl -2 and p53 expression.MaterialWistar rats; Jsc - Ok ultrasound nebulizer; Universal 32R zentrifugen; MetaMorph image analysis system; Ovalbumin; Dexamethasone; Rabbit anti -p53; Rabbit anti -Bcl -2;SABC Histotain TM - Plus Kits;Percoll. .MethodsThirty - six rats were divided into control, asthma and DEX groups. The animals were weighted about 200g at the time of testing. The animals of asthma group and DEX group were immunized by intraperitoneal ( i. p. ) injection of 10% OVA on the first day of study,and challenged with an aerosol of 1% OVA on day 15 after the initial sensitization. The challenge was repeated 3 times. The animals of control group received mock sensitization with i. p. injection of sterile normal salt solution( NS) and were challenged with an aerosol of NS. After challenge, the animals of DEX group received i. p. injection of 0. 5mg/kg DEX every day. The injection was repeated 5 times.The animals were anesthetived and bronchoaveolar lavage ( BAL) of left lungs (5ml ×6) were performed. And the right - upper lobes were fixed with 4% paraformaldehyde via a trachealcannula and immersion in the fixative for a minimum peroid of 1 week. The lung tissues were embedded in paraffin and cut in 5 μm sections. The right - lower lobes were cut and stored - 70℃. 3ml BALF of each animals was counted for total cell numbers and different type of cells. The rest BALF was extracted EOS. The sections were stained with hematoxylin and eosin(HE). Immunohistochemistry was performed to detemine the bronchial expression of p53. Western blot was performed to examine p53 and Bcl -2 in EOS and lungs tissue.Statistical analysis: SPSS was used to deal with all the data. Data were expressed as mean (SE) values, Analyses of variance ( ANOVA) was performed ,using the LSD test to adjust for multiple comparisons, A value of P <0. 05 was considered to indicate a significant difference.ResultsThe HE stained section of asthma group saw bronchial extraction and infiltration of inflammatory cells,which was mainly EOS.The results of BALF: The total number of cell and EOS% in asthma group(47. 67 ±2. 50) × 10~7/L,EOS% (16. 25 ±0. 69)% were higher than those of the control group(30 ± 1.71) × 10~7/L,EOS% (6. 69/ ±0. 37)% (P <0. 01); and the two data of DEX group(33. 33 ± 1. 67) × 10~7L,EOS% (12. 74 ±0. 97 ) % were lower than those of asthma group( P < 0. 01).Bcl -2 in EOS of BALF:IDV in asthma group (14333 ±858) was signifi-candy high compared with that of control group (9867 ± 261 ) and DEX group (10867 ± 98 ) ( P < 0. 05 ). Bcl - 2 in lung: IDV in asthma group(10933 ± 806) increased compared with that of control group (6067 ±261) and DEX group (8067 ±197) (P<0.05).p53 in EOS of BALF:IDV in asthma group (10611 ± 1470) was relativelylow compared with that of control group (21060 ±3494) and DEX group (18549± 598) ( P < 0. 05). p53 in lung: IDV in asthma group(22437 ± 3432) was highcompared with that of control group (6561 ± 1795) and DEX group (9717 ±950)(P<0.05).The expression of p53 in bronchial and cells evidenced by immunohisto-chemistry was high than that of control and DEX group( P <0. 05).DiscussionAsthma is a chronic disorder of airway and EOS is considered to be a major contributor to the chronic inflammation . It has been showed recently that there is a defect in EOS apoptosis in asthma; But the exact mechanism of apoptosis remains unclear. The balance of oncogene Bcl - 2 and anti - cogene p53 plays a potent role in the mechanism of regulating cell apoptosis. Bcl -2 can inhibit cell apoptosis, wheras p53 induce it. Our study showed that EOS expressed there is an increase in Bcl - 2, wheras there is a decrease in p53. The overexpression of Bcl - 2 played an important role in reducing apoptosis of inflammatory cell, which suggested asthma existed in imbalance between Bcl - 2 and p53.Glucocorticoid has been proven to be one of the most effective agents in asthma. We found the total number of cells and percent of EOS in BALF of DEX group dropped signiflcandy, it showed glucocorticoid functioned partly by decreasing the number of EOS. The expression of Bcl - 2 in EOS decreased and expres-...