微管干预剂对大鼠缺氧心肌细胞能量生成的影响

The influence of microtubule intervention drugs on the energy metabolism of myocardial cells after hypoxia

目的了解缺氧条件下微管干预剂对大鼠心肌细胞能量生成的影响。方法常规分离、培养大鼠心肌细胞,分为单纯缺氧组、缺氧+秋水仙碱(微管解聚剂)组及缺氧+5、10、15 mmol/L紫杉醇(微管稳定剂)组。每组细胞加入刺激剂后,缺氧培养0.5、1.0、3.0、6.0、12.0、24.0 h。采用锥虫蓝染色检测细胞死亡率,常规比色法检测细胞肌酸激酶(CK)活性,高效液相色谱法检测细胞腺苷三磷酸(ATP)及腺苷二磷酸(ADP)含量。结果(1)缺氧+秋水仙碱组及缺氧+15 mmol/L紫杉醇组细胞培养1.0～24.0 h死亡率均高于单纯缺氧组(P<0.01);缺氧+5、10 mmol/L紫杉醇组6.0～24.0 h时均低于单纯缺氧组(P<0.05)。(2)缺氧+秋水仙碱组培养1.0～12.0 h时CK活性均高于单纯缺氧组(P<0.01)。0.5～12.0 h时,缺氧+15 mmol/L紫杉醇组CK活性均高于单纯缺氧组(P<0.01);缺氧+5、10 mmoL/L紫杉醇组低于单纯缺氧组(P<0.05或P<0.01)。(3)缺氧+5 mmol/L紫杉醇组培养0.5～6.0 h ATP含量[(49.9±2.8)、(40.7±2.0)、(25.8±1.9)、(19.1±1.2)μg/10~6个细胞]高于单纯缺氧组[(42.9±5.8)、(29.5±1.8)、(18.2±0.9)、(14.1±0.7)μg/10~6个细胞,P<0.05或P<0.01]。0.5～12.0 h时,缺氧+秋水仙碱组低于单纯缺氧组(P<0.01);缺氧+15 mmol/L紫杉醇组低于单纯缺氧组及缺氧+10 mmoL/L紫杉醇组(P<0.01)。各组ADP含量变化趋势与ATP相反。结论微管解聚剂和高浓度微管稳定剂可使缺氧心肌细胞ATP含量锐减。适宜浓度的微管稳定剂在缺氧早期可促进心肌细胞能量生成,对心肌具有保护作用。

To investigate the influence of microtubule intervention drugs on the energy metabolism of myocardial cells after hypoxia. Methods The primary passage of cultured myocardial cells from neonatal rats were divided into A （with hypoxia）, B （with hypoxia and administration of 10 p^mol/ml colchicine） , C （with hypoxia and administration of 5 μmol/ml taxol）, D （with hypoxia and administration of 10 μmol/ml taxol） and E（with hypoxia and administration of 15 p^mol/ml taxol）groups. The creatine kinase （CK） activity and contents of ATP and ADP were assayed with colorimetry and HPLC, respectively, and the vitality of myocardial cells were determined by trypan blue method at 0.5, 1.0, 3.0, 6.0, 12.0, 24.0 post-hypoxia hours（PHH）. Results The mortality was obviously higher in B and E groups than those in A group（ P 〈0.05） at each time-points, but that in C and D groups were markedly lower than those in A group during 6.0 to 24.0 PHH（ P 〈0.01 ）. The CK activity was significantly higher in B group than that in A group during 1.0 to 24.0 PHH, while that in E group was evidently higher, but it was lower in C and D groups than that in A group at each time-points（ P 〈0.05 or 0.01 ）. The ATP contents in C group during 0.5 to 6.0 PHH were [（49.9＋2.8）,（40.7＋2.0）,（25.8 ＋1.9）,（19.1 ＋l.2）μg/10^6 cells, respectively ] , which were obviously higher than those in A group [ （42.9 ＋ 5.8 ） , （ 29.5 ＋ 1.8 ） , （ 18.2 ＋ 0.9 ） , （14.1 ＋ 0.7）μg/10^6 cells,respectively, P 〈 0.05 or P 〈 0.01 ], and those in E group at each time-point were significantly lower than those in A and D groups（ P 〈0.01 ）. The changes in the contents of ADP were on the contrary to the above. Conclusion Microtubule-destabilizing drugs and high concentration microtubule-stabilizing drugs can sharply decrease ATP content in myocardiocytes under hypoxic conditions, while suitable amount of microtubule-stabilizing drugs can nrotect mvocardiocvtes by promoting its energy production.