Epithelial-mesenchymal transition-related long noncoding RNAs in gastric carcinoma.

As an evolutionarily phenotypic conversion program, the epithelial-mesenchymal transition (EMT) has been implicated in tumour deterioration and has facilitated the metastatic ability of cancer cells via enhancing migration and invasion. Gastric cancer (GC) remains a frequently diagnosed non-skin malignancy globally. Most GC-associated mortality can be attributed to metastasis. Recent studies have shown that EMT-related long non-coding RNAs (lncRNAs) play a critical role in GC progression and GC cell motility. In addition, lncRNAs are associated with EMT-related transcription factors and signalling pathways. In the present review, we comprehensively described the EMT-inducing lncRNA molecular mechanisms and functional perspectives of EMT-inducing lncRNAs in GC progression. Taken together, the statements of this review provided a clinical implementation in identifying lncRNAs as potential therapeutic targets for advanced GC.

miR-1273h-5p suppresses CXCL12 expression and inhibits gastric cancer cell invasion and metastasis.

The aim of this study was to verify the biological function of miR-1273h-5p in gastric cancer (GC) and its underlying mechanisms. The differential expression of microRNAs between GC and tumor-adjacent normal tissues was detected using microarrays, miR-1273h-5p, and chemokine (C-X-C motif) ligand 12 (CXCL12) mRNA, and protein levels were evaluated using polymerase chain reaction and Western blotting methods, cell proliferation, apoptosis, migration, and invasion were determined by CCK-8, flow cytometry, and transwell assay. Compared to tumor-adjacent normal tissue and gastric epithelial mucosa cell line cells, miR-1273h-5p was significantly downregulated in tissues and cells of GC. The overexpression of miR-1273h-5p could inhibit cell proliferation, migration, invasion, and promote cell apoptosis; in contrast, inhibition of miR-1273h-5p expression could reverse this process. Moreover, a significant upregulation of CXCL12 was observed when the miR-1273h-5p was downregulated in GC cells. Additionally, miR-1273h-5p significantly reduces tumor volume and weight. Thus, this study suggests that miR-1273h-5p regulates cell proliferation, migration, invasion, and apoptosis during GC progression by directly binding to CXCL12 mRNA 3'-untranslational regions, which may be a novel diagnostic and therapeutic target in GC.

Involvement of 5-HT-BDNF signaling axis in mediating synergistic antidepressant-like effects after combined administration of two oligosaccharide esters.

Potential mechanisms of depression involving herbal medicines and their specific compounds include elevated 5-HT level and downstream BDNF pathway. To identify potentially new combined therapeutic strategies, 3,6'-disinapoylsucrose (DISS) and tenuifoliside A (TFSA) have been observed to show antidepressant-like effects and its related 5-HT-BDNF pathway. We have tried to investigate whether combined administration of DISS and TFSA exerted more effective in the treatment of depression, as assessed through tail suspension test (TST) and forced swimming test (FST). In addition, we also analyzed the expression of three important proteins, cyclic adenosine monophosphate (cAMP) response element binding (CREB), brain-derived neurotrophic factor (BDNF), and cAMP-regulated transcriptional coactivators (CRTC1), which have been shown to be involved in the regulation of the neurotrophic factors in the hippocampus. The DISS and TFSA separately, both at a dose of 5 mg/kg each, displayed small effect in the immobility time. However, combined treatment of these two in multiple doses exhibited better effect. Moreover, combined treatment of DISS and TFSA also demonstrated enhanced levels of 5-hydroxytryptamine (5-HT), and stronger increase in the phosphorylation levels of CREB, BDNF, and CRTC1 proteins in the hippocampus. Overall, our results indicated that coadministration of these two oligosaccharide esters at low dose may induce more pronounced antidepressant activity, in comparison with individual treatment even at high dosage. Thus, the antidepressant properties of both these compounds can be attributed to their ability to influence 5-HT and BDNF pathway, and thereby suggesting that this combination strategy can definitely act as alternative therapy for depression disorder with very limited side effects.

Efficacy and safety of anti-PD-1/anti-PD-L1 antibody therapy in treatment of advanced gastric cancer or gastroesophageal junction cancer: A meta-analysis.

BACKGROUND: Faced with limited and inadequate treatment options for patients with advanced gastric cancer or gastroesophageal junction cancer (GC/GEJC), researchers have turned toward, with the support of promising clinical trials, anti-PD-1/anti-PD-L1 antibody therapy. But there are also different clinical trial results. To better assess its efficacy and safety, we integrated data from 13 eligible studies for a systematic review and meta-analysis. AIM: To comprehensively evaluate the efficacy and safety of anti-PD-1/anti-PD-L1 antibody therapy in the treatment of advanced GC/GEJC patients. METHODS: PubMed, Web of Science, Cochrane Library ,and EMBASE databases were searched to identify eligible articles with outcomes including objective response rate (ORR), disease control rate (DCR), overall survival (OS), progression-free survival (PFS), and adverse events (AEs) of anti-PD-1/anti-PD-L1 antibody therapy. RESULTS: Our study encompassed a total of 13 trials totaling 1618 patients. The outcomes showed a pooled ORR and DCR of 15% (95% confidence interval [CI]: 14%-18%) and 40% (95%CI: 33%-46%), respectively. The pooled 6-mo OS and PFS were 54% (95%CI: 45%-64%) and 26% (95%CI: 20%-32%), respectively, and the 12-mo OS and PFS were 42% (95%CI: 21%-62%) and 11% (95%CI: 8%-13%), respectively. In addition, the incidence of any-grade AEs and grade ≥ 3 AEs was 64% (95%CI: 54%-73%) and 18% (95%CI: 16%-20%), respectively. Most importantly, PD-L1 positive patients exhibited a higher ORR rate than PD-L1 negative patients (odds ratio = 2.54, 95%CI: 1.56-4.15). CONCLUSION: Anti-PD-1/anti-PD-L1 antibody therapy has shown promising anti-tumor efficacy with manageable AEs in advanced GC/GEJC patients, with PD-L1 overexpressing patients exhibiting a higher ORR. What is more, the clinical efficacy of anti-PD-1/PD-L1 combined with traditional chemotherapy drugs is even better, although the occurrence of AEs still causes considerate concerns.

LncRNA COL1A1-014 is involved in the progression of gastric cancer via regulating CXCL12-CXCR4 axis.

BACKGROUND: The aberrant expression of long noncoding RNAs (lncRNAs) is found in various types of cancers and also showed its association with the occurrence and development of gastric cancer (GC). We found lncRNA COL1A1-014 was frequently upregulated in GC. METHODS: This study investigated COL1A1-014 for its biological function at both cellular and animal levels, using MTT, flow cytometry, colony formation and transwell assays. The expression levels of COL1A1-014 and other genes were detected by RT-PCR and western blot. Luciferase reporter assay was used to detect the potential binding of miR-1273h-5p to COL1A1-014 and CXCL12. RESULTS: We found that COL1A1-014 was frequently upregulated in GC tissues as well as cells. COL1A1-014 increased cell proliferation, colony forming efficiency, migration ability, invasion ability, and weight and volume of grafted tumors, while reduced cell apoptosis. Overexpression of COL1A1-014 increased the mRNA expression of chemokine (CXCmotif) ligand (CXCL12) and high levels of CXCL12 and CXCR4 proteins in GC cells. The levels of miR-1273h-5p showed an inverse correlation with COL1A1-014 and CXCL12 in GC cells transfected with miR-1273h-5p. The mRNAs of wild-type COL1A1-014 and CXCL12 showed reduction in HEK293 cells transfected with miR-1273h-5p. This suggested that COL1A1-014 functions as an efficient miR-1273h-5p sponge and as a competing endogenous RNA (ceRNA) to regulate CXCL12. The proliferative activity of COL1A1-014 on GC cells was blocked by CXCL12-CXCR4 axis inhibitor AMD-3100. CONCLUSIONS: These findings demonstrated that COL1A1-014 play an important regulatory role in GC development by functioning as a ceRNA in regulating the CXCL12/CXCR4 axis via sponging miR-1273h-5p.

Sanguinarine inhibits the proliferation of BGC-823 gastric cancer cells via regulating miR-96-5p/miR-29c-3p and the MAPK/JNK signaling pathway.

Sanguinarine (SAN), a quaternary benzophenanthridine alkaloid extracted from the root of Papaveraceae plants, has shown antitumour effects in multiple cancer cells. However, the therapeutic effects and the underlying mechanisms of SAN in gastric cancer (GC) remain elusive. In this study, the in vitro proliferation inhibition effect of SAN in GC cells was determined using CCK-8 assay, the in vivo antitumor effect of SAN was evaluated in mice with xenotransplanted tumor. The mechanism underlying the antitumor activity of SAN was explored by gene microarray assay and bioinformatics analysis. The levels of differentially expressed miRNAs and target genes were verified by real-time RT-PCR and immunohistochemistry. SAN inhibited the proliferation of BGC-823 cells in a concentration-dependent manner in vitro and in vivo. The miR-96-5p and miR-29c-3p were significantly upregulated in untreated BGC-823 cells and significantly downregulated in SAN treated cells. The mRNA and protein expression of their target gene MAP4K4 were upregulated in SAN treated xenotransplanted tumors, and pMEK4 and pJNK1 proteins in the MAPK/JNK signaling pathway were also upregulated by SAN. These indicate that SAN may inhibit the proliferation of BGC-823 cells through the inhibition of miR-96-5p and miR-29c-3p expression, and subsequent activation of the MAPK/JNK signaling pathway.

Protective Effect of JXT Ethanol Extract on Radiation-Induced Hematopoietic Alteration and Oxidative Stress in the Liver.

This study aims at investigating the radioprotective effect of ethanol extract from Ji-Xue-Teng (JXT, Spatholobus suberectus) on radiation-induced hematopoietic alteration and oxidative stress in the liver. Mice were exposed to a single acute γ-radiation for the whole body at the dose of 6.0 Gy, then subjected to administration of amifostine (45 mg/kg) or JXT (40 g crude drug/kg) once a day for 28 consecutive days, respectively. Bone marrow cells and hemogram including white cells, red cells, platelet counts, and hemoglobin level were examined. The protein expression levels of pJAK2/JAK2, pSTAT5a/STAT5a, pSTAT5b/STAT5b, and Bcl-2 in bone marrow tissue; levels of reactive oxygen species (ROS); and the activity of superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione peroxidase (GSH-Px) in serum and liver tissue were determined. At the end of the experiment, the effect of JXT on cell viability and G-CSF and G-CSFR levels in NFS-60 cells were tested by CCK-8 assay, ELISA, and flow cytometry. The results showed that the mice exposed to γ-radiation alone exhibited a typical hematopoietic syndrome. In contrast, at the end of the 28-day experiment, irradiated mice subjected to oral administration of JXT showed an obvious improvement on blood profile with reduced leucopenia, thrombocytopenia (platelet counts), RBC, and hemoglobin levels, as well as bone marrow cells. The expression of pJAK2/JAK2, pSTAT5a/STAT5a, and Bcl-2 in bone marrow tissue was increased after JXT treatment. The elevation of ROS was due to radiation-induced toxicity, but JXT significantly reduced the ROS level in serum and liver tissue, elevated endogenous SOD and GSH-PX levels, and reduced the MDA level in the liver. JXT could also increase cell viability and G-CSFR level in NFS-60 cells, which was similar to exogenous G-CSF. Our findings suggested that oral administration of JXT effectively facilitated the recovery of hematopoietic bone marrow damage and oxidative stress of the mice induced by γ-radiation.

AG36 Inhibits Human Breast Cancer Cells Proliferation by Promotion of Apoptosis In vitro and In vivo.

AG36 is the biotransformation product of triterpenoid saponin from Ardisia gigantifolia stapf. In this study, the antitumor activity and underlying molecular mechanisms of AG36 against human breast MCF-7, MDA-MB-231, and SK-BR-3 cancer cells were investigated. AG36 inhibited the viability of MCF-7, MDA-MB-231, and SK-BR-3 cells in a dose and time-dependent manner, with an IC50 of approximately 0.73, 18.1, and 23.4 µM at 48 h, respectively. AG36 obviously induced apoptosis and G2/M arrest of all the three breast cancer cells. Moreover, AG36 decreased the protein expression of cycle regulatory proteins cyclin B1 or cyclin D1. In MCF-7 and MDA-MB-231 cells, AG36 strongly increased the cleaved caspase-3 and -8 protein expressions, while in SK-BR-3 cells, AG36 only increased the protein expression of cleaved caspase-3. In all the three breast cancer cells, the ratio of Bax/Bcl-2 and cytosolic cytochrome c content increased significantly compared with control group. The death receptor-related proteins Fas/FasL, TNFR1, and DR5 were detected by Western blot, it showed that different breast cancer cells activated the death receptor-mediated extrinsic caspase-8 pathway through different receptors. In addition, the caspase-8 inhibitor z-IETD-fmk could significantly block AG36-triggered MCF-7 cells apoptosis. The in vivo studies showed that AG36 significantly inhibited the growth of MCF-7 xenograft tumors in BALB/c nude mice comparing with control. In conclusion, AG36 inhibited MCF-7, MDA-MB-231, and SK-BR-3 cells proliferation by the intrinsic mitochondrial and the extrinsic death receptor pathways and AG36 might be a potential breast cancer therapeutic agent.

Gan-Dan-Liang-Yi-Tang alleviates p-chlorophenylalanine-induced insomnia through modification of the serotonergic and immune system.

Gan-Dan-Liang-Yi-Tang (GDLYT) is a Traditional Chinese Medicine that has been historically used for the treatment of insomnia. However, investigations into its pharmacological ingredients and the mechanism underlying its sedative and hypnotic effects remain limited. The present study reported the detailed mechanisms underlying the sedative and hypnotic effects of GDLYT. Kunming mice were administered GDLYT at various sub-hypnotic doses, which underwent sodium pentobarbital treatment test, pentetrazole induced convulsant studies and p-chlorophenylalanine (PCPA) induced insomnia model. Potentiated hypnotic and sedative effects in mice was studied, and also the changes in related neurotransmitter and immune factors were evaluated. The results suggested that GDLYT possessed weak sedative effects on pentetrazole-induced convulsive activity in normal mice at a dose of 1.3 mg/kg, with an increase in sleep onset in subhypnotic dose of sodium pentobarbital-treated mice. GDLYT was also able to alleviate insomnia induced by PCPA in the rodent models, and increased 5-hydroxytryptamine levels in the prefrontal cortex, hippocampus, hypothalamus and corpus striatum of PCPA-treated rats. Furthermore, the hypnotic effects of GDLYT were modified, which allowed for PCPA-induced immune system changes, including increased interleukin (IL)-1ß, tumor necrosis factor-α and IL-2 expression levels. The results of the present study indicated that GDLYT induced sedative and hypnotic bioactivity by regulating serotonergic activity in the central nervous system and immune system.

[Anti-radiation effect and mechanism studies of ethanol extracts from Spatholobus suberectus and its active component catechin].

To study the anti-radiation effect and mechanism of ethanol extracts from Spatholobus suberectus and its active component catechin, ICR mice were exposed to 6Gy irradiation and randomly divided into normal group, model group, positive control group (amifostine, 43.6 mg•kg⁻¹, iv 30 min before irradiation), SSD group (10, 20, 40 g•kg⁻¹) and catechin group (50, 100, 200 mg•kg⁻¹). The mice were administered the appropriate drugs once a day after irradiation for 28 consecutive days. Blood samples were collected from the tail end and the number of peripheral blood cells was counted before irradiation and on day 1, 3, 7, 14, 21 and 28 using a microcell counter. Changes of thymus and spleen index of mice on day 7 were observed. The serum SOD, GSH-Px activity and MDA level were detected by the colorimetric method. The colony forming ability of bone marrow hematopoietic progenitor cells on day 7 was detected by semi solid culture method. The HE staining was adopted to observe the pathological changes. The apoptosis of bone marrow cells was detected by flow cytometry. The expression of cleaved caspase-3 and Bax of bone marrow cells were measured separately by western-blotting and immunohistochemistry method. SSD and catechin can both significantly revert the irradiated-induced decline in hematological parameters (RBC, WBC, PLT, Hb), improve thymus and spleen index, significantly enhance serum SOD and GSH-Px activity and decrease the MDA level. The proliferation and differentiation of hematopoietic progenitor cells in bone marrow were promoted, the apoptosis of bone marrow cells was significantly up-regulated and the expression of cleaved caspase-3 and Bax was significantly reduced in SSD and catechin group. SSD and catechin have significant anti-radiation effect and its mechanism may be related to hematopoietic promoting, antioxidant and anti-apoptotic effects.

Enhanced Antitumor Activity of Cetuximab in Combination with the Jak Inhibitor CYT387 against Non-Small-Cell Lung Cancer with Various Genotypes.

Cetuximab, an epidermal growth factor receptor (EGFR) inhibitor, is effective in the treatment of non-small-cell lung cancers (NSCLCs). However, resistance to EGFR inhibitors limits its effectiveness. In this study, we investigated the effectiveness of Jak-2 inhibitor, CYT387, in combination with cetuximab. Xenograft animal models were administered with cetuximab or CYT387 or their combination. It was observed that NSCLC cells exhibited enormous differences in responses to cetuximab; cell lines were more intrinsically resistant to cetuximab. In resistant cell lines (H1975 and H1650), the efficacy of cetuximab was increased when combined with CYT387, whereas CYT387 alone in low doses exhibited little effect on NSCLC cell proliferation. In addition, the antitumor activity of cetuximab was increased in H1975 resistant model in spite of low efficacy of cetuximab treatment alone in. Jak/STAT signaling was suppressed effectively by the combination of cetuximab and CYT387. In summary, our findings indicated that CYT387 has a potent indirect antitumor activity, and it is also synergistic in its activity in combination with cetuximab against NSCLC tumors, especially with cetuximab intrinsic-resistance tumors. These indications were mediated via Janus kinase (Jak)-signal transducer and transcription (STAT) pathway activator. Our results strongly and consistently supported the potential synergism of CYT387 as Jak inhibitor for anti-NSCLC therapy with EGFR-targeting agents.

AG4, a compound isolated from Radix Ardisiae Gigantifoliae, induces apoptosis in human nasopharyngeal cancer CNE cells through intrinsic and extrinsic apoptosis pathways.

3ß-O-{α-L-Pyran rhamnose-(1→3)-[ß-D-xylopyranose-(1→2)]-ß-D-glucopyranose-(1→4)-[ß-D-lucopyranose-(1→2)]-α-L-pyran arabinose}-cyclamiretin A (AG4) is a saponin component obtained from the Giantleaf Ardisia Rhizome (Rhizoma Ardisiae Gigantifoliae). The present study aimed to investigate the antitumor potential of AG4 and its possible mechanisms in human nasopharyngeal carcinoma cells (CNE). We exposed tumor cells to AG4 to investigate which cell line was the most sensitive to AG4. Cell viability was assessed using the MTT reduction assay, and the effects of AG4 on apoptosis, reactive oxygen species (ROS) content, mitochondrial membrane potential (MMP), and cell cycle were detected using a flow cytometer; the glutathione, superoxide dismutase and malondialdehyde activities were measured using colorimetric methods. The relative expressions of Bax, Bad, Bid, Bcl-2, and Fas mRNA were calculated using the (Equation is included in full-text article.)comparative method by real-time PCR studies and protein was detected by western blotting. AG4 markedly inhibited the growth of CNE cells by decreasing cell proliferation, inducing apoptosis, and blocking the cell cycle in the S phase. The release of caspase-3, caspase-8, and caspase-9 was stimulated by AG4 in CNE, and the decreased proliferation induced by AG4 was blocked by the inhibitor of pan caspase (Z-VAD-FMK). Moreover, the MMP was decreased in AG4-treated cells, and AG4-induced cell apoptosis was accompanied by a rapid and lasting increase in ROS, which was abolished by N-acetyl-L-cysteine (NAC); glutathione, superoxide dismutase, and malondialdehyde were regulated by AG4. AG4 inhibited Bcl-2 mRNA and protein expression and stimulated Bax, Bad, Bid, Fas mRNA, and protein expression in CNE cultures, suggesting an effect at the transcriptional and protein level. In addition, both the FasL inhibitor (AF-016) and the Bcl-2 family inhibitor (GX15-070) could prevent the cell apoptosis induced by AG4. The findings suggested that AG4-induced apoptosis in CNE cells involved a death receptor pathway and a Bcl-2 family-mediated mitochondrial signaling pathway by decreasing the MMPs in an ROS-dependent manner and regulating genes and proteins relative to apoptosis; also, regulation of cell cycles may also play a role in the antitumor mechanism of AG4.

[In vitro studies of Raf-CREB, Akt-CREB, and CaMK II -CREB signal transduction pathway regulated by ginsenosides Rb1, Rg1 and Re].

OBJECTIVE: Effects of ginsenoside Rb1, Rg1 and Re on neurotrophic factor signal transduction pathway using liposome-mediated transfection of eukaryotic cells approach. METHOD: The injury model was established by treating SH-SY5Y cells with 0.6 mmol x L(-1) of corticosterone (CORT) by 24 h. SH-SY5Y cell were pretreated with CORT for 30 min followed by co-treated with 120,60 and 20 micromol x L(-1) of Rb1, 120, 80 and 40 micromol x L(-1) of Rg1 and 120, 80 and 40 micromol x L(-1) of Re for 24 h. Cells viability was determined by Cell Counting Kit (CCK) assay. CREB expressing Luciferase reporter gene was constructed and transfected with plasmid containing hRaf, hcAMP, hAkt, hCaMK gene into human embryonic kidney (HEK293) cells using liposornal transfection reagent lipofection 2000. The expression of CREB before and after it addion of Rb1, Rg1 and Re was examined by Luc assay system and Western blotting. RESULT: Compared with normal control group, CORT significantly decreased the viability of SH-SY5Y cells to 67.21% (P < 0.01). CCK results show that Rb1 (60 micromol x L(-1)), Rg1 (80 micromol x L(-1)) and Re (80 micromol x L(-1)) on SH-SY5Y cells have significant protective effect (P < 0.01). Lucassay and Western blotting results show that the gene and protein levels of CREB increased significantly through the pathway of Raf and Akt with Rb1 and Rg1 (P < 0.01), Re can increase significantly the gene and protein levels of CREB through the pathway of Raf and CaMK II. CONCLUSION: Rb1, Rg1 and Re protects SH-SY5Y cells from CORT-induced damage and the neuroprotective mechanism may be associated with the Raf-CREB, Akt-CREB and CaMK II -CREB pathways.

[Spectroscopic studies on binding of beta-elemene to human serum albumin].

Beta-Elemene is an antitumor drug which is isolated from the traditional Chinese medicinal herb Curcumae Phaeocaulis Rhizoma, it is the main component of elemene which is extracted from the plant and delivered via blood circulation after intravenous injection. The antitumor effect of beta-elemene in vitro and in vivo was definite, and beta-elemene could improve the patient immunity and no sever side effect, drug resistance or bone marrow suppression were found during the clinical studies. And human serum albumin (HSA) is a primary extracellular protein which has a high concentration distribution in blood plasma and has many characteristic physiological functions. Therefore, the binding of beta-elemene to protein may be very important for absorption, distribution, metabolism and elimination. Therefore, the study on the interaction of beta-elemene with drug-carrying protein is very important. In this work, molecular binding of beta-elemene to human serum albumin (HSA) was investigated by using spectrofluorometer. the binding constants suggested that a strong interaction and the formation of a complex between beta-elemene and HSA. This clearly implies that beta-elemene can be stored and removed by the proteins in the body. Furthermore, the fluorescence quenching results showed that the HSA fluorescence was quenched by beta-elemene through static quenching mechanism. Thermodynamic parameters showed that hydrophobic interactions play a role in the binding of beta-elemene to HSA. The negative deltaH(0) and positive deltaS(0) in case of beta-elemene therefore showed that electrostatic attraction play a role in the binding of beta-elemene to HSA.

Antitumor activity of triterpenoid saponin-rich Adisia gigantifolia extract on human breast adenocarcinoma cells in vitro and in vivo.

The aim of this study was to explore whether the ethanolic extract of Ardisia gigantifolia rhizomes (AGB-5), a traditional herbal medicine from China, could affect the proliferation of human breast adenocarcinoma (MCF-7) cells in vitro and to explore the antitumor effects of AGB-5 in BALB/c mice engrafted with MCF-7 cells. The results showed that AGB-5 markedly inhibited the proliferation of MCF-7 cells with an IC50 value of 11.89±1.12 µg/mL, increased the S phase and decreased the G2/M phase without influence on G1 phase. MCF-7 cells treated with AGB-5 presented a dose-dependent increase of apoptosis compared with the control group. AGB-5 also significantly increased the activity of caspase-3 and -9 in a dose-dependent manner in MCF-7 cells. Furthermore, in an in vivo model, AGB-5 reduced tumor volume, brought back the red blood cell (RBC) and white blood cell (WBC) count near to normal value, enhanced superoxide dismutase and catalase level of MCF-7 bearing mice. This is the first study to verify the antitumor activity of A. gigantifolia in vivo. The results suggest that AGB-5 may have potential beneficial effects against human breast adenocarcinoma.

Effect of Tenuifoliside A isolated from Polygala tenuifolia on the ERK and PI3K pathways in C6 glioma cells.

Tenuifoliside A (TFSA) is a bioactive oligosaccharide ester component of Polygala tenuifolia Wild, a traditional Chinese medicine which was used to manage mental disorders effectively. The neuroprotective and anti-apoptotic effects of TFSA have been demonstrated in our previous studies. The present work was designed to study the molecular mechanism of TFSA on promoting the viability of rat glioma cells C6. We exposed C6 cells to TFSA (or combined with ERK, PI3K and TrkB inhibitors) to examine the effects of TFSA on the cell viability and the expression and phosphorylation of key proteins in the ERK and PI3K signaling pathway. TFSA increased levels of phospho-ERK and phospho-Akt, enhanced release of BDNF, which were blocked by ERK and PI3K inhibitors, respectively (U0126 and LY294002). Moreover, the TFSA caused the enhanced phosphorylation of cyclic AMP response element binding protein (CREB) at Ser133 site, the effect was revoked by U0126, LY294002 and K252a. Furthermore, when C6 cells were pretreated with K252a, a TrkB antagonist, known to significantly inhibit the activity of brain-derived neurotrophic factor (BDNF), blocked the levels of phospho-ERK, phospho-Akt and phosphor-CREB. Taking these results together, we suggested the neuroprotection of TFSA might be mediated through BDNF/TrkB-ERK/PI3K-CREB signaling pathway in C6 glioma cells.

Effects of (20S*,24R*)-epoxy-9,19-cyclolanstane-3ß,12ß,16ß,25-pentaol-3-O-ß-D-xylopyranoside extracted from rhizoma Beesia on immunoregulation and anti-inflammation.

(20S*,24R*)-epoxy-9,19-cyclolanstane-3ß,12ß,16ß,25-pentaol-3-O-ß-D-xylopyranoside (BC1) is a kind of natural bioactive substance extracted from Beesia calthaefolia (Maxim.)Ulbr. This study was designed to evaluate the effects of BC1 on the proliferation of lymphocytes, phagocytosis of peritoneal macrophage, and cytokine secretion, such as tumor necrosis factor (TNF)-α and interleukin (IL)-1ß, and the foot pad thickness index, which is beneficial for understanding the mechanism of BC1 on immunoregulation and anti-inflammation and also will benefit our further research. The proliferation of splenic lymphocyte induced by mitogen (concanavalin A or lipopolysaccharide (LPS)) was detected using the cell counting kit assay. The neutral red phagocytic test of macrophages was determined by colorimetric method. The gene and protein expressions of TNF-α and IL-1ß were measured by real time RT-PCR and ELISA in serum, spleen, and lymphocytes, respectively. In vitro, our present study has shown that BC1 (31.25-250 µg/ml) could inhibit the proliferation of splenic lymphocyte and phagocytosis of macrophages, and inhibit the increased production of TNF-α and IL-1ß in protein and gene levels. In mice, LPS could increase the gene and protein expressions of TNF-α and IL-1ß, respectively, but BC1 (12.5-50 µg/kg) could recover the increased gene and protein expressions of TNF-α and IL-1ß induced by LPS in the spleen and serum of mice. Treatment of arthritic rats with BC1 (1.5 mg/kg body weight) resulted in a significant reduction in foot pad thickness index and serum TNF-α level comparable to the indomethacin-treated arthritic rats, proving its anti-inflammatory effect. Thus, the function of immunoregulation of BC1 may be accomplished through modulating the gene and protein expressions of TNF-α and IL-1ß.

A new triterpenoid saponin from Ardisia gigantifolia.

A new triterpenoid saponin, named 3-O-ß-d-glucopyranosyl-(1 â†’ 3)-ß-d-xylopyranosyl-(1 â†’ 2)-[α-l-rhamnopyranosyl-(1 â†’ 3)]-ß-d-glucopyranosyl-(1 â†’ 4)-[ß-d-glucopyranosyl-(1 â†’ 2)]-α-l-arabinopyranosyl-3ß,16α,28,30-tetrahydroxy-olean-12-ene (1), along with four known triterpenoids (2-5), was isolated from the rhizomes of Ardisia gigantifolia. Their structures were elucidated by spectroscopic methods. Compounds 1-4 showed cytotoxic activity against Hela, EJ, BCG, and HepG-2 cell lines. The percentage of early apoptotic cells after treatment with 1 was significantly increased compared with control cells (p < 0.05).

Sanguinarine inhibits vascular endothelial growth factor release by generation of reactive oxygen species in MCF-7 human mammary adenocarcinoma cells.

The inhibitory action and the possible mechanism of anticancer compound Sanguinarine (SAN) on vascular endothelial growth factor (VEGF) in human mammary adenocarcinoma cells MCF-7 were evaluated in this study. We exposed MCF-7 to SAN for 24 h, then cell viability was assessed by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay. Human VEGF was measured using a paired antibody quantitative ELISA kit, relative expression of VEGF mRNA was calculated using the real-time PCR studies, and the effect of SAN on the reactive oxygen species (ROS) level was detected by the flow cytometer. Treatment with SAN remarkably inhibited growth of MCF-7 cells and induced cell apoptosis. We found that VEGF release was stimulated by subtoxic concentrations of SAN and inhibited by high dose of SAN, SAN-evoked VEGF release was mimicked by low concentration of H2O2, and SAN-regulated VEGF inhibition was accompanied by increasing of ROS; these changes were abolished by antioxidant. High concentration of SAN inhibited VEGF mRNA expression in MCF-7 cultures, suggesting an effect at transcriptional level, and was also abolished by antioxidant. The present findings indicated that the regulation of VEGF expression and release from MCF-7 cells were possibly through reactive oxygen species evoked by SAN.

JR6, a new compound isolated from Justicia procumbens, induces apoptosis in human bladder cancer EJ cells through caspase-dependent pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Numerous efforts have been conducted in searching for effective agents against cancer, in particular from herbal medicines. Justicia procumbens is a traditional herbal remedy which was produced in the south-western and southern provinces of China and Taiwan province used to treat fever, pain, and cancer. Here, we identified a new compound 6'-hydroxy justicidin A (JR6) from Justicia procumbens, which showed obvious anti-cancer effects. MATERIALS AND METHODS: The cytotoxicity activity was assayed using MTT and SRB. Intracellular ROS visualization and quantification were acquired by using a laser scanning confocal microscopy. Apoptosis was measured using a propidium iodide (PI) apoptosis detection kit by flow cytometry. Activation of caspases (caspase-3, caspase-8, and caspase-9) was evaluated respectively using GloMax luminescence detector and Caspase-Glo 3,8,9 assay kits. Loss of mitochondrial membrane potential was observed by microscopy using JC-1 dye. Quantitative real-time PCR analysis was employed to detect the expression of protein associated with cell death. RESULTS: JR6 remarkably inhibited growth in human bladder cancer EJ cells by decreasing cell proliferation, reduced the SOD activity, increased the content of reactive oxygen species (ROS), and induced apoptosis. Activation of caspase-8, caspase-9, and the subsequent activation of caspase-3 indicated that JR6 may be inducing intrinsic and extrinsic apoptosis pathways. Caspase-3, caspase-8, and caspase-9 inhibition rendered this extract ineffective, thus JR6-induced apoptosis is caspase-dependent. JR6 also disrupted the mitochondrial membrane potential (Δψm) and unregulated the Bax and p53 expressions in EJ cells. CONCLUSION: These observations suggest that JR6 induce apoptosis through caspase-dependent pathway in human bladder cancer EJ cells, emphasizing the importance of this traditional medicine and thus presents a potential novel alternative to bladder cancer therapy.