缺血性脑水肿的AQP-4表达与磁共振成像的相关性研究

目的　研究AQP 4表达与磁共振成像相关性 ,探讨脑水肿磁共振成像的分子生物学机制。方法　将Wistar大鼠 3 4只 ,随机分为 8组 ,即正常组 4只、假手术组 6只和大脑中动脉栓塞 (MCAO)后 15min、3 0min、1h、3h、6h及 2 4h组各 4只 ,分别进行脑部DWI和T2WI扫描 ,观察梗塞区异常信号出现时间 ,测量异常信号强度 (rd)和面积 (rs)、扩散异常区的表观扩散系数 (ADC) ,用免疫组化、原位杂交和图像分析技术检测以上各组的AQP 4蛋白 (rS)及AQP 4mRNA表达 (α) ,并进行统计学分析。结果　rS与α呈明显正相关 (r =0 .949)。栓塞后 15min即在DWI上出现高信号 ,而T2WI未见异常 ;在1h内ADC迅速下降 ,而AQP 4表达与DWI的rd、rs都快速增加。AQP 4mRNA(α)与rs DWI呈明显相关性 (r =0 .914 )。随着时间的延长 ,ADC值逐渐下降 ,于 3h达到最低 [( 2 .1± 0 .6)× 10 -4mm2 /s] ,随后缓慢递增直至 2 4h ,但AQP 4表达、rd、rs仍不断增加。T2WI最早于 3 0min ,平均在 1.4h显示异常高信号 ,在T2WI上的rs值都比相应的DWI上的值低 (P <0 .0 5 )。AQP 4mRNA(α)与rs T2WI无明显相关 (r =0 .116)。结论　AQP 4表达增强是缺血性脑水肿时ADC值下降、DWI上高信号的重要原因 ,与T2WI的信号变化无关。DWI高信号面积可以间接反映AQP 4的表     

脑红蛋白在内毒素脑损伤大鼠模型中的表达变化

Objective To investigate the expression of neuroglobin(Ngb) in frontal cortex, cere-brospinal fluid (CSF) and serum in rats with brain injury induced by LPS and to elucidate its significance. Methods Seventy adult Sprague-Dawley rats were randomly divided into LPS (n = 60, with injection of 0.1 mg/kg LPS into cistern) and control (C, n =10, with injection of equal volume of isotonic saline into cistern) groups, with 10 rats in each group. The plasma, CSF as well as frontal cortex from sacrificed rats were collected in LPS group at 3, 6, 12, 24, 48, 72 post injection hour (PIH), with 10 rats at each time point. The protein expression of Ngb was measured by enzyme-linked immunosorbent assay(ELISA) and Western blot. Expression of Ngb in frontal cortex was determined by SP. Water content of frontal cortex was assessed by drying method. The correlation among Ngb contents in serum, CSF, frontal cortex and water con-tent of frontal cortex was analyzed with multiple comparison. Results Ngb expression ( by ELISA) : Ngb expression of frontal cortex, CSF, and serum in LPS group was higher than that in C group at each time point, and it peaked at 48 PBH (35.4±3.9, 22.7±3.1, 14.4±2.8 ng/mL, respectively, P <0.01). Ngb ex-pression ( by Western blot) : Ngb protein with relative molecular mass of 17×103 was observed in each group. Ngb expression detected by Western blot was similar to that detected by ELISA. Brain water content was significantly greater in LPS group than that of C group at 6-72 PIH ( P<0.01) , and it peaked at 48 PBH ( 83.3±1.9) %. Ngb contents in serum, CSF, frontal cortex, and water content of frontal cortex were positively correlated with muhipie comparison (γ=0.631-0.719, P<0.01). Conclusions Up-regu-lation of Ngb expression in brain injury induced by LPS is correlated with duration after challenge of LPS.     

脑红蛋白在内毒素脑损伤大鼠模型中的表达变化

Objective To investigate the expression of neuroglobin(Ngb) in frontal cortex, cere-brospinal fluid (CSF) and serum in rats with brain injury induced by LPS and to elucidate its significance. Methods Seventy adult Sprague-Dawley rats were randomly divided into LPS (n = 60, with injection of 0.1 mg/kg LPS into cistern) and control (C, n =10, with injection of equal volume of isotonic saline into cistern) groups, with 10 rats in each group. The plasma, CSF as well as frontal cortex from sacrificed rats were collected in LPS group at 3, 6, 12, 24, 48, 72 post injection hour (PIH), with 10 rats at each time point. The protein expression of Ngb was measured by enzyme-linked immunosorbent assay(ELISA) and Western blot. Expression of Ngb in frontal cortex was determined by SP. Water content of frontal cortex was assessed by drying method. The correlation among Ngb contents in serum, CSF, frontal cortex and water con-tent of frontal cortex was analyzed with multiple comparison. Results Ngb expression ( by ELISA) : Ngb expression of frontal cortex, CSF, and serum in LPS group was higher than that in C group at each time point, and it peaked at 48 PBH (35.4±3.9, 22.7±3.1, 14.4±2.8 ng/mL, respectively, P <0.01). Ngb ex-pression ( by Western blot) : Ngb protein with relative molecular mass of 17×103 was observed in each group. Ngb expression detected by Western blot was similar to that detected by ELISA. Brain water content was significantly greater in LPS group than that of C group at 6-72 PIH ( P<0.01) , and it peaked at 48 PBH ( 83.3±1.9) %. Ngb contents in serum, CSF, frontal cortex, and water content of frontal cortex were positively correlated with muhipie comparison (γ=0.631-0.719, P<0.01). Conclusions Up-regu-lation of Ngb expression in brain injury induced by LPS is correlated with duration after challenge of LPS.     

红景天苷对大鼠缺血性脑水肿的影响

目的研究红景天苷(salidroside)对水通道蛋白4(aquaporin 4,AQP4)在大鼠缺血性脑组织中表达的影响。方法采用线拴法制作大鼠局灶性脑缺血模型,实验分为模型组、假手术组、红景天苷组。通过光镜技术观察各组别脑水肿区的病理变化,干湿质量法检测脑含水量,采用原位杂交、免疫组化方法观察AQP4及其mRNA在脑组织中的表达,硫代巴比妥酸法和定磷法测定缺血脑组织中丙二醛(MDA)、Na ,K -ATPase活性。结果缺血后模型组的细胞及血管周围间隙明显扩大,部分细胞坏死,神经胶质细胞增生;大鼠穹隆下器官、脉络丛、大脑皮质、视上核、海马等部位AQP4及其mRNA的表达上调,48～72 h达峰值;脑组织含水量亦在72 h达高峰。红景天苷组脑组织形态结构无明显改变,脑组织含水量、MDA水平、AQP4及其mRNA的表达明显低于模型组;而Na ,K -ATPase活力明显高于模型组。结论红景天苷可抑制缺血性脑水肿组织中AQP4及其mRNA的过表达,防止脑水肿的发生。     

脊髓内去甲肾上腺素能纤维的来源

目的：研究脊髓内去甲肾上腺素(NA)能纤维的来源。方法：选用Wistar大鼠，应用神经递质NA特异性的标志酶——多巴胺β羟化酶(DBH)抗体注入其脊髓单侧，分别于注射后1h-7天灌注大鼠，取出脊髓和脑组织作免疫组化染色。结果：ADBH注入脊髓后48h内依次在腰、胸、颈髓同侧观察到大量DBH阳性纤维，对侧仅少量阳性纤维出现：注射后3—4天，双侧上橄榄核、LC、SC、KF核、臂旁核中出现大量阳性神经元，在延髓内仅见大量阳性纤维，未观察到阳性神经元胞体出现；注射后7天，上述阳性神经元和阳性纤维消失。结论：脊髓内的NA能纤维及终末几乎全部来源于脑桥NA能细胞群。     

单羧酸转运蛋白1、2、4在急性心衰大鼠模型肾上腺的表达变化

目的观察单羧酸转运蛋白(monocarboxylate transporter,MCT)1、2、4在急性心衰大鼠肾上腺的表达变化,以探讨急性心衰肾上腺的能量代谢变化。方法随机将成年健康SD大鼠(雌雄不限,250±20 g)50只,分为3组:正常组(n=10)、假手术组(n=20,2 subgroups,10 in each)和模型组(n=20,2subgroups,10 in each)。用1.5%戊巴比妥钠静脉注射制作大鼠急性心衰模型;注毕,运用BL-420F生物机能实验系统记录左室内压峰值(LVSP)和左室内压最大上升速率(+dp/dtmax)的变化曲线;用IHC和Western Blot检测模型鼠,肾上腺MCT1,2,4的表达变化。结果在急性心衰发生10 min时,肾上腺皮质和髓质MCT1、2、4的表达均有升高,此时假手术组MCT1、2、4也有升高。与正常对照组相比较,两组的MCT1、2、4的表达升高均具有统计学意义(P<0.05);在20 min时,大鼠肾上腺MCT1、2、4的表达较正常组却有所下降,与正常组相比较,仅MCT1的表达下调具有统计学意义(P<0.05),而MCT2、4则无明显差别(P>0.05)。假手术20 min组与10 min组相比较,MCT1、2、4的表达无明显差别(P<0.05)。结论在急性心衰和创伤性应激状态下,肾上腺MCT1、2、4的表达,在十分钟内呈代偿性增加,但随着心衰时间的延长,其表达反而降低,机体的能量代谢出现失代偿,提示MCT1、2、4参与创伤性应激及急性心衰的能量代谢过程。     

大鼠脊髓压迫性损伤后脱髓鞘病变及硫酸软骨素蛋白多糖的表达变化

目的 探讨脊髓压迫性损伤（CSCI）后脱髓鞘病变及硫酸软骨素蛋白多糖(CSPG,NG2)的表达变化。 方法选取健康成年SD大鼠75只,雌雄不限,采用随机数字表法将其分为正常组、假手术组、CSCI 1 d组、CSCI 3 d组和CSCI 7 d组,每组15只。采用自行设计的大鼠脊髓压迫器制作脊髓压迫模型,应用BBB运动功能评分、锇酸染色及透射电镜观察CSCI后1 d、3 d及7 d时大鼠的运动功能情况及白质有髓神经纤维的病理变化,计算脊髓后索有髓神经纤维的数量及髓鞘厚度与轴突直径的比值即G-ratio值,并采用免疫印迹法检测NG2蛋白的表达变化。 结果CSCI后,CSCI 1 d组、CSCI 3 d组和CSCI 7 d组的大鼠后肢BBB评分分别为（1.23±0.45）分、（0.65±0.35）分和（0.00±0.00）分,与正常组（21.00±0.00）分和假手术组（21.00±0.00）分相比,差异有统计学意义（P<0.05）,大鼠的运动功能随受压时间延长而逐渐下降。锇酸染色显示正常组和假手术组大鼠的白质正常,脊髓受压后,CSCI 1 d组、CSCI 3 d组和CSCI 7 d组的有髓神经纤维开始出现水肿、变性、崩解,正常组和假手术组脊髓后索的有髓神经纤维计数分别为 (2771±108）根和（2675±199)根,CSCI 1 d组、CSCI 3 d组和CSCI 7 d组的有髓神经纤维计数分别为（2403±161）根、（1708±70）根和（810±95）根,压迫后有髓神经纤维数目减少,压迫后7 d,有髓神经纤维数目减至最低（P<0.05）,与正常组和假手术组比较,差异有统计学意义（P<0.05）。电镜定量分析结果显示,正常组、假手术组、CSCI 1 d组、CSCI 3 d组和CSCI 7 d组的G-ratio值分别为（18.10±0.4）、（17.70±1.0）、（6.69±0.8）、（5.73±0.4）和（4.95±0.5）,CSCI 1 d组、CSCI 3 d组和CSCI 7 d组的G-ratio值均较正常组和假手术组低,且在压迫后7 d时降至最低,差异有统计学意义（P<0.05）。透射电镜结果显示,正常组和假手术组的轴突、髓鞘板层结构正常;脊髓受压后,轴索出现肿胀,轴浆内细胞器变性、坏死、减少;髓鞘折叠、皱缩,出现“洋葱皮”样变,髓鞘崩解;少突胶质细胞的染色质凝聚;巨噬细胞浸润。NG2蛋白免疫印迹结果显示,脊髓受压后,NG2蛋白表达水平在压迫后第1天升至最高（P<0.05）,且表达水平随压迫时间延长而逐渐下调,但均高于正常组和假手术组,差异有统计学意义（P<0.05）。 结论CSCI后,大鼠运动功能随受压时间延长而逐渐下降,有髓神经纤维发生脱髓鞘病变且数量减少,随着压迫时间延长,溃变呈现出进行性加重趋势;NG2细胞与CSCI后髓鞘的变化情况关系密切,可能增殖分化为少突胶质细胞或其它类型细胞,是脊髓髓鞘内源性修复的机制之一。