In vitro and in vivo effects of isolated fractions of Brazilian propolis on caries development

Recently, two chemically different types of Brazilian propolis (type-3 and -12) were shown to have cariostatic properties. This study aimed to evaluate the influence of their isolated fractions on mutans streptococci viability, glucosyltransferases (GTFs) activity and caries development in rats. The ethanolic extracts of propolis (EEPs) were serially fractionated into hexane (H-fr), chloroform, ethyl acetate, and ethanol. The ability of the four fractions and EEP to inhibit Streptococcus mutans and Streptococcus sobrinus growth and adherence to a glass surface was examined. The effect on GTFs B and C activity was also determined. For the caries study, 60 Wistar rats infected with Streptococcus sobrinus were treated topically twice daily as follows: (1) EEP type-3, (2) H-fr type-3, (3) EEP type-12, (4) H-fr type-12, and (5) control. In general, the H-fr from both types of propolis showed the highest antibacterial activity and GTFs inhibition. Furthermore, the EEP and H-fr type-3 and -12 were equally effective in reducing dental caries in rats. The data suggest that the putative cariostatic compounds of propolis type-3 and -12 are mostly non-polar; and H-fr should be the fraction of choice for identifying further potentially novel anti-caries agents.     

刺果紫玉盘素J的体内外抗肿瘤作用

目的观察刺果紫玉盘素J(calamistrin J,Cal-J)的体内外抗肿瘤活性及其选择性。方法MTT法观察不同质量浓度的Cal-J对人非小细胞肺癌NCI-H460细胞、人肝癌HepG2细胞、人宫颈癌HeLa细胞、人结肠癌Lovo细胞、肉瘤S180细胞5种肿瘤细胞系和正常人胚肺成纤维细胞(HELF)增殖的影响,Hoechst33258染色后在荧光显微镜下观察Lovo细胞凋亡的形态变化。在S180荷瘤小鼠进行整体抑瘤实验,计算Cal-J的抑瘤率。结果Cal-J对NCI-H460、HepG2、HeLa、HELF、S180、Lovo等6种细胞的增殖均有不同程度的抑制作用,其IC50依次为>100、(91±4)、(60.5±2.4)、(39.3±1.7)、(38±4)、(25±4)μg/mL。其中对Lovo细胞株抑制作用最强,且呈剂量、时间双重依赖性。Cal-J作用48h后,荧光显微镜下可见Lovo细胞核固缩,染色质凝聚等典型凋亡形态学特征。Cal-J对荷瘤小鼠S180移植瘤的生长有明显的抑制作用,Cal-J高、中、低(90、30、10mg/kg)剂量组的抑瘤率分别为51%、30%、17%,作用呈剂量依赖性。结论Cal-J对体外培养的多种人类肿瘤细胞和人胚肺成纤维细胞具有不同程度的细胞毒性作用,其中对人结肠癌Lovo细胞作用最强,并具有诱导细胞凋亡的作用。Cal-J还可抑制荷瘤小鼠S180移植瘤的生长。     

In vivo study of propolis supplementation effects on antioxidative status and red blood cells

In vivo study has been conducted on 47 healthy women and men in order to investigate whether daily intake of powdered propolis extract during 30 days has any influence on the following blood parameters: activity of superoxide dismutase, glutathione peroxidase and catalase, concentration of plasma malondialdehyde, total cholesterol, low- and high-density lipoprotein cholesterol, triglycerides, glucose, uric acid, ferritin and transferrin, together with routine red blood cell parameters.

The effect of daily propolis intake seems to be time and gender related. For the men test group after the initial 15 days of propolis treatment, 23.2% (p = 0.005) decrease in concentration of malondialdehyde was observed. After 30 days of treatment, statistically significant (p = 0.010) 20.9% increase in superoxide dismutase activity and change in some of the red blood cell parameters were detected. For the women test group, the propolis treatment did not induce a change in any of the measured parameters.     

Antitumor effects of Pinus massoniana bark extract in murine sarcoma S180 both in vitro and in vivo

Pinus massoniana bark extract (PMBE) is a mixture of flavonoids, and showed a capability of inducing cell apoptosis; however, its properties have not yet been fully investigated. This paper evaluates the antitumor effects of PMBE in murine sarcoma S180 both in vitro and in vivo. In vitro, the growth inhibition of S180 cells was concentration dependent on PMBE as shown by the CCK-8 assay. The AO/EB staining and flow cytometry assay showed that PMBE induced S180 cell apoptosis. Cell cycle analysis revealed that the cells in the S phase were decreased by treatment with PMBE. In vivo, the treatment of 100, 200, and 300 mg/kg PMBE reduced the tumor weight and volume of S180-bearing NIH mice by 9%-67% and 13%-68%, respectively. Peripheral leukocyte count and lymphoproliferation were increased significantly after treatment with PMBE. Our results suggest that PMBE inhibits the tumor cell growth by inducing cell apoptosis and improving lymphoproliferation.     

Immunomodulation by water-soluble derivative of propolis: a factor of antitumor reactivity

The antimetastatic efficacy of a water-soluble derivative of propolis (WSDP) was studied. Tumor was a transplantable mammary carcinoma of CBA mouse. Metastases in the lung were generated by 2 x 10(5) viable tumor cells i.v. WSDP was given intraperitoneally at doses of 50 or 150 mg/kg before or after tumor cell inoculation. Therapies reduced the number of metastases in the lung and tumor growth was suppressed significantly by WSDP. It is likely that antimetastatic activity of the WSDP is mainly mediated by immunomodulatory activity. Changes in several immunological parameters such as production of lymphocyte activating factor by peritoneal macrophages and the efficacy of those macrophages to kill tumor cell in vitro, responses of lymphocytes to mitogen, and weight and cellularity of spleen, respectively, correlated well with antimetastatic properties of the WSDP. Based on results we postulate that the antimetastatic activity of propolis includes a pronounced immunomodulatory activity mainly toward augmentation of nonspecific antitumor resistance in mice via macrophage activation.     

花椒体内外抗肿瘤作用及其机制的初步研究

目的 研究花椒挥发油体内外抗小鼠H22肝癌细胞作用及其免疫学机制.方法 花椒挥发油按不同时间(24,48,72h)及不同浓度(0.5～8 ms/ml)分别处理H22细胞,MTS法分析细胞的增殖速度,用流式细胞仪测定细胞凋亡及细胞周期.建立H22荷瘤小鼠模型,观察小鼠肿瘤的生长情况,计算肿瘤生长抑制率;ELISA法检测花椒挥发油处理后小鼠血清中IL-2和IL-12的活性.结果 4 mg/ml花椒挥发油处理H22细胞72 h可显著地抑制H22细胞增殖,1 mg/ml花椒挥发油处理H22细胞72 h即可导致亚凋亡峰的出现,且G0/G1期细胞增多,S期和G2/M期细胞减少,花椒挥发油对小鼠实体瘤生长具有显著抑制作用,但不能提高荷瘤小鼠血清中IL-2和IL-12的水平.结论 花椒挥发油可抑制H22细胞增殖并激发细胞凋亡,但不能通过提高机体的免疫功能发挥抗肿瘤作用.     

卷丹百合体外对肺腺癌A_(549)细胞抑制作用的实验研究

目的:研究百合对肺腺癌细胞株A549增殖作用的影响及其相关机制。方法:体外培养肺腺癌细胞株A549,用不同浓度的百合水提物分别作用于对数期生长的A549细胞24、48和72h,MTT(甲基噻唑基四唑)比色法检测百合对A549细胞增殖的抑制率,碘化丙啶(PI)染色,荧光显微镜下观察细胞形态改变及凋亡情况。结果:MTT法检测结果显示,百合水提物在1~8mg/mL浓度范围内、24~72h时间范围内对A549的生长有明显抑制作用,且呈浓度和时间依赖关系(P<0.05);PI染色荧光显微镜观察显示百合作用48h后,细胞出现凋亡的形态学改变。结论:百合具有体外抑制肺腺癌细胞株A549增殖的作用,其可能是通过诱导肿瘤细胞凋亡而发挥抑制A549细胞增殖的作用。     

Anti-inflammatory properties of Ajuga bracteosa in vivo and in vitro study and their effects on mouse model of liver fibrosis

Ethnopharmacological relevance

The entire plant of Ajuga bracteosa Wall has been used to treat various inflammatory disorders, including hepatitis, in Taiwan.

Aim

This study evaluated the hepatoprotective ability of Ajuga bracteosa extract (ABE).

Materials and methods

We investigated the inhibitory action of a chloroform fraction of ABE (ABCE) on lipopolysaccharide (LPS)-stimulated RAW264.7 cells and Kupffer cells. Hepatic fibrosis was induced in mice through the administration of CCl₄ twice a week for 8 weeks. Mice in three CCl₄ groups were treated daily with water and ABE throughout the duration of the experiment.

Results

In LPS-stimulated RAW264.7 cells and Kupffer cells, ABCE inhibited the production of NO and/or TNF-α and also blocked the LPS-induced expression of NO synthase. ABCE inhibited the activation of NF-κB induced by LPS, associated with the abrogation of IκBα degradation, with a subsequent decrease in nuclear p65 and p50 protein levels. The phosphorylation of MAPKs in LPS-stimulated RAW264.7 cells was also suppressed using ABCE. In the in vivo study, ABE protected the liver from injury by reducing the activity of plasma aminotransferase, and by improving the histological architecture of the liver. RT-PCR analysis showed that ABE inhibited the hepatic mRNA expression of LPS binding protein, CD14, TNF-α, collagen(α1)(I), and α-smooth actin.

Conclusion

These results indicate that ABE alleviated CCl₄-induced liver fibrosis, and that this protection is probably due to the suppression of macrophage activation.     

甲基丙烯酰紫草素对肝癌的抑制作用研究

目的:研究甲基丙酰紫草素的抗癌作用,并初步探讨其作用机制.方法:MTT法、生长曲线检测甲基丙酰紫草素的体外抗肝癌作用,并计算半数抑制浓度(IC50)值;应用H22荷瘤小鼠模型进行体内抗瘤试验,绘制肿瘤生长曲线并计算抑瘤率;免疫组化方法检测肿瘤组织的凋亡相关蛋白NF-κB p65的表达.结果:甲基丙烯酰紫草素有显著的体外抗癌作用,且与剂量呈正相关,对SMMC-7721的IC50值分别为(21.16±0.96)μg/ml;小鼠移植瘤H22抗瘤试验结果显示,在0.38 mg/kg、0.75mg/kg和1.50 mg/kg的剂量下甲基丙烯酰紫草素的抑瘤率分别为22.98%、39.08%、47.13%.免疫组化结果表明随甲基丙烯酰紫草素浓度增加NF-κB p65蛋白表达下调.结论:甲基丙烯酰紫草素具有较强的体内外抗肝癌作用,其机制可能与抑制肿瘤细胞增殖、诱导细胞凋亡有关.     

An experimental study on antitumor activity of psoralen on mammary cancer cell line EMT6 in vitro and in vivo]

To investigate the antitumor activity of psoralen on mammary cancer cells of EMT6 line, the cytotoxity of the drug against EMT6 cells was tested by MTT assay; an experimental therapy was carried out in 15 nude mice that were subcutaneously injected with EMT6 cells; the morphological changes of tumor tissues were observed under electron microscope; and the DNA content was measured and analysed with a computerized MPV-microspectrophotometer. The result shows that psoralen has a strong antitumor effect on mammary cancer EMT6 cells both in vivo and in vitro, with an IC50 value of 2.23 micrograms/ml. The antitumor effect might be related to the relative decrease of cell DNA content and the degeneration and cavitation of cellular mitochondriums. Therefore psoralen may be an effective agent for the treatment of mammary cancer.     

Differentiation- and apoptosis-inducing activities by pentacyclic triterpenes on a mouse melanoma cell line

In a study to investigate the relationship between the chemical structure and the differentiation-inducing activity of pentacyclic triterpenes, several lupane, oleanane, and ursane triterpenes were prepared and their effects on B16 2F2 melanoma cell differentiation and growth were examined. Eleven lupane triterpenes used in this study acted on the melanoma cells as a melanogen, but no induction of melanogenesis of B16 2F2 cells by oleanane and ursane was detected. The differences at C-17 of the lupane series and acetylation of the OH group at C-3 did not markedly influence their activities. However, the ED(50) value for up-regulation of melanin biosynthesis was markedly decreased by the oxidation of the OH group at C-3 of lupeol (1). Betulinic acid (11), its methyl ester (12), lup-28-al-20(29)-ene-3beta-ol (9), and lup-28-al-20(29)-en-3-one (10) inhibited B16 2F2 cell proliferation by induction of apoptosis. These findings suggested that the carbonyl group at C-17 might be essential for the apoptotic effects of these compounds on B16 2F2 cells.     

沙苑子总黄酮和百草酸的体内外抗肿瘤作用

目的观察沙苑子总黄酮和百草酸的体内外的抗肿瘤作用。方法 110只接种S180肉瘤腹水昆明鼠随机分为模型组、环磷酰胺组、羟基喜树碱组、沙苑子总黄酮组和百草酸组,MTT法观察沙苑子总黄酮和熊果酸对S180肉瘤和人结肠癌细胞SW620的体外抑制作用,RT-PCR法测定百草酸对凋亡基因Bax、Bcl-2及P53的mRNA表达。结果沙苑子总黄酮和百草酸对S180肉瘤小鼠均有抑制作用,百草酸的抑制作用更加明显。两者对胸腺和脾脏均无明显抑制作用;沙苑子总黄酮对人结肠癌细胞SW620无明显抑制作用,而熊果酸却能明显的抑制该肿瘤细胞;熊果酸显著上调促凋亡基因P53和Bax的表达,下调抑凋亡基因Bcl-2的表达。结论百草酸在体内和体外均有抗肿瘤作用,沙苑子总黄酮的抗肿瘤作用尚需进一步研究。     

蜂胶体外抑菌活性的研究

[目的]探讨蜂胶对6种常见皮肤致病菌苏云杆菌、枯草芽孢杆菌、大肠杆菌、藤黄微球菌、白色葡萄球菌、奇异变形杆菌的抑菌效果。[方法]采用琼脂平板扩散法,6种受试菌株分别接种到不同的板上,每块板设置4种不同的浓度及对照组,12 h后观察蜂胶对各致病菌的抑菌圈大小,评价蜂胶对各致病菌的敏感程度及抑菌活性。[结果]不同浓度下,蜂胶对同一种受试菌株的敏感程度不同,抑菌圈大小不同;随着浓度的减小,抑菌圈逐渐减小,抑菌活性减弱。在同一浓度下,蜂胶浓度为10%-20%时,蜂胶对各受试菌株均表现出较高的抑菌活性;蜂胶对各受试菌株抑菌的强弱顺序为:枯草芽孢杆菌>苏云杆菌>藤黄微球菌>奇异变形杆菌>白色葡萄球菌>大肠杆菌。[结论]蜂胶对6种常见致病菌有不同程度的抑菌作用,对苏云杆菌、枯草芽孢杆菌有很强的抑菌作用,为作为外用药等提供依据。     

In vivo and in vitro inhibitory effects of a traditional Chinese formulation on LPS-stimulated leukocyte-endothelial cell adhesion and VCAM-1 gene expression

Aim of the study

A traditional Chinese formulation Huang-Lian-Jie-Du-Tang (HLJDT) exerts anti-inflammatory effects. The present study aimed to investigate the effect of HLJDT on the LPS-stimulated leukocyte–endothelial cell adhesion and VCAM-1 gene expression both in vivo and in vitro.

Materials and methods

HLJDT was extracted from rhizoma coptidis, radix scutellariae, cortex phellodendri and fructus gardeniae in a weight rario of 1:1:1:1. In vivo leukocyte–endothelial cell adhesion was observed in rat lung after LPS stimulation (5 mg/kg, i.p.) with or without HLJDT (350 or 700 mg/kg, i.g.) pretreatment. The protein expression of vascular cell adhesion molecule 1 (VCAM-1) was analyzed by immunohistochemical method. In vitro leukocyte–endothelial cell adhesion was performed by examining the adhesion of THP-1 cells to LPS-stimulated human vascular endothelial cells with or without HLJDT pretreatment. The VCAM-1 expression at the RNA and protein levels was investigated by RT-PCR and western blot analysis, respectively. The activation of NF-κB was examined by the nuclear translocation of NF-κB by immunocytochemical method.

Results

In vivo, HLJDT dose-dependently reduced the number of leukocytes adhered to endothelium and VCAM-1 protein expression in lung venules of LPS-challenged rats. In vitro, HLJDT dose-dependently decreased the number of THP-1 cells adhered to LPS-stimulated endothelial cells and the expression of VCAM-1 both at the RNA and protein levels. The LPS-induced nuclear translocation of NF-kappa B in endothelial cells was also dose-dependently inhibited by HLJDT.

Conclusions

The present study demonstrated an additional mechanism underlying the anti-inflmmatory effect of HLJDT by inhibiting the leukocyte–endothelial cell adhesion and VCAM-1 gene expression. The inhibition of NF-kappa B activation by HLJDT might suggest a profound anti-inflammatory consequences.     

Antiallergic effects of Scutellaria baicalensis on inflammation in vivo and in vitro

Ethnopharmacological relevance

Scutellaria baicalensis (SB) is one of the most widely used medicinal herbs for the treatment of inflammation. In this study, we investigated the antiallergic effect of SB in vivo and in vitro.

Materials and methods

Sprague–Dawley (SD) rats received intradermal injections of anti-DNP IgE at each of three dorsal skin sites. Forty-eight hours later, each rat received an injection of DNP-HSA in saline containing 4% Evans blue through the dorsal vein of the penis. One hour before injection, SB extract was administered orally. The dorsal skin of the rats was removed and the pigment area measured. In addition, rat peritoneal mast cells (RPMCs) were cultured and purified to investigate histamine release. In vitro, human mast cells (HMC-1) were pretreated with SB extract for 30 min before stimulation with phorbol 12-myristate 13-acetate (PMA) plus A23187. The effects on pro-inflammatory cytokine expression and mitogen activated protein (MAP) kinase expression were investigated using TNF-α and IL-8 assays, and Western blotting analysis of HMC-1 cells.

Results and conclusions

SB treatment inhibited the passive cutaneous anaphylaxis reaction compared to the control group, and histamine release decreased significantly following treatment of RPMCs with SB. In HMC-1 cells, SB restored IL-8 and TNF-α expression and inhibited MAP kinase expression in compound 48/80-induced HMC-1 cells. These data suggest that SB may prove to be a useful anti-inflammatory agent through its downregulation of the expression of various inflammatory mediators.     

八正合剂体外抗菌及对动物的解热抗炎作用

目的 探讨八正合剂治疗泌尿系统感染性疾病的作用机制。方法 采用ig给药观察八正合剂对大鼠尿量和角叉菜胶致大鼠发热的影响。及巴豆油致小鼠耳部肿胀和0.6%醋酸致小鼠腹腔毛细血管通透性增高的影响。并用二倍稀释法观察该药对泌尿系8种常见致病菌的体外抑菌作用。结果 八正合剂可明显增加大鼠尿量，减轻角叉菜胶所致大鼠体温升高，巴豆油所致小鼠耳部炎性肿胀和醋酸所致小鼠腹腔毛细血管通透性增高，对大肠杆菌等常见致病菌也有一定抑制作用。结论 八正合剂对泌尿系感染的病因治疗机制主要在于清除尿路致病菌，同时该药还可缓解感染后多种炎性反应症状。     

Analysis of propolis from Baccharis dracunculifolia DC. (Compositae) and its effects on mouse fibroblasts

This paper confirms Baccharis dracunculifolia DC. (Compositae) as the main botanical source of the propolis from southeastern Brazil (state of São Paulo) investigated to ascertain specific biological activity in relation to mouse NIH-3T3 fibroblasts, skin cells directly involved in the cicatrization processes. Flavonoid and total phenolic compounds were determined by spectrophotometry, and chemical composition by HPLC; the chromatographic profile, characterized largely by flavonoids and aromatic acids, was found to be qualitatively similar to that of Baccharis dracunculifolia DC. The adsorption of phenolic compounds in the propolis to skin powder was also investigated, and 68% of these compounds adsorbed to the skin powder. At concentrations from 0.12 to 7.81 μg/ml, the propolis revealed no statistical significant differences from its control solutions; however, at concentrations of 31.25 μg/ml or more, the propolis was toxic to NIH-3T3 cells. Thus, the propolis from Baccharis dracunculifolia DC. (Compositae) presents an in vitro concentration-dependent toxicity on mouse NIH-3T3 fibroblasts.     

Evaluation of radioprotective effects of propolis and its flavonoid constituents: in vitro study on human white blood cells

This in vitro study aimed to evaluate the possible radioprotective effects of the natural substances WSDP, caffeic acid, chrysin and naringin on γ‐irradiated human white blood cells. The effectiveness of tested compounds was evaluated using the alkaline comet assay, the analysis of structural chromosome aberration and the cytokinesis‐block micronucleus assay. The results obtained by the alkaline comet study indicate favourable toxicity profiles of propolis and its polyphenolic components, and confirmed the radioprotective abilities comparable to the chemical radioprotector AET. WSDP and its polyphenolic components were able to reduce the number of necrotic cells. None of tested compounds induced significant genotoxicity, but all of them offered a quite measurable protection against DNA damage. WSDP was found to be the most effective in diminishing the levels of primary and more complex cytogenetic DNA damage in white blood cells. Considering its complex composition, to undoubtedly explain the underlying mechanisms of cyto/radioprotective effects, further studies are needed. Copyright © 2009 John Wiley & Sons, Ltd.     

桂枝汤及其体外消化道模拟产物对于小鼠脑微血管内皮细胞株分泌功能的影响

目的观察桂枝汤及其体外消化道模拟产物对于白细胞介素-1β(IL-1β)介导的小鼠脑微血管内皮细胞株b End.3分泌功能的影响。方法选取小鼠脑微血管内皮细胞株b End.3细胞,分为正常组,对照组,5%,10%和20%桂枝汤含药血清组及5%,10%和20%模拟药组。比较各组前列腺素E2(PGE2)、前列腺素(PGES)、分泌型磷脂酶A2(s PLA2)、15-羟基前列腺素脱氢酶(15-PGDH)和胞浆型磷脂酶A2(c PLA2)表达水平。结果在不同浓度桂枝汤含药血清和模拟药组中,PGE2、PGES、s PLA2及15-PGDH和c PLA2含量均随着血清含药浓度升高而逐渐下降且有统计学意义(P均<0.05);桂枝汤含药血清和模拟药相同药物浓度组中,各指标含量比较差异均无统计学意义(P均>0.05)。结论桂枝汤及其体外消化道模拟产物对于IL-1β介导的小鼠脑微血管内皮细胞株b End.3细胞分泌功能具有相似效果,可为模拟产物应用开发提供支持。