氨基胍对大鼠急性脊髓损伤后脊髓水肿的作用机制研究

目的氨基胍(aminoguanidine,AG)能显著减轻脑外伤及中风动物模型脑水肿,提高神经功能恢复程度。探讨AG对大鼠急性脊髓损伤(spinal cord injury,SCI)后脊髓水肿的作用及相关机制。方法取成年雄性SD大鼠150只(体重230～255 g),分为对照组(A组,25只)、假损伤组(B组,25只)、SCI后未治疗组(C组,25只)和SCI后AG治疗组(75只);AG治疗组按给药剂量分为AG 75 mg/kg组(D组,25只)、AG 150 mg/kg组(E组,25只)和AG300 mg/kg组(F组,25只)。A组未行任何处理,B组仅行椎板切除术但不治疗;C、D、E、F组制备静压型大鼠SCI模型后,C组腹腔注射5%DMSO,D、E、F组腹腔注射相应剂量AG。于造模后0、12、24、48 h用干湿重法检测受损脊髓组织含水量以筛选最佳剂量,进一步用伊文思兰(Evans blue,EB)法评测血-脊髓屏障功能,用RT-PCR检测水通道蛋白4(aquaporins 4,AQP4)mRNA表达,Western blot和免疫组织化学染色检测AQP4蛋白表达。结果脊髓组织含水量检测示,E组在造膜后12、24、48 h对SCI后脊髓组织水肿有明显抑制作用(P<0.05),选择该剂量组用于后续实验。造模后12、24、48 h,E组EB含量明显低于C组(P<0.05),降低血-脊髓屏障通透性。RT-PCR检测结果示造模后12、24、48 h,B、E组AQP4 mRNA表达明显低于C组;Western blot检测示造模后24、48 h,B、E组AQP4蛋白表达明显低于C组;免疫组织化学染色示造模后48 h,B、E组AQP4蛋白表达明显低于C组,差异均有统计学意义(P<0.05);但各指标各时间点B、E组间比较,差异均无统计学意义(P>0.05)。结论急性SCI后大鼠经150 mg/kg AG治疗后,能降低AQP4表达,改善脊髓水肿,减轻损伤。     

DPPH法测定葡萄籽原花青素清除自由基的能力

通过DPPH法测定葡萄籽原花青素抗氧化、清除自由基能力,实验表明:随着葡萄籽原花青素纯化程度的提高,其抗氧化、清除自由基的能力也相应地提高。葡萄籽原花青素纯化物的抗氧化、清除自由基能力大于VC和芦丁,葡萄籽粗提物的抗氧化、清除自由基能力略低于VC,但高于芦丁。GSP、GSPP1、GSPP2、GSPP3、GSPP3-SP、VC、芦丁清除DPPH自由基的半抑制量分别为118,93,88,86,63,110,170 g/kg,其自由基清除能力AE分别为0.84×10-2、1.08×10-2、1.14×10-2、1.16×10-2、1.58×10-2、0.59×10-2、0.59×10-2。     

对大鼠脊髓损伤应用红花注射液的保护作用

[目的]采用重物下降打击法,制备相当于中度大鼠脊髓损伤(spinal cord injury,SCI)的模型,探讨红花注射液对SCI的保护效果。[方法]将72只SD大鼠随机分为假手术组(A组)、脊髓打击损伤组(B组)、甲基强的松龙组(methylprednisolone sodium succinate,MPSS)(C组)、红花溶液组(D组),每组18只。B、C、D组根据改良的重物撞击装置制备脊髓急性打击损伤动物模型,C、D组在脊髓打击损伤后即刻分别腹腔注射MPSS 30 mg/kg和红花溶液100 mg/kg,观察并检测4组大鼠SCI后1、24、48 h 3个不同时间点后肢运动功能评分变化以及脊髓组织病理学、组织含水量、乳酸(lactic acid,LD)含量、乳酸脱氢酶(lactic dehydrogenase,LDH)活性、超氧化物歧化酶(superoxide dismutase,SOD)活性、丙二醛(malondialdehyde,MDA)含量的改变。[结果]术后A组大鼠功能完全恢复;C组或D组与B组相比,大鼠后肢运动功能24、48 h 2个时间点均有所改善,但24 h时已出现明显差异(P<0.05)...     

葡萄籽原花青素对糖尿病大鼠糖基化终产物及肾脏结缔组织生长因子的影响

葡萄籽原花青素（GSPE）是一类在植物界广泛存在的具有抗氧化作用的多酚化合物。本研究观察GSPE对糖尿病大鼠糖基化终产物（AGE）、肾脏结缔组织生长因子（CTGF）的影响,探讨其对糖尿病大鼠肾脏的保护作用及有关机制,为GSPE应用于糖尿病肾病（DN）提供依据。     

葡萄籽原花青素对犬肝动脉栓塞性肝损伤的保护作用

目的观察葡萄籽原花青素（GSPE）对犬碘化油栓塞性肝损伤的保护作用。方法健康家犬12只,随机分为3组：A组经导管向肝动脉注入碘化油10ml;B组经导管向肝动脉注入碘化油10ml＋GSPE饱和溶液5ml;C组经导管向肝动脉注入生理盐水10ml。测定各组犬血清中丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）活性和丙二醛（MDA）、肿瘤坏死因子-α（TNF-α）的含量,同时对肝组织进行病理学检查。结果栓塞后A、B组犬血清ALT、AST值、MDA和TNF-α含量明显高于C组（P均〈0.05）,肝组织的病理损伤明显;栓塞后B组犬血清ALT、AST值、MDA和TNF-α含量明显低于A组（P均〈0.05）,且肝组织病理损伤程度较A组减轻。结论碘化油栓塞犬肝动脉可成功建立犬急性肝损伤模型。GSPE对犬碘化油栓塞性肝损伤具有明显保护作用,其机制可能与其强大的抗氧化作用及抑制炎症反应有关。     

脱脂葡萄籽中低聚原花青素的提取

研究了葡萄籽中低聚原花青素的提取.采用石油醚脱脂的干葡萄籽为原料,以乙醇为溶剂热提取后,通过调节pH除去蛋白质,再用乙酸乙酯萃取分离粗提物,最后用石油醚沉淀纯化产品.     

黄芪注射液对大鼠急性脊髓损伤细胞免疫调节作用影响的实验研究

目的研究并探讨传统中药黄芪对大鼠急性脊髓损伤（spinal cord injury,SCI）细胞免疫调节作用的影响,为其在SCI治疗中的应用提供理论依据。方法SD大鼠48只随机分为4组,每组12只。其中1组为正常对照组,另外3组为脊髓损伤组,采用改良Allen＇s法建立大鼠脊髓损伤模型,随机分为（1）SCI未治疗组;（2）SCI注射生理盐水组（简称SCI＋SAL组）;（3）SCI注射黄芪注射液组（简称SCI＋HQ组）。正常对照组和SCI未治疗组未予以任何药物治疗,SCI＋SAL组和SCI＋HQ组分别在大鼠急性脊髓损伤后12 h、36 h、5～9 d注射生理盐水和黄芪注射液,并用流式细胞仪检测大鼠外周血T淋巴细胞表型CD4和CD8的表达水平。观察胸腺和脾脏的外观形态改变、重量变化及其组织病理学变化。结果 SCI＋HQ组的大鼠伤后36 h外周血T淋巴细胞CD4值较12 h升高,至伤后第5天CD4值明显高于SCI未治疗组（P〈0.01）,与SCI＋SAL组相比,虽无统计学差异,但有明显高于SCI＋SAL组的趋势。伤后5～8 d继续采用黄芪注射液治疗,至伤后第9天,SCI＋HQ组的CD4值明显低于正常对照组（P〈0.05）。停用黄芪注射液,至伤后第14天,SCI＋HQ组的CD4值又逐渐恢复,与正常对照组无统计学差异。SCI＋HQ组的胸腺重量及其占体重的百分比均明显高于SCI＋HQ组（P〈0.01及P〈0.05）。结论黄芪可促使脊髓损伤后外周血T淋巴细胞的CD4值明显升高,对细胞反应低下有显著提高作用,可作为一种免疫增强调节剂纠正免疫反应低下状态;而过大剂量的黄芪也可能是种免疫抑制剂,抑制大鼠的细胞免疫反应。     

汉防己甲素对大鼠急性脊髓损伤的作用及意义

目的：探讨汉防己甲素（Tet）对急性脊髓损伤（ASCI）的保护作用及作用机制。方法：81只大鼠随机分为三组：生盐水对照组（NS组）、汉防己甲素治疗组（Tet组）和甲基强松龙治疗组（MP组），每组27只，用加速型Allen′s打击法制成脊髓急性损伤模型，监测大鼠ASCI后及用药后的血压变化，测定损伤区脊髓Ca^2 、MDA含量；采用氢清除法测定脊髓伤区血流量（SCBF）的变化；连续观测6周ASCI后运动功能评分情况，ASCI后4h,8h、6周取伤区标本进行组织病理检查，结果：ASCI后10s各组动物的MABP显著升高，1min内迅速降至正常水平，之后Tet组舒张压，平均动脉明显降低（P＜0．05），而收缩压降低不明显（P＞0．05）；ASCI后SCBF下降，但Tet组和MP组的SCBF较NS组高（P＜0．05）；损伤区Ca^2 及MDA含量Tet组和MP组均较NS组低，神经功能评分均较NS组高。结论：Tet能改善微循环，防止SCBF的减少；抗脂质过氧化损伤，减少脂质过氧化物MDA的生成；减轻Ca^2 的局部积聚，防止钙超载，阻断继发性损伤的链式反应，减轻组织继发性损伤。对实验性急性脊髓损伤有保护作用。     

复方丹参注射液和甲基强的松龙治疗大鼠急性脊髓损伤的作用及机制

目的探讨联合应用复方丹参注射液和甲基强的松龙(MP)对大鼠急性脊髓损伤的治疗作用和意义.方法采用Allen法将64只SD大鼠制成脊髓中度损伤模型,随机分为4组,每组16只,分别给予复方丹参注射液(丹参组)、MP(MP组)、二药联合应用(联合组)及生理盐水(对照组)治疗,观察比较治疗前后动物不同时间的神经功能评分、斜板试验和体感诱发电位,并定量观察受损伤脊髓组织的原位末端标记(TUNEL)阳性细胞数目变化.结果从伤后3周起,治疗组神经功能评分、斜板试验角度以及体感诱发电位与对照组比较差异均有显著性(P＜0.01),联合治疗组与丹参组及激素组比较差异有显著性(P＜0.05).从伤后1周起,丹参组和对照组比较,胶质细胞TUNEL阳性细胞数目显著性减少(P＜0.05).结论复方丹参注射液对大鼠急性脊髓损伤有治疗作用,联合应用复方丹参注射液与MP治疗脊髓损伤具有协同作用.复方丹参注射液可能通过抑制胶质细胞凋亡发挥其治疗作用.     

实验性大鼠胚胎脊髓移植对损伤脊髓影响的观察

目的 建立了一个胚胎脊髓（ｆｅｔａｌ ｓｐｉｎａｌ ｃｏｒｄ）组织的低温保存方法笔一个损伤移植模式。方法 通过病理学、组织化学和免疫组织化学的研究,选用胎龄１４天的Ｗｉｓｔａｒ大鼠,分离胚胎脊髓,逐步降温,最后置于液氮中保存。１５天后取出,快速复温,并植入损伤大鼠脊椎内。全部动物于伤后８周处死。结果 经过低温保存的移植物能够在异体脊髓内存活并与宿主组织融合,其在损伤移植区的胶质细胞增生少于对照组。     

葡萄籽原花青素对小鼠睾丸扭转复位后生精功能的保护作用

目的:探讨葡萄籽原花青素(GSP)对小鼠睾丸扭转复位后生精功能的保护作用。方法:24只健康雄性昆明小白鼠(8周龄,25～27 g)随机分为3组:对照组、扭转组、治疗组,每组8只。扭转组及治疗组建立单侧睾丸扭转复位动物模型,治疗组于扭转复位前30 min腹腔注射GSP(50 mg/kg),术后采用腹腔注射方式连续给药3 d,每天1次,每次50 mg/kg。扭转组方法同治疗组,治疗同体积生理盐水。术后第4天取扭转侧睾丸,检测组织病理学参数和生精细胞凋亡指数(AI),并检测睾丸组织超氧化物歧化酶(SOD)和丙二醛(MDA)含量。对照组行假手术。结果:治疗组与扭转组相比,Johnsen评分上升[(7.38±0.92)分vs(5.00±1.85)分,P<0.05],生精小管直径略增大[(178.75±1.58)μm vs(176.50±1.60)μm,P>0.05],生精细胞层数增加[(5.75±0.71)层vs(3.75±1.03)层,P<0.05],生精细胞凋亡指数AI明显降低[(16.25±1.67)%vs(40.50±1.60)%,P<0.05)],SOD活性明显上升[(52.67±3.57)U/mg prot vs(29.04±4.46)U/mg prot,P<0.05],MDA含量明显下降[(2.91±0.04)nmol/mg prot vs(4.63±0.05)nmol/mg prot,P<0.05]。结论:GSP对小鼠睾丸扭转复位后生精功能损伤有明显的保护作用,其作用机制可能与其能清除氧自由基、抑制脂质过氧化、提高机体抗氧化能力有关。     

EGb761对急性大鼠脊髓损伤后血脊髓屏障的保护作用及其机制研究

[目的]观察银杏叶提取物(EGb761)对急性大鼠脊髓损伤后炎症反应及血脊髓屏障的保护作用,探讨其对急性脊髓损伤的作用机制.[方法]120只雌性SD大鼠,体重200～220 g,随机分为假手术组(A组)、损伤对照组(B组)、EGb761治疗组(C组),每组40只.A组仅行T9椎板切除术,不致伤脊髓.B、C组用改良Allen's法以25gcf致伤力制作大鼠T9脊髓损伤模型,C组术后至处死前每日经腹腔给予EGb761 100 mg/kg(首次给药在术后30min),A、B组在同一时间给予等量生理盐水.术后3、6、24、72 h分批处死动物,以T9为中心取损伤节段脊髓,采用Evans蓝含量测定法观察SCI后血脊髓屏障(blood cerebrospinal barrier,BSCB)通透性的变化,采用ELISA法测定脊髓组织白介素-1β(interleukin-1β,IL-1β)的含量,免疫组织化学方法检测细胞间粘附分子-1(intercellular adhesion molecular-1,ICAM-1)在脊髓组织中的表达变化.[结果]A组各时间点脊髓中无明显Evan's蓝渗漏,IL-1β含量及ICAM-1表达维持在基础水平;B组各时间点Evan's蓝渗漏、IL-1β含量及ICAM-1表达量均明显高于A组;C组EGb761干预后在6、24、72 h时Evan's蓝通过BSCB漏入脊髓组织量显著低于B组,在各时间点IL-1β、ICAM-1的表达均较B组明显减少.[结论]在Allen's大鼠急性脊髓损伤模型中,EGb761能降低局部脊髓中IL-1β含量,下调ICAM-1的表达,减轻BSCB的破坏.     

Protective effect of deferoxamine on experimental spinal cord injury in rat

Objective

To observe the protective effect of deferoxamine on experimental spinal cord injury (SCI) in rats.

Methods

Sprague-Dawley rats were randomly divided into the following four groups. Control group: rats were performed laminectomy only; SCI group: rats were performed laminectomy with SCI; DFO group: rats were injected intraperitoneally a bolus of 100 mg/kg deferoxamine after SCI; vehicle group: rats were injected intraperitoneally 0.9% saline after SCI. The SCI of animal model was made by using a modified Allen's method on T₁₀. Six rats of each group were sacrificed at 4 h after injured, and the levels of free iron and malondialdehyde (MDA) of involved spinal cord segments were measured by bleomycin assay and the thiobarbituric acid (TBA) separately. The recovery of function was assessed by Modified Tarlov's scale and inclined plane method at 7, 14, 21 d after SCI. The histologic changes of the damaged spinal cord were also examined at 7 d after SCI.

Results

Following SCI, the levels of free iron and MDA were increased significantly and the Modified Tarlov's score and inclined plane angles decreased in SCI group and vehicle group. In DFO group, the levels of free iron and MDA were not increased, but the Modified Tarlov's score and inclined plane angles decreased, the histological findings were improved as well.

Conclusion

Deferoxamine can reduce the levels of free iron and lipid peroxidation, and improve the hind limb functional status of rats with spinal cord injury.     

Protective effect of ketamine on ischemic spinal cord injury in rabbits

We tested our hypothesis that a commonly used anesthetic, ketamine, may offer benefits to protect animals from spinal cord injury, using the ischemia/reperfusion (I/R) injury rabbit model in a randomized controlled study. We used 24 white adult Japanese rabbits from the animal facility at the Medical College of Wuhan University. The rabbits were randomly assigned to one of three groups, eight rabbits per group: group I, sham-operation group; group II, I/R group; group III, I/R with ketamine treatment group. Spinal cord ischemia was induced by infrarenal aortic cross-clamp for 45 min in group II and group III, and ketamine was intravenously infused at 10 mg/kg in 15 mL 0.9% sodium chloride at a speed of 1.5 mL/min to animals in group III, once at 10 min before aortic clamping and once at the onset of reperfusion. Postoperative neurological function, electromyography of rear limbs, histopathology, malondialdehyde (MDA) levels, and superoxide dismutase (SOD) activity in the spinal cord were assessed in all animals. Compared with the control group I, group II showed significant I/R injury-induced changes in neurological function scores, histopathology, and electromyography (p < 0.01). However, group III with ketamine treatment significantly reversed the changes in all these parameters (p < 0.01). At the same time, the I/R-induced increase in MDA content observed in group II was also significantly reduced in group III (p < 0.01), and the I/R-induced decreases in SOD activity were also significantly prevented in group III (p < 0.01). After ketamine treatment, all parameters examined in group III were not significantly different from those obtained in group I. Ketamine showed potent protective effects against spinal cord I/R injury in the rabbit model and protected loss of antioxidant activity in spinal cord tissues.     

Protective effect of methylprednisolone on vascular injury in rat spinal cord injury.

High-dose methylprednisolone (MP) given to patients within 8 h of traumatic spinal cord improved neural function at 6 and 12 months, suggesting a probable secondary injury process that may be amenable to therapeutic intervention. Vascular injury plays an important role in the secondary injury process of CNS trauma. We have examined the effect of MP on vascular changes, including tissue edema, vascular permeability, and polymorphonuclear (PMN) cell infiltration in a rat model of spinal cord impact injury. MP significantly reduced extravasation of fluorescein isothiocyanate dextran (FITC-D), a macromolecular tracer, by 64.3% and 50.7% with trauma forces of 20 and 40 g-cm, respectively, when MP was administered IV immediately after trauma at a bolus of 165 mg/kg, with a subsequent continuous MP infusion at 31.5 mg/kg/h for 23 h. MP reduced the water content in the 40 g-cm traumatic cord lesion to 73.0% compared to the traumatic control (74.3%, p < 0.001) at the same schedule of large dose 24-h infusion. The same doses of MP showed a trend to decrease the extent of neutrophil infiltration as determined by myeloperoxidase (MPO) activity, but the change was not significant. MP had little effect in decreasing FITC-D extravasation and cord edema when given at a lower dose (bolus of 30 mg/kg with continued infusion of 1.3 mg/kg/h for 23 h). MP did not reduce extravasation of FITC-D and edema when administered IV as one bolus injection at high (165 mg/kg) or low (30 mg/kg) doses.(ABSTRACT TRUNCATED AT 250 WORDS)     

Anti-apoptotic and anti-oxidant effects of grape seed proanthocyanidin extract in preventing cyclosporine A-induced nephropathy

Aim: Although the pathogenesis of cyclosporine (CsA) nephropathy is not completely understood, it is attributed to oxidative damage and apoptosis. Grape seed proanthocyanidin extract (GSPE) is a molecule with anti‐oxidant and anti‐apoptotic properties. Our aim was to demonstrate the effects of GSPE in preventing CsA nephropathy. Methods: Twenty‐four Sprague–Dawley rats were divided into four groups. The control, GSPE, CsA and CsA+GSPE groups were given 1 mL olive oil, 100 mg/kg GSPE, 25 mg/kg CsA and 100 mg/kg GSPE+25 mg/kg CsA, respectively. On day 21, blood samples were taken for blood urea nitrogen (BUN), creatinine and CsA levels, and renal tissue was used for total oxidant system (TOS), total anti‐oxidant system (TAS), oxidative stress index (OSI) and malondialdehyde (MDA) measurements. In addition to renal histopathology, apoptosis staining was performed on renal tissue. Results: The BUN, creatinine, TOS, OSI, MDA, histopathological score, and apoptotic index exhibited increases in the CsA group. In the CsA+GSPE group, however, BUN, creatinine, OSI, MDA, renal histopathological score and apoptotic index (AI) decreased and TAS levels increased. In addition, there was no difference between the CsA and CsA+GSPE groups with regard to CsA levels. Conclusion: We demonstrated that GSPE prevents CsA nephropathy and that this effect is achieved by anti‐apoptotic and anti‐oxidant activity. We also achieved a significant recovery in kidney functions without affecting CsA plasma levels.     

Protective effects of oral creatine supplementation on spinal cord injury in rats

STUDY DESIGN: To evaluate a potential protective effect of increased creatine levels in spinal cord injury (SCI) in an animal model. OBJECTIVES: Acute SCI initiates a series of cellular and molecular events in the injured tissue leading to further damage in the surrounding area. This secondary damage is partly due to ischemia and a fatal intracellular loss of energy. Phospho-creatine in conjunction with the creatine kinase isoenzyme system acts as a potent intracellular energy buffer. Oral creatine supplementation has been shown to elevate the phospho-creatine content in brain and muscle tissue, leading to neuroprotective effects and increased muscle performance. SETTING: Zurich, Switzerland. METHODS: Twenty adult rats were fed for 4 weeks with or without creatine supplemented nutrition before undergoing a moderate spinal cord contusion. RESULTS: Following an initial complete hindlimb paralysis, rats of both groups substantially recovered within 1 week. However, creatine fed animals scored 2.8 points better than the controls in the BBB open field locomotor score (11.9 and 9.1 points respectively after 1 week; P=0.035, and 13 points compared to 11.4 after 2 weeks). The histological examination 2 weeks after SCI revealed that in all rats a cavity had developed which was comparable in size between the groups. In creatine fed rats, however, a significantly smaller amount of scar tissue surrounding the cavity was found. CONCLUSIONS: Thus creatine treatment seems to reduce the spread of secondary injury. Our results favour a pretreatment of patients with creatine for neuroprotection in cases of elective intramedullary spinal surgery. Further studies are needed to evaluate the benefit of immediate creatine administration in case of acute spinal cord or brain injury.