Shikonin Isolated from Lithospermum erythrorhizon Downregulates Proinflammatory Mediators in Lipopolysaccharide-Stimulated BV2 Microglial Cells by Suppressing Crosstalk between Reactive Oxygen Species and NF-κB

( Rajapaksha Gedara Prasad ),( Yung Hyun Choi ),( Gi Young Kim ) 한국응용약물학회 2015 Biomolecules & Therapeutics(구 응용약물학회지) Vol.23 No.2

According to the expansion of lifespan, neuronal disorder based on inflammation has been social problem. Therefore, we isolated shikonin from Lithospermum erythrorhizon and evaluated anti-inflammatory effects of shikonin in lipopolysaccharide (LSP)- stimulated BV2 microglial cells. Shikonin dose-dependently inhibits the expression of the proinflammatory mediators, nitric oxide (NO), prostaglandin E2 (PGE2), and tumor necrosis factor-α (TNF-α) as well as their main regulatory genes and products such as inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), and TNF-α in LPS-stimulated BV2 microglial cells. Additionally, shikonin suppressed the LPS-induced DNA-binding activity of nuclear factor-κB (NF-κB) to regulate the key regulatory genes of the proinflammatory mediators, such as iNOS, COX-2, and TNF-α, accompanied with downregulation of reactive oxygen species (ROS) generation. The results indicate that shikonin may downregulate the expression of proinflammatory genes involved in the synthesis of NO, PGE2, and TNF-α in LPS-treated BV2 microglial cells by suppressing ROS and NF-κB. Taken together, our results revealed that shikonin exerts downregulation of proinflammatory mediators by interference the ROS and NF-κB signaling pathway.

Camptothecin enhances c-Myc-mediated endoplasmic reticulum stress and leads to autophagy by activating Ca²⁺-mediated AMPK

Jayasooriya, Rajapaksha Gedara Prasad Tharanga,Dilshara, Matharage Gayani,Karunarathne, Wisurumuni Arachchilage Hasitha Maduranga,Molagoda, Ilandarage Menu Neelaka,Choi, Yung Hyun,Kim, Gi-Young Elsevier 2018 Food and chemical toxicology Vol.121 No.-

<P><B>Abstract</B></P> <P>Camptothecin (CPT) from <I>Camptotheca acuminate</I> was discovered for anticancer drugs, which targets topoisomease I. However, whether CPT regulates c-Myc expression has not been understood in endoplasmic reticulum (ER) stress and autophagy. In this study, we found that CPT enhanced c-Myc expression and that the transient knockdown of <I>c-Myc</I> abrogated reactive oxygen species (ROS) generation, which resulted in the accumulation of ER stress-regulating proteins, such as PERK, eIF2α, ATF4, and CHOP. Moreover, the transfection of <I>eIF2α</I>-targeted siRNA attenuated CPT-induced autophagy and decreased the levels of Beclin-1 and Atg7, which indicated that CPT upregulated ER stress-mediated autophagy. In addition, CPT phosphorylated AMPK in response to intracellular Ca<SUP>2+</SUP> release. Ca<SUP>2+</SUP> chelators, ethylene glycol tetraacetic acid and a CaMKII inhibitor, K252a, decreased CPT-induced Beclin-1 and Atg7, and downregulated AMPK phosphorylation, which suggested that CPT-induced Ca<SUP>2+</SUP> release leads to the activation of autophagy through CaMKII-mediated AMPK phosphorylation. CPT also phosphorylated JNK and activated the DNA-binding activity of AP-1; furthermore, knockdown of <I>JNK</I> abolished the expression level of Beclin-1 and Atg7, which implied that the JNK-AP-1 pathway was a potent mediator of CPT-induced autophagy. Our findings indicated that CPT promoted c-Myc-mediated ER stress and ROS generation, which enhances autophagy via the Ca<SUP>2+</SUP>-AMPK and JNK-AP-1 pathways.</P> <P><B>Highlights</B></P> <P> <UL> <LI> CPT induces c-Myc-mediated ROS formation, leading to CHOP expression. </LI> <LI> c-Myc positively regulates CPT-induced ER stress by increasing ROS generation. </LI> <LI> CPT promotes autophagy formation as a result of ER stress. </LI> <LI> CPT promotes autophagy through increased intracellular Ca2+ release. </LI> <LI> CPT induces JNK-mediated autophagy by enhancing AP-1 activity. </LI> </UL> </P>

Molecular chemotherapeutic potential of butein: A concise review

Jayasooriya, Rajapaksha Gedara Prasad Tharanga,Molagoda, Ilandarage Menu Neelaka,Park, Cheol,Jeong, Jin-Woo,Choi, Yung Hyun,Moon, Dong-Oh,Kim, Mun-Ock,Kim, Gi-Young Elsevier 2018 Food and chemical toxicology Vol.112 No.-

<P><B>Abstract</B></P> <P>Butein is a biologically active flavonoid isolated from the bark of <I>Rhus verniciflua</I> Stokes, which is known to have therapeutic potential against various cancers. Notably, butein inhibits cancer cell growth by inducing G<SUB>2</SUB>/M phase arrest and apoptosis. Butein-induced G<SUB>2</SUB>/M phase arrest is associated with increased phosphorylation of ataxia telangiectasia mutated (ATM) and Chk1/2, and consequently, with reduced cdc25C levels. In addition, butein-induced apoptosis is mediated through the activation of caspase-3, which is associated with changes in the expression of Bcl-2 and Bax proteins. Intriguingly, butein sensitizes cells to tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis via ERK-mediated Sp1 activation, which promotes the transcription of specific death receptor 5. Butein also inhibits the migration and invasion of human cancer cells by suppressing nuclear factor-κB- and extracellular signal-regulated kinases 1/2-mediated expression of matrix metalloproteinase-9 and vascular endothelial growth factor. Additionally, butein downregulates the expression of human telomerase reverse transcriptase and causes a concomitant decrease in telomerase activity. These findings provide the basis for the pharmaceutical development of butein. The aim of this review is to provide an update on the mechanisms underlying the anticancer activity of butein, with a special focus on its effects on different cellular signaling cascades.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Butein induces apoptosis by activating Bax-caspase-3-PARP axis by PI3K/Akt-NF-κB axis through the inhibition of ROS generation. </LI> <LI> Butein induces G<SUB>2</SUB>/M phase cell cycle arrest by inhibiting ROS-mediated ATM-Chk1/2-Cdc25c-cdc2/cyclin B axis. </LI> <LI> Butein enhances TRAIL-mediated apoptosis by increasing DR5 expression through ERK-mediated Sp1 activation. </LI> <LI> Butein suppresses telomerase activity by inhibiting TERT expression and phosphorylation via c-Myc and PI3K/Akt. </LI> <LI> Butein attenuates angiogenesis, invasion/metastasis, and inflammation by suppressing the NF-κB signal pathway. </LI> </UL> </P>

Combined treatment with verrucarin A and tumor necrosis factor-α sensitizes apoptosis by overexpression of nuclear factor-kappaB-mediated Fas

Jayasooriya, Rajapaksha Gedara Prasad Tharanga,Moon, Dong-Oh,Park, Sang Rul,Choi, Yung Hyun,Asami, Yukihiro,Kim, Mun-Ock,Jang, Jae-Hyuk,Kim, Bo Yeon,Ahn, Jong Seog,Kim, Gi-Young Elsevier 2013 Environmental toxicology and pharmacology Vol.36 No.2

<P><B>Abstract</B></P> <P>Verrucarin A (VA) is a member of the family of macrocyclic trichothecenes, which exhibit anti-cancer and immune-modulating activities. However, VA has not yet been demonstrated to be involved in the sensitization of tumor necrosis factor-alpha (TNF-α)-mediated apoptosis. In the present study, we found that VA triggers TNF-α-induced apoptosis in human breast cancer MDA-MB-231 and MCF-7 cells. In particular, activation of caspas-3 and caspase-8 as well as release of cytochrome <I>c</I> were significantly enhanced in response to the combined treatment with VA and TNF-α (VA/TNF-α) and the pan-caspase inhibitor z-VAD-fmk completely reversed the apoptosis, suggesting that caspases are the main effector molecules in VA/TNF-α-induced apoptosis via the intrinsic and extrinsic pathway. Moreover, we confirmed that enhanced Fas expression plays a critical role, because the Fas-blocking antibody partially inhibited VA/TNF-α-induced apoptosis. VA also increased specific DNA-binding activity of nuclear factor-kappaB (NF-κB) via nuclear translocation of p50 and p65. In addition, pretreatment with the NF-κB inhibitor MG132 blocked VA/TNF-α-induced apoptosis by suppression of NF-κB-dependent Fas expression. These results indicated that VA enhances TNF-α-induced apoptosis via NF-κB-dependent Fas overexpression.</P> <P><B>Highlights</B></P> <P> <UL> <LI> VA sensitizes TNF-α-induced apoptosis in human breast cancer cells. </LI> <LI> VA enhances TNF-α-dependent nuclear translocation of p50 and p65. </LI> <LI> VA/TNF-α triggers NF-κB-dependent Fas-induced cell death. </LI> </UL> </P>

Fulvic acid promotes extracellular anti-cancer mediators from RAW 264.7 cells, causing to cancer cell death <i>in vitro</i>

Jayasooriya, Rajapaksha Gedara Prasad Tharanga,Dilshara, Matharage Gayani,Kang, Chang-Hee,Lee, Seungheon,Choi, Yung Hyun,Jeong, Yong Kee,Kim, Gi-Young Elsevier 2016 INTERNATIONAL IMMUNOPHARMACOLOGY Vol.36 No.-

<P><B>Abstract</B></P> <P>Fulvic acid (FA) is known to promote electrochemical balance as a donor or a receptor possessing many biomedical functions. Nevertheless, the effect of FA on the anti-cancer activity has not been elucidated. In the current study, we first isolated FA from humus and investigated whether FA regulates immune-stimulating functions, such as production of nitric oxide (NO), in RAW 264.7 cells. Our data showed that FA slightly enhances cell viability in a dose-dependent manner and secretion of NO from RAW 264.7 cells. It upregulated the protein and mRNA expression of inducible NO synthesis (iNOS). In addition, FA enhanced the DNA-binding activity of nuclear factor-κB (NF-κB) in RAW 264.7 cells; the NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC) effectively attenuated the expression of FA-stimulated iNOS, suggesting that FA stimulates NF-κB to promote iNOS and NO production. Finally, FA-stimulated culture media (FA-CM) from RAW 264.7 cells were collected and MCA-102 fibrosarcoma cells were cultured in this media. The FA-CM augmented MCA-102 fibrosarcoma cell apoptosis; however, an NO inhibitor <I>N</I> <SUP>G</SUP>-monomethyl-<SMALL>L</SMALL>-arginine (NMMA) slightly inhibited the FA-CM-mediated MCA-102 fibrosarcoma cell apoptosis, which was accompanied by low levels of NO. In the present study, we found that FA induces the generation of NO and iNOS in RAW 264.7 cells by inducing NF-κB activation; however, NO did not significantly stimulate MCA-102 fibrosarcoma cell apoptosis in the current study. In addition, FA-CM enhanced cell death in various human cancer cells such as Hep3B, LNCaP, and HL60. Taken together, FA most likely stimulates immune-modulating molecules such as NO and induces cancer cell apoptosis.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Fulvic acid is isolated from humus by acid-base extraction methods. </LI> <LI> Fulvic acid increases proliferation of RAW 264.7 macrophage cells. </LI> <LI> Fulvic acid upregulates the expression of <I>iNOS</I> and NO by inducing NF-κB activity. </LI> <LI> Fulvic acid-stimulated medium of RAW 264.7 macrophages increases apoptosis of MCA-102 fibrosarcoma cells. </LI> </UL> </P>

Glutamine deprivation sensitizes human breast cancer MDA-MB-231 cells to TRIAL-mediated apoptosis

Dilshara, Matharage Gayani,Jeong, Jin-Woo,Prasad Tharanga Jayasooriya, Rajapaksha Gedara,Neelaka Molagoda, Ilandarage Menu,Lee, Seungheon,Park, Sang Rul,Choi, Yung Hyun,Kim, Gi-Young Elsevier 2017 Biochemical and biophysical research communication Vol. No.

<P><B>Abstract</B></P> <P>Tumor cell metabolism is a promising target for various cancer treatments. Apart from aerobic glycolysis, cancer cell growth is dependent on glutamine (Gln) supply, leading to their survival and differentiation. Therefore, we examined whether treatment with TNF-related apoptosis-inducing ligand (TRAIL) sensitizes MDA-MB-231 cells to apoptosis under Gln deprivation condition (TRAIL/Gln deprivation). Gln deprivation decreased cell proliferation as expected, but did not induce remarkable cell death. TRAIL/Gln deprivation, however, significantly increased growth inhibition and morphological shrinkage of MDA-MB-231 cells compared to those induced by treatment with either Gln deprivation or TRAIL alone. Moreover, TRAIL/Gln deprivation upregulated the apoptotic sub-G<SUB>1</SUB> phase accompanied with a remarkable decrease of pro-caspase-3, pro-caspase-9, and anti-apoptotic xIAP, and Bcl-2. Increased cleavage of PARP and pro-apoptotic Bid protein expression suggests that TRAIL/Gln deprivation triggers mitochondrion-mediated apoptosis in MDA-MB-231 cells. Additionally, TRAIL/Gln deprivation upregulated the expression of endoplasmic reticulum (ER) stress markers such as ATF4 and phosphorylated eIF2α, thereby enhancing the C/EBP homologous protein (CHOP) protein level. Transient knockdown of <I>CHOP</I> partically reversed TRAIL/Gln deprivation-mediated apoptosis. Accordingly, TRAIL/Gln deprivation enhanced the expression of death receptor 5 (DR5) and transient knockdown of <I>DR5</I> completely restored TRAIL/Gln deprivation-mediated apoptosis. Taken together, our results suggest that Gln deprivation conditions can be used for the development of new therapies for TRAIL-resistant cancers.</P>