[Effects and mechanisms of Xiao-Ai-Ping injection on angiogenesis].

This study was designed to investigate the effect of Xiao-Ai-Ping injection on cancer angiogenesis. CCK8 assay and Brd U incorporation immunofluorescence assay were used to detect the effect of Xiao-Ai-Ping injection on HUVECs proliferation; wound healing assay and transwell assay were employed to test the effect of Xiao-Ai-Ping injection on HUVECs migration. The anti-angiogenic effect of Xiao-Ai-Ping injection was examined by tube formation assay, rat aortic ring assay and chicken chorioallantoic membrane(CAM) assay. ELISA assay was used to measure the secretion of vascular endothelial growth factor(VEGF); and the activation of vascular endothelial growth factor receptor 2(VEGFR2) protein and its downstream signaling pathways were examined by Western blot. Our data demonstrated that Xiao-Ai-Ping injection inhibited HUVECs proliferation in a time- and dose-dependent manner, and the IC(50) (mg·m L(-1)) values for 24, 48 and 72 h were 48.7 ± 7.14, 29.1 ±2.25 and 22.0 ± 4.53, individually. Xiao-Ai-Ping injection inhibited HUVECs DNA synthesis and migration. Xiao-Ai-Ping injection suppressed HUVECs tube formation, and reduced microvessel sprouting from rat aortic rings and vessel growth in CAMs. Furthermore, Xiao-Ai-Ping injection attenuated the secretion of VEGF, and inhibited the expression of p-VEGFR2 and phosphorylation of protein kinase B(p-AKT). We conclude that Xiao-Ai-Ping injection inhibits angiogenesis by down-regulation of VEGF signaling and AKT pathway.

(+)- and (-)-liriodenol, a pair of novel enantiomeric lignans from Liriodendron hybrid.

(+)- and (-)-liriodenol, a pair of unprecedented enantiomeric lignans bearing a 1,1-disubstituted olefinic group, were isolated from the barks of Liriodendron hybrid. The structure and relative configurations were determined by comprehensive analysis of MS and NMR spectroscopy. The cytotoxicity of these three lignans ((±)-, (+)-, and (-)-liriodenol) was evaluated in vitro against four selected human tumor cell lines, where (+)-liriodenol showed more significant cytotoxic effects than the (±)- and (-)-liriodenol enantiomers.

The extract, LXB-1, from the barks of Liriodendron × hybrid, induced apoptosis via Akt, JNK and ERK1/2 pathways in A549 lung cancer cells.

The effect of LXB-1, an extract from Liriodendron × hybrid, was determined on A549 human lung adenocarcinoma cell lines. Growth inhibition of LXB-1 was analyzed by MTT assay. Cancer cell cycle was measured by flow cytometry. To verify the apoptosis effect of LXB-1 on A549 cells, annexin V/PI double staining assay was performed. The expression levels of proapoptotic proteins were also measured by western blot. The potential mechanisms of LXB-1 inducing apoptosis - the expression and phosphorylation of ERK, p38, JNK and Akt - were investigated by western blot. The IC50 values of LXB-1 on A549 for 24, 48 and 72 h treatment were determined to be 12.97±1.53 µg/mL, 9.55±1.42 µg/mL, and 5.90±0.74 µg/mL, respectively. LXB-1 induced an obvious G2/M cell cycle arrest in A549 cells and resulted in significant cell apoptosis. LXB-1 also increased the cleavage of both caspase-3 and caspase-9, and greatly decreased the protein levels of Bcl-2. Moreover, LXB-1 increased the expression of phosphorylated JNK but decreased the levels of phosphorylated ERK1/2 and Akt. These results suggest that that LXB-1 induced apoptosis through JNK, ERK1/2, and Akt pathways in A549 cells.

Topotecan inhibits cancer cell migration by down-regulation of chemokine CC motif receptor 7 and matrix metalloproteinases.

AIM: The aim of this study was to investigate the effect of topotecan (TPT) on cancer cell migration. METHODS: Growth inhibition of TPT was analyzed by MTT assay, and cancer cell migration was measured by transwell double chamber assay. To verify the effect of TPT on the chemokine receptors CXCR4 and CCR7, quantitative PCR, semi-quantitative PCR and Western blot analysis were performed. The secretion of MMP-2 and MMP-9 was detected by enzyme-linked immunosorbent assay (ELISA) and gelatin zymography. To evaluate possible contributions of CCR7 to MMP secretion, the overexpression vectors pcDNA3.1(+)-CCR7 and CCR7 siRNA were transiently transfected into MDA-MB-435 cells. RESULTS: TPT inhibited cancer cell migration in a dose-dependent manner. Additionally, TPT significantly decreased the expression of CCR7 in both MDA-MB-435 and MDA-MB-231 cells and moderately reduced the expression of CXCR4 in MDA-MB-435 cells. The secretion of MMPs (MMP-2, MMP-9) was also inhibited by TPT. Overexpression of CCR7 increased the secretion of MMP-2/9 and cancer cell migration, whereas knockdown of CCR7 reduced active MMP-2/9 production and migration of MDA-MB-435 cells. CONCLUSION: TPT inhibited cancer cell migration by down-regulation of CCR7 and MMPs (MMP-2 and MMP-9).

Nuclear Magnetic Resonance-Based Metabolomics Approach to Evaluate the Prevention Effect of Camellia nitidissima Chi on Colitis-Associated Carcinogenesis.

Colorectal cancer (CRC) is one of the most common malignant tumors worldwide, occurring in the colon or rectum portion of large intestine. With marked antioxidant, anti-inflammation and anti-tumor activities, Camellia nitidissima Chi has been used as an effective treatment of cancer. The azoxymethane/dextran sodium sulfate (AOM/DSS) induced CRC mice model was established and the prevention effect of C. nitidissima Chi extracts on the evolving of CRC was evaluated by examination of neoplastic lesions, histopathological inspection, serum biochemistry analysis, combined with nuclear magnetic resonance (NMR)-based metabolomics and correlation network analysis. C. nitidissima Chi extracts could significantly inhibit AOM/DSS induced CRC, relieve the colonic pathology of inflammation and ameliorate the serum biochemistry, and could significantly reverse the disturbed metabolic profiling toward the normal state. Moreover, the butanol fraction showed a better efficacy than the water-soluble fraction of C. nitidissima Chi. Further development of C. nitidissima Chi extracts as a potent CRC inhibitor was warranted.

Repression of telomerase reverse transcriptase mRNA and hTERT promoter by gambogic acid in human gastric carcinoma cells.

OBJECTIVES: To investigate the effects and potential mechanisms of gambogic acid (GA), a naturally occurring anticancer agent, on the expression and regulation of telomerase in human gastric carcinoma cells. METHODS: GA-induced inhibition of cell proliferation was evaluated by the commonly employed MTT assay on two human gastric carcinoma cell lines, MGC-803 and SGC-7901. Telomerase activity and hTERT mRNA expression were determined by telomeric repeat amplication protocol-polymerase chain reaction and reverse transcription-polymerase chain reaction, respectively. The hTERT promoter activity was measured by luciferase assay. The expression of c-MYC, an apoptotic gene that modulates the expression of hTERT promoter, was quantified by Western blotting. RESULTS: The proliferation of human gastric carcinoma cell lines, MGC-803 and SGC-7901, was significantly inhibited with GA treatment. Both telomerase activity and hTERT mRNA expression were notably decreased in cells treated with GA. The activity of hTERT promoter and the expression of c-MYC were also remarkably decreased in GA-treated cells. CONCLUSION: This study demonstrated that GA treatment of human gastric carcinoma cell lines, MGC-803 and SGC-7901, significantly reduced the expression of c-MYC in a time- and concentration-dependent manner accompanied with the down-regulation of the hTERT transcription and the ultimate reduction in telomerase activity. Our results indicate that the hTERT is a target of c-MYC activity and identify a feasible mechanism of GA's potent anticancer activity.

Inhibition of human telomerase reverse transcriptase gene expression by gambogic acid in human hepatoma SMMC-7721 cells.

The activation of human telomerase, a process regulated by the human telomerase reverse transcriptase (hTERT), is a crucial step during cellular immortalization and malignant transformation. We have reported that gambogic acid (GA), a natural product isolated from the gamboge resin of Garcinia hanburyi tree, is an effective telomerase inhibitor and thus displays potent anticancer activity both in vitro and in vivo. Here we present the direct interaction of GA with oncogene c-MYC, a ubiquitous transcription factor involved in the control of cell proliferation and differentiation, as the molecular mechanism of GA's inhibitory effect on telomerase activity. Consistent with the recently reported association between c-MYC overexpression and induction of telomerase activity, we find here that GA treatment of a human hepatoma cell line SMMC-7721 significantly reduced the expression of c-MYC in a time- and concentration-dependent manner accompanied with the down-regulation of the hTERT transcription and the ultimate reduction in telomerase activity. Our results indicate that the hTERT is a target of c-MYC activity and identify a feasible mechanism of GA's potent anticancer activity.

Marsdenia tenacissima extract suppresses A549 cell migration through regulation of CCR5-CCL5 axis, Rho C, and phosphorylated FAK.

Marsdenia tenacissima, a traditional Chinese medicine, is long been used to treat various diseases including asthma, cancer, trachitis, tonsillitis, pharyngitis, cystitis, and pneumonia. Although Marsdenia tenacissima has been demonstrated to have strong anti-tumor effects against primary tumors, its effect on cancer metastasis remains to be defined, and the molecular mechanism underlying the anti-metastatic effect is unknown. In the present study, we investigated the effects of XAP (an extract of Marsdenia tenacissima) on A549 lung cancer cell migration and explored the role of CCR5-CCL5 axis in the anti-metastatic effects of XAP. Our resutls showed that XAP inhibited A549 lung cancer cell migration and invasion in a dose-dependent manner. The protein levels of CCR5, but not CCR9 and CXCR4, were decreased by XAP. The secretion of CCL5, the ligand of CCR5, was reduced by XAP. XAP down-regulated Rho C expression and FAK phosphorylation. In conclusion, XAP inhibited A549 cell migration and invasion through down-regulation of CCR5-CCL5 axis, Rho C, and FAK.

DT-13 attenuates human lung cancer metastasis via regulating NMIIA activity under hypoxia condition.

Cancer metastasis plays a major role in tumor deterioration. Metastatic processes are known to be regulated by hypoxic microenvironment and non-muscle myosin IIA (NMIIA). DT-13, a bioactive saponin monomer isolated from Ophiopogon japonicus, has been reported to inhibit various cancer metastasis, but whether NMIIA is involved in the anti-metastatic activity of DT-13 under hypoxia remains to be determined. Thus, this study aims to clarify the role of DT-13 in regulating 95D cell metastasis under hypoxic microenvironment and to further investigate whether NMIIA is involved in the anti-metastatic mechanism of DT-13. We found that DT-13 significantly inhibited 95D cells metastasis in vitro and in vivo. Furthermore, hypoxia significantly inhibited the expression of NMIIA and redistributed NMIIA to the cell periphery, whereas DT-13 reversed the hypoxic effects by upregulating the expression of NMIIA. Moreover, DT-13 treatment redistributed NMIIA to the nuclear periphery and reduced the formation of F-actin in 95D cells. In addition, we found that the Raf-ERK1/2 signaling pathway is involved in regulation of NMIIA by DT-13. Collectively, these findings support NMIIA as a target of DT-13 to prevent lung cancer metastasis.

Marsdenia tenacissima extract induces G0/G1 cell cycle arrest in human esophageal carcinoma cells by inhibiting mitogen-activated protein kinase (MAPK) signaling pathway.

Marsdenia tenacissima extract (MTE, trade name: Xiao-Ai-Ping injection) is an extract of a single Chinese plant medicine. It has been used for the treatment of cancer in China for decades, especially for esophageal cancer and other cancers in the digestive tract. In the present study, the potential mechanism for MTE's activity in esophageal cancer was explored. The effects of MTE on the proliferation of human esophageal cancer cells (KYSE150 and Eca-109) were investigated by the MTT assay, the BrdU (bromodeoxyuridine) incorporation immunofluorescence assay, and flow cytometric analysis. MTE inhibited cell proliferation through inducing G0/G1 cell cycle arrest in KYSE150 and Eca-109. Western blot analysis was employed to determine protein levels in the MTE treated cells. Compared with the control cells, the expression levels of the cell cycle regulatory proteins cyclin D1/D2/D3, cyclin E1, CDK2/4/6 (CDK: cyclin dependent kinase), and p-Rb were decreased significantly in the cells treated with MTE at 40 mg·mL(-1). In addition, MTE had an inhibitory effect on the MAPK (mitogen-activated protein kinase) signal transduction pathway, including ERK (extracellular signal-regulated kinase), JNK (c-Jun N-terminal kinase), and p38MAPK. Moreover, MTE showed little additional effects on the regulation of cyclin D1/D3, CDK4/6, and p-Rb when the ERK pathway was already inhibited by the specific ERK inhibitor U0126. In conclusion, these data suggest that MTE inhibits human esophageal cancer cell proliferation through regulation of cell cycle regulatory proteins and the MAPK signaling pathways, which is probably mediated by the inhibition of ERK activation.

Potent anti-angiogenic activity of B19--a mono-carbonyl analogue of curcumin.

AIM: The compound B19 (C21H22O5) is a newly synthesized, mono-carbonyl analog of curcumin that has exhibited potential antitumor effects. This present study was performed to identify the anti-angiogenic activity of this compound. METHODS AND RESULTS: B19 inhibited migration and tube formation of human umbilical vein endothelial cells, and arrested microvessel outgrowth from rat aortic rings. In addition, B19 suppressed the neovascularization of chicken chorioallantoic membrane. Mechanistic studies revealed that B19 suppressed the downstream protein kinase activation of vascular endothelial growth factor (VEGF) by decreasing phosphorylated forms of serine/threonine kinase Akt, extracellular signal-regulated kinase, and p38 mitogen-activated protein kinase, with or without stimulating vascular endothelial growth factor (VEGF). CONCLUSIONS: B19 exerted anti-angiogenic activity in vitro and ex vivo, which suggests that it merits further investigation as a promising anticancer angiogenesis compound.

The saponin DT-13 inhibits gastric cancer cell migration through down-regulation of CCR5-CCL5 axis.

AIM: To investigate the effect of DT-13 on gastric cancer cell migration, and to explore the possible mechanisms underlying the anti-metastasis activity of DT-13. METHODS: Growth inhibition of DT-13 was analyzed by the MTT assay. Cell migration was measured by the scratch-wound assay and transwell double chamber assay. To investigate the possible mechanisms underlying the anti-metastasis activity of DT-13, chemokine receptors that are involved in cancer metastasis (CCR2, CCR5, CCR7, CXCR4, and CXCR6) were detected by conventional PCR. The effect of DT-13 on CCR5 and CXCR4 expression was further evaluated by quantitative PCR and Western blot, respectively. The secretion of CCL5 (ligand of CCR5) and SDF-1 (ligand of CXCR4) were detected by enzyme-linked immunosorbent assay (ELISA). RESULTS: DT-13 inhibited BGC-823 and HGC-27 cell growth in a dose dependent manner, and the estimated IC50 value for 24 h treatment was 23.5 ± 5.1 µmol·L(-1) for BGC-823 cells and 35.6 ± 7.6 µmol·L(-1) for HGC-27 cells. DT-13 also significantly decreased gastric cancer cell migration. DT-13 significantly decreased the gene expression of CCR5 in both BGC-823 and HGC-27 gastric cancer cells, and moderately reduced the expression of CXCR4. Similar to the results of gene expression, significant down-regulation of CCR5 protein was observed, but CXCR4 protein levels were much less affected. CCL5 secretion, but not SDF-1 production, was inhibited by DT-13. CONCLUSION: DT-13 inhibited gastric cancer cell migration by down-regulation of the CCR5-CCL5 axis.