Da-Hye Lee

Pharmacokinetics, Tissue Distribution, and Anti-Lipogenic/Adipogenic Effects of Allyl-Isothiocyanate Metabolites

Allyl-isothiocyanate (AITC) is an organosulfur phytochemical found in abundance in common cruciferous vegetables such as mustard, wasabi, and cabbage. Although AITC is metabolized primarily through the mercapturic acid pathway, its exact pharmacokinetics remains undefined and the biological function of AITC metabolites is still largely unknown. In this study, we evaluated the inhibitory effects of AITC metabolites on lipid accumulation in vitro and elucidated the pharmacokinetics and tissue distribution of AITC metabolites in rats. We found that AITC metabolites generally conjugate with glutathione (GSH) or N-acetylcysteine (NAC) and are distributed in most organs and tissues. Pharmacokinetic analysis showed a rapid uptake and complete metabolism of AITC following oral administration to rats. Although AITC has been reported to exhibit anti-tumor activity in bladder cancer, the potential bioactivity of its metabolites has not been explored. We found that GSH-AITC and NAC-AITC effectively inhibit adipogenic differentiation of 3T3-L1 preadipocytes and suppress expression of PPAR-γ, C/EBPα, and FAS, which are up-regulated during adipogenesis. GSH-AITC and NAC-AITC also suppressed oleic acid-induced lipid accumulation and lipogenesis in hepatocytes. Our findings suggest that AITC is almost completely metabolized in the liver and rapidly excreted in urine through the mercapturic acid pathway following administration in rats. AITC metabolites may exert anti-obesity effects through suppression of adipogenesis or lipogenesis.

γ-Oryzanol Enhances Adipocyte Differentiation and Glucose Uptake

Recent studies show that brown rice improves glucose intolerance and potentially the risk of diabetes, although the underlying molecular mechanisms remain unclear. One of the phytochemicals found in high concentration in brown rice is γ-oryzanol (Orz), a group of ferulic acid esters of phytosterols and triterpene alcohols. Here, we found that Orz stimulated differentiation of 3T3-L1 preadipocytes and increased the protein expression of adipogenic marker genes such as peroxisome proliferator-activated receptor gamma (PPAR-γ) and CCAAT/enhanced binding protein alpha (C/EBPα). Moreover, Orz significantly increased the glucose uptake in insulin-resistant cells and translocation of glucose transporter type 4 (GLUT4) from the cytosol to the cell surface. To investigate the mechanism by which Orz stimulated cell differentiation, we examined its effects on cellular signaling of the mammalian target of rapamycin complex 1 (mTORC1), a central mediator of cellular growth and proliferation. The Orz treatment increased mTORC1 kinase activity based on phosphorylation of 70-kDa ribosomal S6 kinase 1 (S6K1). The effect of Orz on adipocyte differentiation was dependent on mTORC1 activity because rapamycin blocks cell differentiation in Orz-treated cells. Collectively, our results indicate that Orz stimulates adipocyte differentiation, enhances glucose uptake, and may be associated with cellular signaling mediated by PPAR-γ and mTORC1.

Nutrikinetic study of genistein metabolites in ovariectomized mice

This study was designed to evaluate the effect of ovariectomy on nutrikinetics of genistein metabolites. To characterize the time-dependent changes in genistein metabolite concentrations, we identified 13 genistein metabolites using ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry. The nutrikinetics of the individual metabolites at different time points were analyzed. Nutrikinetic analysis showed that genistein, genistein 4′-glucuronide, genistein 7-glucuronide, 3-hydroxygenistein, and hippuric acid showed relatively high bioavailability in the sham group compared to that in the ovariectomy group, suggesting that ovariectomy likely results in lower genistein bioavailability. These results may be related to alteration of gut microbiota by ovariectomy. The relative abundance of species of the Parabacteroides, Dorea, and Butyricimonas genera, and Desulfovibrionaceae_unclassified, Lachnospiraceae_unclassified, and Rikenellaceae_unclassified families increased in the ovariectomy group while the relative abundance of 523_7_unclassified and Y52_unclassified_unclassified increased in the sham group. These results suggest that gut microbiota alteration by ovariectomy may affect genistein bioavailability.

3-Decylcatechol induces autophagy-mediated cell death through the IRE1α/JNK/p62 in hepatocellular carcinoma cells

The natural, phenolic lipid urushiol exhibits both antioxidant and anticancer activities; however, its biological activity on hepatocellular carcinoma (HCC) has not been previously investigated. Here, we demonstrate that an urushiol derivative, 3-decylcatechol (DC), induces human HCC Huh7 cell death by induction of autophagy. DC initiates the autophagic process by activation of the mammalian target of rapamycin signaling pathway via Unc-51-like autophagy activating kinase 1, leading to autophagosome formation. The autophagy inhibitor, chloroquine, suppressed autolysosome formation and cell death induction by DC, indicating an autophagic cell death. Interestingly, DC also activated the endoplasmic reticulum (ER) stress response that promotes autophagy via p62 transcriptional activation involving the inositol-requiring enzyme 1α/c-Jun N-terminal kinase/c-jun pathway. We also show that cytosolic calcium mobilization is necessary for the ER stress response and autophagy induction by DC. These findings reveal a novel mechanism by which this urushiol derivative induces autophagic cell death in HCC.The natural, phenolic lipid urushiol exhibits both antioxidant and anticancer activities; however, its biological activity on hepatocellular carcinoma (HCC) has not been previously investigated. Here, we demonstrate that an urushiol derivative, 3-decylcatechol (DC), induces human HCC Huh7 cell death by induction of autophagy. DC initiates the autophagic process by activation of the mammalian target of rapamycin signaling pathway via Unc-51-like autophagy activating kinase 1, leading to autophagosome formation. The autophagy inhibitor, chloroquine, suppressed autolysosome formation and cell death induction by DC, indicating an autophagic cell death. Interestingly, DC also activated the endoplasmic reticulum (ER) stress response that promotes autophagy via p62 transcriptional activation involving the inositol-requiring enzyme 1α/c-Jun N-terminal kinase/c-jun pathway. We also show that cytosolic calcium mobilization is necessary for the ER stress response and autophagy induction by DC. These findings reveal a novel mechanism by which this urushiol derivative induces autophagic cell death in HCC.

Pharmacokinetics of Tyrosol Metabolites in Rats

Tyrosol is considered a potential antioxidant; however, little is known regarding the pharmacokinetics of its metabolites. To study the pharmacokinetics of tyrosol-derived metabolites after oral administration of a single dose of tyrosol, we attempted to identify tyrosol metabolites in rat plasma by using ultra-performance liquid chromatography and quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). Two tyrosol metabolites (M1 and M2) were detected in the plasma. M1 was identified as tyrosol-4-sulfate (T4S) with an [M − H]− ion at m/z 217. While M2 showed an [M − H]− ion at m/z 151.0, its metabolite was not identified. Pharmacokinetic analysis of T4S and M2 showed rapid uptake after oral administration of tyrosol within 1 h. The metabolites were rapidly distributed in most organs and tissues and eliminated within 4 h. The greatest T4S deposition by tissue weight was observed in the liver, followed by the kidney and spleen, while M2 was most concentrated in the kidney followed by the liver and spleen. These findings indicate that T4S and M2 were distributed mainly in tissues with an abundant blood supply and were rapidly excreted in urine.

Nutrikinetics of Isoflavone Metabolites After Fermented Soybean Product (Cheonggukjang) Ingestion in Ovariectomized Mice

Scope Cheonggukjang (CGJ) is a soybean‐based quick‐fermented food popular in Korea that contains a variety of biologically active compounds including isoflavones and saponins. Isoflavone bioavailability may be important for the bone health of postmenopausal women; therefore, the aim of this study is to evaluate the influence of fermentation on the isoflavone metabolite nutrikinetic profile after single dose CGJ or unfermented soybean administration in ovariectomized (OVX) and sham mice. Methods and results We identify 34 isoflavone metabolites using UPLC–QTOF‐MS and analyze their nutrikinetics at different time points (0.25, 0.5, 1, 2, 4, 8, 16, and 24 h) to understand their fermentation‐ and OVX‐mediated time‐dependent concentration changes. Nutrikinetics analysis shows that genistein, daidzein, genistein 4′‐sulfate, dihydrodaidzein sulfate, equol 4′‐sulfate, and equol‐7‐glucuronide are present at high concentrations in all groups based on area‐under‐the‐curve analysis. OVX mice appear to show lower isoflavone bioavailability than mice in the sham group. CGJ enhances various isoflavone metabolite bioavailability including genistein, 3‐hydroxygenistein, and equol 7‐glucuronide, compared to the unfermented soybean‐treated group. Among these metabolites, intact isoflavones, 3‐hydroxygenistein, genistein 4′‐sulfate, and equol 7‐glucuronide promote osteoblastogenesis and inhibit osteoclast formation. Conclusions CGJ has good isoflavone bioavailability and may be beneficial for the bone health of postmenopausal women.

Effects of yuja peel extract and its flavanones on osteopenia in ovariectomized rats and osteoblast differentiation.

SCOPE Yuja (Citrus junos Tanaka) possesses various health benefits, but its effects on bone health are unknown. In this study, the preventative effects of yuja peel ethanol extract (YPEE) on osteopenia were determined in ovariectomized (OVX) rats, and the mechanisms by which YPEE and its flavanones regulate osteoblastogenesis were examined in vitro. METHODS AND RESULTS The effects of YPEE on osteoblastogenesis were investigated in MC3T3-E1 cells. YPEE promoted alkaline phosphatase (ALP) activity, mineralization, and the expression of osteoblast differentiation marker genes, such as ALP, runt-related transcription factor 2 (Runx2), and osteocalcin. YPEE and its flavanones promoted osteoblast differentiation via BMP-2-mediated p38 and the Smad1/5/8 signaling pathway. YPEE supplementation significantly decreased body weight and increased uterine weight and bone mineral density in OVX rats. Based on a micro-CT analysis of femurs, YPEE significantly attenuated osteopenia and increased trabecular volume fraction, trabecular separation, and trabecular number (p < 0.05). CONCLUSION Dietary YPEE has a protective effect on OVX-induced osteopenia. YPEE and its flavanones promote osteoblastogenesis via the activation of the BMP/p38/Smad/Runx2 pathways. These results extend our knowledge of the beneficial effects of YPEE and provide a basis for the development of novel therapies for osteoporosis.

A Pilot Study on Characteristics of Metabolomics and Lipidomics according to Sasang Constitution

Although classification of an individuals Sasang constitution is a key step in the prescription of traditional Korean medicine, the classifying process is complex and not objective. Identification of metabolic-based biomarkers could allow the development of a reliable and sensitive classification technique and even therapeutic management. Our pilot study investigated whether metabolites in plasma are characteristic of Sasang constitutions. Ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry-based metabolic analysis was conducted against 15 Soyangin (SY), 15 Taeeumin (TE), and 18 Soeumin (SE) individuals, as classified according to the Questionnaire for Sasang Constitution Classification II (QSCC II) and specialist diagnosis. Metabolomics data showed that the TE group was significantly separated from the SY and SE groups. Nine canonical pathways related to constitution; phenylalanine metabolism, aminoacyl-tRNA, tyrosine, and tryptophan biosynthesis were activated in the TE group as compared with the other groups. Similar to the results of the metabolomics analysis, the TE group was also significantly separated from the other two groups by lipidomic analysis. On the other hand, the intensity of lipid metabolites was higher in the SY group than in the other groups. Our findings suggest that the combined analysis of metabolomics and lipidomics can provide useful information for characteristics of Sasang constitutions.