Résumé
Objective:To detect the expression level of RAB39B gene and the effect of RAB39B on autophagy and α-synuclein, and then investigate the role of RAB39B gene mutation c.536dupA in the pathogenesis of Parkinson′s disease.Methods:Based on the novel RAB39B gene c.536 dupamutation identified in the previous work, the recombinant expression plasmid (pcDNA3.1-HA-RAB39B-536) of RAB39B gene with this mutation and wild-type recombinant expression plasmid (pcDNA3.1-HA-RAB39B) of RAB39B gene were constructed, and the recombinant expression plasmid was transfected into N2a cells with liposome as experimental group. The control group was made up with N2a cells transfected with plasmid pcDNA3.1-HA-RAB39B. Real-time polymerase chain reaction, Western blotting, immunofluorescence and immunoprecipitation techniques were used to detect the expression level of mutant RAB39B gene and the effects of RAB39B on autophagy and α-synuclein.Results:In the N2a cell model, the transcription level of mutant RAB39B was about twice that of wild type RAB39B, while the protein level of mutant RAB39B (0.30±0.00) was significantly lower than that of wild type (1.50±0.25, t=8.313, P<0.05). After adding proteasome inhibitor MG132, the protein level of mutant RAB39B increased (0.70±0.10, t=6.925, P<0.05); the level of microtubule-associated protein 1 light chain 3 BⅡ/Ⅰ of mutant RAB39B (3.11±0.30) was significantly lower than that of wild type (7.03±0.20, t=18.831, P<0.05); overexpression of wild type and mutant RAB39B did not affect the level of endogenous α-synuclein; overexpression of wild-type RAB39B resulted in elevated level of exogenous wild-type (p.A53T; from 0.60±0.11 to 1.25±0.08, t=8.254, P<0.05) and mutant (from 0.55±0.08 to 1.15±0.08, t=9.293, P<0.05) α-synuclein. Conclusions:The stability of the RAB39B protein decreased with the appearance of c.536 dupA mutation, the mutant protein may be degraded through the ubiquitin-proteasome pathway, and this mutation may affect the autophagy level of cells. RAB39B protein may interact with α-synuclein in vivo and may be involved in the maintenance of the stable level of α-synuclein.
Résumé
Objective To investigate the effects of sevoflurane wash-in during cardiopulmonary bypass (CPB) on myocardial injury in patients undergoing coronary artery bypass grafting(CABG).Methods Forty ASA Ⅱ or Ⅲ patients aged 50-64 yr,weighing 53-90 kg undergoing scheduled for CABG under CPB were randomly divided into 2 groups (n =20): control group (group C) and sevoflurane group(group S).Anesthesia was maintained with propofol 3-5 mg·kg-1 ·h-1 and sufentanil 0.5-1.0 μg·kg-1 ·h-1 in both groups.Sevoflurane 1％-2％ was washed into extracorporeal circuit during CPB in group S.Blood samples were taken from central vein after the induction of anesthesia (T0,baseline) and at 6,12 and 24 h (T1-3) after operation for determination of plasma cardiac troponin I(cTnI) concentration and creatine kinase-MB (CK-MB) activity.Myocardial specimens were obtained from right auricle before aortic cross-clamping and at the end of CPB for ultrastructure examination.The severity of mitochondria injury was assessment and scored (0 =normal,4 =impaired inner mitochondrial membrane integrity).Results CPB significantly increased plasma cTnI concentration at T1-3 as compared with the baseline values at T0 before CPB.Plasma cTnI concentration was significantly lower at T2 and T3 in group S than in group C.Mitochondrial injury index was significantly lower at the end of CPB in group S than in group C.There was no significant difference in plasma CK-MB activity between the 2 groups.Conclusion Wash-in of sevoflurane during CPB can attenuate myocardial injury in patients undergoing CABG.