钙蛋白酶介导的酪氨酸激酶受体B截断对剖腹探查术后小鼠认知功能的影响

目的 观察海马酪氨酸激酶受体B（TrkB）在小鼠术后认知功能障碍（POCD）中的表达变化，并探讨钙蛋白酶活化是否参与其调控。
方法 16月龄雄性C57BL/6小鼠54只，体重28~30 g，随机分为三组，每组18只：对照组（C组）、七氟醚麻醉+剖腹探查组（P组）和七氟醚麻醉+剖腹探查+钙蛋白酶抑制剂MDL28170组(PM组)。C组吸入纯氧；P组采用吸入3%七氟醚+剖腹探查术，建立POCD动物模型；PM组在七氟醚麻醉前即刻腹腔注射钙蛋白酶抑制剂MDL28170（20 mg/kg），其后每天注射1次直至术后第5天。C组和P组腹腔注射生理盐水20 mg/kg。术后第5天进行旷场试验，记录小鼠探索路程及停留在中央格的时间，第6天进行条件性恐惧试验训练，第7天进行场景性条件性恐惧试验测试，记录小鼠5 min内僵直反应（除呼吸运动外无其他运动的状态）时间。于术后24 h每组取6只小鼠海马组织，采用Western blot法检测钙蛋白酶活性；行为学测试完成后2 h每组取6只小鼠海马组织，采用Western blot法检测全长型TrkB受体（TrkB-FL）、截断形式的TrkB受体（TrkB-T₁）、突触后致密蛋白-95（PSD95）的含量。
结果 旷场试验中，三组小鼠总的探索路程和中心区域停留时间差异无统计学意义。与C组比较，P组僵直反应时间百分比明显降低，海马TrkB-FL、PSD95含量明显降低，海马钙蛋白酶活性明显增高、TrkB-T₁含量明显升高（P＜0.05）；与P组比较，PM组在场景性恐惧测试中僵直反应时间百分比明显增高，海马TrkB-FL、PSD95含量明显升高，海马钙蛋白酶活性降低、TrkB-T₁含量明显降低（P＜0.05）。
结论 钙蛋白酶活化截断TrkB-FL，导致TrkB-T₁表达增多，损害小鼠术后认知功能。

Effect of calpain-mediated truncation of tyrosine kinase receptor B on the postoperative cognitive dysfunction of aging mice after laparotomy surgery

Ojective To observe the levels of tyrosine kinase receptor B(TrkB) in postoperative cognitive dysfunction(POCD) in aging mice and to investigate whether calpain activation was involved in the underlying mechanisms.
Methods Fifty-four male C57BL/6 mice,16 month olds, weighing 28-30 g, were equally randomized into three groups (n = 18): control group (group C), sevoflurane anesthesia+ surgery group (group P), sevoflurane anesthesia+ surgery+ MDL28170 group (group PM). Group C was given 100% oxygen inhalation. Group P adopts POCD animal mode which was established by inhaling 3% sevoflurane anesthesia and exploratory laparotomy. In the group PM, mice were intraperitoneally injected with MDL28170 (20 mg/kg) before anesthesia and once a day until 5 d after the operation. The animals in the rest of groups received the equal volume of saline. Open field test was performed 5 d after the surgery to record the exploration distance and the time in center. Fear-conditioned training was performed 6 d after the surgery, and fear-conditioned test was performed 7 d after the surgery to record the freezing time within 5 min (without respiratory movement). The hippocampus tissues of 6 mice in each group were collected 24 h after surgery to detect the activation of calpain by Western blot method (n = 6). The hippocampus tissues of the rest were collected 2 h after the behavioral tests(n = 6). The content of TrkB-FL, TrkB-T₁, and PSD95 were detected by Western blot.
Results In the open field test, no significant difference was observed in the total travel distance and the time spent in the center of arena. Compared with group C, the percentage of freezing time significantly reduced in group P, and the expression levels of TrkB-FL and PSD95 were significantly decreased, activation of calpain and the expression levels of TrkB-T₁ both were significantly increased in group P (P < 0.05). Compared with group P, the percentage of freezing time and the expression levels of TrkB-FL and PSD95 were significantly increased, the activation of calpain was limited, and the expression levels of TrkB-T₁ were significantly decreased in group PM (P < 0.05).
Conclusion Calpain-mediated truncation of TrkB-FL could lead to the levels of TrkB-T₁ increase, and may be involved in the pathophysiology of POCD in aging mice by damaging postoperative cognitive competence.