Suppression of hypoxia and inflammatory pathways by Phyllanthus niruri extract inhibits angiogenesis in DMBA-induced breast cancer mice.

BACKGROUND AND PURPOSE: Angiogenesis has been one of the hallmarks of cancer. In recent years, Phyllanthus niruri extract (PNE) was reported to inhibit angiogenesis by decreasing the levels of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α) in breast cancer. However, the experimental results were confirmed in cancer cell lines only, whereas the anti-angiogenic activity in animal models has not been demonstrated. In this study, we tried to examine the anti-angiogenic activity of PNE on BALB/c strain mice models that were induced for breast cancer using the carcinogenic substance 7,12- dimethylbenz[a]anthracene (DMBA). EXPERIMENTAL APPROACH: Experimental animals were divided into five different groups; vehicle, DMBA, PNE 500 mg/kg, PNE 1000 mg/kg; and PNE 2000 mg/kg. Mammary carcinogenesis was induced using a subcutaneous injection of 15 mg/kg of DMBA for 12 weeks. Afterward, oral PNE treatment was given for the following 5 weeks. VEGFA and HIF-1α were observed using immunohistochemistry. Endothelial cell markers CD31, CD146, and CD34 were observed using the fluorescent immunohistochemistry method. The levels of interleukin-6 (IL-6), IL-17, and C-X-C motif chemokine (CXCL12) were measured using flow cytometry. FINDINGS/RESULTS: The survival analysis indicated that PNE increased the survival rate of mice (P = 0.043, log-rank test) at all doses. The PNE treatment decreased the immunoreactive score of angiogenic factors (VEGF and HIF-1α), as well as the endothelial cell markers (CD31, CD146, and CD34). The PNE- treated groups also decreased the levels of inflammatory cytokines (IL-6, IL-17, and CXCL12) at all doses. CONCLUSION AND IMPLICATIONS: This finding suggests that PNE may inhibit the progression of angiogenesis in breast cancer mice by targeting the hypoxia and inflammatory pathways.

Elicited soybean extract attenuates proinflammatory cytokines expression by modulating TLR3/TLR4 activation in high-fat, high-fructose diet mice.

BACKGROUND: The high-fat, high-fructose diet (HFFD) provokes overnutrition and inflammation directly, mainly through Toll-like receptors (TLRs). Soybean (Glycine max L.) contains isoflavone that can be transformed into glyceollin by microbial and physical stimuli. Glyceollin possesses many beneficial effects on health. OBJECTIVE: This study evaluates the beneficial effect of soybean extract elicited by Saccharomyces cerevisiae and light (ESE) on dendritic cells (DCs) profile and naïve T cells in HFFD mice. MATERIALS AND METHODS: Female Balb/C mice were fed with HFFD for 24 weeks then orally administered with simvastatin 2.8 mg/kg BW or ESE 78, 104, and 130 mg/kg BW at the last four weeks. The expression of splenic CD11c+TLR3+, CD11c+TLR4+, NFκB+, CD11c+IL-17+, CD11c+TNF-α+, CD4+CD62L+, and CD8+CD62L+ subsets was measured by flow cytometry. The molecular docking has been measured using Pyrx 0.8, displayed in PyMol and Biovia Discovery Studio. RESULT: HFFD significantly increased CD11c+TLR3+, CD11c+TLR4+, NFκB+, CD11c+IL-17+, CD11c+TNF-α+ expression and decreased CD4+CD62L+ and CD8+CD62L+ (p < 0.05) compared to normal diet (ND) groups. ESE reduced CD11c+TLR3+, CD11c+TLR4+, thereby decreasing NFκB+, as well as decreased the CD11c+IL-17+, CD11c+TNF-α+, and restores CD4+CD62L+ and CD8+CD62L+ subsets in HFFD mice. Glyceollin II exhibited the best binding affinity with an average energy of -7.3 kcal/mol to TLR3 and -7.9 kcal/mol to TLR4. CONCLUSION: The bioactive compound in ESE act synergistically to modulate TLR3/TLR4 activation, reduced NFκB, IL-17, and TNF-α, and restores naïve T cells expression in HFFD mice. ESE was a favorable candidate to mitigate chronic inflammation.

Flow Cytometric Analysis of Pro-inflammatory Cytokine Production in Hyperglycemic Mouse Model.

OBJECTIVE: The sharp increase of pro-inflammatory cytokine becomes a severe problem in some degenerative diseases. This study aimed to investigate the effect of Cherry Leaf Extract (CLE) on pro-inflammatory cytokine production in hyperglycaemic mouse model. A recent patent mentioned that Muntinga calabura is a flavonoid-containing plant which is capable of reinforcing the stomach, appetizing, helping digestion (CN 201611138374), and improving stomach distention (CN106071017A). The efficacy of this miracle plant has inspired this experiment. METHODS: In this study, we induced hyperglycemia with high-fat diet and 20% fructose drink. Hyperglycemic mice were orally administered with CLE with the doses 420, 700, and 28000 mg/kg BW for two weeks. After treatment, the pro-inflammatory molecule was analyzed by flow cytometry from splenic cells. RESULTS: This study showed that CLE decreased the production of pro-inflammatory molecules such as IL-6, NF-κB, TNF-α, and IFN γ. In hyperglycemic mice, we found CD68+IL-6+, NK1.1+IL- 6+, B220+NFκB+, CD4+TNF-α+, and CD4+IFN γ + increased significantly (p < 0.05) compared to control mice without manipulation. After treatment with 420, 700, or 2800 mg/kg BW of CLE, the pro-inflammatory molecules decreased significantly (p < 0.05), and reached a normal physiology level. CONCLUSION: CLE can suppress pro-inflammatory cytokines in hyperglycemic mouse model, suggesting this medicinal herb may be beneficial as a potential strategy for future therapeutic interventions in degenerative diseases or a disease involving cell activation.

1,4-butanediyl-bismethanethiosulfonate (BMTS) induces apoptosis through reactive oxygen species-mediated mechanism.

Although methane sulfonate compounds are widely used for the protein modification for their selectivity of thiol groups in proteins, their intracellular signaling events have not yet been clearly documented. This study demonstrated the methane sulfonate chemical 1,4-butanediyl-bismethanethiosulfonate (BMTS)-induced cascades of signals that ultimately led to apoptosis of Jurkat cells. BMTS induced apoptosis through fragmentation of DNA, activation of caspase-9 and caspase-3, and downregulation of Bcl-2 protein with reduction of mitochondrial membrane potential. Moreover, BMTS intensely and transiently induced intracellular reactive oxygen species (ROS) production and ROS produced by BMTS was mediated through mitochondria. We also found that a reducing agent dithiothreitol (DTT) and an anti-oxidant N-acetyl cysteine (NAC) inhibited BMTS-mediated caspase-9 and -3 activation, ROS production and induction of Annexin V/propidium iodide double positive cells, suggesting the involvement of ROS in the apoptosis process. Therefore, this study further extends our understanding on the basic mechanism of redox-linked apoptosis induced by sulfhydryl-reactive chemicals.

Protective effect of hyperpigmented skin on UV-mediated cutaneous cancer development.

Recently, we crossed an original haired RET-transgenic mouse of line 242 with a hairless mouse and established a hairless RET-(HL/RET)-transgenic mouse line (242-hr/hr) with hyperpigmented skin but no tumors. In this study, we examined the effect of hyperpigmented skin in HL/RET-transgenic mice on UV irradiation-mediated cutaneous cancer development. UV irradiation to this mouse line never induced melanoma despite the presence of melanoma-inducible transgenic RET oncogenes. On the contrary, the hyperpigmented skin efficiently protected UV-mediated squamous carcinoma development in the skin. Probably underlying this result, hyperpigmentation protected the skin from damage and blocked the accompanying signal transduction for tyrosine phosphorylation of multiple cellular proteins and activation/phosphorylation of extracellular signal-regulated, c-Jun N-terminal, and p38 kinases. Thus, we demonstrated hyperpigmentation-mediated in vivo protection against UV irradiation-induced skin cancer.

mRNA expression of inducible nitric oxide synthase, endothelial nitric oxide synthase and vascular endothelial growth factor in esophageal mucosa biopsy specimens from patients with reflux esophagitis.

BACKGROUND/AIMS: Nitric oxide (NO) production is elevated in the intestine and may contribute to intestinal injury during inflammation. However, how the expression of inducible NO synthase (iNOS) mRNA and endothelial NO synthase (eNOS) mRNA in the esophageal mucosa contribute to mucosal damage caused by reflux esophagitis remains unknown. Since vascular endothelial growth factor (VEGF) exerts its action on microcirculation, contributing to angiogenesis and inflammation, we examined the role of VEGF together with iNOS and eNOS on development of reflux esophagitis. METHODOLOGY: The mRNA expression levels of iNOS, eNOS and VEGF were measured in biopsy specimens from 25 patients with reflux esophagitis, using TaqMan PCR and reverse transcription PCR. RESULTS: The expression of iNOS mRNA in the esophageal mucosa increased parallel to the severity of the esophagitis. There were no significant differences between both eNOS and VEGF mRNA expression levels and the severity of the esophagitis. The existence of gastric mucosal atrophy, hiatus hernia, therapy and Helicobacter pylori infection did not affect the levels of mRNA expression. CONCLUSIONS: The accumulation of NO, produced by iNOS, was considered to be related to the exacerbation of reflux esophagitis. Therapeutic intervention that reduces NO production may thus be of use in preventing development of esophageal mucosal injury in patients with reflux esophagitis.

[A case of advanced gastric cancer with peritoneal dissemination effectively treated by combined chemotherapy of paclitaxel (TXL) and TS-1].

The patient was a 49-year-old woman. Chemotherapy was conducted combining paclitaxel (TXL) and TS-1 under the diagnosis of non-resectable advanced gastric cancer with peritoneal dissemination. The administration schedule was as follows: 60 mg/m2 of TXL on days 1, 8 and 15 intravenously and 120 mg/day of TS-1/on days 1 5, 8-12, and 15-19 orally. One cycle lasted for 5 weeks. Grade 1 peripheral neuropathy was noted, but no other serious adverse reaction occurred. Ascites fluid was reduced after completion of the 1st cycle, and the therapeutic efficacy was rated as PR. Abdominal fullness was relieved shortly after starting the treatment, making it possible to conduct treatment on an ambulatory basis in the 2nd and subsequent cycles. At present, 6 months after starting chemotherapy, there is no evidence of relapse or adverse reactions that require intervention. Chemotherapy is being continued on an ambulatory basis. Combination of TXL and TS-1 is expected to show good therapeutic efficacy and improve patients' QOL in patients with gastric cancer associated with peritoneal dissemination.

Paeoniflorin induces apoptosis of lymphocytes through a redox-linked mechanism.

Paeoniflorin (PF), isolated from paeony root, has been used as a herbal medicine for more than 1,200 years in China, Korea, and Japan for its anti-allergic, anti-inflammatory, and immunoregulatory effects. In this study, we found that PF induces apoptosis in both murine T-lineage cells and human T-cell leukemia Jurkat cells. This apoptosis was mediated through the reduction of mitochondrial membrane potential, activation of caspase, and fragmentation of DNA. Interestingly, PF induced generation of reactive oxygen species (ROS) and a reducing agent, dithiothreitol (DTT), and a ROS scavenger, N-acetyl cysteine (NAC), successfully attenuated the PF-induced apoptosis. Additionally, PF induced the phosphorylation of three mitogen-activated protein (MAP) family kinases, extracellular signal-regulated kinase, c-Jun amino-terminal kinase (JNK), and p38 MAP kinase. Curcumin, an anti-oxidant and JNK inhibitor, inhibited PF-induced apoptosis, suggesting the possible involvement of curcumin-sensitive JNK or other redox-sensitive elements in PF-induced apoptosis. These results partially explain the action mechanism of PF-containing paeony root as a herbal medicine.

Caspase activation is accelerated by the inhibition of arsenite-induced, membrane rafts-dependent Akt activation.

Renewed interest in arsenic has been shown recently due to its dual nature of being a potent toxin and a drug for treatment of acute promyelocytic leukemia (APL) because of its ability to trigger caspase activation. Here, we found that sodium arsenite (NaAsO(2)) also triggers the signal for activation of Akt and downstream glycogen synthase 3beta (GSK3beta). Such Akt/GSK3beta activation was abrogated completely by wortmannin, an inhibitor of PI-3 kinase, and greatly by pertussis toxin, a G-protein inhibitor. Arsenite-induced Akt phosphorylation also was inhibited by sequestrating membrane cholesterol with beta cyclodextrin. Reducing reagents/reactive oxygen species (ROS) scavengers reduced arsenite-induced Akt phosphorylation and beta cyclodextrin reduced arsenite-mediated ROS production, suggesting that arsenite-induced G-protein/Akt/GSK3beta pathway is membrane raft dependent and redox linked. We also found that a combination of a low concentration (1 microM) of arsenite and wortmannin triggers the signal for caspase activation, whereas neither of these elements alone did so. These results suggested that selective blockade of the arsenite-provoked PI-3 kinase/Akt pathway can promote the arsenite-triggered pathway for caspase activation, and this may open a new study area for wider applications of arsenic as a drug for treating various kinds of leukemia.

Clinical characteristics of Japanese reflux esophagitis patients as determined by Los Angeles classification.

BACKGROUND: Recent studies have shown that the number of patients with reflux esophagitis is increasing in Japan, but the prevalence and risk factors associated with reflux esophagitis in Japanese patients are not well defined. METHODS: By using all endoscopic records in the Katta General Hospital from April through to September 1999, we identified 392 patients. We examined the Los Angeles classification, peptic ulcer, gastric mucosal atrophy, hiatal hernia and other medical variable factors for their contribution to esophagitis in the patients. RESULTS: Patients (13.8%) were diagnosed as having reflux esophagitis with a mucosal break. In a multivariate analysis, reflux esophagitis was associated with hiatal hernia (odds ratio (OR) 2.276, 95% confidence interval (CI) 1.164-4.450), with patients over 65 years of age (OR 2.521, 95% CI 1.238-5.134) and the open type of gastric mucosal atrophy (OR 0.420, 95% CI 0.225-0.785). There was no significant difference between esophagitis and Helicobacter pylori infection and peptic ulcer. CONCLUSIONS: We observed that age, hiatal hernia and a lower rate of gastric mucosal atrophy were associated with the proportion of mucosal breaks accompanying esophagitis.