Puerarin attenuates hepatic steatosis via G-protein-coupled estrogen receptor-mediated calcium and SIRT1 signaling pathways.

Puerarin, the major bioactive ingredient isolated from the root of Pueraria lobata (Willd.), attenuates body weight gain and reduces lipid levels in high-fat diet-induced obese mice; however, the underlying mechanism responsible for regulating lipid metabolism remains unclear. This study investigated the molecular mechanism(s) underlying the role of puerarin in regulating lipogenesis and lipolysis in human HepG2 cells. In this study, puerarin strongly inhibited the expression of fatty acid synthase (FASN) and sterol regulatory element binding protein 1c (SREBP-1c). Moreover, puerarin significantly induced the expression of adipose triglyceride lipase (ATGL), which is responsible for triacylglycerol hydrolase activity in cells. Puerarin enhanced 5' AMP-activated protein kinase (AMPK) activity, which is a central regulator of hepatic lipid metabolism. Furthermore, this AMPK activation could be mediated by sirtuin 1 (SIRT1) and calcium signaling pathways involved in G protein-coupled estrogen receptor (GPER) signaling. GPER blockage significantly reversed the effect of puerarin on lipid accumulation and the related signaling pathways. Docking studies showed that puerarin could bind in the GPER in a similar manner as GPER agonist G1. Our results suggest that puerarin can improve hepatic steatosis by activating GPER; it's signaling cascade sequentially induced calcium and SIRT1 signaling pathways. Thus, puerarin may be a potential therapeutic agent for the treatment of non-alcoholic fatty liver disease.

Rutaecarpine Protects against Acetaminophen-Induced Acute Liver Injury in Mice by Activating Antioxidant Enzymes.

Rutaecarpine, an indolopyridoquinazolinone alkaloid isolated from the unripe fruit of Evodia rutaecarpa, is used to treat hypertension, postpartum hemorrhage, dysentery, and amenorrhea as a traditional medicine in Asia. We investigated the effect of rutaecarpine on acetaminophen-induced hepatotoxicity in mice. Rutaecarpine was administered orally daily for seven consecutive days, followed by intraperitoneal injection of acetaminophen in mice on day seven to induce hepatotoxicity. Rutaecarpine pretreatment significantly decreased acetaminophen-induced serum alanine aminotransferase (ALT)/aspartate aminotransferase (AST) activities and hepatic malondialdehyde content and prevented acetaminophen-induced hepatic glutathione depletion. Furthermore, CYP2E1 expression was decreased by rutaecarpine pretreatment in a dose-dependent manner. Rutaecarpine pretreatment inhibited acetaminophen-induced expression of inflammatory cytokines by inhibiting NF-κB activation by JNK1/2. Also, rutaecarpine pretreatment promoted Nrf2-mediated activation of the antioxidant enzymes GCLC, HO-1, and NQO1. This indicates that the protective effect of rutaecarpine during acetaminophen-induced acute liver injury is mediated by the activation of antioxidant enzymes. Therefore, rutaecarpine has a protective effect of APAP-induced liver damage.

Suppression of PMA-induced human fibrosarcoma HT-1080 invasion and metastasis by kahweol via inhibiting Akt/JNK1/2/p38 MAPK signal pathway and NF-κB dependent transcriptional activities.

Coffee is one of the widely sales beverage worldwide and contains numerous phytochemicals that are beneficial to health. Kahweol acetate (KA), a coffee-specific diterpene, exhibits anti-tumoric properties in human tumoric cells. However, the effect of KA on the metastasis and invasion of cancer cells and the underlying mechanisms remain unclear. The objectives of this study were to estimate the anti-tumor activity of KA and reveal the possible molecular mechanisms. KA markedly inhibited the cell proliferation enhanced by phorbol 12-myristate 13-acetate (PMA) in human fibrosarcoma cells. As well as, KA attenuated PMA-induced cell migration and invasion in a concentration-dependent manner. KA suppressed PMA-enhanced activation of matrix metalloproteinase-9 (MMP-9) through suppression of nuclear factor kappa B (NF-κB) activation. KA repressed the PMA-induced phosphorylation of Akt, c-Jun N-terminal kinase (JNK) 1/2, and p38 MAPK, which are signaling molecules upstream of MMP-9 expression. In summary, we demonstrated that the anti-tumor effects of KA might occur through the inhibition of Akt/JNK1/2/p38 MAPK phosphorylation and downregulation of NF-κB activation, leading to a decrease in MMP-9 expression. Thus, KA is a useful chemotherapeutic agent that may contribute to prevent to the metastatic tumor.

Pseudoshikonin I enhances osteoblast differentiation by stimulating Runx2 and Osterix.

Pseudoshikonin I (PSI), a novel biomaterial isolated from Lithospermi radix, has been recognized as an herbal medicine for the treatment of infectious and inflammatory diseases. Bone remodeling maintains a balance through bone resorption (osteoclastogenesis) and bone formation (osteoblastogenesis). Bone formation is generally attributed to osteoblasts. However, the effects of PSI on the bone are not well known. In this study, we found that the ethanol extracts of PSI induced osteoblast differentiation by increasing the expression of bone morphogenic protein 4 (BMP 4). PSI positively regulates the transcriptional expression and osteogenic activity of osteoblast-specific transcription factors such as Runx2 and Osterix. To identify the signaling pathways that mediate PSI-induced osteoblastogenesis, we examined the effects of serine-threonine kinase inhibitors that are known regulators of Osterix and Runx2. PSI-induced upregulation of Osterix and Runx2 was suppressed by treatment with AKT and PKA inhibitors. These results suggest that PSI enhances osteoblast differentiation by stimulating Osterix and Runx2 via the AKT and PKA signaling pathways. Thus, the activation of Runx2 and Osterix is modulated by PSI, thereby demonstrating its potential as a treatment target for bone disease.

Evaluation of Estrogenic Activity of Extract from the Herbal Mixture Cynanchum wilfordii Hemsley, Phlomis umbrosa Turczaninow, and Angelica gigas Nakai.

Hormone replacement therapy (HRT) consists of highly effective prescription medications for treating menopausal symptoms; however, these agents have exhibited side effects including the risk of estrogen-induced carcinogenesis. Therefore, interest in phytotherapy-based materials as a natural source of alternatives to estrogen therapy has increased. However, some of these herbal medicines have been reported to increase the risk of estrogen-induced cancer. Herbal formulations composed of a combination of Cynanchum wilfordii Hemsley (CW), Phlomis umbrosa Turczaninow (PU), and Angelica gigas Nakai (AG) extracts (CPAE) have been used for treating menopausal symptoms. Therefore, in this study, we aimed to examine the safety of CPAE by determining its potential adverse estrogenic activity using the Organization for Economic Cooperation and Development (OECD) test guideline 455 (TG455) in a stably transfected transcriptionally activated human estrogen receptor α (hERα)-HeLa9903 cell model. We found that CPAE did not how any estrogenic activity or stimulate promoters containing estrogen response elements in MCF-7 cells. In addition, CPAE showed no significant selective activity against hERα and hERß, non-selective activity against the ER, or effects on ER target gene expression. Furthermore, CPAE did not significantly induce MCF-7 cell proliferation and uterine weight increase in ovariectomized rats. These results demonstrate that CPAE can be used as beneficial herbal drug for prevention and therapeutic intervention of estrogen carcinogenesis in menopausal women.

Saponins from the roots of Platycodon grandiflorum ameliorate high fat diet-induced non-alcoholic steatohepatitis.

Platycodon grandiflorum has been healthy effects due to its various nutritious compounds and is considered as a functional food. Platycodon grandiflorum root-derived saponins (CKS) have been reported to show a variety of effects including anti-inflammatory and anti-oxidative activity. Although CKS have been studied on various bioactivities, the inhibitory effect of CKS on non-alcoholic steatohepatitis (NASH) is not examined. In this study, the inhibitory effects on HFD-induced NASH by CKS were determined. CKS suppressed HFD-induced hepatic lipid peroxidation level, collagen deposition, pro-fibrogenic and pro-inflammatory cytokines expression. CKS treatment suppressed HFD-induced COX-2 expression via inhibition of NF-κB p65 nuclear translocation and IκBα degradation. CKS treatment restored HFD-reduced Nrf2-mediated antioxidant enzymes expression. Furthermore, CKS treatment reinstated HFD-reduced peroxisomal proliferator-activated receptor alpha (PPARα)-regulated acyl-coA oxidase and carnitine-palmitoyl-coA transferase-1 expression. These findings suggest that CKS reduces HFD-induced NASH by up-regulation of Nrf2-mediated anti-oxidant enzymes and PPARα-regulated fatty acid oxidation.

Ethanol extract of Lithospermum erythrorhizon Sieb. et Zucc. promotes osteoblastogenesis through the regulation of Runx2 and Osterix.

Bone remodeling and homeostasis are largely the result of the coordinated action of osteoblasts and osteoclasts. Osteoblasts are responsible for bone formation. The differentiation of osteoblasts is regulated by the transcription factors, Runx2 and Osterix. Natural products of plant origin are still a major part of traditional medicinal systems in Korea. The root of Lithospermum erythrorhizon Sieb. et Zucc. (LR), the purple gromwell, is an herbal medicine used for inflammatory and infectious diseases. LR is an anti-inflammatory and exerts anticancer effects by inducing the apoptosis of cancer cells. However, the precise molecular signaling mechanisms of osteoblastogenesis as regards LR and osteoblast transcription are not yet known. In this study, we investigated the effects of ethanol (EtOH) extract of LR (LES) on the osteoblast differentiation of C2C12 myoblasts induced by bone morphogenetic protein 4 (BMP4) and the potential involvement of Runx2 and Osterix in these effects. We found that the LES exhibited an ability to induce osteoblast differentiation. LES increased the expression of the osteoblast marker, alkaline phosphatase (ALP), as well as its activity, as shown by ALP staining and ALP activity assay. LES also increased mineralization, as shown by Alizarin Red S staining. Treatment with LES increased the protein levels (as shown by immunoblotting), as well as the transcriptional activity of Runx2 and Osterix and enhanced osteogenic activity. These results suggest that LES modulates osteoblast differentiation at least in part through Runx2 and Osterix.

Saponins, especially platyconic acid A, from Platycodon grandiflorum reduce airway inflammation in ovalbumin-induced mice and PMA-exposed A549 cells.

We investigated the inhibitory effects of Platycodon grandiflorum root-derived saponins (Changkil saponins: CKS) on ovalbumin-induced airway inflammation in mice. CKS suppressed leukocytes number, IgE, Th1/Th2 cytokines, and MCP-1 chemokine secretion in bronchoalveolar lavage fluid. Also, ovalbumin-increased MUC5AC, MMP-2/9, and TIMP-1/-2 mRNA expression, NF-κB activation, leukocytes recruitment, and mucus secretion were inhibited by CKS treatment. Moreover, the active component of CKS, platyconic acid A (PA), suppressed PMA-induced MUC5AC mRNA expression (from 2.1 ± 0.2 to 1.1 ± 0.1) by inhibiting NF-κB activation (from 2.3 ± 0.2 to 1.2 ± 0.1) via Akt (from 3.7 ± 0.3 to 2.1 ± 0.2) (p < 0.01) in A549 cells. Therefore, we demonstrate that CKS or PA suppressed the development of respiratory inflammation, hyperresponsiveness, and remodeling by reducing allergic responses, and they may be potential herbal drugs for allergen-induced respiratory disease prevention.

Platycodon grandiflorum root-derived saponins attenuate atopic dermatitis-like skin lesions via suppression of NF-κB and STAT1 and activation of Nrf2/ARE-mediated heme oxygenase-1.

PURPOSE: The consequences of precipitously rising allergic skin inflammation rates worldwide have accelerated the risk of atopic dermatitis (AD). Natural product-based agents with good efficacy and low risk of side effects offer promising prevention and treatment strategies for inflammation-related diseases. We have already reported that Platycodon grandiflorum root-derived saponins (Changkil saponins, CKS) have many pharmacological effects, including anti-inflammatory and anti-allergic effects, but its influence on AD remains unclear. Therefore, we evaluated the inhibitory effect of CKS, mainly platycodin D, on AD-like skin symptoms in mice and the possible mechanisms in cells. METHODS: Mice were sensitized and challenged with 2,4-dinitrochlorobenzene (DNCB). Four weeks after challenge, mice were treated with oral administration of CKS for 4 weeks. In addition, cells were used to evaluate the effect of CKS, mainly platycodin D, on the TARC expression regulated mechanism. RESULTS: CKS attenuated DNCB-induced dermatitis severity, serum levels of IgE and TARC, and mRNA expression of TARC, TNF-α, IFN-γ, IL-4, IL-5, and IL-13 in mice. Histopathological examination showed reduced thickness of the epidermis/dermis and dermal infiltration of inflammatory cells and mast cells in the ears. Moreover, CKS and platycodin D inhibited TNF-α/IFN-γ-induced TARC expression through the suppression of NF-κB and STAT1 and induction of Nrf2/ARE-mediated hemeoxygenase-1 (HO-1) expression in cells. CONCLUSION: We suggest that CKS and platycodin D inhibited the development of AD-like skin symptoms by regulating cytokine mediators and may be an effective alternative therapy for AD-like skin symptoms.

Saponins from the roots of Platycodon grandiflorum suppresses TGFß1-induced epithelial-mesenchymal transition via repression of PI3K/Akt, ERK1/2 and Smad2/3 pathway in human lung carcinoma A549 cells.

Transforming growth factor ß (TGFß) is a multifunctional cytokine that induces growth arrest, tissue fibrosis, and epithelial-mesenchymal transition (EMT) through activation of Smad and non-Smad signaling pathways. EMT is the differentiation switch by which polarized epithelial cells differentiate into contractile and motile mesenchymal cells. Our previous studies have shown that saponins from the roots of Platycodon grandiflorum (CKS) have antiinflammatory, antioxidant, antimetastatic, and hepatoprotective effects. In this study, we investigated the inhibitory effect of CKS on TGFß1-induced alterations characteristic of EMT in human lung carcinoma A549 cells. We found that CKS-treated cells displayed inhibited TGFß1-mediated E-cadherin downregulation and Vimentin upregulation and also retained epithelial morphology. Furthermore, TGFß1-increased Snail expression, a repressor of E-cadherin and an inducer of the EMT, was reduced by CKS. CKS inhibited TGFß1-induced phosphorylation of Akt, ERK1/2, and glycogen synthase kinase-3ß (GSK-3ß). Inhibition of PI3K/Akt and ERK1/2 also blocked TGFß1-induced GSK-3ß phosphorylation and Snail activation. Furthermore, TGFß1-increased Snail expression was reduced by selective inhibitors of Akt and ERK1/2. Moreover, CKS treatment attenuated TGFß1-induced Smad2/3 phosphorylation and upregulated Smad7 expression. These results indicate that pretreatment with the CKS inhibits the TGFß1-induced EMT through PI3K/Akt, ERK1/2, GSK-3ß and Smad2/3 in human lung carcinoma cells.

Cultivated ginseng inhibits 2,4-dinitrochlorobenzene-induced atopic dermatitis-like skin lesions in NC/Nga mice and TNF-α/IFN-γ-induced TARC activation in HaCaT cells.

Ginseng contains many bioactive constituents, including various ginsenosides that are believed to have anti-allergic, anti-oxidant, and immunostimulatory activities; however, its effects on atopic dermatitis (AD) remain unclear. In the current study, we hypothesized that cultivated ginseng (CG) would inhibit 2,4-dinitrochlorobenzene (DNCB)-induced AD-like skin lesions in NC/Nga mice by regulating the T helper (Th)1/Th2 balance. Also, CG inhibits TNF-α/IFN-γ-induced thymus- and activation-regulated chemokine (TARC) expression through nuclear factor-kappa B (NF-κB)-dependent signaling in HaCaT cells. CG ameliorated DNCB-induced dermatitis severity, serum levels of IgE and TARC, and mRNA expression of TARC, TNF-α, IFN-γ, IL-4, IL-5, and IL-13 in mice. Histopathological examination showed reduced thickness of the epidermis/dermis and dermal infiltration of inflammatory cells in the ears. Furthermore, CG suppressed the TNF-α/IFN-γ-induced mRNA expression of TARC in HaCaT cells. CG inhibited TNF-α/IFN-γ-induced NF-κB activation. These results suggest that CG inhibited the development of the AD-like skin symptoms by modulating Th1 and Th2 responses in the skin lesions in mice and TARC expression by suppressing TNF-α/IFN-γ-induced NF-κB activation in keratinocytes, and so may be a useful tool in the therapy of AD-like skin symptoms.

Platycodi Radix attenuates dimethylnitrosamine-induced liver fibrosis in rats by inducing Nrf2-mediated antioxidant enzymes.

The purpose of this study was to investigate the anti-fibrotic effects of the aqueous extract of the Platycodi Radix root (Changkil: CK) on dimethylnitrosamine (DMN)-induced liver fibrosis in rats. DMN treatment for 4 weeks led to marked liver fibrosis as assessed by serum biochemistry, histopathological examination, and hepatic lipid peroxidation and collagen content. CK significantly inhibited DMN-induced increases in serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, fibrosis score, and hepatic malondialdehyde and collagen content. CK also inhibited DMN-induced reductions in rat body and liver weights. Reverse transcription polymerase chain reaction (RT-PCR) and western blot analyses revealed that CK inhibited DMN-induced increases in matrix metalloproteinase-13 (MMP-13), tissue inhibitor of metalloproteinase-1 (TIMP-1), and tumor necrosis factor-α (TNF-α) mRNA, and collagen type I and α-smooth muscle actin protein. DMN-induced cyclooxygenase-2 (COX-2) expression and nuclear factor-kappa B (NF-κB) activation was reduced by CK treatment. Furthermore, CK induced activation of nuclear erythroid 2-related factor 2 (Nrf2)-mediated antioxidant enzymes such as γ-glutamylcysteine synthetase (γ-GCS), heme oxygenase-1 (HO-1), NAD(P)H quinone oxidoreductase 1 (NQO1), and glutathione-S-transferase (GST) in HepG2 cells. These results demonstrated that CK attenuates DMN-induced liver fibrosis through the activation of Nrf2-mediated antioxidant enzymes.

Inhibitory effect of Psidium guajava water extract in the development of 2,4-dinitrochlorobenzene-induced atopic dermatitis in NC/Nga mice.

Atopic dermatitis (AD) is a chronic, relapsing, and inflammatory skin disease associated with eczematous symptoms and IgE hyperproduction. Psidium guajava is an important food crop and medicinal plant with anti-oxidant, anti-inflammatory, and anti-allergic activities, supporting its traditional uses. Our previous studies have shown that P. guajava extract inhibits Th2 chemokine expression by suppressing the activation of NF-κB and STAT1 co-stimulated with TNF-α and INF-γ. In this study, we investigated the inhibitory effect of P. guajava water extract (PGW) on 2,4-dinitrochlorobenzene (DNCB)-induced AD-like skin lesions in NC/Nga mice. Treatment of cream containing PGW onto DNCB-induced AD-like skin lesions in NC/Nga mice ameliorated lesion intensity scores, levels of IgE, thymus and activation-regulated chemokine (TARC), TNF-α, and IL-4 in serum and ears. In contrast, PGW increased level of the immunosuppressive cytokine IL-10. Histological analyses demonstrated decreased thickening of the epidermis/dermis as well as dermal infiltration by inflammatory cells. These results suggest that cream containing PGW may be a potential therapeutic modality for AD and adjunctive agent to control pruritus in AD.