RÉSUMÉ
Objective To evaluate dexmedetomidine-induced cardioprotection in a mouse model of lung ischemia-reperfusion (I/R) and the relationship with endoplasmic reticulum stress.Methods Forty healthy SPF male C57BL/6J mice,weighing 20-24 g,aged 8-10 weeks,were divided into 4 groups (n=10 each) using a random number table:sham operation group (Sham group),lung I/R group (I/R group),dexmedetomidine group (Dex group) and dexmedetomidine plus atipamezole (specific α2-adrenergic receptor antagonist) group (DA group).The model of lung I/R injury was established by clamping the left hilum of lung for 30 min followed by 180 min of reperfusion.In group Sham,only sternotomy was performed,the hilum of lung was not clamped,and the mice were mechanically ventilated for 210 min.In Dex and DA groups,dexmedetomidine 20 μg/kg and dexmedetomidine 20 μg/kg plus atipamezole 250 μg/kg were injected intraperitoneally,respectively,at 30 min before establishment of the model.At 180 min of reperfusion,blood samples were collected from the orbit for determination of creatine kinase-MB (CK-MB) and lactic dehydrogenase (LDH) activities in serum.The animals were then sacrificed,and hearts were removed for determination of apoptosis in cardiomyocytes (by TUNEL) and expression of phosphorylated c-Jun N-terminal kinase (p-JNK),caspase-12,CCAAT/enhancer-binding protein homologous protein (CHOP) and glucose-regulated protein 78 (GRP78) in myocardial tissues (by Western blot),and expression of JNK,caspase-12,CHOP,GRP78 mRNA in myocardial tissues (by real-time polymerase chain reaction).Apoptosis index was calculated.Results Compared with Sham group,the serum CKMB and LDH activities and apoptosis index were significantly increased,the expression of p-JNK,JNK mRNA,and caspase-12,CHOP and GRP78 protein and mRNA was up-regulated in I/R,Dex and DA groups (P＜0.01).Compared with I/R group,the serum CK-MB and LDH activities and apoptosis index were significantly decreased,the expression of p-JNK,JNK mRNA,and caspase-12 and CHOP protein and mRNA was down-regulated,the expression of GRP78 protein and mRNA was up-regulated in group Dex,and the expression of GRP78 protein and mRNA was significantly up-regulated (P＜0.01),and no significant change was found in the other parameters in group DA (P＞0.05).Compared with DEX group,the serum CK-MB and LDH activities and apoptosis index were significantly increased,the expression of pJNK,JNK mRNA,and caspase-12 and CHOP protein and mRNA was up-regulated (P＜0.01),and no significant change was found in the expression of GRP78 protein and mRNA in DA group (P＞0.05).Conclusion Dexmedetomidine can reduce myocardial injury induced by lung I/R,and the mechanism may be related to activation of α2-adrenergic receptors and inhibition of endoplasmic reticulum stress in myocardial cells of mice.
RÉSUMÉ
Objective To observe the effects of Yiqi Huoxue Tongluo Jiedu fang (YHTJF) on pneumocyte apoptosis after lung ischemia/reperfusion (I/R) injury (LIRI) in mice and to investigate whether c-Jun N-terminal protein kinase (JNK) is involved in the mechanism of apoptosis.Methods Seventy C57BL/6J male mice were randomly divided into seven groups:normal control group (C group),carboxyl methyl cellulose-Na+normal control group (CMC-Na+C group),CMC-Na+sham group (CMC-Na+S group),CMC-Na+I/R group (CMC-Na+I/R group) and CMC-Na+YHTJF-low,-middle,-high dose groups (CMC-Na+YL,CMC-Na+YM,CMC-Na+YH groups).C group did not undergo any processing;in CMC-Na+S group,only was chest opened without clipping the lung hilum;in the rest of the four groups,they all underwent opening of the chest and clipping the lung hilum for 30 minutes,then the clipping of artery was relieved and left lung reperfusion was carried out for 3 hours.After operation,the mice were sacrificed,the lung tissues were harvested.Under light and electron microscopes,the lung morphological and ultra-structural changes were observed,and the changes of index of quantitative evaluation for alveolar damage (IQA) were determined.The terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) was applied to evaluate the apoptosis index (AI) of the lung tissues.The protein and mRNA expressions of JNK and glucose regulating protein 78 (GRP78) in lung tissues were detected by Western Blot and reverse transcription-polymerase chain reaction (RT-PCR);the correlations between lung AI and the expressions of mRNA and protein of JNK and GRP78,IQA were analyzed.Results Compared with CMC-Na+S group,IQA,AI and mRNA and the protein expressions of JNK and GRP78 in CMC-Na+I/R group were obviously higher [IQA:(74.00 ± 7.31)％ vs.(7.00 ± 1.23)％,AI:(64.40 ± 11.97)％ vs.(5.60 ± 1.14)％,JNK mRNA (gray value):1.143 ± 0.284 vs.0.152 ± 0.128,GRP78 mRNA (gray value):0.897 ± 0.129 vs.0.284 ± 0.044,JNK protein (A value):0.428 ± 0.074 vs.0.073 ± 0.052,GRP78 protein (A value):1.075 ± 0.145 vs.0.589 ± 0.060].Compared with CMC-Na+I/R group,the IQA,AI,protein and mRNA expressions of JNK and GRP78 in CMC-Na+YL,CMC-Na+YM,CMC-Na+YH groups were all lower,and the degree of reduction in group CMC-Na+YM was the most remarkable,greater than that in CMC-Na+YL or CMC-Na+YH group [IQA:(26.20 ± 3.35)％ vs.(34.00±5.34)％,(41.20±9.18)％,AI:(29.40±3.05)％ vs.(48.20±3.83)％,(39.20±6.14)％,JNK mRNA (gray value):0.681 ± 0.130 vs.0.804 ± 0.153,0.938 ± 0.11,GRP78 mRNA (gray value):0.450 ± 0.105 vs.0.747 ± 0.231,0.566 ± 0.115,JNK protein (A value):0.188 ± 0.049 vs.0.261 ± 0.065,0.209 ± 0.063,all P ＜ 0.01],compared with the CMC-Na+I/R group,the expression of GRP78 protein was obviously higher in CMC-Na+YH,CMC-Na+YL,CMC-Na+YM groups and the most remarkably high was in CMC-Na+YH group (A value:1.429 ±0.226 vs.1.130±0.169,1.128 ±0.177,all P ＜ 0.01).The apoptosis of each group was mainly in the pulmonary vascular endothelial cells and alveolar epithelial cells,and brown particles were positive cells under light microscope.Under transmission electron microscope:nuclear pyknosis and margination under the nuclear membrane,cytoplasm condensed,lamellar bodies decreased and emptying increased,cell membrane microvilli decreased or disappeared,mitochondria swelling,inflammatory cells increased in alveolar septum and adhering onto the capillary walls could be seen in CMC-Na+I/R group.Compared with CMC-Na+I/R group,the lung tissue ultrastructural damage alleviated,ultrastructure of alveoli clearly seen,nuclear chromatin relatively uniform,cytoplasm increased,type Ⅱ alveolar epithelial cell surface microvilli relatively plenty,lamellar corpuscle number increased,mitochondria swelling ameliorated in CMC-Na+YH,CMC-Na+YL,CMC-Na+YM groups and the most remarkable one was CMC-Na+YM group.AI was significantly positive correlated with the mRNA and protein expressions of JNK,GRP78 and IQA (r =0.907,0.928,0.880,0.712,0.911,all P ＜ 0.01).Conclusions YHTJF may effectively alleviate the cell apoptosis in mice LIRI,and its mechanism may be related to the inhibition of JNK pathway.
RÉSUMÉ
AIM:To investigate the effects and c-Jun N-terminal kinase(JNK) expression of dexmedetomidine (DEX) on A549 cells with hypoxia/reoxygenation (H/R) injury.METHODS:A549 cells were cultivated and were randomly divided into four groups (n =10):control group (N),DEX group (D),hypoxia/reoxygenation injury group (H),hypoxia/reoxygenation injury + DEX interfere group (HD).After all models were completed,the morphological changes of A549 cells were observed under the inverted microscope.Cell activity was detected by CCK-8 and the apoptosis index (AI) was detected by in situ end labeling (TUNEL) method.The expression of GRP78,p-JNK,caspase-3 at protein levels and GRP78,JNK mRNA were detected by Western blot and RT-PCR.RESULTS:Compared with N group,the number of adherent cells in H group decreased significantly,and cell morphology changed.The expression of OD value in H group decreased obviously (P ＜ 0.01),the expression of AI value,GRP78,p-JNK,caspase-3 protein and GRP78,JNK mRNA were significantly increased (P＜ 0.01).HD group compared with H group,the cell damage alleviated,the expression of OD value was increased (P ＜ 0.01),the number of apoptosis cells and the AI value in HD group were significantly decreased (P ＜ 0.01).Dramatically decreased the expression of p-JNK,caspase-3 protein and JNK mRNA (P ＜ 0.01).CONCLUSION:DEX can effectively alleviate A549 cells damage induced by H/R injury,which may be related to inhibition of the JNK pathway.