维生素D对创伤性脑损伤大鼠学习记忆及脑组织溶酶体钙通道蛋白Mcoln-1的影响

Effects of vitamin D on learning and memory and expression of Mcoln-1 in brain tissue of rats with traumatic brain injury

目的:研究大鼠创伤性脑损伤(traumatic brain injury,TBI)后给予维生素D(Vitamin D,VD)治疗对大鼠认知功能、海马组织与皮层组织内溶酶体活性及其钙通道蛋白Mcoln-1的影响。方法:45只SD大鼠随机分为3组(每组15只):空白对照组(Sham组)、模型组(TBI组)和钙三醇(活性VD,Calcitriol)治疗组(Calcitriol组)。TBI组和Calcitriol组大鼠采用电子脑皮质撞击仪进行TBI造模。Calcitriol组于TBI造模后30 min、24 h、48 h经腹腔注射给予1μg/kg的Calcitriol。造模3 d后,Western blot检测海马组织和皮层组织中Mcoln-1、LAMP-1和cathepsin-B的蛋白表达水平,免疫荧光检测Mcoln-1与神经元的共定位情况。三组大鼠均于第8、9、10天进行Morris水迷宫测试。结果:Morris水迷宫结果显示,与Sham组[(19.54±3.54)s;(18.64±4.63)s;(17.64±5.88)s]比较,在造模后第8、9、10天TBI大鼠寻找平台逃避潜伏期[(58.75±6.65)s;(50.64±5.56)s;(42.64±5.87)s]分均显著增加(t=18.042,14.325,10.117;均P<0.05)。造模后第8、9、10天Calcitriol组逃避潜伏期[(44.54±3.75)s,(30.74±4.74)s,(24.43±4.75)s]均较TBI组降低(t=6.539,8.909,7.369;均P<0.05)。Western blot结果显示,TBI组Mcoln-1在皮层和海马组织内的表达与Sham组相比均差异无统计学意义(均P>0.05)。与TBI组大鼠比较,Calcitriol组大鼠皮层和海马组织Mcoln-1蛋白表达均升高,差异有统计学意义(t=18.862,17.336;均P<0.05)。免疫荧光结果显示,Calcitriol组大鼠的皮层和海马组织内Mcoln-1与NeuN(神经元标记物)的表达存在共定位。结论:VD能够改善大鼠TBI后学习记忆功能障碍,其机制可能与激活Mcoln-1钙通道有关。

Objective:To explore the effects of vitamin D(VD)on lysosome activity and calcium channel protein Mcoln-1 in hippocampus and cortex of rats after traumatic brain injury(TBI).Methods:Forty-five SD rats were randomly divided into control group(Sham group),TBI model group(TBI group)and calcitriol treatment group(Calcitriol group).TBI models were established by electronic cortical impactor in TBI group and Calcitriol group.The rats in Calcitriol group were given of calitriol(1μg/kg)by intraperitoneal injection at 30 min,24 h and 48 h after TBI.Western blot was used to detect the expression of Mcoln-1,LAMP-1 and cathepsin-B in hippocampus and cortex region of rats.Immunofluorescence was used to detect the co-localization of Mcoln-1 and neurons three days later.Morris water maze test was performed on the 8th,9th and 10th day after surgery in all groups.Results:Morris water maze results showed that compared with sham group((19.54±3.54)s,(18.64±4.63)s,(17.64±5.88)s),the latency of seeking platform escape((58.75±6.65)s,(50.64±5.56)s,(42.64±5.87)s)were significantly increased in TBI group(t=18.042,14.325,10.117;all P<0.05).On the 8th,9th and 10th day after modeling,the escape latency of Calcitriol group((44.54±3.75)s,(30.74±4.74)s,(24.43±4.75)s)were significantly lower than those of TBI group(t=6.539,8.909,7.369,all P<0.05).Western blot results showed that the expression of Mcoln-1 in cortex and hippocampus of TBI group were not significantly different from those of Sham group(both P>0.05).Compared with TBI group,the expression of Mcoln-1 protein in cortex and hippocampus of rats in Calcitriol group were significantly increased(t=18.862,17.336,both P<0.05).Immunofluorescence showed that the expression of Mcoln-1 and NeuN(neuron marker)co-located in the cortex and hippocampus of rats in the Calcitriol group.Conclusion:VD can improve learning and memory dysfunction after TBI in rats,and its mechanism may be related to the activation of Mcoln-1 calcium channel.