| Objective: To Observe the effects of all-trans retinoic acid (ATRA) onMMP-9/TIMP-1balance and TGF-β1in asthmatic rats. In the pathogenesis ofasthma, a variety of cellular components and cytokines were involved,MMP-9/TIMP-1and TGF-β1play an important role in the regulation of cellmatrix formation and degradation, airway fibrosis, airway vascularremodeling, so we want to explore the effects of ATRA on airway remodelingin asthmatic rats by measuring MMP-9, TIMP-1and TGF-β1levels and mayfind a new drug way for the clinical treatment of bronchial asthma.Methods: Thirty6-week-old SPF SD male rats, weighing between100-120g, were randomly divided into normal control group, asthma modelgroup, ATRA intervention group(10rats in each group). Both the asthmamodel group and the ATRA intervention group were sensitized byintraperitoneal injection of OVA100mg and aluminum hydroxide100mg(mixed in0.9%sodium chloride solution2ml) on first and8thday. And thenthe rats were placed in a non-fully enclosed self-made simple atomizationchamber OVA inhalation challenge test solution since the15thday, theconcentration of1%,1.5%,2%,2.5%,3%OVA solution in ascendingconcentration order, increasing concentration once every four times inhalation,the chanllenge test was carried out every other day, that lasting30minuteseach time, and total of20times. The control group was given0.9%sodiumchloride to replace OVA, the challenge procedure as same as OVA group.ATRA intervention group was gavaged with ATRA (20mg/kg) mixed with0.9%sodium chloride30minutes before each challenge. After the24hourslast challenge the rats were sacrificed with Applications1%sodiumpentobarbital anesthesia. The BALF, blood serum and lung tissue of the ratswere collected. the number of total cells and neutrophils, eosinophils, lymphocytes in BALF were Detected. The concentration of TGF-β1in serumwas detected through ELISA method. The MMP-9、TIMP-1levels of the lungtissue were detected by immunohistochemistry qualitative analysis. Part of thelung tissue were prosessed with hematoxylin-eosin staining, the bronchialwall thickness (wall area/wall circumference) and bronchial smooth musclethickness of the lung tissue were measured under pathological microscopymagnify200times, the relative expression of MMP-9and TIMP-1werecaculated with the computer image analysis software.Results:1Rats bronchial asthma modelBy intraperitoneal injection of OVA sensitization and repeatedatomization inhalation chanllenge, we successfully established the ratbronchial asthma model. Asthmatic rats in the experimental process were seenagitating, shortness of breath, oral cyanosis, abdominal breathing, abdominalcramps, unstable activity performance,etc.2The results of total number of cells in BALF and cell classification（106/L）The total number of cells in BALF of asthma group (33.91±3.67)higher than that of ATRA group (12.88±2.72), there is a significantdifference between the groups.(P <0.01); And the two groups were higherthan in normal control group (9.68±2.71), there are significant differenceamong the groups(F=183.370, P <0.01).The eosinophils in BALF of asthma group (5.38±0.63) higher than thatof ATRA group (2.19±0.53), there is a significant difference between thegroups (P <0.01); And the two groups were higher than that in normal controlgroup (1.17±0.48), there are significant difference among the groups (F=160.003, P <0.01).The neutrophils in the BALF of asthma group (1.14±0.20) higher thanthat of ATRA group (0.65±0.18), there is a significant difference betweenthe groups (P <0.01); And the two groups were higher than that in normalcontrol group (0.28±0.12), there are significant difference among the groups(F=63.937, P <0.01). The lymphocyte in the BALF of asthma group (2.99±0.53) higher thanthat of ATRA group (2.28±0.49), there is a significant difference betweenthe groups (P <0.01); and the two groups were higher than in normal controlgroup (1.27±0.36), there are significant difference among the groups (F=34.568, P <0.01).3The pathological image analysis results:The airway Wall thickness and the thickness of the airway smoothmuscle in asthma group were98.78±2.86、45.49±2.40, significantly higherthan the normal control group（70.87±1.80,17.68±1.14），(P <0.01, P <0.01);ATRA group that was86.20±2.36,33.53±1.36, compared with the asthmagroup,they are decreased, but still higher than the normal control group (P<0.01), there was a significant difference among the three groups (F=345.002,P <0.01;F=345.002, P <0.01).4The expression of MMP-9、TIMP-1and MMP-9/TIMP-1by immunohistochemical method:MMP-9: asthma group (0.63±0.02) was significantly higher than thecontrol group (0.22±0.01)(P<0.01), ATRA group (0.41±0.02) wassignificantly lower than the asthma group but still higher than the normalcontrol group (0.22±0.01), there was a significant difference (F=1391.446,P <0.01) between the three groups.TIMP-1: asthma group (0.77±0.03) was significantly higher than thecontrol group (0.21±0.02)(P<0.01), ATRA group (0.44±0.03) wassignificantly lower than the asthma group but still higher than the normalcontrol group (0.21±0.02), there was a significant difference (F=1326.128,P <0.01) between the three groups.MMP-9/TIMP-1: asthma group (0.82±0.01) was significantly lower thanthe control group (1.05±0.02)(P <0.01), ATRA group (0.92±0.01)compared with the asthma group was increased, but still low in the controlgroup (1.05±0.02), there was a significant difference (F=527.533, P <0.01)between the three groups.5The contents of TGF-β1(ng/ml) in serum by ELISA method: Asthma group (28.50±3.72) was significantly higher than the controlgroup (12.87±1.82)(P <0.01), ATRA group (19.18±3.65) was significantlylower than the asthma group but still higher than the normal control group(12.87±1.82)(P <0.01), there was a significant difference (F=60.763, P<0.01) between the three groups.6The result of correlation analysis:The results showed that MMP-9were significantly positively correlatedwith TIMP-1(r=0.999, P<0.01)、TGF-β1(r=0.887, P<0.01) and airway wallthickness(r=0.990, P<0.01); TIMP-1was significantly positively correlatedwith airway wall thickness(r=0.984,P<0.01); MMP-9/TIMP-1wassignificantly negatively correlated with airway wall thickness(r=-0.993, P<0.01); TGF-β1was significantly positively correlated with smooth musclethickness (r=0.876, P<0.01).Conclusions:1The asthma group rats showed obvious infiltration of inflammatory cellsaround the airways,the tube cavity filled a large number of inflammatoryexudate, Pathological image analysis system showed significant airwayremodeling in asthmatic rats,suggesting that we have successfully made thebronchial asthma rat model. Application of ATRA can significantly reduceairway inflammation, suggesting that ATRA has certain anti-inflammatoryeffects.2MMP-9and TIMP-1, TGF-beta1level and airway wall thickness arepositive correlation between four and participate in the airway remodeling.3ATRA has reduced TGF-β1, MMP-9, TIMP-1expression, coordinatedMMP-9/TIMP-1balance,and can slow down the airway remodelingsuggesting that ATRA has the role of anti-airway remodeling. |
PDF Full Text Request