黄芪提取物免疫调节活性的体外实验研究

目的：研究黄芪提取物（Astragalus membranaceus extract,AME)对人的外周血免疫细胞（PBIC）免疫功能的调节作用。方法：采用^3H-TdR掺入法分析AME对外周血单个核细胞（PBMC）的增殖活性及对外周血粘附单核细胞（PBAM）吞噬肿瘤细胞的影响；采用^51Cr释放法测定AME对杀伤性T细胞（CTL）杀伤肿瘤细胞的影响；用ELISA法和生物学法研究AME对外周血B细胞（PBBC）产生IgG及对PBAM产生细胞因子的影响；采用SDS－PAGE分析AME的蛋白组成。结果：AME能促进PBMC的增殖；提高CTL对肿瘤细胞的杀伤活性；增强PBAM对肿瘤细胞的吞噬和产生细胞因子的功能；促进了PBBC产生IgG;AME含有多种蛋白成分。结论：AME对人免疫功能有增强作用，提高了抗肿瘤免疫效应，可应用于临床调节免疫功能和治疗肿瘤等疾病。     

Immunomodulatory activities of Trichilia glabra leaf aqueous extracts

Human peripheral blood monocytes and polymorphonuclear leukocytes treated with leaf aqueous extracts of the Meliaceae tree Trichilia glabra showed a diminution in both their phagocytic and respiratory burst activities. A decrease in the haemolytic capacity of the human complement was also observed. The significance of the inhibitory effect observed over some components of the human immune system closely related with the inflammatory process is discussed. © 1998 John Wiley & Sons, Ltd.     

Immunomodulatory activities of Cedrela tubiflora leaf aqueous extracts

Human peripheral blood monocytes and polymorphonuclear leukocytes treated with leaf aqueous extracts of the Meliaceae tree Cedrela tubiflora showed a diminution in both their phagocytic and respiratory burst activities. Besides, the extract inhibited the proliferation of Concanavalin A stimulated lymphocytes. A decrease in the hemolytic capacity of the human complement was also observed. The significance of the inhibitory effect observed over some components of the human immune system closely related with the inflammatory process is discussed.     

Neem leaf preparation induces apoptosis of tumor cells by releasing cytotoxic cytokines from human peripheral blood mononuclear cells

A neem leaf preparation (NLP) was investigated for its role in the induction of tumor cell apoptosis to elucidate the mechanism of NLP mediated immunoprophylaxis in tumor growth restriction. As NLP did not induce direct apoptosis of human tumor cell lines KB, MCF7 and K562, it was used instead to stimulate human peripheral blood mononuclear cells (PBMC) for 72 h. The PBMC derived culture supernatant (NLP-CS) was observed to induce the restriction of tumor cell proliferation as well as apoptosis. An enzyme linked immunosorbant assay revealed the presence of cytotoxic cytokines, IFN-gamma and TNF-alpha, in the NLP-CS. The inhibition of secretion of IFN-gamma and TNF-alpha in NLP-CS caused a significant decrease in tumor cell apoptosis. Furthermore, stimulation of these tumor cells with NLP-CS resulted in upregulation of the caspase 3 and downregulation of the Bcl 2 and cyclin D1. These observations suggested that NLP could induce tumor cellular apoptosis by releasing cytotoxic cytokines from human PBMC.     

枸杞多糖对人外周血巨噬细胞抗肿瘤作用的影响

目的研究枸杞多糖(LBP)对正常人外周血巨噬细胞免疫调节功能的影响。方法无菌分离人外周单核细胞,加入PHA和不同质量浓度的LBP,MTT法检测巨噬细胞对肝癌细胞株Bel-7402生长的影响;ELISA法检测LBP作用后巨噬细胞培养上清液中肿瘤坏死因子-α(TNF-α)水平。结果用LBP处理后的巨噬细胞对肿瘤细胞的杀伤功能增强,80 mg/L1、00 mg/L LBP刺激后,巨噬细胞的吞噬作用明显增强(P均<0.01)。80 mg/L、100 mg/L LBP组肿瘤细胞与巨噬细胞培养上清液中TNF-α的量明显增高(P均<0.01),并证实了该作用可能与LBP促进巨噬细胞分泌TNF-α有关。结论LBP能增强人外周血巨噬细胞的免疫功能,发挥抗肿瘤作用。     

超抗原SEA对人外周血单个核细胞杀伤乳腺癌细胞效力的诱导作用

[目的]研究超抗原葡萄球菌肠毒素A(SEA)对外周血单个核细胞(PBMC)杀伤人乳腺癌细胞(MCF-7)效力的增强作用.[方法]淋巴细胞分离液梯度离心法从人外周血中分离出PBMC,在其中加入终浓度分别为0.01、0.1、1、10mg/L的SEA,与MCF-7在96孔板中共同培养.24、48、72h后用酶标仪检测吸光度值.计算杀伤率.[结果]SEA的促杀伤作用随着时间的增加而增强.SEA与PBMC相互作用72 h后,SEA浓度10 mg/L时,可达到最大杀伤效率.[结论]SEA能够提高PBMC对癌细胞的杀伤作用.     

灵芝提取物对人外周血细胞免疫调节作用的研究

 目的 研究灵芝子实体的水提取物、醇提取物单独及与诱导物一起使用对人外周血吞噬细胞和淋巴细胞的免疫调解作用。方法 以吞噬细胞化学发光、淋巴细胞化学发光和淋巴细胞增殖的方法,观察灵芝提取物的免疫调节作用。结果 单独灵芝提取物不能引起细胞免疫反应,与诱导物一起使用后,低浓度的灵芝提取物对细胞免疫功能有激活作用,高浓度则有免疫抑制作用。结论 与诱导物一起使用后,灵芝水提取物和醇提取物对人外周血吞噬细胞和淋巴细胞均具有双向调节作用。     

枸杞多糖对淋巴细胞功能的影响

目的研究枸杞多糖(LBP)对正常人外周血淋巴细胞免疫调节功能的影响。方法无菌分离人外周血淋巴细胞,加入植物凝集素(PHA)和不同质量浓度的LBP,MTT法检测其对淋巴细胞增殖的影响;ELISA法检测LBP作用后淋巴细胞培养上清液中肿瘤坏死因子-α(TNF-α)的表达情况。结果 50~100 mg/L LBP能明显增强PHA活化的人外周血淋巴细胞的增殖;LBP能促进淋巴细胞产生TNF-α,并呈现浓度依赖性。结论 LBP能增强人外周血淋巴细胞的免疫功能,发挥抗肿瘤作用。     

Immunomodulatory effect of Antrodia camphorata mycelia and culture filtrate

Aim of the study

Antrodia camphorata, a precious folkloric medicinal mushroom, has been used to treat tumorigenic diseases in Taiwan. This study was to investigate the innate immunity augmentation effects of different fractions prepared from hot water extracts of submerged cultured Antrodia camphorata (AC).

Materials and methods

The cytokine induction potency of AC fraction in diluted peripheral blood culture was measured by ELISA. The effects of AC fraction on phagocytic activity and CD11b expression were measured by the ingestion of FITC-labeled Escherichia coli and by labeling with PE-labeled CD11b monoclonal antibody, respectively, using flow cytometry. The molecular mass of hot water-soluble polysaccharides and content of adenosine in AC fraction were determined by gel permeation chromatography (GPC) and HPLC, respectively.

Results

The mycelia fraction, Fr. M II, and culture filtrate fractions, Fr. E II and Fr. E III, showed the strongest TNF-α and IL-6 induction effect as a function of their concentration. These fractions (20 μg/ml) also showed marked activity in enhancing phagocytosis in human polymorphonuclear neutrophils (PMN) and monocytes. In parallel, the expression of CD11b, an early marker of PMN activation, was also up-regulated dose-dependently. Composition analysis suggested that immunomodulatory effect of mycelia is mainly attributed to the 10–20 kDa polysaccharides and adenosine.

Conclusions

These results provide evidences that Antrodia camphorata can modulate innate immunity and may serve as an adjuvant for tumor treatment.     

复圣散对血管内皮细胞活性及其与中性粒细胞粘附率的影响

探讨中性粒细胞与血管内皮细胞的粘附特性的影响因素，以及中药复圣散的干预作用。方法；以培养人脐静脉内皮细胞作为靶细胞，观察在高脂，高糖，肿瘤坏死因子及缺氧再氧化条件下，对培养的ＶＥＣ活性以及ＰＭＮ与ＶＥＣ相互粘附的影响。结果：高脂，高糖，ＴＮＦ－α和缺氧再氧化等均可抑制ＶＥＣ活性，并显著增强ＰＭＮ与ＶＥＣ的相互粘附作用，其粘附率正常对照组为１００％，高脂组则为１６７．７％，缺氧再氧化组为１５１．４％     

黄芪注射液对法洛四联症患儿根治术后免疫功能的影响

目的 观察黄芪注射液对法洛四联症(tetralogy of Fallot,TOF)患儿根治术后免疫功能的影响。方法 40例TOF术后患儿分为对照组(20例,给予常规治疗)和黄芪组(20例,在常规治疗的基础上加用黄芪注射液15ml静脉滴注,每天2次,共2周);并观察两组患儿免疫球蛋白(IgG和IgM)、补体(C3、C4)、淋巴细胞表型和细胞因子的变化。结果 应用黄芪注射液后,异常升高的IgG、IgM、G、C4、CD8a^+和CD19^+’于治疗后第1～2周开始下降,且于第3～4周基本恢复到正常组水平;而异常下降的IgA、CD3^+、CD4^+、CD4^+／CD8^+、CD3^+/LA-DR^+和CD3^+／CD16^+-CD56^+于治疗后1周开始逐渐升高,于治疗后2～3周恢复到正常组水平。血浆和外周血单核细胞体外产生的白细胞介素-6和肿瘤坏死因子-α于治疗后1周开始逐渐下降．于治疗后3～4周恢复到正常组水平,上述指标治疗前后差值治疗组均优于对照组(P<0．05或P<0．01)。结论 黄芪注射液可明显改善TOF患儿术后的免疫功能。     

补肾方对慢性乙型肝炎T细胞亚群及其功能的影响

目的观察补肾方对慢性乙型肝炎患者免疫功能状态的调节作用。方法收集HBeAg阳性ALT异常者30例,给予补肾方治疗,用药6个月,于用药前、用药3个月、6个月时分别检测下列指标。采用罗氏荧光定量PCR法检测HBVDNA,外周血单个核细胞(peripheral blood mononuclear cell,PBMC)加入HBeAg、HBcAg与植物血凝素(Phytohemagglutinin,PHA)培养48h,检测培养上清液中IL-10、IFN-γ,血液中CD4^+、CD8^+、CD8^+CD28^+、CD8^+CD28^-和CD28^+T细胞的比例及加入HBeAg、HBcAg与PHA培养48h后培养细胞中CD8^+CD28^+的比例。结果补肾方治疗有效组在治疗3个月后,PBMC培养中IFN-γ较治疗前明显上升,IL-10明显下降(P<0．05,P<0．01);CD8^+CD28^+T、CD28^+T、培养后CD8^+CD28^+T细胞较治疗前明显上升,CD8^+CD28^-T细胞明显下降(P<0．05或P<0．01)。结论补肾方能明显增强Th1型细胞因子的表达,降低Th2型细胞因子的表达,促进CD8^+(Cytotoxic T lymphocyte,CTL)的表达,这可能是使HBVDNA复制得到抑制的机制之一.     

黄芪多糖对有核细胞分泌造血细胞因子的影响

目的 研究注射用黄芪多糖对分离的人外周血单个核细胞(PBMC)分泌造血细胞因子的影响，以支持黄芪多糖的造血活性。方法 取正常人浓缩白细胞，分离出人PBMC单细胞悬液，进行细胞培养，采用酶联免疫吸附方法(EUSA)，测定上清液中的粒细胞集落刺激因子(G—CSF)、粒细胞巨噬细胞集落刺激因子(CM—CSF)等造血细胞因子。结果 注射用黄芪多糖能刺激人PBMC产生大量的G—CSF和CM—CSF，表现出明显的量效关系和时效关系。结论 注射用黄芪多糖在体外能刺激人PBMC产生G—CSF和CM—CSF造血细胞因子，且呈明显的量效关系和时效关系，提示其有升高白细胞的作用，展示其临床应用前景。     

Mountain grown ginseng induces apoptosis in HL-60 cells and its mechanism have little relation with TNF-alpha production

The root of ginseng is one of the most popular natural tonics in Oriental countries. Ginseng grown in the wild, deep in the mountains, is known as Sansam (mountain grown ginseng, MGG). MGG belongs to Araliaceae and Panax. In this study, we investigated the effects of MGG on the cytotoxicity, induction of apoptosis and the putative pathways of its actions in human promyelocytic leukemia cells, HL-60. Using apoptosis analysis, we found that MGG is a potent inducer of apoptosis, but it has less effect on human peripheral blood mononuclear cells. Caspase-3 activation and subsequent apoptotic cell death in MGG-treated cells were partially blocked by the caspase-3 inhibitor, Z-DEVD-FMK. MGG also inhibited the caspase-8 activity. To determine whether MGG-induced apoptosis is involved in tumor necrosis factor-alpha (TNF-alpha) secretion, TNF-alpha secretion was quantified by enzyme-linked immunosorbent assay (ELISA) method. Unexpectedly, MGG significantly decreased the TNF-alpha secretion compared to the control. These results suggest that MGG-induced cytotoxicity have little relation with the secretion of TNF-alpha in HL-60 cells. Furthermore, MGG with rIFN-gamma synergistically increased nitric oxide (NO) production in mouse peritoneal macrophages. Taken together, our data indicate that MGG is a potent inducer of apoptosis on HL-60 cells and these abilities could be used clinically for the treatment of cancer.     

慢性ITP外周血单个核细胞Th1/Th2亚群反应状态及生血灵干预研究

目的探讨辅助性T淋巴细胞亚群Th1、Th2在慢性特发性血小板减少性紫癜(ITP)发病中的作用,观察生血灵对慢性ITP Th1、Th2亚群功能状态的影响。方法以酶联免疫吸附法(ELISA法)检测慢性ITP患者外周血单个核细胞(PBMC)培养上清中Th1型细胞因子IL-2和Th2型细胞因子IL-10的水平。结果慢性ITP患者PBMC培养上清中IL-2和IL-10的含量均较正常对照组明显减低(P均<0.001),生血灵对其具有明显提升作用(P<0.01)。结论慢性ITP患者Th亚群免疫功能低下。生血灵治疗慢性ITP能够增强Th亚群免疫功能,恢复其平衡。     

莲必治注射液(穿心莲内酯)对免疫功能的调节作用

目的 :研究穿心莲内酯对免疫功能的影响 ,探索其发挥治疗作用的免疫机制。方法 :采用生物活性法和ELISA法检测用有效成分为穿心莲内酯的莲必治注射液处理人外周血单个核细胞上清中的IFN-α ,IFN-γ ,TNF-α,IL-8含量 ;用单核巨噬细胞吞噬鸡红细胞来研究其促吞噬功能及用LDH释放法检测其对NK细胞杀伤活性的影响。结果 :和空白对照组比 ,莲必治注射液作用外周血单个核细胞能显著诱导IFN-α ,IFN γ ,TNF-α产生 (P<0.05) ,但对IL-8诱生无明显影响 ,甚至轻度抑制。莲必治注射液能提高 豚鼠腹腔单核巨噬细胞对鸡红细胞的吞噬百分率 (P<0.05)及提高人外周血中NK细胞杀伤K562 细胞的活性 (P<0.05)。结论 :穿心莲内酯是一种具有调节机体非特异免疫功能的免疫刺激剂 ,通过对NK ,MΦ及细胞因子分泌的影响而发挥免疫调节作用。     

黑木耳多糖对外周血单个核细胞增殖和细胞因子表达的影响

目的观察黑木耳多糖(APP)对外周血单个核细胞(PBMCs)增殖和细胞因子表达的影响.方法采用水提、乙醇沉淀、DEAE-纤维素柱层析分离纯化APP,并作用于体外培养的PBMCs,以Cell Counting Kit-8(CCK-8)试剂检测细胞增殖;酶联免疫法(ELISA)测定PBMCs培养上清液中TNF-α和IFN-γ的浓度.结果从黑木耳中分离的3个多糖组分在100～800 μg/ml剂量范围内对PBMCs均有不同程度的促进增殖作用;其中组分AAP2能显著增加PBMCs细胞培养上清液中TNF-α和IFN-γ的浓度(P＜0.05或P＜0.01),并显示剂量依赖效应.结论APP具有免疫促进作用,并能通过细胞因子TNF-α和IFN-γ发挥免疫调节功能.     

丹参酮对原发性高血压病人外周血单核细胞激活的抑制作用

 目的研究丹参酮对原发性高血压(EH)病人病程早期外周血单核细胞(PBMC)激活的作用。方法选择病程<5年且无心、脑、肾、血管等靶器官损害的EH病人30例,并设正常对照,通过体外细胞培养,血管紧张素Ⅱ(AngⅡ)刺激以及丹参酮干预,采用显微镜-计算机下细胞计数方法及双抗体夹心酶联免疫吸附实验(ELISA)和逆转录-聚合酶链反应(RT-PCR)等技术,检测PBMC的黏附活性及其分泌和表达炎性细胞因子的水平。结果EH病人PBMC与内皮细胞的黏附数及其分泌和表达肿瘤坏死因子α(TNF-α)、白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)等炎性细胞因子的浓度和mRNA水平在基础状态下同对照组相当,经AngⅡ刺激后却显著高于对照组,丹参酮干预可使2组PBMC的黏附及分泌活性降至同一水平。结论EH病人病程早期PBMC处于预激活状态,丹参酮能抑制预激活的PBMC进一步激活。     

丁公藤注射液雾化吸入对大鼠呼吸道和全身免疫功能的影响

目的：研究丁公藤注射液雾化吸入对大鼠呼吸道局部和全身性免疫功能的影响。方法：将体重２００～２５０ｇ雄性Ｗｉｓｔａｒ大鼠２０只，随机分为２组。丁公藤组吸入雾化丁公藤注射液，对照组吸入雾化生理盐水。每天１次，连续给药１４天后活杀，检测免疫指标。结果：丁公藤注射液能显著提高呼吸道Ｔ淋巴细胞比例和肺泡巨噬细胞吞噬率（Ｐ＜００５，Ｐ＜００１）；使外周血Ｔ淋巴细胞数和脾脏特异性抗体形成细胞数明显增加（Ｐ＜００１，Ｐ＜００５）；血液Ｔ淋巴细胞释放淋巴因子功能和中性白细胞吞噬功能也有提高倾向。结论：从呼吸道吸入雾化丁公藤注射液不仅能兴奋呼吸道局部免疫，而且可以兴奋全身性免疫功能     

多棘海盘车多糖对正常小鼠免疫功能的影响

 目的 研究多棘海盘车多糖（Asterias amurensis polysaccharides, AAP）对正常小鼠免疫功能的影响。方法 昆明种小鼠灌胃给予低、中、高剂量AAP（50, 100, 200 mg·kg-1）,连续14 d后,处死小鼠,测定免疫器官指数来考察AAP对小鼠免疫器官的影响;测定白细胞数目、吞噬活性及血清溶菌酶含量来考察AAP对小鼠非特异性免疫的影响;测定肝脏细胞中谷胱甘肽含量来考察AAP对小鼠肝脏解毒功能的影响;采用刀豆球蛋白A（ConA）诱导小鼠淋巴细胞增殖实验考察AAP对小鼠细胞免疫的影响。结果 AAP能显著提高小鼠的免疫器官指数、血液中中性粒细胞吞噬能力、血清溶菌酶含量、肝脏细胞中谷胱甘肽含量及淋巴细胞增殖能力,但对小鼠血液中白细胞数目无显著影响。结论 AAP对正常小鼠的非特异性免疫和细胞免疫有显著的增强作用。