Hiroomi Tamura

Anti-inflammatory activity of flavonoids in Nepalese propolis is attributed to inhibition of the IL-33 signaling pathway

Propolis has been used in folk medicine to improve health and prevent inflammatory diseases; however, the components that exhibit its anti-inflammatory activity remain unknown. We herein investigated the effects of flavonoids isolated from Nepalese propolis on the IL-33 signaling pathway to clarify the anti-inflammatory mechanism involved. Of the 8 types of flavonoids isolated from Nepalese propolis, 4 types of compounds, such as 3,4-dihydroxy-4-methoxydalbergione, 4-methoxydalbergion, cearoin, and chrysin, markedly inhibited the IL-33-induced mRNA expression of inflammatory genes including IL-6, TNFα and IL-13 in bone marrow-derived mast cells (BMMC). These four flavonoids also inhibited the IL-33-induced activation of nuclear factor κB (NF-κB), which was consistent with their inhibitory effects on cytokine expression. The effects of these flavonoids are attributed to inhibition of IL-33-induced activation of IKK, which leads to the degradation of IκBα and nuclear localization of NF-κB. On the other hand, other flavonoids isolated from Nepalese propolis, such as isoliquiritigenin, plathymenin, 7-hydroxyflavanone, and (+)-medicarpin, had no effect on the IL-33 signaling pathway or cytokine expression. Therefore, these results indicate that 3,4-dihydroxy-4-methoxydalbergione, 4-methoxydalbergion, cearoin, and chrysin are the substances responsible for the anti-inflammatory activity of Nepalese propolis.

New diterpenoids with estrogen sulfotransferase inhibitory activity from Leonurus sibiricus L.

Four new diterpenoids (1–4), along with eight known diterpenoids (5–12) were isolated from the acetone extract of Leonurus herb (aerial parts of Leonurus sibiricus L.). The structures of the compounds were determined by spectroscopic methods. Among the isolated compounds, compounds 2, 3, 8, 9 and 10 showed inhibitory activity against human liver cytosol estrogen sulfotransferase (E-ST), which plays a key role in the maintenance of cellular estrogen levels. Compound 2 showed the strongest activity with an IC50 value of 7.9xa0μM, which is comparable to the activity of the positive control, meclofenamic acid (IC50 5.4xa0μM).

Nepetaefuran and leonotinin isolated from Leonotis nepetaefolia R. Br. potently inhibit the LPS signaling pathway by suppressing the transactivation of NF-κB

Leonotis nepetaefolia R. Br., also known as Klip Dagga or Lions Ear, has traditionally been used as a folk medicine to treat inflammatory diseases such as rheumatism, bronchitis, and asthma; however, the components that exhibit its anti-inflammatory activity have not yet been identified. In the present study, we investigated the effects of three types of diterpenoids, nepetaefuran, leonotinin, and leonotin, which were isolated from L. nepetaefolia R. Br., on the LPS signaling pathway in order to elucidate the anti-inflammatory mechanism involved. Nepetaefuran more potently inhibited the LPS-induced production of NO and CCL2 than leonotinin by suppressing the expression of iNOS mRNA and CCL2 mRNA. On the other hand, leonotin failed to inhibit the production of NO and CCL2 induced by LPS. Although nepetaefuran and leonotinin had no effect on the LPS-induced degradation of IκBα or nuclear translocation of NF-κB p65, they markedly inhibited the transcriptional activity of NF-κB. Nepetaefuran and leonotinin also inhibited the transcriptional activity of the GAL4-NF-κB p65 fusion protein. On the other hand, nepetaefuran, leonotinin and leonotin did not affect the LPS-induced activation of MAP kinase family members such as ERK, p38, and JNK. In addition, inhibitory effect of nepetaefuran and leonotinin on NF-κB activation is well correlated with their ability to induce activation of Nrf2 and ER stress. Taken together, these results demonstrated that nepetaefuran and leonotinin could be the components responsible for the anti-inflammatory activity of L. nepetaefolia R. Br. by specifically inhibiting the LPS-induced activation of NF-κB.

Human keratinocyte caspase-14 expression is altered in human epidermal 3D models by dexamethasone and by natural products used in cosmetics

Caspase-14 is a cysteinyl-aspartate-specific proteinase that is specifically expressed in epidermal keratinocytes. Dysregulation of caspase-14 expression is implicated in impaired skin barrier formation. To elucidate the regulation of caspase-14 in differentiated keratinocytes, we characterized the expression of caspase-14 in normal human epidermal keratinocytes (NHEKs) and two types of three-dimensional (3D) human epidermis culture models, EPI-200 and EPI-201, via RT-PCR and immunoblot analyses. Caspase-14 expression was absent in subconfluent NHEKs, but was present in confluent NHEKs as well as those induced to differentiate by calcium. Caspase-14 expression levels in the 3D epidermis models were almost equal to that in the Ca2+-treated differentiated NHEKs. Despite the presence of caspase-14 expression in these models, caspase-14 activity was found only in the mature 3D skin model, EPI-200. This was confirmed by detection of a 17xa0kDa cleaved fragment of caspase-14 present only in the EPI-200 model. Since glucocorticoid (GC) receptor is required for skin barrier competence, we investigated whether the GC dexamethasone (Dex) and various natural components of common skin moisturizers affect caspase-14 expression in keratinocytes. Dex decreased caspase-14 expression in undifferentiated, but not differentiated, NHEKs. Conversely, Dex increased caspase-14 expression in both 3D skin models, although it did not alter caspase protease activity. Similar to treatment with Dex, treatment of the premature 3D skin mode, EPI-201 with a Galactomyces ferment filtrate markedly increased expression of caspase-14. Further, these results suggest that the effect of Dex, or lack thereof, on caspase-14 expression is dependent on the stage of keratinocyte differentiation.

Coffee extract inhibits adipogenesis in 3T3-L1 preadipocyes by interrupting insulin signaling through the downregulation of IRS1

Although epidemiological data have indicated that a strong negative association exists between coffee consumption and the prevalence of obesity-associated diseases, the molecular mechanisms by which coffee intake prevents obesity-associated diseases has not yet been elucidated. In this study, we found that coffee intake significantly suppressed high-fat diet (HFD)-induced metabolic alternations such as increases in body weight and the accumulation of adipose tissue, and up-regulation of glucose, free fatty acid, total cholesterol and insulin levels in the blood. We also found that coffee extract significantly inhibited adipogenesis in 3T3-L1 preadipocytes. In the early phase of adipogenesis, 3T3-L1 cells treated with coffee extract displayed the retardation of cell cycle entry into the G2/M phase called as mitotic clonal expansion (MCE). Coffee extract also inhibited the activation of CCAAT/enhancer-binding protein β (C/EBPβ) by preventing its phosphorylation by ERK. Furthermore, the coffee extract suppressed the adipogenesis-related events such as MCE and C/EBPβ activation through the down-regulation of insulin receptor substrate 1 (IRS1). The stability of the IRS1 protein was markedly decreased by the treatment with coffee extract due to proteasomal degradation. These results have revealed an anti-adipogenic function for coffee intake and identified IRS1 as a novel target for coffee extract in adipogenesis.

Coffee induces vascular endothelial growth factor (VEGF) expression in human neuroblastama SH-SY5Y cells.

Recent evidence indicates that hypoxia-inducible vascular endothelial growth factor (VEGF) has neurotrophic and neuroprotective effects on neuronal and glial cells. On the other hand, recent epidemiological studies showed that daily coffee consumption has been associated with a lower risk of several neuronal disorders. Therefore, we investigated the effect of coffee on VEGF expression in human neuroblastoma SH-SY5Y cells. We found that even low concentration of coffee (<2%) strongly induced VEGF expression via an activation of HIF-1α. The activation of HIF-1α by coffee was attributed to the coffee-dependent inhibition of prolyl hydroxylation of HIF1α, which is essential for proteolytic degradation of HIF-1α. However, no inhibition was observed at the catalytic activity in vitro. Coffee component(s) responsible for the activation of HIF-1α was not major constituents such as caffeine, caffeic acid, chlorogenic acid, and trigonelline, but was found to emerge during roasting process. The active component(s) was extractable with ethyl acetate. Our results suggest that daily consumption of coffee may induce VEGF expression in neuronal cells. This might be related to protective effect of coffee on neural disorders such as Alzheimers disease and Parkinsons disease.

Effects of Coffee on Estrogen Sulfation in Human Colon Carcinoma Caco-2 Cells

Abstract Recent epidemiological studies have shown that moderate coffee consumption is associated with a lower risk of certain types of cancers, particularly colon cancer in postmenopausal women. To elucidate the molecular basis of the preventive action of coffee toward colon cancer, we focused on the effects of coffee on the metabolism of estrogens by sulfation, a reaction that inactivates estrogens. Estrogens are sulfonated by estrogen sulfotransferase (SULT1E1), and the sulfonated estrogens are reversely hydrolyzed by steroid sulfatase (STS). Sulfonated estrogens are pumped out of the cells by an ABC transporter, breast cancer resistance protein (BCRP). Intracellular concentrations of active estrogens may be determined by the balance of these activities. In human colon cancer Caco-2 cells, coffee decreases the expression of SULT1E1 and STS, whereas it increases BCRP expression. Activities of these three proteins change according to the expression level of each gene. Active constituents of coffee extract can be generated during the roasting process of coffee beans at different times. Modulation of the expression of these genes is likely to be mediated via activation of transcription factors such as NF-κB (BCRP) and stimulating protein 1 (SULT1E1). These results suggest that coffee consumption may modulate the inactivation of estrogens by sulfation in human colon cells.