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目的探讨多药耐药蛋白(P 糖蛋白,P-glycoprotein,P-gp)对 FK506透过血脑屏障进入脑内浓度及其脑保护作川的影响。方法用腔内线栓法制造小鼠脑缺血再灌注动物模型(MCAO);用甲酚紫染色法显示脑梗死区;用 Western blot 方法观察 P-gp 的表达;用 ELISA 法检测 FK506浓度;用 TUNEL 法观察凋亡细胞。结果 MCAO 30 min 再灌注3 h 纹状体 P-gp 的表达开始显著升高,持续至24 h,MCAO 90 min 组 P-gp 的表达升高幅度更大。脑缺血前后,P-gp 抑制剂 tariquidar(TQD)对血中 FK506浓度的影响差异无统计学意义;使用 TQD 后,缺血侧大脑半球 FK506含量显著升高。单独使用 FK506时,只有较大剂量(5 mg/kg)才有抗细胞凋亡作用。单独使用 TQD 无抗细胞凋亡和减少脑梗死体积的作用,但当 TQD 与 FK506合用时,FK506较低剂量(1 mg/kg)即可发挥抗细胞凋亡作用,使脑梗死体积从(113.5±11.1)mm~3显著降低至(70.6±10.2)mm~3(P<0.01)。结论 TQD 抑制P-gp 活性,增加 FK506进入脑内的浓度,降低 FK506脑保护作用的阀值(从5 mg/kg 降低至1 mg/kg),从而加强 FK506的神经保护作用。
Objective To investigate the effect of multidrug resistance protein (P-glycoprotein, P-gp) on the concentration of FK506 in the brain through the blood-brain barrier and its protective effect on brain. Methods The cerebral ischemia-reperfusion animal model (MCAO) was established by intraluminal suture method. The cerebral infarction area was observed by cresyl violet staining. The expression of P-gp was observed by Western blot. The concentration of FK506 was determined by ELISA. TUNEL method was used to observe apoptotic cells. Results The expression of P-gp in striatum began to increase 30 min after reperfusion for 30 min in MCAO, and the expression of P-gp in MCAO 90 min increased more significantly for 24 h. Before and after cerebral ischemia, P-gp inhibitor tariquidar (TQD) had no significant effect on the concentration of FK506 in blood. After using TQD, the content of FK506 in the ischemic hemisphere was significantly increased. FK506 alone had anti-apoptotic effects only at higher doses (5 mg / kg). TQD alone did not have anti-apoptotic effects and decreased infarct volume. However, when TQD was administered in combination with FK506, anti-apoptotic effects of FK506 at a lower dose (1 mg / kg) increased the infarct volume from (113.5 ± 11.1) mm ~ 3 decreased to (70.6 ± 10.2) mm ~ 3 (P <0.01). Conclusions TQD can inhibit the P-gp activity, increase the concentration of FK506 into the brain and decrease the threshold of FK506 brain protection (from 5 mg / kg to 1 mg / kg), thereby enhancing the neuroprotective effect of FK506.