Protective effect of sesquiterpene lactone parthenolide on LPS-induced acute lung injury

장유진,백문정,Zhicheng Fu,이주현,원종훈,하해찬,이혜경,장지민,최종민,김대경 대한약학회 2016 Archives of Pharmacal Research Vol.39 No.12

Acute lung injury (ALI) is a respiratory failuredisease and the major source of mortality in the critically illpatients. The main pathological changes involved in ALIinclude the excessive recruitment and activation of neutrophilsby increased pro-inflammatory mediators. However,any specific therapy for ALI has not been developed. The objective of this study was to investigate protectiveeffects of parthenolide, a sesquiterpene lactone produced infeverfew, on LPS-induced lung injury. In the present study,parthenolide treatment reduced infiltration of inflammatorycells, airway permeability and production of pro-inflammatorycytokines in LPS-induced ALI mouse model. Further,LPS-stimulated phosphorylation of NF-jB, the keyregulatory transcription factor in ALI, was inhibited byparthenolide treatment in lung epithelial BEAS-2B cellsand alveolar macrophage MH-S cells. These results suggestthat parthenolide may provide a beneficial therapeuticstrategy for ALI.

Paraquat Induces Apoptosis through a Mitochondria-Dependent Pathway in RAW264.7 Cells

장여진,원종훈,백문정,Zhicheng Fu,장지민,하해찬,홍승범,장민선,김대경 한국응용약물학회 2015 Biomolecules & Therapeutics(구 응용약물학회지) Vol.23 No.5

Paraquat dichloride (N,N-dimethyl-4-4′-bipiridinium, PQ) is an extremely toxic chemical that is widely used in herbicides. PQ generates reactive oxygen species (ROS) and causes multiple organ failure. In particular, PQ has been reported to be an immunotoxic agrochemical compound. PQ was shown to decrease the number of macrophages in rats and suppress monocyte phagocytic activity in mice. However, the effect of PQ on macrophage cell viability remains unclear. In this study, we evaluated the cytotoxic effect of PQ on the mouse macrophage cell line, RAW264.7 and its possible mechanism of action. RAW264.7 cells were treated with PQ (0, 75, and 150 μM), and cellular apoptosis, mitochondrial membrane potential (MMP), and intracellular ROS levels were determined. Morphological changes to the cell nucleus and cellular apoptosis were also evaluated by DAPI and Annexin V staining, respectively. In this study, PQ induced apoptotic cell death by dose-dependently decreasing MMP. Additionally, PQ increased the cleaved form of caspase-3, an apoptotic marker. In conclusion, PQ induces apoptosis in RAW264.7 cells through a ROS-mediated mitochondrial pathway. Thus, our study improves our knowledge of PQ-induced toxicity, and may give us a greater understanding of how PQ affects the immune system.

Development of a Label-Free LC-MS/MS-Based Glucosylceramide Synthase Assay and Its Application to Inhibitors Screening for Ceramide-Related Diseases

Zhicheng Fu,윤소윤,원종훈,백문정,장지민,하해찬,이해경,신인철,김주은,김희수,김대경 한국응용약물학회 2019 Biomolecules & Therapeutics(구 응용약물학회지) Vol.27 No.2

Ceramide metabolism is known to be an essential etiology for various diseases, such as atopic dermatitis and Gaucher disease. Glucosylceramide synthase (GCS) is a key enzyme for the synthesis of glucosylceramide (GlcCer), which is a main ceramide metabolism pathway in mammalian cells. In this article, we developed a liquid chromatography-tandem mass spectrometry (LCMS/ MS) method to determine GCS activity using synthetic non-natural sphingolipid C8-ceramide as a substrate. The reaction products, C8-GlcCer for GCS, could be separated on a C18 column by reverse-phase high-performance liquid chromatography (HPLC). Quantification was conducted using the multiple reaction monitoring (MRM) mode to monitor the precursor-to-product ion transitions of m/z 588.6 → 264.4 for C8-GlcCer at positive ionization mode. The calibration curve was established over the range of 0.625-160 ng/mL, and the correlation coefficient was larger than 0.999. This method was successfully applied to detect GCS in the human hepatocellular carcinoma cell line (HepG2 cells) and mouse peripheral blood mononuclear cells. We also evaluated the inhibition degree of a known GCS inhibitor 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP) on GCS enzymatic activity and proved that this method could be successfully applied to GCS inhibitor screening of preventive and therapeutic drugs for ceramide metabolism diseases, such as atopic dermatitis and Gaucher disease.

Age-Related Effects of Sodium Arsenite on Splenocyte Proliferation and Th1/Th2 Cytokine Production

Yuri Cho,김대경,Kyong Hoon Ahn,백문정,최종민,Jung Eun Ji,Jong Hoon Won,Zhicheng Fu,Ji Min Jang 대한약학회 2012 Archives of Pharmacal Research Vol.35 No.2

Aging is associated with immune dysfunction and conditions such as inflamm-aging and immunosuppression. Arsenic, an environmental contaminant distributed worldwide, affects the immune system. This study tested the hypothesis that arsenic has distinct effects on T cell proliferation and the production of cytokines by activated T cells. Murine splenocytes from young (2 months) and aged (24-26 months) C57BL/6 mice were exposed to arsenite (As3+), the most toxic form of inorganic arsenic, and stimulated with concanavalin A (Con A) or anti-CD3 antibody. T cell proliferation decreased significantly in response to Con A and anti-CD3 at subtoxic doses of arsenite in splenocytes from both young and aged mice. Arsenite, added concurrently with Con A or anti-CD3, significantly inhibited the production of interleukin-2 (IL-2), interferon-γ (IFN-γ), and interleukin-4 (IL-4) by splenocytes from young mice and significantly reduced the production of IL-10 by splenocytes from aged mice. In contrast, the production of IL-2 and IL-4 by splenocytes from aged mice was only slightly affected by arsenite. The results show that arsenic exposure reduces the immune response in splenocytes. Moreover,this effect may be influenced by aging.

3, 5, 30-Triiodothyroacetic acid (TRIAC) is an anti-inflammatory drug that targets toll-like receptor 2

하해찬,장지민,ZHOU DAN,김한결,백문정,신인철,윤소윤,Yongwei Piao,최종민,원종훈,김대경 대한약학회 2018 Archives of Pharmacal Research Vol.41 No.10

Drug repositioning is a strategy that explores new pharmaceutical applications of previously launched or failed drugs, and is advantageous since it saves capital and time. In this study, we examined the inhibition of TLR2 signaling by drug candidates. HEK-BlueTM-hTLR2 cells were pretreated with drugs and stimulated using the TLR2 ligand, Pam3CSK4. Among the drugs that inhibited TLR2 signaling, we selected TRIAC, which is yet to be patented. Pretreatment with TRIAC decreased the TLR2 level and the phosphorylation of Akt and MAPKs in HEK-BlueTM- hTLR2 cells. Since TLR2 is overexpressed in patients with acute hepatitis, we confirmed that TRIAC alleviates necrosis in a mouse model of Con A-induced acute hepatitis. The serum AST and ALT levels are indicators of liver damage, and are increased in Con A-induced hepatitis. Additionally, TLR2 and inflammatory cytokine levels are increased following administration of Con A and lead to liver damage. TRIAC decreased the serum levels of AST and ALT, and reduced liver tissue necrosis in mice with Con A-induced acute fulminant liver damage, by reducing the levels of inflammatory cytokines. In conclusion, TRIAC alleviates inflammation in mouse models of Con A-induced hepatitis by inhibiting the phosphorylation of Akt and MAPKs, the sub-mechanisms underlying TLR2 signaling.

C6-Ceramide Enhances Phagocytic Activity of Kupffer Cells through the Production of Endogenous Ceramides

최종민,So Jung Chu,Kyong Hoon Ahn,Seok Kyun Kim,Jung Eun Ji,Jong Hoon Won,김형철,백문정,김대경 한국분자세포생물학회 2011 Molecules and cells Vol.32 No.4

Ceramide has been suggested to be not only a tumor-suppressive lipid but also a regulator of phagocytosis. We examined whether exogenous cell-permeable C_6-ceramide enhances the phagocytic activity of Kupffer cells (KCs) and affects the level of cellular ceramides. Rat KCs were isolated by collagenase digestion and differential centrifugation, using Percoll system. Phagocytic activity was measured by FACS analysis after incubating KCs with fluorescence-conjugated latex beads, and the level of cellular ceramide was analyzed by liquid chromatography tandem-mass spectrometry (LC-MS/MS). In this study we found that permeable C6-ceramide increases the cellular levels of endogenous ceramides via a sphingosine-re-cycling pathway leading to enhanced phagocytosis by KCs.