The Role Of MicroRNA-23a In Cigarette-induced Mice Models Of Emphysema

Objective:In this study,a mouse model of emphysema was constructed by intraperitoneal injection of cigarette smoke extract(CSE)combined with cigarette smoke(CS).The expression level of Micro RNA-23a(mi RNA-23a),the concentration of Tumor necrosis factor-α(TNF-α),and the apoptosis index of alveolar septum cells were measured to investigate the role of mi RNA-23 a in mouse model of emphysema and its relationship with TNF-α and apoptosis of alveolar septal cells to provide theoretical basis for clinical research on the pathogenesis of emphysema and seek new treatment.Methods: Twenty-three male C57BL/6J mice were numbered and randomly divided into three groups: emphysema 4 weeks group,emphysema 8 weeks group and normal control group.There were 8 mice in each model group,and 7 mice in normal control group.The mice of emphysema 4 weeks group were exposed to CS for four weeks.They were intraperitoneally injected with(CSE+phosphate buffer(PBS))on the third day,the fourteenth day,and the twenty-fifth day during this period time.The mice of emphysema 8weeks group were exposed to CS for eight weeks.They were intraperitoneally injected with(CSE+PBS)on the third day,the fourteenth day,and the twenty-fifth day during this period time.The mice of normal control group were intraperitoneally injected with PBS on the third day,the fourteenth day,and the twenty-fifth day.Blood,Bronchoalveolar Lavage Fluid(BALF)and lung tissue of mice in the three groups were collected within 24 hours after completion of molding.The following tests were completed in time:(1)The expression level of mi RNA-23 a in lung tissue of mice was detected by quantitative real-time PCR(QRT PCR).(2)The pathological changes of lung tissues after Hematoxylin-eosin(HE)staining were observed and calculated.Mean Linear Intercept(MLI)and Mean alveolar Numbers(MAN)were calculated.(3)The concentration of TNF-α in BALF was determined by Enzyme Linked Immunosorbent Assay(ELISA).The total number of cells in BALF was calculated by Nexcelom Cellometer.After BALF centrifugation,the precipitates were prepared and stained and inflammatory cells(leukocytes,macrophages and neutrophils)were counted.(4)Apoptosis index of alveolar septal cells was determined by Terminal deoxynucleotidyl trans ferasemediated d UTP Nick end labeling(TUNEL).Results:(1)Comparison of the general situation of mice in the three groups: mice in the normal control group were agile and had normal diet.After smoke exposure,the emphysema model group gradually showed easy hair loss,shortness of breath,increased oral secretions,increased nasal scratching and slow reaction.(2)Comparison of mi RNA-23 a in lung tissues of mice in the three groups:the emphysema 4 weeks group > the emphysema8 weeks group > the normal control group.The difference among the two model groups and the normal control group was statistically significant(P < 0.05).(3)The comparison of MLI in lung tissues of mice in three group was as follows: the normal control group <the emphysema 4 weeks group <the emphysema 8 weeks group,and the differences among all groups were statistically significant(P < 0.05).The comparison of MAN in lung tissues of mice in the three groups was as follows: the normal control group >the emphysema 4weeks group > the emphysema 8 weeks group,and there was statistical significance among all groups(P < 0.05).(4)The number of white blood cells,neutrophils and macrophages in BALF of mice in three group was in the order of the emphysema 8 weeks group >the emphysema 4 weeks group > the normal control group,with statistical significance(P <0.05).The concentration of TNF-α in BALF of three groups: the emphysema 8 weeks group >the emphysema 4 weeks group > the normal control group,and there was significant difference among three groups(P < 0.05).(5)Comparison of alveolar septum apoptosis index among all groups: the emphysema 8 weeks group > the emphysema 4weeks group >the normal control group,and the difference was statistically significant among all groups(P< 0.05).(6)The level of mi RNA-23 a in the emphysema 4 weeks group was positively correlated with the apoptosis index of alveolar septal cells(r =0.806,P<0.05).TNF-α concentration was positively correlated with the apoptosis index of alveolar septal cells(r =0.73,P < 0.05).There was a positive correlation between mi RNA-23 a level and TNF-α concentration in lung tissue(r =0.85,P < 0.05).(7)The level of mi RNA-23 a was positively correlated with the apoptosis index of alveolar septa in the emphysema 8 weeks group mice(r =0.743,P < 0.05).TNF-α concentration was positively correlated with the apoptosis index of alveolar septal cells(r =0.89,P < 0.05).There was a positive correlation between mi RNA-23 a level and TNF-α concentration in lung tissue(R =0.74,P < 0.05).Conclusion:(1)Intraperitoneal injection of CSE combined with fumigation induced apoptosis of alveolar septum cells and emphysema in mice.(2)The expression of mi RNA-23 a was positively correlated with the concentration of TNF-α and the apoptosis of alveolar septum cells.It is speculated that mi RNA-23 A may be involved in the formation of emphysema by regulating the function of TNF-α to induce the apoptosis of alveolar septal cells.(3)Airway inflammation and apoptosis of alveolar septal cells may be further increased with the extension of fumigation time.