人工虫草多糖对免疫抑制小鼠免疫调节作用的研究

目的:研究人工虫草多糖对环磷酰胺(CTX)所诱导的免疫抑制小鼠的免疫调节作用。方法:将50只昆明小鼠分为空白组、模型组和人工虫草多糖高、中、低剂量组。采取腹腔注射环磷酰胺诱导造模方式,连续灌胃给药20d。观察各组体质量等一般情况的变化,末次给药后检测脾脏组织中乳酸脱氢酶(LDH)和酸性磷酸酶(ACP)的活性、腹腔巨噬细胞吞噬功能、胸腺及脾脏指数、血清溶血素及小鼠血清细胞因子IL-2、TNF-α含量的变化。结果:与模型组比较,人工虫草多糖各剂量组一般情况均有不同程度的改善,中、高剂量组LDH和ACP活性提高,腹腔巨噬细胞的吞噬指数及胸腺、脾脏指数提升(P0.01),血清溶血素水平及血清IL-2、TNF-α含量均有所提高(P0.05)。结论:人工虫草多糖能够拮抗环磷酰胺的免疫抑制作用,进而有效提高机体免疫功能。     

藏药忍冬果对小鼠腹腔巨噬细胞吞噬功能及分泌细胞因子的影响

目的:研究藏药忍冬果水提物(FLM)对小鼠腹腔巨噬细胞吞噬功能及分泌细胞因子的影响。方法:运用吞噬中性红试验观察FLM对小鼠腹腔巨噬细胞吞噬功能的影响,用小鼠腹腔巨噬细胞分泌物对小鼠胸腺细胞增殖及对L929细胞杀伤作用的影响分别测定IL-1及TNF-α的活性,用RT-PCR技术观察药物对小鼠腹腔巨噬细胞TNF-αmRNA和IFN-γmRNA表达的影响。结果:FLM在1～100μg.mL^-1剂量内对小鼠腹腔巨噬细胞吞噬功能有明显促进作用,同时能诱导巨噬细胞分泌IL-1及TNF-α,并促进细胞因子TNF-αmRNA和IFN-γmRNA的表达。结论:FLM对小鼠腹腔巨噬细胞吞噬功能及分泌细胞因子都有促进作用。     

灵芝多糖对小鼠M1巨噬细胞活性的研究

目的研究灵芝多糖对小鼠M1巨噬细胞活化及对T细胞增殖的影响。方法吉姆沙染色巨噬细胞后,显微镜观察巨噬细胞吞噬现象;ELISA法检测小鼠腹腔巨噬细胞分泌IL-10、IL-12和TNF的水平;MTT法检测M1巨噬细胞诱导T细胞的增殖作用。结果灵芝多糖能促进小鼠M1巨噬细胞对金色葡萄球菌的吞噬作用;体外实验显示,100μg/ml灵芝多糖可以诱导小鼠巨噬细胞分泌IL-10和TNF-α;灵芝多糖介导的巨噬细胞和T细胞的联合培养与脾脏细胞混合培养有显著性差别。结论灵芝多糖能诱导小鼠巨噬细胞活化,产生M1型细胞因子,介导T细胞活化增殖。     

茯苓酸性多糖调节免疫功能活性研究

目的:探讨茯苓酸性多糖提取物调节免疫功能作用。方法:采用氢化可的松皮下注射液注射复制小鼠免疫功能低下模型,灌胃茯苓酸性多糖提取物,检测小鼠腹腔内吞噬细胞功能,测定小鼠血清中IL-2、TNF-α、INF-γ含量以及免疫器官脾脏和胸腺指数。结果:茯苓酸性多糖能增强氢化可的松致免疫低下小鼠巨噬细胞吞噬功能,显著提高小鼠细胞免疫因子IL-2、TNF-α、INF-γ的含量,显著增加脾脏和胸腺指数。结论:茯苓酸性多糖对氢化可的松致免疫低下小鼠具有调节免疫功能的作用。     

芦荟及其复方提取物对RAW264.7细胞炎症因子的影响

目的:观察芦荟及其复方提取物对小鼠巨噬细胞炎症因子的影响.方法:RAW264.7细胞按2×105/mL稀释,100μL/孔接种入96孔板,药物质量浓度为250,125,62.5 g?L-1下采用噻唑蓝比色法(MTT法)作用4h,检测芦荟及其复方提取物对小鼠巨噬细胞的毒性作用;药物质量浓度为50,25,12.5 g?L-1下采用硝酸还原酶法作用24 h,测定小鼠巨噬细胞一氧化氮(N0)含量,并用酶联免疫吸附法(ELISA法)测定其在单纯药物及脂多糖(LPS)1 mg?L-1诱导情况下小鼠巨噬细胞肿瘤坏死因子-α(TNF-α)、白介素-10(IL-10)的含量.结果:芦荟及其复方提取物具有促进巨噬细胞释放NO的作用,并与药物浓度成正相关(P＜0.01);芦荟有促进TNF-α分泌(P＜0.01)和抑制IL-10分泌(P＜0.05)的作用,其复方在LPS诱导下则对TNF-α和IL-10都有抑制作用(P＜0.01).结论:芦荟在250～7.8 g?L-1下对臣噬细胞无细胞毒性,其复方则有一定毒性;芦荟及其复方对小鼠巨噬细胞炎症因子的释放有一定影响.     

少棘蜈蚣抗菌肽粗品对小鼠巨噬细胞的体外激活作用

分别用62.5、125、250、500和1000μg/ml的少棘蜈蚣抗菌肽粗品,体外诱导对小鼠腹腔巨噬细胞(PMΦ)及肺泡巨噬细胞(AMΦ)48 h,测定巨噬细胞内酸性磷酸酶(ACP)和乳酸脱氢酶(LDH)的活力,并进一步观察巨噬细胞摄取中性红的能力。结果显示,少棘蜈蚣水提取物使正常小鼠PMΦ、AMΦ内ACP、LDH活力显著升高,并能显著增强小鼠PMΦ、AMΦ的吞噬能力,表明少棘蜈蚣水提取物对腹腔及肺泡巨噬细胞具有激活作用。     

短葶山麦冬多糖对小鼠腹腔巨噬细胞功能的影响

目的观察短葶山麦冬多糖(LMP)对小鼠腹腔巨噬细胞免疫功能的影响。方法分离纯化小鼠腹腔巨噬细胞用不同质量浓度LMP(62.5、125、250、500μg/m L)共同培养,检测巨噬细胞吞噬中性红试验,趋化作用和分泌TNF-α、IL-6的功能。结果不同质量浓度的LMP能显著增强巨噬细胞吞噬能力和趋化作用,促进TNF-α、IL-6释放。结论 LMP能增强小鼠腹腔巨噬细胞免疫功能。     

当归醇沉物及其中性组份体外对巨噬细胞分泌TNF-α和IL-1的影响

目的观察当归醇沉物(ESA)及其中性组分(ESA-1)体外对小鼠腹腔巨噬细胞分泌TNF-α、IL-1的影响.方法L929细胞株细胞毒法测TNF-α;L929细胞增殖法测IL-1.结果ESA、ESA-1与小鼠腹腔巨噬细胞(Mφ)共同培养可显著增强其TNF-α、IL-1的分泌.在5～20μg/mL浓度范围,ESA的这种作用呈剂量依赖性;ESA-1虽有明显的作用,但未表现出剂量依赖.结论当归醇沉物及其中性组分体外可增强小鼠腹腔巨噬细胞分泌TNF-α、IL-1.     

九节龙皂苷Ⅰ对环磷酰胺模型小鼠免疫功能的影响

 目的研究九节龙皂苷Ⅰ(AR-Ⅰ)对环磷酰胺(CTX)模型小鼠免疫功能的影响。方法以CTX(100 mg·kg^-1)制造小鼠免疫低下模型,同时灌胃给AR-Ⅰ8 d,测定小鼠巨噬细胞吞噬率、血清50%溶血值、自然杀伤细胞和淋巴因子激活的杀伤细胞活性以及小鼠血清IL-2,TNF-α和TFN-γ含量,并观察二硝基氟苯诱导的小鼠迟发性超敏反应。结果AR-Ⅰ可增强CTX所致免疫低下小鼠巨噬细胞吞噬功能和迟发性超敏反应,促进血清溶血素形成,提高自然杀伤细胞和淋巴因子激活的杀伤细胞的杀伤活性,增加血清细胞因子IL-2,TNF-α和IFN-γ的含量。结论AR-Ⅰ能明显改善CTX抑制的小鼠免疫功能,这可能是其抗肿瘤活性的作用机制之一。     

骨碎补醇提物对免疫抑制小鼠免疫功能的影响

目的研究骨碎补乙醇提取物(Rhizoma Drynaria ethanol extract,RDEE)对免疫抑制小鼠免疫功能的影响及其可能的作用机制。方法腹腔注射环磷酰胺(CTX)建立免疫抑制小鼠模型,给药20天后,取血,处死小鼠,采集肝脏、脾、胸腺。检测小鼠胸腺、脾脏指数;巨噬细胞吞噬功能;外周血白细胞(WBC)计数、红细胞(RBC)、血红蛋白(HGB)、淋巴细胞百分率(LYNPH%)和IL-2、TNF-α、IFN-γ水平;比色法测定脾脏乳酸脱氢酶(LDH)和酸性磷酸酶(ACP)活性。结果与模型组比较,RDEE各剂量组的胸腺指数、吞噬指数α、WBC、RBC、HGB、LYNPH%、LDH和ACP活性明显提高;中、高剂量组的脾脏指数、IL-2、IFN-γ含量明显提高;高剂量组的廓清指数K和TNF-α含量明显提高。结论 RDEE对环磷酰胺导致的免疫抑制小鼠的免疫功能有调节作用,可能是通过保护吞噬细胞和体液因子的非特异性免疫功能而发挥作用。     

Modulatory effects of the acid polysaccharide fraction from one of anamorph of Cordyceps sinensis on Ana-1 cells

Ethnopharmacological relevance

Cordyceps sinensis has been used as a precious herbal medicine for thousands of years in China. Its polysaccharide fraction has been confirmed possessing immunomodulatory function and we have reported the acid polysaccharide fraction (APSF), from an anamorph of C. sinensis, has stimulating activity on macrophages. The mechanism still needs to be further elucidated.

Materials and methods

In order to investigate the effects of APSF on macrophage's phenotypes, Ana-1 mouse macrophages were polarized to M2 phenotype by culturing the cells with culture supernatant of H22 cells. M2 phenotype was determined by measuring the expression of TNF-α and checking cell surface markers mannose receptor (MR) and scavenger receptor (SR). After cultured with H22 supernatant for 72 h, the TNF-α level of Ana-1 cells was decreased while the SR and MR expressions were up-regulated, suggesting that Ana-1 cells were polarized towards M2 macrophages. Then the effects of APSF on M2 macrophages were investigated by measuring mRNA levels of TNF-α, inducible nitric oxide synthase (iNOS), IL-12 and IL-10. Nuclear NF-κB was detected by Western blotting.

Results

APSF treatment increased the expressions of TNF-α, IL-12 and iNOS, and reduced the expression of IL-10 of Ana-1 cells. Besides, the expressions of SR and MR were down-regulated by APSF. And the result of Western blotting showed NF-κB level was decreased in M2 macrophages and up-regulated after APSF treatment.

Conclusions

APSF may convert M2 macrophages to M1 phenotype by activating NF-κB pathway.     

蝉拟青霉多糖对老年大鼠免疫功能的调节作用

目的:研究蝉拟青霉水提物多糖对老龄大鼠免疫功能的调节作用。方法:以生理盐水处理的10~12周龄SD青年大鼠、18月龄SD老龄大鼠为对照组,实验组以不同剂量蝉拟青霉水提物多糖(50,100,200 mg.kg-1.d^-1)处理老龄大鼠3周,观察对照组、实验组大鼠腹腔、肺泡巨噬细胞对葡萄球菌的吞噬作用;以MTT法检测大鼠脾组织细胞分别在刀豆蛋白A(ConA),磷酸脂多糖(LPS)诱导下的淋巴细胞增殖活性;同时检测大鼠脾组织细胞内酸性磷酸酶(ACP),乳酸脱氢酶(LDH),精氨酸酶(ARG)等酶活力,并于透射电镜下观察大鼠脾组织超微结构。结果:与青年组比较,老龄组大鼠巨噬细胞的噬菌能力及脾组织淋巴细胞的增殖反应显著降低(P<0.01),脾细胞内ACP,LDH,ARG等酶活力也显著低下(P<0.01)。各种剂量蝉拟青霉多糖作用3周后老龄大鼠的这些指标均有改善,脾组织超微结构显示线粒体、内质网结构明显且数量增多。结论:蝉拟青霉水提物多糖能提高老龄大鼠腹腔、肺泡巨噬细胞吞噬功能和脾细胞免疫功能及增殖反应能力,使老龄大鼠低下的免疫功能得以改善,可能具有一定的抗衰老作用。     

EseroS-GS对脂多糖诱导小鼠腹腔巨噬细胞炎性因子生成的调节

 目的探讨EseroS-GS对脂多糖(LPS)诱导小鼠腹腔巨噬细胞核因子-kB(NF-kB)活化及炎性细胞因子肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)、白细胞介素-6(IL-6)基因表达的调节,为Eseros-GS的临床运用提供理论依据。方法分别用LPS或Eseros-GS+LPS处理体外培养的小鼠巨噬细胞,采用蛋白质印迹分析和电泳迁移率改变分析法(EMSA)检测细胞中NF-kB活性,用逆转录-聚合酶链反应(RT-PCR)和酶联免疫吸附法(ELISA)检测细胞中TNF-α,IL-1β、IL-6 mRNA和蛋白的表达。结果LPS组NF-kB活性和TNF-α、IL-1β、IL-6含量在刺激后2～12h明显高于正常对照组(P<0.01),而EseroS-GS+LPS组NF-kB活性和TNF-α、IL-1β、IL-6含量均显著低于LPS组(P<0.01)。结论结果提示,LPS可诱导巨噬细胞NF-kB活化,导致TNF-α、IL-1β、IL-6基因表达增强,而EseroS-GS能抑制NF-kB活化而调节TNF-α、IL-1β、IL-6基因的表达。     

Polypeptides of Aloe barbadensis miller

Major polypeptide species of proteins have been identified and analysed by sodium dodecyl sulphate polyacrylamide gel electrophoresis in fresh extracts of the whole leaf, leaf gel, root and stalk portions of Aloe barbadensis Miller plants of immature, young and mature ages. Extracts of the fresh Aloe plant portions were prepared by dissection, tissue disruption, differential centrifugation and gel filtration methods. Extracted plant portions analysed by separation electrophoresis were also assayed by biochemical and immunological techniques for the presence of lectin associated reactions, i.e. agglutination or mitogenicity. Results of the separation electrophoresis analysis of extracts prepared from fresh whole leaves and leaf gel of mature Aloe barbadensis Miller plants revealed 23 identifiable different polypeptides. Molecular weights of these polypeptides, calculated from sets of molecular weight reference standards, ranged from 70 000 for the largest to 3000 for the smallest. Electrophoresis profiles of commercially processed and freshly processed Aloe barbadensis Miller and Aloe saponaria Haw leaf gel extracts revealed similar patterns for major peptides. Treatment of mature whole Aloe leaf extracts with acidic and alkaline conditions revealed distinct changes in pH stability of ten peptides. Comparisons of separation electrophoresis profiles of fresh extracts of Aloe whole leaves and of leaf gel portions revealed marked differences in both molecular weights and concentrations of peptides found in extracts from mature, young or immature plants. This report is the first to describe the nature and types of polypeptides detected in extracts of whole leaf, leaf gel, stalk and root portions of immature, young and mature Aloe plants. Accordingly, information in this report may be of considerable value in helping to identify and characterize Aloe substances present during processing in extracts and in products.     

Effect of Aloe barbadensis Miller Juice on oxidative stress biomarkers in aerobic cells using Artemia franciscana as a model

This study reports on the induction of oxidative stress in aerobic cell systems by Aloe barbadensis Miller (Aloe vera) juice using the salt water crustacean Artemia franciscana as a model. A consistent pattern was observed in which Artemia franciscana nauplii responded to Aloe vera juice exposure with a decrease in the overall activity of redox related enzymes. Exposure of Artemia franciscana to sub‐lethal levels of Aloe vera juice resulted in a decreased activity of thioredoxin reductase, glutathione reductase and glutathione peroxidase by 34% (66% enzymatic activity), 79% (21% enzymatic activity) and 90% (10% enzymatic activity), respectively. Similarly apparent was the trend whereby the co‐exposure of the nauplii to vitamin E counteracted this effect. For each of the biomarker enzymes tested, vitamin E co‐exposure resulted in enzyme activities closer to the control value (78%, 56% and 32% of control enzymatic activities for thioredoxin reductase, glutathione reductase and glutathione peroxidase activity, respectively). These results indicate that exposure to sub‐lethal doses of Aloe vera juice induces alterations in the cellular redox status of Artemia franciscana and that the addition of vitamin E helps the Artemia franciscana nauplii to overcome/block the juice induced oxidative stress. Copyright © 2009 John Wiley & Sons, Ltd.     

芦荟粗多糖对人表皮细胞体外分泌细胞因子的影响

目的:观察芦荟粗多糖对体外培养表皮细胞分泌生长因子(EGF,TGF-α,TGF-β₁)、白细胞介素(IL-1β,IL-6,IL-8)及肿瘤坏死因子(TNF)的影响。方法:用不同剂量(75,150,300,600,1 200 mg.L^-1)芦荟粗多糖作用于表皮细胞,采用ELISA和放射免疫法(RIA)测定细胞培养上清液中EGF,TGF-α,TGF-β₁,IL-1β,IL-6,IL-8,TNF的水平,并以等体积的细胞培养液处理为对照组。结果:经芦荟粗多糖作用后,表皮细胞培养液中EGF,TGF-α,TGF-β₁,IL-1β,IL-6,IL-8,TNF水平呈不同程度升高,其中EGF,TGF-α,IL-1β,IL-6,IL-8水平与对照组比较其差异具有显著性意义(P<0.05,P<0.01),且呈一定的量效关系。结论:芦荟粗多糖对表皮细胞分泌EGF,TGF-α,IL-1β,IL-6,IL-8具有促进作用。     

大黄附子汤对BALB/c小鼠腹腔巨噬细胞功能的影响

目的:研究大黄附子汤对BALB/c小鼠腹腔巨噬细胞功能的影响.方法:制备小鼠腹腔巨噬细胞,纯化后以5×105/mL细胞接种于96孔培养板或6孔板中,分为:细胞对照组、脂多糖(lipopolysaccharide,LPS)模型组(10 mg·L-1)、大黄附子汤组(终质量浓度为25,50,100,200,400,800 mg·L-1),在细胞贴壁过夜后,正常组给予不含血清的培养液,其余各组给予LPS(10 mg·L-1),大黄附子汤组同时给予不同浓度药物,48 h后测细胞存活率,ELISA法测肿瘤坏死因子-α(TNF-α),白介素-6(IL-6)水平,黄嘌呤氧化酶法(羟氨法)测细胞上清液中超氧化物歧化酶(SOD)活力,硫代巴比妥酸法测丙二醛(MDA)含量,Griess法测一氧化氮(NO)水平.结果:在25 ～ 400 mg·L-内,大黄附子汤对正常细胞代谢MTT活力无显著影响;剂量达50 mg·L-1即能抑制LPS诱导NO的生成(P＜0.01),100 mg·L-1时能降低MDA含量(P＜0.01),增强SOD酶活力(P＜0.05),并能显著抑制LPS诱导TNF-α,IL-6等细胞因子的合成(P＜0.01).结论:大黄附子汤能有效调节BALB/c小鼠腹腔巨噬细胞免疫及抗氧化功能,改善LPS对BALB/c小鼠腹腔巨噬细胞的诱导作用.     

血小板激活因子受体拮抗剂SY0916抑制巨噬细胞血管生成作用的研究

 目的 考察血小板激活因子（platelet activating factor,PAF受体拮抗剂SY0916对巨噬细胞释放血管内皮生成相关因子的影响,探讨SY0916抗血管生成的相关分子机制。方法 采用Boyden Chamber法检测小鼠腹腔巨噬细胞的趋化反应;放射性免疫分析法检测巨噬细胞U937上清中白细胞介素（IL-1β含量;L929生物测定法分析肿瘤坏死因子（TNF-α分泌;ELISA法测定血管内皮细胞生长因子（VEGF含量;明胶酶谱法检测间质金属蛋白酶（MMP-9活性;Western Blot法检测MMP-9蛋白表达;RT-PCR法观察IL-1β、TNF-α、VEGF mRNA的表达;凝胶电泳迁移率变更法（EMSA分析核转录因子（NF-κB活性。结果 SY0916对PAF诱导的小鼠腹腔巨噬细胞趋化反应具有显著的抑制作用;SY0916可呈剂量依赖性地抑制PMA刺激人U937巨噬细胞引起的IL-1β、TNF-α、VEGF的生成以及IL-1β、TNF-α、VEGF的mRNA表达;SY0916能显著抑制U937细胞MMP-9的活性及蛋白表达;SY0916明显抑制NF-κB的活化。结论 PAF受体拮抗剂SY0916可能通过抑制巨噬细胞NF-κB的活性而下调促血管生成因子和MMP的表达发挥抗血管生成的作用。     

Dichroa febrifuga Lour. inhibits the production of IL-1β and IL-6 through blocking NF-κB, MAPK and Akt activation in macrophages

Aim of the study

The roots of Dichroa febrifuga Lour. have been used as a traditional antimalarial drug and also used in the treatment of productive cough and unstable fever caused by infection in China and Korea. In this study, we evaluated the anti-inflammatory effect and underlying molecular mechanism of aqueous extract of Dichroa febrifuga (AEDF) in C57BL/6 mouse peritoneal macrophages.

Materials and methods

The effect of AEDF on proinflammatory cytokine (IL-1β and IL-6) production was analyzed by ELISA and real-time RT-PCR. The effects of AEDF on NF-κB/IκB-α/IKK were measured by reporter assay (in RAW 264.7 cells), EMSA, Western blotting and kinase assay. The effects of AEDF on Akt and MAPKs activity were assayed by Western blotting.

Results

AEDF inhibited the production of IL-1β and IL-6, NF-κB activation, IκB-α degradation, and IKK, Akt, ERK1/2 and JNK activities in LPS-stimulated mouse peritoneal macrophages.

Conclusions

These results suggest that AEDF inhibits proinflammatory cytokine (IL-1β and IL-6) production in LPS-stimulated mouse peritoneal macrophages, and that these effects are mediated by the inhibition of the activity of IKK/IκB/NF-κB and the phosphorylation of Akt, ERK1/2, and JNK. Our results provide a molecular basis for understanding the inhibitory effects of Dichroa febrifuga roots on endotoxin-mediated inflammation.