外周神经源性疼痛的实验模型研究进展

神经源性疼痛是严重困扰人类的临床问题之一。为了探索其发病机理及治疗方法,研究人员建立了多种神经损伤性疼痛模型,其中大鼠部分脊神经结扎,慢性压迫性神经损伤和L5／L6脊神经结扎的外周神经损伤的模型应用最为广泛。细胞模型也显示出了很好的应用前景,原代和永久性感觉神经元,作为研究疼痛发生和发展的分子机理,特别是神经递质的释放和信号转导的研究非常有用。     

大鼠心肌缺血再灌注模型构建的改良

目的改良大鼠心肌缺血再灌注模型的构建方法,增强实用性。方法32只SD雄性大鼠随机分为四组（A组：伪手术组;B组：缺血组;C组：缺血30 min再灌注组;D组：缺血60 min再灌注组）。缺血过程在人工机械通气条件下完成,术前术中监测心电图变化,模型成功72 h后抽血进行心肌酶谱分析,并观察左室压变化。结果造模成功率为91%。B、C、D组各心肌酶含量明显高于A组（P〈0.05）,C、D组低于B组（P〈0.05）。B、C、D组左室压值低于A组（P〈0.05）,C、D组高于B组（P〈0.05）。结论改进后的模型制备方法简便易行,成功率高,评价指标符合临床应用。     

大鼠心肌缺血再灌注损伤模型的实验研究

目的 制备一种新型的心肌急性缺血再灌注损伤模型,以探讨一种更符合临床实际需求的实验方法.方法 将20只雌性SD(Sprague-Dawley)大鼠随机分成2组(对照组、实验组),采用结扎主动脉根部引起心肌缺血5min再灌注30 min建立心肌急性缺血再灌注模型;通过应用透射电镜观察心肌细胞超微结构的改变,同时检测心肌组织匀浆丙二醛(Maleic Dialdehyde,MDA)含量、超氧化物歧化酶(Superoxide Dismutase,SOD)活力.结果 透射电镜下超微结构显示实验组较对照组明显加重了心肌组织结构和线粒体的损害;实验组心肌组织MDA明显高于对照组(P<0.01),而SOD明显低于对照组(P<0.01).结论 本实验成功建立了方法简便、易于操作、取材范围广泛的心肌缺血再灌注损伤模型,为心肌缺血再灌注损伤研究提供了一种更为可行的模型.     

伤害性刺激对大鼠感觉皮质NOS阳性神经元影响的实验研究

研究选用Wistar大鼠18只,分为正常对照组6只,损伤致痛组12只,分别在损伤后30分钟和3小时迅速断头处死。取大脑顶叶,用NOS组织化学法和图像定量分析观察了损伤致痛条件下大鼠感觉皮质NOS阳性神经元的变化。结果显示,损伤致痛后NOS阳性神经元NOS反应产物较对照组增多,图像定量分析结果经统计学处理,实验组与对照组比较有极显性差异（P＜0.001）,实验组30分钟组与3小时组比较有显性差异（P＜0.05）。结果表明NO参与感觉皮质对痛觉信息的中枢调制。     

大鼠心肌梗死模型构建和评价方法的改良

目的 优化和改良大鼠心肌梗死模型的构建和评价方法,提高模型的可靠性和稳定性.方法 取雄性SD大鼠结扎左冠状动脉前降支建立心肌梗死模型,在模型的构建过程中从麻醉、插气管、保温、手术操作、术后护理等环节进行优化和改进,并观察不同的麻醉方法和术后时间对心肌梗死程度的影响,用不同的染色方式进行心肌梗死模型的评价.结果 对比大鼠心肌梗死模型构建过程中各组大鼠麻醉时间、术后恢复以及心肌梗死面积的结果,戊巴比妥钠是更合适的麻醉药;结扎手术后时间对模型心肌梗死范围无明显影响（P＆gt;0.05）,但心肌缺血危险区面积随术后时间的延长明显减少（P〈0.01）;TTC与依文思蓝双重染色相对TTC染色能明显观察到心肌缺血危险区和梗死区范围.结论 优化和改进后的大鼠心肌梗死模型,提高了动物福利,制备和评价方法更加客观准确.     

贻贝粘蛋白对痔病大鼠模型治疗作用的实验研究

目的 本研究基于直肠局部用药要求建立巴豆油诱导的痔病大鼠模型,探究贻贝粘蛋白(MAP)的抗痔疗效和黏附性,并探讨其作用机制。方法 以巴豆油为致炎剂建立痔病大鼠模型,将40只SPF级SD大鼠按体重随机分为4组,造模后1 h各组大鼠予以相应治疗。D4取材,处死前30 min将1%伊文思蓝(EBD)注入大鼠尾静脉,计算肛肠系数,评价血管通透性,Western Blot法检测黏蛋白Muc2、Muc4的蛋白相对表达量,ELISA法检测大鼠血清TNF-α、IL-6、MDA表达量。通过在体实验和离体实验比较异硫氰酸荧光素(FITC)标记的MAP在大鼠直肠的荧光成像,观察比较MAP在直肠的黏附性。结果 MAP可减少大鼠肛门直肠肿胀、分泌物和渗出情况,肛肠系数结果提示MAP可降低肛门直肠肿胀程度(P<0.01),EBD含量提示MAP可降低局部血管通透性(P<0.01),增加黏蛋白Muc2、Muc4的相对表达量(P<0.05),降低血清TNF-α、IL-6、MDA表达量(P<0.01),荧光成像结果显示MAP在直肠环境内表现出良好的黏附性和稳定性。结论 海洋生物材料贻贝粘蛋白具有良...     

大鼠铜梳烧伤模型建立方法的改良

目的:改良大鼠铜梳烧伤模型的建立方法,对比不同方法建立的烧伤模型的烧伤间隙区初始面积及坏死程度的差异,探索更加理想的大鼠铜梳烧伤模型的建立方法。方法:48只SD大鼠被随机分为A组(原方法)、B组(改良法1)及C组(改良法2)。将铜梳烧伤器加热后,即刻置于各组大鼠背部中线两侧的皮肤上,A组铜梳接触皮肤时,除自身重力以外,不施加任何压力;B组铜梳接触皮肤时施加压力,使皮肤凹陷约0.5 cm;C组使用铜梳烧伤器嵌合聚乙烯泡沫塑料模型后再接触皮肤,并施加压力,使皮肤凹陷约0.5 cm。测量各组伤后即刻的烧伤间隙区面积;伤后6、12、24、48小时的间隙区组织坏死面积;各个时间点取材后,通过HE染色观察间隙区组织坏死程度并测量坏死深度。结果:伤后即刻C组的烧伤间隙区面积介于A组与B组之间,且与理论值最接近;B组间隙区组织坏死进程最快,伤后24小时已基本坏死。A组坏死进程最慢,间隙区组织基本坏死时间超过48小时。C组间隙区组织基本坏死时间是伤后48小时,与理论上的时间最接近。结论:通过铜梳烧伤器嵌合聚乙烯泡沫塑料模型后再接触皮肤,并施加压力,使皮肤凹陷约0.5 cm,是建立大鼠铜梳烧伤模型更加理想的方法。     

不同剂量野百合碱诱导大鼠肺动脉高压模型的实验研究

目的:肺动脉高压患者的预后非常差并且目前对肺动脉高压的治疗缺乏有效的方法.因此,对肺动脉高压的研究十分迫切,而建立稳定、操作简便、稳定的肺动脉高压模型是研究肺动脉高压的基础.本实验的目的就是采用不同剂量野百合碱(monocrotaline,MCT)诱导建立SD大鼠肺动脉高压模型,探讨肺动脉高压模型的制作方法及其稳定性.方法:清洁级SD雄性大鼠75只,随机分为C组、M1、M2、M3和M4组,每组各15只.其中M1～M4组大鼠分别一次性腹腔注射MCT 30、40、50和60 mg/kg,诱导制作肺动脉高压动物模型;C组为对照组,给予相同剂量溶剂腹腔注射.腹腔注射4周后比较各组大鼠的生存率、检测平均肺动脉压力(mPAP),肺动脉收缩压(PASP),右心肥大指数(RVHI),并取肺组织行苏木素-伊红染色,观察肺的病理改变,采用肺小动脉形态计量学指标综合判断肺动脉高压模型的稳定性.结果:C组无死亡,M1～M4组大鼠的生存率分别为87％,87％,67％,40％.M1～M4各组SD大鼠平均肺动脉压、肺动脉收缩压、右心肥大指数依次增大(P＜0.05),肺小动脉形态计量学指标检测显示肺血管中膜厚度百分比依次增大(P＜0.05),而M3与M4组各测量结果无明显差异.结论:腹腔注射50 mg/kg、60 mg/kg剂量野百合碱均可引起大鼠肺动脉压力升高和肺血管重构,均可诱导稳定的肺动脉高压模型,而50 mg/kg剂量有更高的生存率.所以50mg/kg剂量野百合碱腹腔注射是成功诱导大鼠肺动脉高压模型的合适剂量.     

咖啡因对大鼠杏仁核点燃模型的作用研究

目的:观察兴奋药咖啡因对点燃癫痫模型的作用.方法:建立大鼠杏仁核点燃模型,观察不同剂量的咖啡因及与抗癫痫药物联用对杏仁核点燃大鼠的后放电时程(ADD)和Racine's分级的影响.结果:咖啡因提高点燃大鼠的ADD,但联用抗苯妥英钠和苯巴比妥可降低ADD(P＜0.01)及Racine's分级(P＜0.01).结论:小剂量的咖啡因与抗癫痫药物能产生协同作用.     

The effect of a nitroxide antioxidant on ischemia-reperfusion injury in the rat in vivo hind limb model

Microsurgical procedures such as free tissue transfer or replantations of amputated digits involve an obligatory ischemic period leading to regional tissue oedema, rhabdomyolysis, systemic acidosis, hypercalcemia and multiple organ dysfunction syndrome reflecting ischemia-reperfusion (I/R) injury. Since nitroxide stable radicals act as antioxidants their potential protective effects were tested. Anaesthetized Sabra rats were subjected to regional ischemia of the hind limb for 2 h using a tourniquet. Upon reperfusion rats were injected with 4-OH-2,2,6,6-tetramethylpiperidine-1-oxyl (TPL). Systemic I/R-induced damage was assessed by sampling blood for differential count, lactate dehydrogenase (LDH) and creatine phosphokinase (CPK) serum levels. Regional injury was evaluated by analysing excised muscle samples for oedema (tissue water content) and inflammatory infiltrate (number of cell nuclei in histomorphometric analysis). I/R-induced changes of biomarkers reflecting systemic damage peaked about 8 h following the start of reperfusion and fully disappeared as the biomarkers relaxed to their pre-ischemic values after 24 h. TPL facilitated the recovery of some of these parameters and partially affected release of cellular CPK and LDH. The parameters of I/R-induced regional tissue injury did not demonstrate any recovery and were not inhibited by TPL.     

The effect of a nitroxide antioxidant on ischemia-reperfusion injury in the rat in vivo hind limb model

Microsurgical procedures such as free tissue transfer or replantations of amputated digits involve an obligatory ischemic period leading to regional tissue oedema, rhabdomyolysis, systemic acidosis, hypercalcemia and multiple organ dysfunction syndrome reflecting ischemia-reperfusion (I/R) injury. Since nitroxide stable radicals act as antioxidants their potential protective effects were tested. Anaesthetized Sabra rats were subjected to regional ischemia of the hind limb for 2 h using a tourniquet. Upon reperfusion rats were injected with 4-OH-2,2,6,6-tetramethylpiperidine-1-oxyl (TPL). Systemic I/R-induced damage was assessed by sampling blood for differential count, lactate dehydrogenase (LDH) and creatine phosphokinase (CPK) serum levels. Regional injury was evaluated by analysing excised muscle samples for oedema (tissue water content) and inflammatory infiltrate (number of cell nuclei in histomorphometric analysis). I/R-induced changes of biomarkers reflecting systemic damage peaked about 8 h following the start of reperfusion and fully disappeared as the biomarkers relaxed to their pre-ischemic values after 24 h. TPL facilitated the recovery of some of these parameters and partially affected release of cellular CPK and LDH. The parameters of I/R-induced regional tissue injury did not demonstrate any recovery and were not inhibited by TPL.     

The protective efficacy of magnolol in hind limb ischemia-reperfusion injury

We investigated the protective effects of magnolol, an active antioxidant and free radical scavenger extracted from Magnolia officinalis, in a hind limb ischemic-reperfusion animal model. Adult male Spraque-Dawley rats were subjected to hind limb ischemic insult for 2 hours and were intravenously treated with magnolol at 0.01 mg/kg (n=8), 0.3 mg/kg (n=8) mg/kg or 1 mg/kg (n=8) mg/kg, or vehicle (n=8). At 24 h post-insult, the levels of nitrite/nitrate (NOX), malondialdehyde (MDA) and myeloperoxidase (MPO), as well as the degree of muscle damage, were assessed. Relative to controls, animals treated with magnolol (0.3 and 1 mg/kg) had attenuated muscular inflammation, edema and damage. Magnolol (0.3–1 mg/kg) also effectively reduced postischemic rises in the MDA, NOx and MPO levels (p<0.05, respectively). Magnolol administrated at 0.01 mg/kg, however, failed to protect against the ischemic-perfusion limb injury. In addition, magnolol (0.01–1 mg/kg) did not affect local muscular blood reperfusion or other physiological parameters, including hematocrit, glucose, arterial blood gases and mean arterial blood pressure. Thus, intravenous administration with magnolol at 0.3–1 mg/kg protects against ischemic limb damage in rats. This cytoprotection may be attributed to its antioxidant, anti-nitrosative and anti-inflammatory actions.     

Pathway to carrageenan-induced inflammation in the hind limb of the rat

A sequential 43-step pathway scheme for the inflammatory response of the rat to interdermal injection of carrageenan (C) was devised. It consisted of a nonphagocytic inflammatory response (NPIR) followed by a phagocytic inflammatory response (PIR) in the dermis and an epidermal NPIR. The dermal NPIR comprised edema, hyperemia, and hyperalgesia followed by hypoalgesia. Antiserotonin agents inhibited the hypoalgesia and part of the edema. These findings and histological observations suggested that dermal mast cells were injured by C. The hyperalgesia and part of the edema were sensitive to arachidonate cyclooxygenase inhibitors (AACOIs). It is speculated that injured mast cells metabolize arachidonic acid and reactive intermediates, not prostaglandins, mediate the NPIR hyperalgesia and part of the edema. The dermal PIR consisted of mobilization of neutrophils, edema, hyperalgesia, mobilization of monocytes, and proliferation of fibroblasts and vascular tissue. Selective drug actions revealed that the edema, hyperalgesia, and monocyte mobilization of the PIR depended on the mobilization of neutrophils. After the mobilization of neutrophils, AACOIs reduced edema formation and hyperalgesia. Arachidonic acid metabolism by neutrophils is speculated to produce the mediators of phagocytic inflammatory (PI) edema and hyperalgesia. Monocyte function was associated with cessation of PI edema formation and phagocytosis of neutrophils and cellular debris. Interleukin 1 is speculated to mediate the adherence of neutrophils to injured dermal endothelium. The epidermal NPIR consisted of edema, hyperplasia, and hyperkeratosis. These parameters were not studied mechanistically. There was no evidence for histamine, bradykinin, platelets, clotting factors, or complement mediating any events in the pathway.     

Mast cell amines and inosineinduced vasoconstriction in the rat hind limb

Under certain circumstances injected inosine causes a net vasoconstrictive effect on the arterioles, which has been attributed to 5-hydroxytryptamine (5HT) released in response to adenosine type 3 (A(3)) receptor stimulation of mast cells residing in the adventitia. We have sought further evidence for this hypothesis using blood vessels of the rat hind limb perfused in vitro at constant rate with a gelatin-containing physiological salt solution. Injection of inosine (2.7 mg) caused a rise in perfusion pressure, which was only slightly increased by inclusion of N-nitro-L-arginine methyl ester (100 muM) in the perfusate. Inclusion in the perfusate of cyproheptadine (1 muM), compound 48 80 (1 mug ml), 8-phenyltheophylline (1 muM) or 8-cyclopentyl-1,3 dipropylxanthine (0.1 muM) greatly reduced the pressor response to inosine. The pressor effect of injected 5HT (400 mug) was abolished by pre-treatment with cyproheptadine, but not by pre-treatment with compound 48 80. These results suggest that the net pressor response to injected inosine was mainly the result of an A(1) receptor-mediated release of 5HT, most probably from mast cells. No evidence was found for an involvement of A(3) receptor stimulation.     

Protective effects of mitochondrion-targeted peptide SS-31 against hind limb ischemia-reperfusion injury

Hind limb ischemia-reperfusion injury is an important pathology in vascular surgery. Reactive oxygen species are thought to be involved in the pathogenesis of hind limb ischemia-reperfusion injury. SS-31, which belongs to a family of mitochondrion-targeted peptide antioxidants, was shown to reduce mitochondrial reactive oxygen species production. In this study, we investigated whether the treatment of SS-31 could protect hind limb from ischemia-reperfusion injury in a mouse model. The results showed that SS-31 treatment either before or after ischemia exhibited similar protective effects. Histopathologically, SS-31 treatment prevented the IR-induced histological deterioration compared with the corresponding vehicle control. SS-31 treatment diminished oxidative stress revealed by the reduced malondialdehyde level and increased activities and protein levels of Sod and catalase. Cellular ATP contents and mitochondrial membrane potential increased and the level of cytosolic cytC was decreased after SS-31 treatment in this IR model, demonstrating that mitochondria were protected. The IR-induced increase of levels of inflammatory factors, such as Tnf-α and Il-1β, was prevented by SS-31 treatment. In agreement with the reduced cytosolic cytC, cleaved-caspase 3 was kept at a very low level after SS-31 treatment. Overall, the effect of SS-31 treatment before ischemia is mildly more effective than that after ischemia. In conclusion, our results demonstrate that SS-31 confers a protective effect in the mouse model of hind limb ischemia-reperfusion injury preventatively and therapeutically.