Anti-inflammatory activity of Dasyphyllum brasiliensis (Asteraceae) on acute peritonitis induced by beta-glucan from Histoplasma capsulatum

The tea prepared from leaves and thorns of Dasyphyllum brasiliensis (Asteraceae) is used in the traditional medicine in Brazil for the treatment of oral and oropharyngeal diseases. In this study, we investigated the anti-inflammatory activity of this plant. The aqueous crude extract (ACE), the methanol-water (MeOH-H(2)O) fraction obtained by solvent partition and its fractionation products were evaluated for their anti-inflammatory activities on acute peritonitis induced by beta-glucan from the cell walls of Histoplasma capsulatum. The antiedematogenic activity was also tested using the carrageenan-induced paw edema assay in mice. Oral administration of 100 and 300mg/kg of the ACE in mice caused a significant reduction of neutrophil and eosinophil recruitment in the acute peritonitis assay. In addition, ACE at 300mg/kg inhibited the number of mononuclear cells recruitment. The MeOH-H(2)O fraction and its fractionation products (all at 100mg/kg) also presented anti-inflammatory activities, confirmed by the inhibition of cells recruited to the peritoneal cavity. ACE at 100mg/kg did not show any significant reduction of the edema in the mice paw injected with carrageenan. These data together suggest that Dasyphyllum brasiliensis presents significant anti-inflammatory activity, thus supporting the popular use of the tea in the treatment of inflammatory diseases.     

Role of IKK and oscillatory NFkappaB kinetics in MMP-9 gene expression and chemoresistance to 5-fluorouracil in RKO colorectal cancer cells

Nuclear factor kappa B (NFkappaB) is a central participant in the metastasis and chemoresistance of colorectal cancer (CRC). However, it is not fully understood to what extent NFkappaB contributes to induction of the metastasis-associated matrix metalloprotease-9 (MMP-9) gene and sensitivity to the commonly used chemotherapeutic 5-fluorouracil (5-Fu) in CRC. Using the RKO human CRC cell line and two NFkappaB signaling deficient RKO mutants, we investigated NFkappaB's role in the induction of MMP-9 and 5-Fu sensitivity in RKO CRC cells. NFkappaB plays a predominant role in MMP-9 gene induction in RKO cells, as evidenced by the failure of tumor necrosis factor alpha (TNFalpha) to induce MMP-9 in either of the NFkappaB signaling mutants. RKO cells exhibit a robust, oscillatory NFkappaB activity in response to TNFalpha not seen in either of the NFkappaB mutant cell lines, which instead demonstrate diminished, nonoscillatory NFkappaB activation. Analysis of TNFalpha-induced phosphorylation and MMP-9 promoter recruitment of the p65 NFkappaB subunit revealed a significant reduction in p65 phosphorylation as well as reduced and altered recruitment of p65 to the MMP-9 gene promoter in the mutants compared to the parental RKO cell line. 5-Fu only activated NFkappaB in the parental RKO cells through induction of IkappaB-kinase (IKK) activity and increased sensitivity to 5-Fu is observed in both NFkappaB mutant lines. Our results suggest that TNFalpha-dependent induction of MMP-9 gene expression is tightly regulated by oscillatory/cumulative activation of NFkappaB and that 5-Fu stimulates NFkappaB and RKO CRC cell survival through induction of IKK activity.     

Molecular mechanisms for apigenin-induced cell-cycle arrest and apoptosis of hormone refractory human prostate carcinoma DU145 cells

Development of effective agents for treatment of hormone-refractory prostate cancer has become a national medical priority. We have reported recently that apigenin (4',5,7-trihydroxyflavone), found in many common fruits and vegetables, has shown remarkable effects in inhibiting cell growth and inducing apoptosis in many human prostate carcinoma cells. Here we demonstrate the molecular mechanism of inhibitory action of apigenin on androgen-refractory human prostate carcinoma DU145 cells that have mutations in the tumor suppressor gene p53 and pRb. Treatment of cells with apigenin resulted in a dose- and time-dependent inhibition of growth, colony formation, and G1 phase arrest of the cell cycle. This effect was associated with a marked decrease in the protein expression of cyclin D1, D2, and E and their activating partner, cyclin-dependent kinase (cdk)2, 4, and 6, with concomitant upregulation of WAF1/p21, KIP1/p27, INK4a/p16, and INK4c/p18. The induction of WAF1/p21 and its growth inhibitory effects by apigenin appears to be independent of p53 and pRb status of these cells. Apigenin treatment also resulted in alteration in Bax/Bcl2 ratio in favor of apoptosis, which was associated with the release of cytochrome c and induction of apoptotic protease-activating factor-1 (Apaf-1). This effect was found to result in a significant increase in cleaved fragments of caspase-9, -3, and poly(ADP-ribose) polymerase (PARP). Further, apigenin treatment resulted in downmodulation of the constitutive expression of nuclear factor-kappaB (NF-kappaB)/p65 and NF-kappaB/p50 in the nuclear fraction that correlated with an increase in the expression of IkappaB-alpha (IkappaBalpha) in the cytosol. Taken together, we concluded that molecular mechanisms during apigenin-mediated growth inhibition and induction of apoptosis in DU145 cells was due to (1) modulation in cell-cycle machinery, (2) disruption of mitochondrial function, and (3) NF-kappaB inhibition.     

核因子κB在大鼠胃缺血再灌注损伤中的表达及意义

目的研究核因子κB(NF-κB)在大鼠胃缺血再灌注损伤中的表达及意义。方法采用大鼠胃缺血再灌注(gastric ischemia/reperfusion,GI/R)模型(夹闭腹腔动脉30 min后再灌注),分别于再灌注0.5、1、32、4 h取胃,计算胃黏膜损伤指数。应用免疫组化、Western blot方法检测胃黏膜NF-κB p65。结果大鼠GI/R后引起胃黏膜损伤,再灌注1 h时损伤最明显,随后降低,24 h接近正常。GI/R后胃黏膜NF-κB阳性细胞数和蛋白表达量增多,与胃黏膜损伤变化规律一致。结论NF-κB在大鼠GI/R损伤过程中发挥着重要作用。     

辛伐他汀对动脉粥样硬化家兔血管壁NF—κB—DNA结合活性与MCP-1表达的影响

目的观察辛伐他汀对兔动脉粥样硬化斑块中核因子-κB（NF-κB）．DNA结合活性与单核细胞趋化因子-1（MCP-1）表达的影响，探讨辛伐他汀降脂效应以外的抗动脉粥样硬化（AS）作用机制。方法36只雄性新西兰大耳白兔被随机分为低脂对照组（LC）、高脂对照组（HC）和辛伐他汀组（HC＋S）。实验中动态观察血清总胆固醇（TC）、甘油三酯（TG）和低密度脂蛋白胆固醇（LDL-C）的变化；实验结束时，用电泳移动迁移技术（EMSA）检测三组兔主动脉组织中NF-κB-DNA结合活性；用免疫组化技术观察各组血管组织中MCP-1的表达；显微镜下测定各组主动脉内膜厚度与粥样斑块面积。结果实验结束时，HC＋S与LC组的TC、TG、LDL-C水平、NF-κB-DNA结合活性、MCP-1表达、主动脉内膜厚度和粥样斑块面积均明显小于HC组（P〈0．05）；HC＋S组的的TC、TG和LDL-C水平与LC组相比虽无明显差异（P〉0．05），但其NF-κB-DNA结合活性、MCP-1表达、内膜厚度和粥样斑块面积均小于LC组（P〈0．05）。结论辛伐他汀可以通过抑制NF-κB-DNA结合活性、减弱MCP-1表达而减轻AS的形成。     

氢溴酸高乌甲素对神经病理性疼痛大鼠的影响

目的研究氢溴酸高乌甲素对大鼠慢性缩窄性损伤(CCI)模型神经病理性疼痛的镇痛作用。方法24只大鼠建立CCI模型,分为四组,于术后第7天腹腔分别注射不同剂量的氢溴酸高乌甲素2、4和6mg/kg及生理盐水,给药后进行辐射热痛阈值的测定,测后取大鼠脊髓、脑组织标本采用EMSA进行核因子-κB(NF-κB)的活性测定。结果术后大鼠结扎侧足的热痛阈值明显缩短,氢溴酸高乌甲素4mg/kg组和6mg/kg组与生理盐水组相比较热痛觉过敏反应明显缓解(P<0·01)。NF-κB在生理盐水组中表达最高,2mg/kg组表达最低。结论氢溴酸高乌甲素能明显缓解CCI大鼠的热痛觉过敏现象,达到镇痛效果,同时能使NF-κB的表达减少。     

参附注射液对大鼠肝缺血再灌注损伤保护作用的实验研究

目的:探讨参附注射液对大鼠肝缺血再灌注损伤的保护作用及其机制。方法:24只大鼠随机分为肝缺血再灌注组(IR组,n=12)和参附注射液加肝缺血再灌注组(SF组,n=12)。SF组给予参附注射液10ml/kg,腹腔注射,每日1次,连续给药6d,第6天于手术前30min给药。IR组大鼠同样方法给予相同剂量的生理盐水。两组均于第6天手术,两组均采用Pringle’s法阻断肝门缺血15min再灌注1h、3h,测定血浆血栓素B2(TXB2)和6-酮-前列腺素F1α(6-keto-PGF1α)和肝组织匀浆Na+-K+-ATP酶、Ca2+-Mg2+-ATP酶和SOD、GSH变化,肝组织NF-κBp65表达及肝组织形态学改变。结果:NF-κBp65阳性细胞为细胞核或细胞浆染成棕黄色或有棕黄色颗粒沉积,肝脏缺血再灌注后肝细胞深染,枯否细胞亦可见表达。肝缺血15min再灌注1h,SF组的阳性细胞百分数较IR组显著减低(P<0.05),SOD活性明显高于IR组(P<0.05),还原型GSH水平高于IR组但无显著性差异(P>0.05)。再灌注1h、3hSF组肝组织中Na+-K+-ATP酶、Ca2+-Mg2+-ATP酶水平均显著高于IR组(P<0.05)。再灌注3h后SF组血浆TXB2浓度较IR组低(P>0.05),而6-keto-PGF1a浓度升高(P>0.05),TXB2/6-keto-PGF1a比值显著降低(P<0.05)。SF组肝实质细胞和线粒体损伤明显减轻。结论:参附注射液通过抑制NF-κB活化、提高SOD和ATPase活性、改善TXA2/PGI2平衡保护肝脏缺血再灌注损伤。     

葛根素对脑缺血再灌注后核因子kappaB表达的影响

目的：研究大鼠局灶性脑缺血再灌注后核因子kappaB(NF-κB)在时间和空间上表达的变化过程及葛根素对其的影响；方法：线栓法建立右侧大脑中动脉闭塞大鼠模型,分别于缺血90 min后再灌注2,6,12,24,72 h,缺血前1 h及随后每间隔6 h腹腔内给药。大鼠在各时间点断头取脑,测算脑梗死体积,免疫组织化学方法和免疫印迹法检测脑组织内NF-κB的表达。结果：大鼠缺血再灌注后,免疫组化分析提示NF-κB从细胞浆向细胞内移位,核内表达水平在6 h开始明显升高,在24 h达到高峰,72 h下降,其脑梗死体积随着再灌注时间延长而增加。葛根素明显降低NF-κB在缺血再灌注后24,72 h的表达及减小脑梗死体积。结论：脑缺血再灌注后,脑梗死周边区域出现NF-κB从胞浆至胞核的转位并伴表达增加,葛根素可能通过抑制NF-κB的表达,减轻缺血再灌注损伤。