Anti-angiogenic effect of furanodiene on HUVECs in vitro and on zebrafish in vivo

Ethnopharmacological relevance

Furanodiene is an active ingredient of the traditional Chinese medicine, Rhizoma Curcumae, commonly used for the treatment of cancer in China.

Aim of the study

To investigate the anti-cancer property of Rhizoma Curcumae, this study describes the anti-angiogenic activities of furanodiene in human umbilical vein endothelial cells (HUVECs) in vitro and in zebrafish in vivo.

Materials and methods

HUVECs were treated with different doses of furanodiene in the presence or absence of vascular endothelial growth factor (VEGF). The anti-proliferative effect of furanodiene was measured using the XTT assay. The anti-migration and anti-invasion activities of this compound were investigated with a wound-healing migration model and a three-dimensional cell invasion model, respectively. The effects of furanodiene on HUVEC differentiation were assessed by in vitro tube formation in Matrigel™. The expression of related proteins was detected by Western blot. Morphological observations of zebrafish were evaluated in transgenic Tg (fli1: EGFP) zebrafish embryos.

Results

Our results showed that furanodiene exposure could significantly inhibit the proliferation of HUVECs in a dose-dependent manner and inhibit VEGF-induced proliferation at a low dose. Relative to the VEGF-induced control, the number of invading and migrating cells was significantly reduced in the furanodiene-treated groups. Furanodiene also dramatically suppressed tube formation and p-Akt (Ser473), p-Erk 1/2 (Thr202/Tyr204), ICAM-1, p-p85 (Ser428) as well as p85 protein expression. Furthermore, exposure to furanodiene inhibited angiogenesis in the zebrafish model.

Conclusions

This study demonstrated that furanodiene exposure exhibits a potential anti-angiogenic effect through suppression of endothelial cell growth, invasion, migration and tube formation via regulation of the PI3K pathway. This potential anti-angiogenic effect of furanodiene may play an important role in the anti-tumor activity of the traditional Chinese medicine, Rhizoma Curcumae.     

Triptolide: Progress on research in pharmacodynamics and toxicology

Ethnopharmacological relevance

Tripterygium wilfordii Hook. f. (Tripterygium wilfordii), also known as Huangteng and gelsemium elegan, is a traditional Chinese medicine that has been marketed in China as Tripterygium wilfordii glycoside tablets. Triptolide (TP), an active component in Tripterygium wilfordii extracts, has been used to treat various diseases, including lupus, cancer, rheumatoid arthritis and nephritic syndrome. This review summarizes recent developments in the research on the pharmacodynamics, pharmacokinetics, pharmacy and toxicology of TP, with a focus on its novel mechanism of reducing toxicity. This review provides insight for future studies on traditional Chinese medicine, a field that is both historically and currently important.

Materials and methods

We included studies published primarily within the last five years that were available in online academic databases (e.g., PubMed, Google Scholar, CNKI, SciFinder and Web of Science).

Results

TP has a long history of use in China because it displays multiple pharmacological activities, including anti-rheumatism, anti-inflammatory, anti-tumor and neuroprotective properties. It has been widely used for the treatment of various diseases, such as rheumatoid arthritis, nephritic syndrome, lupus, Behcet?s disease and central nervous system diseases. Recently, numerous breakthroughs have been made in our understanding of the pharmacological efficacy of TP. Although TP has been marketed as a traditional Chinese medicine, its multi-organ toxicity prevents it from being widely used in clinical practice.

Conclusions

Triptolide, a biologically active natural product extracted from the root of Tripterygium wilfordii, has shown promising pharmacological effects, particularly as an anti-tumor agent. Currently, in anti-cancer research, more effort should be devoted to investigating effective anti-tumor targets and confirming the anti-tumor spectrum and clinical indications of novel anti-tumor pro-drugs. To apply TP appropriately, with high efficacy and low toxicity, the safety and non-toxic dose range for specific target organs and diseases should be determined, the altered pathways and mechanisms of exposure need to be clarified, and an early warning system for toxicity needs to be established. With further in-depth study of the efficacy and toxicity of TP, we believe that TP will become a promising multi-use drug with improved clinical efficacy and safety in the future.     

莪术油在Caco-2细胞模型中的吸收机制研究

 目的研究莪术油及其部分活性单体在Caco-2体外细胞吸收模型中的吸收机制,并比较各成分以单体形式及以总油形式在细胞模型上透过的异同。方法应用Caco-2体外细胞吸收模型,分别考察莪术油(Zedoary Turmeric oil,ZTO)及其部分单体,包括莪术二酮(curdione)、吉马酮(germacrone)、呋喃二烯(furanodiene)、莪术醇(curcumol)在不同方向,不同浓度下的转运情况,并比较各单体与挥发油中相应成分在同一浓度下的转运情况。采用高效液相色谱法测定药物浓度,计算其表观渗透系数。结果莪术二酮、吉马酮、莪术醇有很高的表观渗透系数,且不随转运方向及给药浓度影响(P>0.05);而呋喃二烯及挥发油中的其他高脂溶性成分[包括呋喃二烯、莪术烯(curzerene)、β-榄香烯(β-elemene)]在Caco-2细胞模型上不透过;挥发油中其他成分不影响莪术二酮、吉马酮在细胞模型上的透过(P>0.05);莪术油及其活性单体透过Caco-2单层细胞膜后并无代谢产物可被检测。结论莪术油及其部分单体在Caco-2细胞模型中的转运机制为被动扩散,莪术油中高脂溶性成分及呋喃二烯由于被单层细胞膜摄取而不透过Caco-2细胞,挥发油中其他成分对单体成分的透过无影响。     

Fighting fire with fire: poisonous Chinese herbal medicine for cancer therapy

Ethnopharmacological relevance

Following the known principle of “fighting fire with fire”, poisonous Chinese herbal medicine (PCHM) has been historically used in cancer therapies by skilled Chinese practitioners for thousands of years. In fact, most of the marketed natural anti-cancer compounds (e.g., camptothecin derivatives, vinca alkaloids, etc.) are often known in traditional Chinese medicine (TCM) and recorded as poisonous herbs as well. Inspired by the encouraging precedents, significant researches into the potential of novel anticancer drugs from other PCHM-derived natural products have been ongoing for several years and PCHM is increasingly being recognized as a gathering place for promising anti-cancer drugs. The present review aimed at giving a rational understanding of the toxicity of PCHM and, especially, providing the most recent developments on PCHM-derived anti-cancer compounds.

Materials and methods

Information on the toxicity and safety control of PCHM, as well as PCHM-derived anti-cancer compounds, was gathered from the articles, books and monographs published in the past 20 years.

Results

Based on an objective introduction to the CHM toxicity, we clarified the general misconceptions about the safety of CHM and summarized the traditional experiences in dealing with the toxicity. Several PCHM-derived compounds, namely gambogic acid, triptolide, arsenic trioxide, and cantharidin, were selected as representatives, and their traditional usage and mechanism of anti-cancer actions were discussed.

Conclusions

Natural products derived from PCHM are of extreme importance in devising new drugs and providing unique ideas for the war against cancer. To fully exploit the potential of PCHM in cancer therapy, more attentions are advocated to be focused on their safety evaluation and mechanism exploration.     

Effect of Rhizoma Polygonati on 12-O-tetradecanoylphorbol-acetate-induced ear edema in mice

Ethnopharmacological relevance

Rhizoma Polygonati is originated from the dried rhizomes of Polygonatum sibircum Red. It has long been used in traditional Chinese medicine for the treatment of inflammatory disorders.

Aim of the study

The present study aims to investigate the anti-inflammatory effect of aqueous extract of Rhizoma Polygonati (ERP) in a mouse model of inflammation induced by 12-O-tetradecanoylphorbol-acetate (TPA).

Materials and methods

The anti-inflammatory effect was evaluated by measuring the ear thickness and activity of myeloperoxidase (MPO). The anti-inflammatory mechanism was explored by determining the protein and mRNA levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6.

Results

The results showed that ERP significantly decreased the ear thickness and MPO activity in mouse model of inflammation induced by TPA. In addition, ERP also remarkably inhibited the protein and mRNA levels of iNOS, COX-2, TNF-α, IL-1β, and IL-6.

Conclusions

These results indicate that ERP has potential anti-inflammatory effect on TPA-induced inflammatory in mice, and the anti-inflammatory effect may be mediated, at least in part, by inhibiting the mRNA expression of a panel of inflammatory mediators including iNOS, COX-2, TNF-α, IL-1β, and IL-6.     

Chemopreventive and anti-cancer properties of the aqueous extract of flowers of Butea monosperma

Ethnopharmacological relevance

Butea monosperma (Lam.) (Fabaceae) popularly known as ‘flame of the forest’ has been widely used in the traditional Indian medical system of ‘Ayurveda’ for the treatment of a variety of ailments including liver disorders.

Aim of the study

To evaluate the antioxidative, anti-inflammatory, hepatoprotective and anti-cancer activities of the aqueous extract of Butea monosperma flowers.

Materials and methods

Dried flowers of Butea monosperma were extracted with water. The extract was tested for its anti-proliferative, pro-apoptotic and anti-carcinogenic effects in hepatoma cell lines. The chemopreventive and anti-angiogenic effects of the extract were evaluated by its daily oral administration in a HBV-related X15-myc mouse model of hepatocellular carcinoma (HCC).

Results

Treatment with the aqueous extract inhibited cell proliferation and accumulation of cells in G1 phase. This was accompanied by a marked reduction in the levels of activated Erk1/2 and SAPK/JNK and induction of apoptotic cell death. Oral administration of the extract in transgenic mice conferred hepatoprotection as is evident from normal serum ALT levels and improved liver histopathology and lowered serum VEGF level.

Conclusions

The ability of aqueous extract of Butea monosperma flowers to impose growth arrest and trigger pro-apoptotic death in cell culture strongly correlated with its strong chemopreventive effect in vivo when given orally.     

Huangkui capsule,an extract from Abelmoschus manihot (L.) medic,ameliorates adriamycin-induced renal inflammation and glomerular injury via inhibiting p38MAPK signaling pathway activity in rats

Ethnopharmacological relevance

Abelmoschus manihot (L.) medic (AM) is a natural medicinal plant used for the treatment of inflammatory diseases in China. Huangkui capsule (HKC), an extract from AM, has been proved clinically effective in improving renal inflammation and glomerular injury in chronic kidney disease (CKD). However, the dose-effects and the mechanisms involved in vivo are still unclear.

Aim of the study

This study was performed to examine the dose-effects of HKC on renal inflammation and glomerular lesion in adriamycin-induced nephropathy (ADRN), then to clarify the mechanisms in vivo of HKC by investigating its actions on modulating the activation of p38 mitogen-activated protein kinase (p38MAPK) signaling pathway.

Materials and methods

The rats with chronic ADRN, created by the unilateral nephrectomy and twice adriamycin injections (ADR, 4 mg/kg and 2 mg/kg) within 4 weeks, were divided into four groups, a Sham group, a Vehicle group, a high-dose HKC group, and a low-dose HKC group, and that, sacrificed at the end of the 4th week after the administration. The rat's general status, renal morphological appearance, proteinuria, blood biochemical parameters, glomerular morphological changes, podocyte shape, and macrophage (ED1⁺ and ED3⁺ cells) infiltration in glomeruli were examined, respectively. The protein expressions of inflammatory cytokines including tumor necrosis factor (TNF)-α and interleukin (IL)-2, as well as p38MAPK signaling molecules such as transforming growth factor (TGF)-β1, p38MAPK, and phosphorylated-p38MAPK (p-p38MAPK), were also evaluated individually.

Results

HKC at high dose of 2 g/kg/d not only significantly ameliorated the rat's general status, renal morphological appearance, proteinuria, albumin, and glomerulosclerosis, but also obviously reduced the infiltrated ED1⁺ and ED3⁺ macrophages in glomeruli and TNF-α protein expression in the kidney, in addition to these, evidently down-regulated TGF-β1 and p-p38MAPK protein expressions in ADRN rats, but had no influence on podocyte shape and renal function.

Conclusion

HKC could dose-dependently ameliorate renal inflammation and glomerular injury in ADRN rats, by way of reducing the infiltration and the activation of macrophages in glomeruli, and TNF-α protein expression in the kidney, as well as inhibiting p38MAPK signaling pathway activity via the down-regulation of p-p38MAPK and TGF-β1 protein expressions in vivo.     

Biological studies on Brazilian plants used in wound healing

Aim of the study

n-Hexanic and ethanolic extracts from twelve plants (Brugmansia suaveolens Brecht. et Presl., Eupatorium laevigatum Lam., Galinsoga parviflora Cav., Iresine herbstii Hook., Kalanchöe tubiflora Hamet-Ahti, Petiveria alliacea L., Pluchea sagittalis (Lam.) Cabrera, Piper regnellii DC., Schinus molle L., Sedum dendroideum Moç et Sessé ex DC., Waltheria douradinha St. Hill., Xanthium cavanillesii Schouw.) used in traditional South Brazilian medicine as wound healing agents were investigated in various biological assays, targeting different aspects in this complex process.

Materials and methods

The extracts were investigated on NF-κB DNA binding, p38α MAPK, TNF-α release, direct elastase inhibition and its release as well as on caspase-3. Fibroblasts migration to and proliferation into the wounded monolayers were evaluated in the scratch assay, the agar diffusion test for antibacterial and the MTT assay for cytotoxic effects.

Results

The hydrophilic extracts from Galinsoga parviflora, Petiveria alliacea, Schinus molle, Waltheria douradinha and Xanthium cavanillesii as well as the lipophilic extract of Waltheria douradinha turned out to be the most active ones.

Conclusions

These results increase our knowledge on the wound healing effects of the investigated medicinal plants. Further studies are necessary to find out the effective secondary metabolites responsible for the observed effects.     

Anti-melanoma activity of Cynanchi atrati Radix is mediated by regulation of NF-kappa B activity and pro-apoptotic proteins

Ethnopharmacological relevance

Cynanchi atrati Radix has been traditionally prescribed for patients with inflammatory fever or chronic tumoral disorders. Melanoma is one of the most devastating cancer types, in which overexpression of nuclear factor kappa B (NF-κB) enables the cancer to survive without apoptosis. To identify a potential anti-melanoma candidate, we evaluated the apoptotic activity of an ethanol extract of Cynanchi atrati Radix (CAE) on melanoma and its underlying mechanisms.

Materials and methods

Sixty C57BL/6 N mice with melanoma were orally administrated CAE (100 or 200 mg/kg) or distilled water for 10 days. Survival, tumor weight and volume were monitored and measured. Intratumoral apoptotic change was measured using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. To confirm the pro-apoptotic activity of CAE (10, 50 or 100 μg/mL) compared to positive drug (10 μg/mL of IKK-2 inhibitor IV), cell proliferation, caspase-3/7 activity, flow cytometric analysis, TUNEL and DAPI staining, immunoblotting and gene expression analyses for apoptosis-associated genes were conducted using B16F10 cell line.

Results

CAE administration remarkably improved survivability with a significant reduction in tumor weight (p<0.01) and volume (p<0.01), as well as increased apoptotic bodies in melanoma tissue. The CAE treatment significantly inhibited proliferation of B16F10 cells (p<0.001), but increased caspase-3/7 activity (p<0.01 or 0.001) and apoptotic population. The CAE partially blocked nuclear translocation of NF-κB but activated the p53-associated apoptotic pathway.

Conclusion

These results indicate that the CAE has anti-melanoma potential, and the underlying mechanisms involve inhibition of the activities of NF-κB and its target proteins as well as promoting the activities of pro-apoptotic proteins.     

Aloe-emodin from rhubarb (Rheum rhabarbarum) inhibits lipopolysaccharide-induced inflammatory responses in RAW264.7 macrophages

Ethnopharmacological relevance

Rheum rhabarbarum (rhubarb) has long been used for the treatment of inflammation in China and other Asian countries. However, the mechanism underlying the anti-inflammatory activity of this medicinal plant is not fully understood. The present study was designed to investigate the anti-inflammatory effects of anthraquinones, the major constituents in rhubarb, and the molecular mechanism involved in their anti-inflammatory effects.

Materials and methods

RAW264.7 cells were stimulated by lipopolysaccharide (LPS) in the presence or absence of the compounds examined. The proliferation of RAW264.7 cells was assayed by the Alamar-Blue method. The quantity of nitric oxide (NO) was determined by Griess assay. The expression of pro-inflammatory cytokines was determined by enzyme-linked immunosorbent assay (ELISA) and quantitative real-time PCR. Inducible nitric oxide synthase (iNOS), inhibitor of nuclear factor κBα (IκBα), extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (MAPK), c-Jun NH2-terminal kinase (JNK), and Akt/phosphoinositide 3-kinase (PI3K) protein expression levels were determined by Western blotting.

Results

Aloe-emodin markedly suppressed the production of NO, interleukin-6 (IL-6), and interleukin-1β (IL-1β) in LPS-stimulated RAW264.7 cells with no apparent cytotoxicity. The mRNA expression levels of iNOS, IL-6, and IL-1β genes were also significantly inhibited by aloe-emodin. Western blot analysis showed that aloe-emodin suppressed LPS-induced iNOS protein expression, IκBα degradation, and the phosphorylation of ERK, p38, JNK, and Akt.

Conclusions

These results demonstrate that aloe-emodin is the bioactive component of rhubarb that confers an anti-inflammatory effect through a likely mechanism involving a decrease in pro-inflammatory cytokine production in LPS-induced RAW264.7 macrophages via inhibition of NF-κB, MAPK, and PI3K pathways.     

Insulin-sensitizing and insulin-mimetic activities of Sarcopoterium spinosum extract

Ethnopharmacological relevance

Sarcopoterium spinosum is an abundant plant in Israel, used by Bedouin medicinal practitioners for the treatment of diabetes. In our previous study we validated the anti-diabetic activity of Sarcopoterium spinosum. The aim of this study was to further clarify its mechanism of action.

Materials and methods

In-vivo studies were performed on KK-a/y mice given the extract for 6 weeks. Insulin tolerance test was performed, and relative pancreatic islets area was measured. Mechanisms of action were investigated in L6 myotubes using protein array, Western blot analysis and confocal microscopy. Glucose uptake assays were performed in 3T3-L1 adipocytes.

Results

Sarcopoterium spinosum extract reduced fasting blood glucose and improved insulin sensitivity in treated mice. Hypertrophic islets were detected in diabetic, but not in Sarcopoterium spinosum-treated mice. Sarcopoterium spinosum phosphorylated PTEN on ser380 and thr382/383, which are known inhibitory sites. PKB was not phosphorylated by Sarcopoterium spinosum, however, translocation of PKB from cytoplasm to the membrane and nucleus was detected. Target proteins of PKB were regulated by Sarcopoterium spinosum; GSK3β was phosphorylated and cytosolic localization of FoxO was increased. Glucose uptake was increased in a PI3K and AMPK-independent mechanism.

Conclusions

We suggest that Sarcopoterium spinosum inhibited PTEN and activated PKB by a mechanism which is independent of ser473 and thr308 phosphorylation. Other post translation modifications might be involved and should be analyzed further in order to understand this unique PKB activation. Identifying the active molecules in the extract, may lead to the development of new agents for the treatment of insulin resistance.     

Isolation and characterization of an anticancer catechol compound from Semecarpus anacardium

Ethnopharmacological relevance

The fruits and seeds of Semecarpus anacardium are used widely for the treatment of human cancers and other diseases in the Ayurvedic and Sidda systems of medicine in India.

Aim of the study

The principal aim of this investigation was to isolate and characterize the anticancer compound from the kernel of Semecarpus anacardium nut.

Materials and methods

The bioactivity-tailored isolation and detailed chemical characterization were used to identify the active compound. Cytotoxicity, apoptosis, cell cycle arrest as well as synergism between the identified anticancer compound and doxorubicin in human tumor cell lines were analyzed.

Results

GC/MS, IR, proton NMR, carbon NMR and collisionally induced dissociation (CID) spectra analysis showed that the isolated active compound is 3-(8′(Z),11′(Z)-pentadecadienyl) catechol (SA-3C). SA-3C is cytotoxic to tumor cell lines with IC₅₀ values lower than doxorubicin and even multidrug resistant tumor cell lines were equally sensitive to SA-3C. SA-3C induced apoptosis in human leukemia cell lines in a dose-dependent manner and showed synergistic cytotoxicity with doxorubicin. The cell cycle arrest induced by SA-3C at S- and G₂/M-phases correlated with inhibition of checkpoint kinases.

Conclusion

SA-3C isolated from the kernel of Semecarpus anacardium can be developed as an important anticancer agent for single agent and/or multiagent cancer therapy.     

Preventive and therapeutic effects of Danhong injection on lipopolysaccharide induced acute lung injury in mice

Ethnopharmacological relevance

Danhong injection (DHI), a Chinese Materia Medica standardized product extracted from Radix et Rhizoma Salviae Miltiorrhizae (Salvia miltiorrhiza Bge., Labiatae, Danshen in Chinese) and Flos Carthami (Carthamus tinctorius L., Compositae, Honghua in Chinese), has been reported to have anti-inflammatory, anti-oxidative and anti-fibrinolytic properties, which is used extensively for the treatment of cardiovascular diseases in clinic.

Aim of this study

The present study aimed to investigate the preventive and therapeutic effects of DHI on lipopolysaccharide (LPS) induced acute lung injury (ALI) in mice.

Materials and methods

Lung injury was induced by intranasal instillation with 10 μg LPS. Mice were randomly divided into four groups:Control group; LPS group; LPS+5 ml/kg DHI group and LPS+10 ml/kg DHI group. The effects of DHI on LPS-induced neutrophils influx, inflammatory cytokines release, protein leakage, myeloperoxidase (MPO) and superoxide dismutase (SOD) activities, malondialdehyde (MDA) level were examined. In addition, the NF-κB activation in lung tissues was detected by Western blot.

Results

In LPS challenged mice, DHI significantly reduced the infiltration of activated neutrophils and decreased the levels of TNF-α and IL-6 in bronchoalveolar lavage fluid (BALF). DHI also inhibited protein extravasation in BALF, attenuated edema and the pathological changes in the lung. In addition, DHI markedly prevented LPS-induced elevation of MDA and MPO levels, as well as reduction of SOD activity. Further study demonstrated that DHI effectively inhibited the NF-κB activation in lung tissues.

Conclusion

DHI has been demonstrated to protect mice from LPS induced acute lung injury by its anti-inflammatory and anti-oxidant activities.