NF-κB/IκBα在MNU致实验性大鼠视网膜损伤中的变化

目的观察核因子κappaB(NF-κB)在N-甲基-N-亚硝脲(MNU)诱导大鼠视网膜光感受器细胞凋亡中的的变化,探讨MNU损伤视网膜的机制。方法给生后50d的雌性SD大鼠一次腹腔注射MNU60mg/kg,分别在MNU处理不同时间后处死动物。光学显微镜观察视网膜的形态学变化;TUNEL试剂盒检测光感受器细胞凋亡;Westernblotting分析NF-κB。结果MNU处理后24h,视网膜光感受器细胞核固缩和光感受器细胞层外节部定向障碍;7d后,外颗粒层和光感受层几乎完全消失。光感受器细胞凋亡在MNU处理后24h达高峰。在凋亡发生的过程中,仅在MNU作用12h和24h后,胞核内有低水平的p65蛋白。相反,胞浆和胞核内的κB蛋白水平呈时间依赖性地显著增加。结论NF-κB/IκBα信号通路的激活可能介导了MNU所致的视网膜损伤。

马齿苋多糖对溃疡性结肠炎大鼠肠组织IL-6/STAT3及NF-κB的影响

目的:采用TNBS(2,4,6-三硝基苯磺酸)复制溃疡性结肠炎大鼠模型,探索马齿苋多糖对溃疡性结肠炎大鼠肠组织IL6/STAT3及NF-κB的影响,明确IL-6/STAT3信号通路与慢性炎症性肠病发病的关系,为慢性溃疡性结肠炎的治疗寻找新靶点。方法:将40只SD大鼠随机分为对照组、模型组、美沙拉嗪组和马齿苋组(n=10)。采用TNBS诱导复制结肠炎模型,造模成功后第3天开始灌胃给药:美沙拉嗪组剂量为每次10 mg/kg,每日1次,连续3周;马齿苋组给予马齿苋多糖,每次10 ml/kg,每日1次,连续3周;模型组和对照组大鼠给予等体积生理盐水灌胃,每日1次,连续3周。收集大鼠结肠内容物称重,干燥后再次称重,取结肠组织作病理切片。采用ELISA试剂盒检测血清IL-6、IL-1β、TNF-α和核转录因子-kappa B(NF-κB)含量;免疫组化染色法测定结肠髓过氧化物酶(MPO);RT-PCR法检测信号转导和转录激活因子(STAT3)、IL-6的mRNA。结果:与模型组、美沙拉嗪组比较,马齿苋组大鼠排便状态明显改善,肠粘膜水肿减轻;血清IL-6、s IL-6Rα、gp130,肠组织MPO、NF-κB含量均降低(P<0.01)。与模型组比较,马齿苋组STAT3、IL-6mRNA的表达水平明显降低(P<0.01)。与对照组比较,上述指标无显著性差异(P>0.05)。结论:马齿苋多糖通过降低大鼠血清IL-6、s IL-6Rα、gp130含量及肠组织MPO、NF-κB水平,减轻s IL-6Rα与IL-6形成复合物所致的炎症反应;经IL-6/STAT3信号通路下调大鼠肠组织STAT3和IL-6的mRNA水平,从而抑制炎症的发生。

Hhcy及B族维生素干预对大鼠脑血管ICAM-1表达的影响

目的:探讨高同型半胱氨酸血症(Hhcy)及叶酸、维生素B12和维生素B6干预对大鼠脑皮层血管细胞间黏附分子-1(ICAM-1)表达的影响。方法:将40只雄性SD大鼠随机分为对照组、Hhcy组、治疗组和预防组,采用不同饮食结构及干预手段制备动物模型。采用高效液相色谱(HPLC)荧光检测技术测定不同时期各组大鼠血浆总同型半胱氨酸(tHcy)水平,并于第12周末通过免疫组化法观察ICAM-1在大鼠脑皮层血管的表达情况。结果:第4周末大鼠Hhcy模型制备成功,而第12周末对照组、治疗组和预防组血浆tHcy水平差异无统计学意义;预防组大鼠脑皮层ICAM-1阳性血管数与对照组差异无统计学意义,余各组比差异有统计学意义(P<0.05或P<0.01)。结论:Hhcy可以促进大鼠皮层脑血管ICAM-1表达,B族维生素可以有效降低血浆tHcy水平,并且对ICAM-1表达有一定的干预作用。

白屈菜碱对活化后大鼠肝星状细胞CFSC-8B的增殖、胶原合成及TGF-β1受体的影响

目的:考察白屈菜碱对活化后大鼠肝星状细胞CFSC-8B的增殖、胶原合成及转化生长因子β(1 TGF-β1)受体的影响。方法:取对数生长期的CFSC-8B细胞,分为正常对照组、模型组、溶剂组(乙醇)、阳性对照组(1μg/mL秋水仙碱乙醇溶液)和白屈菜碱低、中、高浓度组(2.1、4.2、8.4μg/mL白屈菜碱乙醇溶液)。除正常对照组外,其余各组细胞利用20μg/L的TGF-β1作用24h进行活化,后5组细胞给予相应药物干预24h。采用CCK-8法检测各组细胞增殖情况,酶消化法检测细胞上清液中羟脯氨酸(Hyp)含量,酶联免疫吸附测定法(ELISA)检测细胞上清液中Ⅰ型胶原蛋白(Col-Ⅰ)和Ⅲ型胶原蛋白(Col-Ⅲ)水平,Western blot法检测细胞中TGF-β1Ⅰ型受体(TβR-Ⅰ)和TβR-Ⅱ蛋白的表达,实时荧光定量-聚合酶链式反应法检测细胞中α-平滑肌肌动蛋白(α-SMA)、TβR-Ⅰ和TβR-ⅡmRNA的表达。结果:与正常对照组比较,模型组细胞增殖率、Hyp含量、Col-Ⅰ和Col-Ⅲ水平、TβR-Ⅰ和TβR-Ⅱ的蛋白表达水平以及α-SMA、TβR-Ⅰ、TβR-Ⅱ的mRNA表达水平均显著升高(P<0.05)。与模型组比较,溶剂组细胞的上述指标均无明显差异(P>0.05);白屈菜碱低浓度组细胞增殖率无明显差异(P>0.05);阳性对照组和白屈菜碱高浓度组细胞的上述指标均显著降低(P<0.05);白屈菜碱中浓度组细胞除Hyp和Col-Ⅲ水平降低不显著外,上述其余指标均显著降低(P<0.05)。与白屈菜碱中浓度组比较,白屈菜碱高浓度组细胞增殖率、Col-Ⅰ水平、TβR-Ⅰ和TβR-Ⅱ的蛋白及mRNA表达水平均显著降低(P<0.05)。结论:白屈菜碱能抑制活化后CFSC-8B细胞的增殖、胶原合成以及TβR-Ⅰ、TβR-Ⅱ蛋白和mRNA的表达。

Effects of Shuang Dan Ming Mu Capsule on Expression of VEGF-a, VEGF-b, VEGF-c and the VEGF receptor, Flk-1, in Diabetic Retinopathy Rats

Objective The effects of Shuang Dan Ming Mu capsule on expression of VEGF-a,VEGF-b,VEGF-c and the VEGF receptor,Flk-1,were examined in a diabetic retinopathy rats model.Methods Forty Sprague-Dawley(SD)rats were randomized into Groups A(blank),B(model),C(Shuang Dan Ming Mu)and D(positive control)group,with each group containing10rats and20eyes.Rats from groups B,C and D were administered one dose of50mg/kg streptozotocin(STZ)by tail vein injection to establish a diabetic rat model.One week after model preparation,medication was continuously administered by gavage.After gavage for8weeks,the animals were sacrificed and retinal expression of VEGF-a,VEGF-b,VEGF-c and Flk-1was quantified by immunohistochemical analysis.Results At week8of drug administration after model preparation,the average protein expression grayscale values for VEGF-a,VEGF-b,VEGF-c and Flk-1in the rats model,Shuang Dan Ming Mu and positive control groups were all lower than those in the normal group,while the mean optical density values were higher than those in the normal group.When the model group was compared to the normal group,the difference was extremely significant(P<0.01).The mean grayscale values of VEGF-a,VEGF-b,VEGF-c and Flk-1in the Shuang Dan Ming Mu and positive control groups were all higher than those in the model group,while the mean optical density values were lower than those in the model group(P<0.05or0.01).Conclusion Shuang Dan Ming Mu capsule can significantly decrease the expression of VEGF-a,VEGF-b,VEGF-c and Flk-1in the retinas of diabetic model rats and exhibit some protective effects in their retinas.

补阳还五汤对失神经肌萎缩大鼠NF-κB/MuRFl表达的影响

为了研究补阳还五汤(黄芪、当归、白芍、地龙、川芎、红花、桃仁)对失神经肌萎缩大鼠NF-κB/MuRFl信号通路的影响,探讨其对失神经肌萎缩的保护机制,该实验建立大鼠腓总神经夹伤模型。将造模后的60只SPF级SD大鼠随机分为:假手术组、补阳还五汤高、中、低剂量组、弥可保组与模型组,术后每日灌胃给药。10 d与21 d后qRT-PCR与Western blot检测NF-κB,MuRFl基因与蛋白的差异表达。结果显示:与模型组相比,补阳还五汤中、高剂量组NF-κB,MuRFl mRNA与蛋白表达均显著降低(P<0.05或P<0.01)。提示:补阳还五汤可能通过拮抗NF-κB/MuRFl信号通路,抑制蛋白酶体系统导致的蛋白质降解来防治失神经肌萎缩的发生。

Effect of moxibustion on tumor necrosis factor-α and nuclear transcription factor kappa B in ankle joints of rats with rheumatoid arthritis

Objective： To observe the effect of moxibustion on the expressions of tumor necrosis factor-α（TNF-α） and nuclear transcription factor kappa B（NF-κB） proteins in ankle joints of rats with rheumatoid arthritis（RA）, and to explore the anti-inflammatory mechanism of moxibustion in the treatment of RA. Methods： Adjuvant arthritis（AA） rat models were induced and used as rat models of RA. Rats were randomly divided into a normal group, a model group, a moxibustion group and a saline group. Rats in the normal and model groups were not treated; rats in the moxibustion group accepted treatment by moxibustion at Zusanli（ST 36） and Shenshu（BL 23） after modeling; rats in the saline group were injected with 0.15 mL saline to the bottom of the left hind paw and had no other treatments. Hematoxylin-eosin（HE） staining was used to observe the histopathological changes of rats＇ ankle joints under light microscope. Immunohistochemistry was used to observe the expressions of TNF-α and NF-κB in ankle joints of rats. Results： Compared with the normal group, the rats＇ ankle joints in the model group showed disorganization, joint surface defect, and significantly increased mean optical density（MOD） of TNF-α and NF-κB（all P〈0.05）. After moxibustion treatment, rats in the moxibustion group showed repaired ankle tissues, smooth joint surface without defects, decreased MOD of TNF-α and NF-κB than those in the model group（all P〈0.05）. Rats in the saline group had no damage to ankle joints, while TNF-α and NF-κB were significantly different from those in the model group（P〈0.05）. Conclusion： Moxibustion could down-regulate the expressions of NF-κB and TNF-α protein in ankle joints of RA rats, and produce an anti-inflammatory effect to promote tissue repair.

Inhibition of hypoxia and serum deprivation-induced apoptosis by salvianolic acid in rat mesenchymal stem cells

OBJECTIVE:To investigate the influence and mechanism of salvianolic acid B(SalB) on apoptosis inhibition in rat bone marrow-derived mesenchymal stem cells(BMSCs) induced by hypoxia and serum deprivation(hypoxia/SD).METHODS:SalB concentration of 0.1,1,10 or 100 mg/L(drug groups) were investigated for their ability to inhibit apoptosis in rat BMSCs.BMSCs in both the apoptosis model and drug groups were cultured under hypoxic conditions for 6 h,after which cell apoptosis and change in mitochondrial membrane potential(MMP) were detected using flow cytometry.Activation of caspase-3 was detected using western blot analysis.RESULTS:Hypoxia/SD induced apoptosis in rat BMSCs.The early apoptosis rate was lower in the drug groups compared to the apoptosis model group(P<0.05).SalB was found to inhibit the reduction in MMP and decrease the activation of caspase-3.CONCLUSION:0.1,1 and 10 mg/L of SalB inhibits activation of caspase-3 and early apoptosis of rat BMSCs induced by hypoxia/SD and could therefore enhance the survival rate of grafted stem cells.