Purple perilla extracts with α-asarone enhance cholesterol efflux from oxidized LDL-exposed macrophages.

The cellular accumulation of cholesterol is critical in the development and progression of atherosclerosis. ATP-binding cassette (ABC) transporters play an essential role in mediating the efflux of excess cholesterol. In the current study, we investigated whether purple Perilla frutescens extracts (PPE) at a non-toxic concentration of 1-10 µg/ml stimulate the induction of the ABC transporters, ABCA1 and ABCG1, and cholesterol efflux from lipid-laden J774A.1 murine macrophages exposed to 50 ng/ml oxidized low-density lipoprotein (LDL). Purple perilla, an annual herb in the mint family and its constituents, have been reported to exhibit antioxidant and cytostatic activity, as well as to exert anti-allergic effects. Our results revealed that treatment with oxidized LDL for 24 h led to the accumulation of lipid droplets in the macrophages. PPE suppressed the oxidized LDL-induced foam cell formation by blocking the induction of scavenger receptor B1. However, PPE promoted the induction of the ABC transporters, ABCA1 and ABCG1, and subsequently accelerated cholesterol efflux from the lipid-loaded macrophages. The liver X receptor (LXR) agonist, TO-091317, and the peroxisome proliferator-activated receptor (PPAR) agonist, pioglitazone, increased ABCA1 expression and treatment with 10 µg/ml PPE further enhanced this effect. PPE did not induce LXRα and PPARγ expression per se, but enhanced their expression in the macrophages exposed to oxidized LDL. α-asarone was isolated from PPE and characterized as a major component enhancing the induction of ABCA1 and ABCG1 in macrophages exposed to oxidized LDL. α-asarone, but not ß-asarone was effective in attenuating foam cell formation and enhancing cholesterol efflux, revealing an isomeric difference in their activity. The results from the present study demonstrate that PPE promotes cholesterol efflux from macrophages by activating the interaction of PPARγ-LXRα-ABC transporters.

Purple perilla extracts allay ER stress in lipid-laden macrophages.

There is a growing body of evidence that excess lipids, hypoxic stress and other inflammatory signals can stimulate endoplasmic reticulum (ER) stress in metabolic diseases. However, the pathophysiological importance and the underlying mechanisms of this phenomenon remain unknown. The current study investigated that 50 ng/ml oxidized LDL promoted unfolded protein response (UPR) and ER stress in J774A1 murine macrophages, which was blocked by extracts (PPE) of purple Perilla frutescens, a plant of the mint family Lamiaceae. The ER stressor tunicamycin was employed as a positive control. Treating 1-10 µg/ml oxidized LDL for 24 h elicited lipotoxic apoptosis in macrophages with obvious nuclear condensation and DNA fragmentation, which was inhibited by PPE. Tunicamycin and oxidized LDL activated and induced the UPR components of activating transcription factor 6 and ER resident chaperone BiP/Grp78 in temporal manners and such effects were blocked by ≥5 µg/ml PPE. In addition, PPE suppressed the enhanced mRNA transcription and splicing of X-box binding protein 1 (XBP1) by tunicamycin and oxidized LDL. The protein induction and nuclear translocation of XBP1 were deterred in PPE-treated macrophages under ER stress. The induction of ATP-binding cassette transporter A1 (ABCA1), scavenger receptor-B1 (SR-B1) and intracellular adhesion molecule-1 (ICAM-1) was abolished by the ER stressor in activated macrophages. The protein induction of ABCA1 and ICAM1 but not SR-B1 was retrieved by adding 10 µg/ml PPE to cells. These results demonstrate that PPE inhibited lipotoxic apoptosis and demoted the induction and activation of UPR components in macrophages. PPE restored normal proteostasis in activated macrophages oxidized LDL. Therefore, PPE was a potent agent antagonizing macrophage ER stress due to lipotoxic signals associated with atherosclerosis.

Blockade of Airway Inflammation by Kaempferol via Disturbing Tyk-STAT Signaling in Airway Epithelial Cells and in Asthmatic Mice.

Asthma is characterized by bronchial inflammation causing increased airway hyperresponsiveness and eosinophilia. The interaction between airway epithelium and inflammatory mediators plays a key role in the asthmatic pathogenesis. The in vitro study elucidated inhibitory effects of kaempferol, a flavonoid found in apples and many berries, on inflammation in human airway epithelial BEAS-2B cells. Nontoxic kaempferol at ≤20 µ M suppressed the LPS-induced IL-8 production through the TLR4 activation, inhibiting eotaxin-1 induction. The in vivo study explored the demoting effects of kaempferol on asthmatic inflammation in BALB/c mice sensitized with ovalbumin (OVA). Mouse macrophage inflammatory protein-2 production and CXCR2 expression were upregulated in OVA-challenged mice, which was attenuated by oral administration of ≥10 mg/kg kaempferol. Kaempferol allayed the airway tissue levels of eotaxin-1 and eotaxin receptor CCR3 enhanced by OVA challenge. This study further explored the blockade of Tyk-STAT signaling by kaempferol in both LPS-stimulated BEAS-2B cells and OVA-challenged mice. LPS activated Tyk2 responsible for eotaxin-1 induction, while kaempferol dose-dependently inhibited LPS- or IL-8-inflamed Tyk2 activation. Similar inhibition of Tyk2 activation by kaempferol was observed in OVA-induced mice. Additionally, LPS stimulated the activation of STAT1/3 signaling concomitant with downregulated expression of Tyk-inhibiting SOCS3. In contrast, kaempferol encumbered STAT1/3 signaling with restoration of SOCS3 expression. Consistently, oral administration of kaempferol blocked STAT3 transactivation elevated by OVA challenge. These results demonstrate that kaempferol alleviated airway inflammation through modulating Tyk2-STAT1/3 signaling responsive to IL-8 in endotoxin-exposed airway epithelium and in asthmatic mice. Therefore, kaempferol may be a therapeutic agent targeting asthmatic diseases.

Novel Danshen methoxybenzo[b]furan derivative antagonizing adipogenic differentiation and production of inflammatory adipokines.

Many benzo[b]furan lignans are known to be biologically active in nature. 2-(3'-Methoxy-4'-hydroxy-phenyl)-5-(3-hydroxypropyl)-7-methoxy-benzo[b]furan-3-carbaldehyde (XH-14) is found as a bioactive component isolated from the plant Salvia miltiorrhiza, commonly known as Danshen, which is a traditional Chinese medicine that is used as a cardiovascular medication. This study examined whether 3 different XH-14 derivatives can inhibit adipocyte differentiation and induction of the adipokines visfatin and resistin in 3T3-L1 adipocytes. Adipocytes were cultured and differentiated in Dulbecco's modified Eagle medium containing fetal bovine serum, 3-isobytyl-1-methylxanthine, dexamethasone, and insulin for 6-8d in the absence and presence of 1-25µM XH-14-derived benzo[b]furan derivatives. Nontoxic 2-(3'-methoxy-4'-hydroxy-phenyl)-6-(3-hydroxypropyl)-5-methoxy-benzo[b]furan (5-MBF) at ≥5µM attenuated cellular lipid accumulation and down-regulated induction of peroxisome proliferator activated receptors γ (PPARγ) and CCAAT enhancer binding protein α (C/EBPα) in a dose-dependent manner, as evidenced by Oil Red O staining and Western blot analysis. Such inhibition of PPAR( and C/EBP( by 5-MBF was achieved at transcriptional mRNA levels. However, 2-(3'-methoxy-4'-hydroxy-phenyl)-5-(3-hydroxypropyl)-7-methoxy-benzo[b]furan (7-MBF) and 2-(3'-methoxy-4'-hydroxy-phenyl)-5-(3-hydroxypropyl)-7-methoxy-benzo[b]furan (6-MBF) had minimal effects on adipogenic differentiation, suggesting a structure-activity relationship of methoxybenzo[b]furan derivatives as an inhibitor of adipogenic differentiation. Furthermore, ≥5µM 5-MBF retarded protein and mRNA expression of proinflammatory and insulin resistance-enhancing adipokines of visfatin and resistin in differentiated adipocytes. Induction of visfatin and resistin was, at least in part, mediated via adipocyte differentiation-associated activation of PPARγ signal targeting adipocyte protein 2 and stearoyl-CoA desaturase. These results demonstrate that the 2-(3'-methoxy-4'-hydroxy-phenyl)-3-hydroxypropyl benzo[b]furan lignan, with a methoxy group at the 5-position on the benzene ring, may be a promising agent for disturbance of adipogenic differentiation and for blockage of obesity-associated inflammatory and metabolic diseases.