Hairong Cheng

Anti-fatigue activity of the water-soluble polysaccharides isolated from Panax ginseng C. A. Meyer

AIM OF THE STUDY Panax ginseng C. A. Meyer (ginseng) is a well-known Chinese herb often used in Asian countries for physical strength development. Ginseng polysaccharides are its active component and have a lot of pharmaceutical activities. However, anti-fatigue activity of ginseng polysaccharides has not yet been tested. The current study was designed to evaluate the anti-fatigue activity of ginseng polysaccharides (WGP) in an animal test for fatigue and compare the activities between the neutral (WGPN) and acidic (WGPA) portion in an attempt to determine whether the medicinal uses are supported by pharmacological effects. MATERIALS AND METHODS WGP, WGPN and WGPA were orally administrated to mice once daily for 15 days. Anti-fatigue activity was assessed using the forced swim test (FST) and serum biochemical parameters were determined by autoanalyzer and commercially available kits. RESULTS While all compounds were found to reduce immobility in the FST, the effect of WGPA was demonstrated in lower doses compared with WGP and WGPN. Moreover, the FST-induced reduction in glucose (GLU) and glutathione peroxidase (GPx) and increase in creatine phosphokinase (CK), lactic dehydrogenase (LDH) and malondialdehyde (MDA) levels, all indicators of fatigue, were inhibited by the corresponding doses of WGP, WGPN and WGPA. CONCLUSIONS Ginseng polysaccharides have anti-fatigue activity, also reflected in the effects on the physiological markers for fatigue. The acidic polysaccharide is more potent than the neutral polysaccharide.

Comparative studies of the antiproliferative effects of ginseng polysaccharides on HT-29 human colon cancer cells

Ginseng polysaccharide has anticancer activity. However, the structure–activity relationship and the activity mechanism are still unclear. Therefore, it is necessary to study the anticancer activity of structurally different ginseng polysaccharide fractions and their potential mechanisms. Ginseng polysaccharide fractions and their temperature-modified products were assayed for their effects on HT-29 cell proliferation by MTT assay, on cell cycle progression by flow cytometry, and on caspase-3 activation by western blot analysis. The HG-rich ginseng pectin inhibited cell proliferation and induced cell cycle arrest in the G2/M phase. The temperature-modified HG-rich pectin had dramatically increased antiproliferative effect and induced apoptosis accompanied by the activation of caspase-3. Starch-like glucan and arabinogalactan of ginseng exhibited no antiproliferative effects. Even after temperature modification, their inhibitory effects either remained unchanged or increased slightly. The HG-rich pectin exerts its antiproliferative effect via cell cycle arrest and the temperature modification markedly increased the antiproliferative effect.

The inhibitory effects and mechanisms of rhamnogalacturonan I pectin from potato on HT-29 colon cancer cell proliferation and cell cycle progression

Pectin is an important dietary component of all fruits and vegetables. Some pectins have been shown to inhibit cancer cell growth, but the effective structures and mechanisms have remained unclear. In this study, we investigated the effects of four structurally distinct pectins on human colon cancer HT-29 cells and the possible mechanisms accounting for the actions. The proliferation inhibitory effect was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Flow cytometry was used to visualize the cell cycle distribution. An reverse transcription polymerase chain reaction (RT-PCR)-based assay was utilized to detect mRNA levels of the proteins related to cell cycle arrest. The data showed that the rhamnogalacturonan I domain-rich pectin from potato inhibited the proliferation of HT-29 cells and induced significant G2/M cell cycle arrest. This inhibitory effect was due to the down-regulation of cyclin B1 and cyclin-dependent kinase 1 expression, but not p21WAF1/CIP1 expression. The results suggested that the rhamnogalacturonan I domain might relate to the anticancer activity of pectin.

Ginsenoside compound K sensitizes human colon cancer cells to TRAIL-induced apoptosis via autophagy-dependent and -independent DR5 upregulation.

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a potent cancer cell-specific apoptosis-inducing cytokine with little toxicity to most normal cells. However, acquired resistance of cancer cells to TRAIL is a roadblock. Agents that can either potentiate the effect of TRAIL or overcome resistance to TRAIL are urgently needed. This article reports that ginsenoside compound K (CK) potentiates TRAIL-induced apoptosis in HCT116 colon cancer cells and sensitizes TRAIL-resistant colon cancer HT-29 cells to TRAIL. On a cellular mechanistic level, CK downregulated cell survival proteins including Mcl-1, Bcl-2, surviving, X-linked inhibitor of apoptosis protein and Fas-associated death domain-like IL-1-converting enzyme-inhibitory protein, upregulated cell pro-apoptotic proteins including Bax, tBid and cytochrome c, and induced the cell surface expression of TRAIL death receptor DR5. Reduction of DR5 levels by siRNAs significantly decreases CK- and TRAIL-mediated apoptosis. Importantly, our results indicate, for the first time, that DR5 upregulation is mediated by autophagy, as blockade of CK-induced autophagy by 3-MA, LY294002 or Atg7 siRNAs substantially decreases DR5 upregulation and reduces the synergistic effect. Furthermore, CK-stimulated autophagy is mediated by the reactive oxygen species–c-Jun NH2-terminal kinase pathway. Moreover, we found that p53 and the C/EBP homologous (CHOP) protein is also required for DR5 upregulation but not related with autophagy. Our findings contribute significantly to the understanding of the mechanism accounted for the synergistic anticancer activity of CK and TRAIL, and showed a novel mechanism related with DR5 upregulation.

Comparative studies on the anti-tumor activities of high temperature- and pH-modified citrus pectins

High temperature and pH modification could produce functional pectins. In this study, high temperature-modified (HTCP) and pH-modified (MCP) citrus pectins were prepared for studying their anti-tumor activities in eight cancer cell lines and a mouse Sarcoma-180 (S-180) tumor model. HTCP inhibited the proliferation of these cancer cells and induced a caspase-3-dependent cell apoptosis and cell cycle arrest at G2/M phase. It also inhibited the growth of S-180 tumor to 49% of the control at the dose of 200 mg kg(-1) d(-1) and extended the survival time of the tumor-bearing mice. MCP had no anti-proliferative effects on these cancer cells and no anti-tumor effect in the mouse model. The anti-tumor activity of HTCP in the mouse tumor model was not correlated with immunomodulation and galectin-3 inhibition, but correlated well with proliferation inhibition. HTCP might be exploited as a functional food for cancer prevention and/or treatment.

Gefitinib enhances human colon cancer cells to TRAIL-induced apoptosis of via autophagy- and JNK-mediated death receptors upregulation

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a potent cancer cell-specific apoptosis-inducing cytokine with little toxicity to most normal cells. Here, we report that gefitinib and TRAIL in combination produce a potent synergistic effect on TRAIL-sensitive human colon cancer HCT116 cells and an additive effect on TRAIL-resistant HT-29 cells. Interestingly, gefitinib increases the expression of cell surface receptors DR4 and DR5, possibly explaining the synergistic effect. Knockdown of DR4 and DR5 by siRNA significantly decreases gefitinib- and TRAIL-mediated cell apoptosis, supporting this idea. Because the inhibition of gefitinib-induced autophagy by 3-MA significantly decreases DR4 and DR5 upregulation, as well as reduces gefitinib- and TRAIL-induced apoptosis, we conclude that death receptor upregulation is autophagy mediated. Furthermore, our results indicate that death receptor expression may also be regulated by JNK activation, because pre-treatment of cells with JNK inhibitor SP600125 significantly decreases gefitinib-induced death receptor upregulation. Interestingly, SP600125 also inhibits the expression CHOP, yet CHOP has no impact on death receptor expressions. We also find here that phosphorylation of Akt and ERK might also be required for TRAIL sensitization. In summary, our results indicate that gefitinib effectively enhances TRAIL-induced apoptosis, likely via autophagy and JNK- mediated death receptor expression and phosphorylation of Akt and ERK.

The inhibitory effect of ginseng pectin on L-929 cell migration

We tested the effects of ginseng pectin prepared by enzymatic hydrolysis of ginseng polysaccharides on cell migration. Ginseng pectin impaired the migration of L-929 cells and reduced their migration speed by up to 50% of control in the presence or absence of serum, suggesting it worked on both serum-dependent and serum-independent migration pathways. Ginseng pectin impaired cell migration via decreased cell spreading. These findings represent a significant contribution towards understanding the bioactivities of ginseng polysaccharides and applying them to health food and medicine.

Tunicamycin enhances human colon cancer cells to TRAIL-induced apoptosis by JNK-CHOP-mediated DR5 upregulation and the inhibition of the EGFR pathway

Tumor necrosis factor related apoptosis-inducing ligand (TRAIL) is a cytokine that selectively induces apoptosis in many tumor cells while leaving normal cells intact and is thus an attractive candidate for antitumor therapies. This paper reports that the combination of tunicamycin plus TRAIL produced a strong synergistic effect in TRAIL-sensitive human colon cancer HCT116 cells and TRAIL-resistant HT-29 cells. On a cellular mechanistic level, tunicamycin-enhanced TRAIL-induced apoptosis by death receptor (DR) 5 upregulation and DR4 deglycosylation. Knockdown of DR5 but not DR4 expression by specific shRNAs or siRNAs significantly increased tunicamycin-mediated and TRAIL-mediated cell viability. DR5 induction was regulated by C/EBP homologous protein (CHOP) and JNK as CHOP siRNA or JNK inhibitor SP600125 considerably abolished the DR5 induction. In addition, tunicamycin inhibited epidermal growth factor receptor glycosylation and the downstream signaling pathways, Akt and extracellular signal–regulated kinases activation, which might also be required for TRAIL sensitization by tunicamycin. In summary, tunicamycin effectively enhanced TRAIL-induced apoptosis might through JNK-CHOP-mediated DR5 upregulation and the inhibition of the epidermal growth factor receptor pathway.

The N-terminal tail coordinates with carbohydrate recognition domain to mediate galectin-3 induced apoptosis in T cells

Galectin-3 is a galectin with a unique flexible N-terminal tail (NT) connected to the conserved carbohydrate recognition domain (CRD). Galectin-3 is associated with tumor immune tolerance and exhibits an ability to induce T cell apoptosis. We used Jurkat, Jurkat E6-1 and CEM T-cell lines and human peripheral blood mononuclear cells (PBMCs) to investigate the specific roles of the CRD and NT in inducing T cell apoptosis. Galectin-3 triggered sustained extracellular signal-regulated kinase (ERK) phosphorylation that induced apoptosis. ERK was situated upstream of caspase-9 and was independently activated by reactive oxygen species (ROS) and protein kinase C (PKC). The first twelve NT residues had no role in the apoptosis. Residues 13-68 were essential for activating ROS, but did not activate PKC. However, residues 69-110 were required for activation of PKC. An NT fragment and a NT-specific antibody antagonized the apoptosis triggered by full-length galectin-3 further supporting our findings. These findings indicate the CRD and NT play important roles during induction of T cell apoptosis, which suggests their potential as therapeutic targets for reversing cancer immune tolerance.

Structural characterization and macrophage activation of a hetero-galactan isolated from Flammulina velutipes

We isolated and purified a new polysaccharide (WFVP-N-b1) with a molecular weight of 20 kDa from Flammulina velutipes. Results showed that WFVP-N-b1 is composed of an α (1 → 6)-linked D-galactan backbone and branched at the O-2 of its Galp residues by an α-D-(1 → 6)-linked Manp attached to t-β-D-Glcp or t-α-D-Fucp side chains. WFVP-N-b1 can significantly induce cytokines secretion and release of toxic molecules. On a cellular level, WFVP-N-b1 is recognized by Toll-like receptor 4 (TLR4). Thereby, the hetero-galactan increased the phosphorylation of mitogen-activated protein kinases (MAPKs) and Akt, promoted degradation of IκB-α and the nuclear translocation of the NF-κB p65 subunit. Importantly, our results indicate that WFVP-N-b1 activated macrophage is mediated by autophagy, as blockade of WFVP-N-b1-induced autophagy by Baf-A1 significantly decreases macrophage activation. This is the first report that hetero-galactan-induced macrophage activation is mediated by autophagy. Collectively, WFVP-N-b1 activated RAW264.7 cells through MAPKs, autophagy, and Akt/NF-κB signaling pathways via TLR4 receptor.