大鼠脊髓损伤后诱导型一氧化氮合酶基因表达变化的实验研究

目的探讨大鼠脊髓损伤后诱导型一氧化氮合酶(iNOS)mRNA表达的变化规律.方法SD大鼠48只,随机分为8组,采用Allen's脊髓损伤打击模型,以逆转录-聚合酶链反应(RT-PCR)法测定伤段脊髓组织iNOS mRNA表达情况.结果iNOS mRNA在脊髓损伤前即有表达,损伤后早期无明显变化,伤后72h开始升高,1周时达到高峰.结论脊髓继发性损害的持续时间可能大于传统观念,针对继发性损害所作的治疗应持续至脊髓损伤后较长的一段时间.     

大鼠脊髓损伤后诱导型—氧化氮合酶基因表达变化的实验研究

目的：探讨大鼠脊髓损伤后诱导型-氧化氮合酶(iNOS)mRNA表达的变化规律。方法：SD大鼠48只，随机分为8组，采用Allen’s脊髓损伤打击模型，以逆转录-聚合酶链反应(RT-PCR)法测定伤段脊髓组织iNOS mRNA表达情况。结果：iNOS mRNA在脊髓损伤前即有表达，损伤后早期无明显变化，伤后72h开始升高，l周时达到高峰。结论：脊髓继发性损害的持续时间可能大于传统观念，针对继发性损害所作的治疗应持续至脊髓损伤后较长的一段时间。     

大鼠脊髓损伤后诱导型-氧化氮合酶基因表达变化的实验研究

目的 :探讨大鼠脊髓损伤后诱导型一氧化氮合酶 (iNOS)mRNA表达的变化规律。方法 :SD大鼠 48只 ,随机分为 8组 ,采用Allen’s脊髓损伤打击模型 ,以逆转录 -聚合酶链反应 (RT -PCR)法测定伤段脊髓组织iNOSmRNA表达情况。结果 :iNOSmRNA在脊髓损伤前即有表达 ,损伤后早期无明显变化 ,伤后 72h开始升高 ,1周时达到高峰。结论 :脊髓继发性损害的持续时间可能大于传统观念 ,针对继发性损害所作的治疗应持续至脊髓损伤后较长的一段时间     

大鼠脊髓损伤后单核细胞趋化蛋白-1、巨噬细胞炎症蛋白-2α基因表达的变化

目的：探讨大鼠脊髓损伤后单核细胞趋化蛋白-1(MCP-1)、巨噬细胞炎症蛋白-1α(MIP-1α)mRNA表达的变化规律。方法：SD大鼠42只，随机分为7组，采用改良Allen’s脊髓损伤打击模型，以逆转录-聚合酶链反应(RT—PCR)法测定伤段脊髓MCP-1、MIP-1α mRNA表达情况。结果：正常脊髓组织内存在MCP-1、MIP-1α mRNA的表达，脊髓损伤后MCP-1、MIP-1αmRNA表达逐渐增强，MCF-1在伤后24h达到高峰，MIP-1α伤后6h达到高峰。结论：MCP—1、MIP-1α存在于正常的脊髓组织内，脊髓损伤后MCP-1、MIP-1α表达迅速增强，提示参与了继发性脊髓损伤过程，并可能是损伤因素。     

大鼠脊髓损伤后单核细胞趋化蛋白-1、巨噬细胞炎症蛋白-2α基因表达的变化

目的:探讨大鼠脊髓损伤后单核细胞趋化蛋白-1(MCP-1)、巨噬细胞炎症蛋白-1α(MIP-1α)mRNA表达的变化规律.方法:SD大鼠42只,随机分为7组,采用改良Allens脊髓损伤打击模型,以逆转录-聚合酶链反应(RT-PCR)法测定伤段脊髓MCP-1、MIP-1α mRNA表达情况.结果:正常脊髓组织内存在MCP-1、MIP-1α mRNA的表达,脊髓损伤后MCP-1、MIP-1α mRNA表达逐渐增强,MCP-1在伤后24h达到高峰,MIP-1α伤后6h达到高峰.结论:MCP-1、MIP-1α存在于正常的脊髓组织内,脊髓损伤后MCP-1、MIP-1α表达迅速增强,提示参与了继发性脊髓损伤过程,并可能是损伤因素.     

大鼠坐骨神经损伤后脊髓及坐骨神经中睫状神经营养因子及其受体mRNA的表达

目的　探讨不同年龄大鼠坐骨神经损伤再生过程中 ,脊髓及坐骨神经睫状神经营养因子 (CNTF)及其受体α(CNTFRα)mRNA变化。方法　幼年、成年及老年大鼠各 96只 ,随机分为正常组 (1 2只 )和实验组 (84只 ) ,实验组大鼠切除右侧长 6mm的坐骨神经 ,分为伤后 1、3d和 1、2、4、8、1 2周组。利用逆转录 聚合酶链式反应 (RT PCR)及原位杂交法检测脊髓及坐骨神经中CNTF、CNTFRαmRNA的表达变化。结果　原位杂交及RT PCR结果显示 ,各年龄组大鼠伤后损伤侧脊髓和伤侧神经CNTFR及CNTFRαmRNA表达均较正常组及未伤侧显著升高 (P <0 .0 5) ,4周达最高峰 ;伤侧近、远端神经中CNTFmRNA 1d～ 1周表达较正常组及未伤侧减低 (P <0 .0 5) ,2周后开始升高 ,4周达高峰 (近端 :1 .59± 0 .1 3、1 .1 9± 0 .1 7和 0 .69± 0 .0 4 ,远端 :1 .1 3±0 .1 6、0 .84± 0 .0 6和 0 .57± 0 .0 3 ,P <0 .0 5)。其中幼年鼠变化最大 ,成年鼠次之 ,老年鼠最弱 ,近端较远端神经有变化大的趋势。结论　不同年龄大鼠坐骨神经损伤后脊髓、神经中CNTF及CNTFRαmRNA表达显著升高。     

一氧化氮对大鼠肝缺血-再灌注损伤中肝细胞凋亡的影响

目的探讨一氧化氮对肝缺血-再灌注损伤及肝细胞凋亡的影响。方法在一氧化氮合酶（NOS）增强剂和抑制剂条件下,观察大鼠肝缺血-再灌注后1、3、6、24h肝的血清门冬氨酸氨基转移酶（AST）和丙氨酸氨基转移酶（ALT）变化,同时进行肝细胞胞浆诱导型一氧化氮合酶（iNOS）表达及肝细胞凋亡的免疫组织化学分析。结果在再灌注6h点AST、ALT、肝细胞凋亡水平上,L-精氨酸（L-arg）组值分别为（341.88±111.24）IU/L、（311.75±139.41）IU/L和2.80±1.79,显著低于L-硝基精氨酸甲酯（L-NAME）组（2080.88±241.98）IU/L、（1153.00±110.92）IU/L和5.50±0.71（P值均＜0.05）。而L-arg组的肝细胞内iNOS表达灰度值152.07±3.46显著高于L-NAME组灰度值180.45±4.46（P＜0.05）。结论一氧化氮可减少肝缺血-再灌注损伤后肝酶的释放,抑制肝细胞的凋亡,改善肝缺血-再灌注损伤。     

大鼠脊髓损伤后白细胞介素-1β、肿瘤坏死因子-α基因表达的变化

目的 探讨大鼠脊髓损伤后白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)mRNA表达的变化规律。方法 SD大鼠42只，随机分为7组，采用改良Allen's脊髓损伤打击模型，以逆转录-聚合酶链反应(RT-PCR)法测定伤段脊髓组织IL-1β、TNF-α mRNA的表达情况。结果 正常脊髓组织内存在IL-1β、TN-αmRNA的表达，脊髓损伤后IL-1β、TNF-αmRNA表达迅速增强，在伤后1h达到高峰。结论 IL-1β、TNF-α存在于正常的脊髓组织内，脊髓损伤后IL-1β、TNF-α表达迅速增强，提示协同参与了继发性脊髓损伤过程，并可能是损伤性因素。     

大鼠脊髓损伤后白细胞介素-1β、肿瘤坏死因子-α基因表达的变化

目的 探讨大鼠脊髓损伤后白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)mRNA表达的变化规律。方法SD大鼠42只,随机分为7组,采用改良Allen's脊髓损伤打击模型,以逆转录-聚合酶链反应(RT-PCR)法测定伤段脊髓组织IL-1β、TNF-αmRNA的表达情况。结果 正常脊髓组织内存在IL-1β、TNF-αmRNA的表达,脊髓损伤后IL-1β、TNF-αmRNA表达迅速增强,在伤后1h达到高峰。结论 IL-1β、TNF-α存在于正常的脊髓组织内,脊髓损伤后IL-1β、TNF-α表达迅速增强,提示协同参与了继发性脊髓损伤过程,并可能是损伤性因素。     

脊髓损损后一氧化氮合酶活性变化的动态观察及其基因表达

目的：观察一氧化氮合酶在脊髓损伤中的变化及其机制。方法：用液压致伤装置造成大鼠脊髓中、重度损伤，伤后不同时间分别测量损伤区NOS活性。结果：显示脊髓损伤后早期NOS活性一过性升高，而后又迅速降低，其升高、降低幅度与损伤严重程度相一致，提示一氧化氮参与继发性脊损伤病理过程NOSmRNA原位杂交法，发现脊髓中度损伤后30minNOSmRNA的表达水平没有明显变化。结论：提示脊髓损伤早期NOS活性增高主要由于刺激因素如钙离子等使没有发挥活性的NOS激活，使酶蛋白总量增加引起的。     

Changes in nitric oxide synthase expression in young and adult rats after spinal cord injury

OBJECTIVE: To examine the clinical meaning of the changes in nitric oxide synthase (NOS) expression and activity after spinal cord injury (SCI) according to the age of the experiment animal. MATERIAL AND METHOD: Ten 5- and 16-week-old Sprague-Dawley rats were laminectomized at T10 and SCI induced at this level using a New York impactor. Outcome measures to assess SCI utilized the Basso-Beatti-Bresnahan scale to quantitate hind limb motor dysfunction as a functional outcome measure. NOS isoforms (nNOS, neuronal NOS; iNOS, inducible NOS; and eNOS, endothelial NOS) were also immunolocalized in sections of control and spinal cord injury in the two sample groups using specific monoclonal antibodies. Student's t-test evaluated the difference between the young and adult rats, and P<0.05 was considered as significant value. RESULT: As the expression of nNOS on the spinal gray matter of the adult rat decreased, eNOS activity increased. Different from the adult rat, expression of the nNOS in the young rat was maintained until 1 day after SCI, and compared with the adult rat; eNOS activity was increased in the vessels from the damaged gray matter area after 7 days of SCI. iNOS expression was maintained until the 7th day of SCI on the adult rat, but iNOS expression after 7 days of SCI on young rat decreased. The young rat showed relatively less motor disability on the hind limb when compared with the adult rat, and had a rapid recovery. CONCLUSION: Neural protective eNOS activity increased after SCI in the young rat, and neural destructive iNOS expression was more remarkable in the adult rat.     

Changes in nitric oxide and expression of nitric oxide synthase in spinal cord after acute traumatic injury in rats

The aim of this study was to observe the time course of NO production and NOS expression in the spinal cord following acute traumatic injury. Rat spinal cord was injured by extradural static weight-compression, which resulted in an incomplete transverse spinal cord lesion with paralysis of the lower extremities. Using this model, measurement of NO by microdialysis and Griess reaction and histological and immunohistochemical examinations using polyclonal antibodies to nNOS and iNOS were performed from immediately to 14 days after injury. In injured cord, the amount of NO markedly increased immediately after injury and gradually decreased between 1 and 12 h after injury. A second wave of increase in NO level was observed at 24 h and 3 days after injury. Histologically, hematomas and necrotic changes were observed after injury and demyelination of nerve fibers increased with time in the compressed segment. Immunohistochemically, the number of cells with expression of nNOS was increased immediately to 12 h after injury. Expression of iNOS was observed from 12 h to 3 days after injury. These findings suggested that the initial maximal increase of NO production might be caused mainly by nNOS and that the second wave of increase in NO might be due mainly to iNOS.     

Gene expression of inducible nitric oxide synthase in injured spinal cord tissue

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神经源性膀胱组织中一氧化氮合酶表达的特点和意义

目的 探讨神经元型一氧化氮合酶(nNOS)、诱导型NOS(iNOS)和内皮型NOS(eNOS)在神经源性膀胱组织中表达状况,探讨一氧化氮在神经源性膀胱组织中产生及作用特点.方法 神经源性膀胱患儿30例.男18例,女12例.年龄(6.3±3.1)岁.30例均行手术治疗,术中留取膀胱组织标本,采用免疫组织化学方法检测膀胱组织中nNOS、iNOS和eNOS表达状况,10例正常膀胱组织标本作对照. 结果 正常膀胱体部组织nNOS阳性表达,走行在平滑肌纤维之间,分布于平滑肌细胞表面,膀胱基质也有表达,组织化学评分(HS)为2.8～4.0和1.2～2.7;平滑肌细胞iNOS阴性表达,平滑肌细胞基质有少量稀疏表达,HS为0～0.4和0～0.1;eNOS表达分布于血管内皮细胞中,分布稀疏,平滑肌细胞无表达.膀胱颈部组织表达高于膀胱体部组织,以nNOS表达为主.神经源性膀胱组织中以iNOS表达为主,nNOS表达明显减少;eNOS主要分布于膀胱基质的内皮细胞,膀胱平滑肌、成纤维细胞阴性表达;病变膀胱组织血管稀疏,微血管密度100倍视野下可见到(6.8±3.2)个血管灶,低于正常膀胱组织的(16.7±6.3)个(P＜0.01).结论 正常膀胱组织nNOS主要分布在膀胱颈中,NO合成主要受nNOS调节.神经源性膀胱患者膀胱组织中氮能神经元分布稀疏,nNOS表达减少,iNOS表达上调,NO的合成与调节可能主要来源于iNOS,受iNOS水平调节,eNOS表达下降,提示膀胱组织血供不良.     

大鼠睾丸内一氧化氮合酶的表达

目的 :阐明一氧化氮合酶 (NOS)在睾丸内的表达 ,探讨一氧化氮 (NO)在睾丸生殖功能中的作用。　方法 :取雄性SD大鼠睾丸于 4 %多聚甲醛中固定 ,常规石蜡切片。用免疫组化ABC法检测成年大鼠睾丸NOS的表达和定位。　结果 :内皮型NOS(eNOS)、神经型NOS(nNOS)、诱导型NOS(iNOS)在睾丸间质细胞内均有表达 ,精曲小管周围肌样细胞、血管内皮细胞和平滑肌细胞内仅为eNOS免疫反应阳性 ,而血管外膜纤维内仅有nNOS表达。免疫反应物质主要定位于细胞质 ,胞核为阴性。在生精细胞内 3种NOS免疫反应均为阴性。　结论 :3种NOS在睾丸内均有表达 ,不同的NOS分布部位有一定区别     

Modulation of nitric oxide synthase isoenzymes inreperfused skeletal muscle

Objective:To investigate the modulation of nitric oxide synthase(NOS)isoenzymes in skeletal muscle during 3h ischemia/reperfusion (I/R,3h ischemia followed by 3h reperfusion).Methods:The extensor digitorum longuses(EDLs) from 20 adult rats were divided into 4 groups:the normal,the sham operation,the ischemia(3h),and the ischemia/reperfusion group.One normal EDL from each rat was used as the non-operated control,and the opposite ones are distributed into the 3 remaining groups.All the samples were studied with Western blotting technique and immunohistochemistry staining.Results:Three sizes or protein bands verified with the proteins of relative molecule to be of 155000,140000 and 135000,were detected in the EDL homogenate by Western blotting,which were comparable with the positive controls for nNOS,eNOS and iNOS,respectively.Immunostaining demonstrated that nNOS was present in the muscle fiber,with a similar location of the muscle stria,eNOS was found apparently in microvascular endothelia,but not found in muscle fibers,and iNOS was found in the leukocytes around the muscle fiber and some endothelia cells.Immunostaining paralleled the Western blotting results.Conclusions:It suggests that the constitutive nNOS and eNOS protein can be regulated by I/R,and I/R results in a down regulation of nNOW and up-regulation of eNOS and iNOS in reperfused skeletal muscle.The fact that nNOS is present around stria suggests that nNOS may have a close relationship with muscle function.The localization of eNOS in endothelial cell indicates its role in regulating blood supply of the muscle.Based on these findings,it is possible that No produced by distinct NOS may play a different role in I/R injury.