Phenethyl isothiocyanate inhibits hypoxia-induced accumulation of HIF-1α and VEGF expression in human glioma cells

Phenethyl isothiocyanate (PEITC), a natural dietary isothiocyanate, inhibits angiogenesis but the molecular mechanisms that underlie this effect are not known. In this study, under hypoxic conditions (1% O₂), we examined the effect of PEITC on the intracellular level of the hypoxia inducible factor (HIF-1α) and extracellular level of the vascular endothelial growth factor (VEGF) in a variety of human cancer cell lines. Surprisingly, we observed that PEITC suppressed the HIF-1α accumulation during hypoxia in human glioma U87, human prostate cancer DU145, colon cancer HCT116, liver cancer HepG2, and breast cancer SkBr3 cells. PEITC treatment also significantly reduced the hypoxia-induced secretion of VEGF. Suppression of HIF-1α accumulation during treatment with PEITC in hypoxia was related to PI3K and MAPK pathways. Taken together, these results suggest that PEITC inhibits the HIF-1α expression through inhibiting the PI3K and MAPK signalling pathway and provide a new insight into a potential mechanism of the anticancer properties of PEITC.     

Phenethyl isothiocyanate sensitizes human cervical cancer cells to apoptosis induced by cisplatin

Scope: Naturally‐occurring chemopreventive agent phenethyl isothiocyanate (PEITC), derived primarily from watercress, has been shown to inhibit cell growth and induce apoptosis in cancer cells. In this study, we examined the potential of PEITC in enhancing cisplatin‐induced apoptosis in cervical cancer cells and its mechanisms. Methods and results: HeLa cells were exposed to PEITC, cisplatin or both. Pretreatment of cells with PEITC strongly enhanced cisplatin‐induced cytotoxicity. PEITC activated the mitogen‐activated protein kinases, including c‐Jun N‐terminal kinase (JNK), extracellular signal‐related kinase (ERK), and p38. Caspase‐3 activity assay demonstrated that the synergistic induction of apoptosis was significantly attenuated by MEK1/2 inhibitor U0126, but not by JNK or p38 inhibitor, suggesting that ERK activation is responsible for the synergistic effect. We found that NF‐κB signaling pathway is not involved in the synergistic effect. Sulforaphane and benzyl isothiocyanate, two other members of the isothiocyanate family, also sensitize HeLa cells to apoptosis induced by cisplatin. Furthermore, we found that the synergistic effect was also observed in cervical cancer C33A and breast cancer MCF‐7 cells but not in normal mammary epithelial MCF‐10A cells. Finally, we demonstrated that Noxa induction was associated with apoptosis induced by PEITC plus cisplatin. Conclusion: Taken together, this study shows that PEITC can sensitize cancer cells to apoptosis induced by cisplatin and this effect is mediated through ERK activation, suggesting the potential of PEITC to be used as an adjuvant with cisplatin in combination therapeutic treatments.     

Phenethyl isothiocyanate decreases thymic stromal lymphopoietin‐induced inflammatory reactions in mast cells

Phenethyl isothiocyanate (PEITC) has been reported to have anti‐inflammatory and anti‐carcinogenic properties. However, the anti‐inflammatory effects of PEITC on thymic stromal lymphopoietin (TSLP)‐induced inflammatory responses are uncertain. This study evaluates pharmacological activities of PEITC on inflammatory reactions in TSLP‐stimulated mast cells. Human mast cell line HMC‐1 was treated with PEITC (0.04, 0.4, and 4 µM) and subjected to inflammation by TSLP. Our results showed that PEITC significantly attenuated IL‐13 and TNF‐α levels increased by TSLP in HMC‐1. PEITC significantly decreased TSLP‐promoted HMC‐1 proliferation and Ki67 mRNA expression. Protein levels of MDM2 and pSTAT6 increased by TSLP were significantly suppressed by PEITC in HMC‐1. In addition, PEITC significantly enhanced protein levels of cleaved poly ADP‐ribose polymerase and p53 decreased by TSLP. Based on the effects of PEITC on inflammation and proliferation in this study, it is possible that PEITC is a potential candidate to treat mast cells‐mediated inflammatory disorders.

Practical applications

This report provides strong evidence that Phenethyl isothiocyanate (PEITC) which is a dietary constituent derived from cruciferous vegetables, may be considered an alternative agent for treatment of mast cells‐mediated inflammatory disorders.     

Phytate hydrolysate induces circumferential F‐actin ring formation at cell–cell contacts by a Rho‐associated kinase‐dependent mechanism in colorectal cancer HT‐29 cells

Phytate (inositol hexa‐phosphate or IP6) possessing anticancer activity is hydrolyzed by phytase in intestinal microbes and the metabolites are distributed throughout the colon. Cellular circumferential F‐actin rings, which are involved in cell polarity and structure, are lost early during tumorigenesis. We investigated F‐actin ring formation by the phytate hydrolysate in colorectal cancer HT‐29 cells to explore the novel mechanisms underlying the phytate‐mediated anticancer function. The phytate hydrolysate, but not inositol or phytate, induced F‐actin ring formation with a peak at 10 min in the cells and was associated with phosphorylation of myosin regulatory light chain. F‐actin ring formation and myosin regulatory light chain phosphorylation by the phytate hydrolysate were suppressed by inhibitors of Rho‐associated kinase (ROCK), Janus kinase (JAK), c‐Jun N‐terminal kinase (JNK), and protein kinase Cδ (PKCδ). Activation of ROCK and JAK, but not JNK or PKCδ, was observed at 10 min and/or earlier after stimulation with the phytate hydrolysate. Altogether, the phytate hydrolysate induces circumferential F‐actin ring formation through a ROCK‐dependent myosin II activation in the HT‐29 cells, which requires JAK activation and basal activities of JNK and PKC. Hydrolysis products of phytate in the intestine may contribute to anticancer function of phytate.     

雌马酚及大豆活性成分对人前列腺癌细胞DU145的生长影响

目的:观察大豆异黄酮代谢产物雌马酚对人前列腺癌细胞DU145的生长影响,并在生理浓度下比较其与大豆异黄酮主要成分金雀异黄素、大豆素的体外抗增殖活性。方法:采用MTT法检测不同浓度的雌马酚、金雀异黄素及大豆素对DU145细胞增殖的影响,并以流式细胞术分析细胞周期分布情况。结果:雌马酚在1×10-6～1×10-5mol/L表现为抑制作用,金雀异黄素在1×10-7～1×10-6mol/L时表现为促增殖效应,达到5×10-5mol/L时出现抑制作用,大豆素在生理浓度下则主要表现为促增殖效应;雌马酚在1×10-6mol/L时使细胞在G0/G1期阻滞,在5×10-6～1×10-5mol/L时使S期及G2/M期细胞增多,金雀异黄素在5×10-6mol/L出现G2/M期阻滞,大豆素使S期及G2/M期细胞增多。结论:雌马酚在体外具有一定的抗前列腺癌活性;在生理浓度下,其抗癌活性可能强于金雀异黄素及大豆素。     

Phenethyl isothiocyanate suppresses nitric oxide production via inhibition of phosphoinositide 3‐kinase/Akt‐induced IFN‐γ secretion in LPS‐activated peritoneal macrophages

Phenethyl isothiocyanate (PEITC), a constituent of many cruciferous vegetables, is well known to have versatile physiological activities, including chemopreventive effects. On the other hand, its anti‐inflammatory effects are poorly reported. Nitric oxide (NO) is associated with a wide variety of inflammatory diseases. In this study, we investigated the effects of PEITC on NO production in LPS‐activated peritoneal macrophages from ICR mice. The signaling pathway of LPS‐induced NO production was examined using neutralizing antibodies [anti‐interferon (IFN)‐γ and anti‐interleukin (IL‐12)] and specific protein kinase inhibitors, as well as others. The activity of PEITC toward NOx production was assessed in mice that received LPS via intraperitoneal administration. The neutralizing antibody of anti‐IFN‐γ, but not anti‐IL‐12, suppressed LPS‐induced NO production by 90%. LY294002, a specific inhibitor of phosphoinositide‐3‐kinase, suppressed Akt and IFN‐γ mRNA expression up‐regulated by LPS, whereas PEITC exhibited a similar inhibition profile. Furthermore, oral administration of PEITC significantly suppressed the serum concentration of NOx in ICR mice. Our results suggest that PEITC suppresses LPS‐induced NO production via inhibition of Akt activation and the resultant decrease in expression of IFN‐γ. This is one of the first reports to demonstrate a marked anti‐inflammatory effect of PEITC following its oral administration.     

可溶性青梅果胶的表征及其对脂多糖诱导的RAW246.7巨噬细胞炎症的抑制作用

本研究从青梅果汁中提取可溶性果胶（WSP）,通过高效液相色谱（HPLC）、高效凝胶色谱（HPGFC）、傅里叶变换红外光谱（FT-IR）、扫描电镜（SEM）及X-射线衍射（XRD）等对其组成和结构进行了表征,并用酶联免疫法（ELISA）对脂多糖（LPS）诱导的RAW246.7细胞的抗炎活性进行评价。结果表明,WSP的酯化度为18.89%,属于低酯果胶,其半乳糖醛酸（GalA）含量为69.72%,主要中性多糖是半乳糖（Gal）（18.36%）,重均分子量（Mw）为353.73 kDa,属于非晶型结构。WSP可显著抑制LPS诱导的RAW264.7细胞中肿瘤坏死因子α（TNF-α）(P<0.05)、干扰素-γ（IFN-γ）(P<0.05)、白介素6（IL-6）（P<0.001）的释放量,增加白介素10（IL-10）(P<0.01）的分泌,起到抗炎作用。为了探究其抗炎机理,通过实时荧光定量聚合酶链式反应法(Real-time qPCR)和蛋白印迹法（Western blot）研究了WSP对Janus 激酶/信号转导和转录激活因子信号通路（JAK/STAT信号通路）的影响。与LPS模型组相比,WSP显著上调了与炎症密切相关的JAK1（P<0.01）,JAK2（P<0.05）,JAK3（P<0.001）和STAT1（P<0.001）,STAT2（P<0.001）和STAT3（P<0.05）的mRNA表达量,且使JAK1和STAT3的磷酸化水平显著降低（P<0.05）,该结果表明WSP可通过干预JAK/STAT信号通路,抑制与炎症信号相关的JAK和STAT家族的磷酸化水平来缓解LPS诱导的RAW264.7细胞炎症。本研究可为评价青梅果的健康促进作用和提高青梅果加工附加值提供参考。     

Nutritional Sources and Anticancer Potential of Phenethyl Isothiocyanate: Molecular Mechanisms and Therapeutic Insights

Phenethyl isothiocyanate (PEITC), a compound derived from cruciferous vegetables, has garnered attention for its anticancer properties. This review synthesizes existing research on PEITC, focusing on its mechanisms of action in combatting cancer. PEITC has been found to be effective against various cancer types, such as breast, prostate, lung, colon, and pancreatic cancers. Its anticancer activities are mediated through several mechanisms, including the induction of apoptosis (programmed cell death), inhibition of cell proliferation, suppression of angiogenesis (formation of new blood vessels that feed tumors), and reduction of metastasis (spread of cancer cells to new areas). PEITC targets crucial cellular signaling pathways involved in cancer progression, notably the Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-κB), Protein Kinase B (Akt), and Mitogen-Activated Protein Kinase (MAPK) pathways. These findings suggest PEITC's potential as a therapeutic agent against cancer. However, further research is necessary to determine the optimal dosage, understand its bioavailability, and assess potential side effects. This will be crucial for developing PEITC-based treatments that are both effective and safe for clinical use in cancer therapy.     

乌贼墨多肽诱导人前列腺癌DU-145细胞凋亡的机制研究

本文探讨了乌贼墨多肽(SHP)诱导DU-145细胞凋亡机制。采用CCK-8法检测SHP对DU-145细胞增殖的影响;采用HE染色和AO/EB荧光染色观察DU-145细胞的形态学的变化;采用流式细胞术检测细胞早期凋亡率;并通过Western Blotting检测细胞中p53、Bcl-2、Bax、Caspase-3、VEGF的蛋白表达变化。结果表明,SHP对DU-145细胞的增殖具有明显的抑制作用且呈现剂量和时间依赖性;SHP作用后的DU-145细胞出现凋亡的形态学特征;流式细胞术结果显示,随着SHP浓度和作用时间的增加,DU-145细胞的早期凋亡率从12.25%增加到34.20%;Western Blotting结果显示,当SHP作用24 h后,VEGF、Bcl-2蛋白表达量降低,p53、Bax、Caspase-3蛋白表达量增加。综上可知,SHP能够诱导DU-145细胞凋亡,其机制有可能是通过激活抑癌基因p53,下调Bcl-2/Bax比例;下调VEGF,激活凋亡蛋白酶Caspase-3,诱发凋亡级联反应来实现的。     

Quercetin Suppresses Intracellular ROS Formation,MMP Activation,and Cell Motility in Human Fibrosarcoma Cells

Cell metastasis is a major cause of death from cancer and can arise from excessive levels of oxidative stress. The objective of this study was to investigate whether the natural flavonoid quercetin can inhibit matrix metalloproteinase (MMP)‐2 and ‐9 activities through the attenuation of reactive oxygen species (ROS) formation, an event expected to lead to the inhibition of cell motility. To induce sustained ROS formation, cells were treated with phenazine methosulfate (PMS; 1 μM). Noncytotoxic concentrations of quercetin inhibited PMS‐induced increases in cell motility in HT1080 human fibrosarcoma (HT1080) cells. While nearly 100% of cells were observed to migrate after 24 h of PMS treatment, quercetin significantly (P < 0.01) suppressed this effect. We also found that quercetin, up to 10 μg/mL, attenuated PMS‐induced MMP‐2 activation. We then investigated whether the decreased levels of MMP‐2 activation could be attributable to lower levels of ROS formation by quercetin. We found that quercetin treatments significantly attenuated PMS‐induced ROS formation (P < 0.01) and resulted in decreased cell motility associated with a reduction in MMP‐2 and ‐9 activitiy in HT1080 cells, even in the absence of PMS treatment. Collectively, these results suggest that quercetin inhibits cell motility via the inhibition of MMP activation in HT1080 cells in the presence and absence of PMS. This is likely to be a result of the suppression of intracellular ROS formation by quercetin.     

Cytotoxic effects of 7-O-butyl naringenin on human breast cancer MCF-7 cells

The effect of 7-O-butyl naringenin (BN), a chemically synthesized derivative of naringenin, was tested on the proliferation of human breast cancer MCF-7 cells. BN inhibited the proliferation of MCF-7 cells in dosedependent manner (IC₅₀: 67.5±2.1 μM), resulting in an increase in the sub-G1 phase cell population. BN induced the generation of intracellular reactive oxygen species (ROS), which were reduced by pretreatment with N-acetylcysteine (NAC). BN also increased the phosphorylation of stress-activated protein kinase/c-Jun NH₄-terminal kinase 1/2 (SAPK/JNK1/2), c-Jun, and p38. However, the phosphorylation of extracellular-regulated kinase 1/2 (Erk1/2) was decreased in BN-treated cells. Pretreatment of cells with the specific inhibitors SP600125 and SB203580 diminished the BN-induced activation of SAPK/JNK1/2 and p38, respectively. These results indicate that the BN-induced cytotoxicity of MCF-7 cells is mediated by the generation of ROS as well as through the p38, SAPK/JNK1/2, and c-Jun activation signaling pathways. BN may therefore possess chemotherapeutic potential as an anti-proliferative agent.     

Olive Oil Phenolics Prevent Oxysterol‐Induced Proinflammatory Cytokine Secretion and Reactive Oxygen Species Production in Human Peripheral Blood Mononuclear Cells,Through Modulation of p38 and JNK Pathways

1 Scope

The aim of the present study was to investigate the ability of extra virgin olive oil (EVOO) polyphenols to counteract the proinflammatory effects induced by dietary and endogenous oxysterols in ex vivo immune cells.

2 Methods and results

Peripheral blood mononuclear cells (PBMCs), separated from the whole blood of healthy donors, were utilized and were stimulated with an oxysterols mixture, in the presence of physiologically relevant concentrations of the EVOO polyphenols, hydroxytyrosol, tyrosol, and homovanillic alcohol. Oxysterols significantly increased the production of proinflammatory cytokines, interleukin‐1β, regulated on activation, normal T‐cell expressed and secreted and macrophage migration inhibitory factor in ex vivo cultured PBMCs. Increased levels of reactive oxygen species (ROS) were also detected along with increased phosphorylation of the p38 and JNK. All phenolic compounds significantly reduced cytokine secretion induced by the oxysterols and inhibited ROS production and mitogen activated protein kinase phosphorylation.

3 Conclusions

These results suggest that extra virgin olive oil polyphenols modulate the immune response induced by dietary and endogenous cholesterol oxidation products in human immune cells and may hold benefit in controlling chronic immune and/or inflammatory processes.     

MALDI‐TOF MS characterization of proanthocyanidins from cranberry fruit (Vaccinium macrocarpon) that inhibit tumor cell growth and matrix metalloproteinase expression in vitro

Proanthocyanidin‐rich extracts were prepared by fractionation of the fruit of the North American cranberry (Vaccinium macrocarpon). In vitro growth inhibition assays in eight tumor cell lines showed that selected fractions inhibited the growth of H460 lung tumors, HT‐29 colon and K562 leukemia cells at GI₅₀ values ranging from 20 to 80 µg ml^?1. Matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS) of one of these fractions found it to be composed of polyflavan‐3‐ols, which are primarily tetramers through heptamers of epicatechin containing one or two A‐type linkages. Whole cranberry extract and the proanthocyanidin fractions were screened for effect on the expression of matrix metalloproteinases in DU 145 prostate carcinoma cells. The expression of MMP‐2 and MMP‐9 was inhibited in response to whole cranberry extract and to a lesser degree by the proanthocyanidin fractions. Copyright © 2005 Society of Chemical Industry     

Evaluation of the Anti-Proliferative Activity of Rare Aldohexoses against MOLT-4F and DU-145 Human Cancer Cell Line and Structure-Activity Relationship of D-Idose

D-Allose (D-All), a C-3 epimer of D-glucose (D-Glc), is a naturally rare monosaccharide, which shows anti-proliferative activity against several human cancer cell lines. Unlike conventional anticancer drugs, D-All targets glucose metabolism and is non-toxic to normal cells. Therefore, it has attracted attention as a unique “seed” compound for anticancer agents. However, the anti-proliferative activities of the other rare aldohexoses have not been examined yet. In this study, we evaluated the anti-proliferative activity of rare aldohexoses against human leukemia MOLT-4F and human prostate cancer DU-145 cell lines. We found that D-All and D-idose (D-Ido) at 5 mM inhibited cell proliferation of MOLT-4F cells by 46 % and 60 %, respectively. On the other hand, the rare aldohexoses at 5 mM did not show specific anti-proliferative activity against DU-145 cells. To explore the structure–activity relationship of D-Ido, we evaluated the anti-proliferative activity of D-sorbose (D-Sor), 6-deoxy-D-Ido, and L-xylose (L-Xyl) against MOLT-4F cells and found that D-Sor, 6-deoxy-D-Ido, and L-Xyl showed no inhibitory activity at 5 mM, suggesting that the aldose structure and the C-6 hydroxy group of D-Ido are important for its activity. Cellular glucose uptake assay and western blotting analysis of thioredoxin-interacting protein (TXNIP) expression suggested that the anti-proliferative activity of D-Ido is induced by inhibition of glucose uptake via TXNIP-independent pathway.     

Innovative Soaking and Grinding Methods and Cooking Affect the Retention of Isoflavones,Antioxidant and Antiproliferative Properties in Soymilk Prepared from Black Soybean

This study's objective was to characterize the effect of traditional and 3 newly devised (soaking+grinding) methods combined with cooking on the content and composition of phenolic substances, antioxidant, and antiproliferative properties of soymilk prepared from black soybean. Phenolic substances and antioxidant profile were characterized and antiproliferation of prostate cancer DU145 cells was conducted using a cell culture assay. Results indicated Grinding Method 4 produced significantly (P < 0.05) higher total phenolic content (TPC), total flavonoid content (TFC), condensed tannin content (CTC), and total isoflavone content in both raw and cooked black soymilk as compared to Method 1. Cooking soymilk reduced 23% to 38% of total phenolic substances. Raw black soymilk produced by Method 4 displayed the highest antioxidant capability, which was determined using ORAC, FRAP, and DPPH assays, and a higher antiprostate cell proliferation ability. Cooking only slightly reduced the potency to inhibit DU145 prostate cancer cells as IC₅₀ value was increased from the average of about 4.0 mg/mL of raw soymilk extracts to 5.5 mg/mL of cooked soymilk extracts of all grinding methods. Overall, total isoflavone content was the only component that was negatively correlated with IC₅₀ value (r = –0.93, P < 0.05) which indicates the ability to inhibit prostate cancer cell is associated with the increase in total isoflavone content, not with any other phenolic substances or antioxidant properties.