Gα₁₂ gep oncogene deregulation of p53-responsive microRNAs promotes epithelial–mesenchymal transition of hepatocellular carcinoma

Yang, Y M,Lee, W H,Lee, C G,An, J,Kim, E-S,Kim, S H,Lee, S-K,Lee, C H,Dhanasekaran, D N,Moon, A,Hwang, S,Lee, S J,Park, J-W,Kim, K M,Kim, S G Macmillan Publishers Limited 2015 Oncogene Vol.34 No.22

Hepatocellular carcinoma (HCC) has a poor prognosis owing to aggressive phenotype. Gα<SUB>12</SUB> gep oncogene product couples to G-protein-coupled receptors, whose ligand levels are frequently increased in tumor microenvironments. Here, we report Gα<SUB>12</SUB> overexpression in human HCC and the resultant induction of zinc-finger E-box-binding homeobox 1 (ZEB1) as mediated by microRNA deregulation. Gα<SUB>12</SUB> expression was higher in HCC than surrounding non-tumorous tissue. Transfection of Huh7 cell with an activated mutant of Gα<SUB>12</SUB> (Gα<SUB>12</SUB>QL) deregulated microRNA (miRNA or miR)-200b/a/429, -194-2/192 and -194-1/215 clusters in the miRNome. cDNA microarray analyses disclosed the targets affected by Gα<SUB>12</SUB> gene knockout. An integrative network of miRNAs and mRNA changes enabled us to predict ZEB1 as a key molecule governed by Gα<SUB>12</SUB>. Decreases of miR-200a/b, -192 and -215 by Gα<SUB>12</SUB> caused ZEB1 induction. The ability of Gα<SUB>12</SUB> to decrease p53 levels, as a result of activating protein-1 (AP-1)/c-Jun-mediated mouse double minute 2 homolog induction, contributed to transcriptional deregulation of the miRNAs. Gα<SUB>12</SUB>QL induced ZEB1 and other epithelial–mesenchymal transition markers with fibroblastoid phenotype change. Consistently, transfection with miR-200b, -192 or -215 mimic prevented the ability of Gα<SUB>12</SUB>QL to increase tumor cell migration/invasion. In xenograft studies, sustained knockdown of Gα<SUB>12</SUB> decreased the overall growth rate and average volume of tumors derived from SK-Hep1 cell (mesenchymal-typed). In HCC patients, miR-192, -215 and/or -200a were deregulated with microvascular invasion or growth advantage. In the HCC samples with higher Gα<SUB>12</SUB> level, a correlation existed in the comparison of relative changes of Gα<SUB>12</SUB> and ZEB1. In conclusion, Gα<SUB>12</SUB> overexpressed in HCC causes ZEB1 induction by deregulating p53-responsive miRNAs, which may facilitate epithelial–mesenchymal transition and growth of liver tumor. These findings highlight the significance of Gα<SUB>12</SUB> upregulation in liver tumor progression, implicating Gα<SUB>12</SUB> as an attractive therapeutic target.

Rapid and label-free bioanalytical method of alpha fetoprotein detection using LSPR chip

Kim, D.,Kim, J.,Kwak, C.H.,Heo, N.S.,Oh, S.Y.,Lee, H.,Lee, G.W.,Vilian, A.T.E.,Han, Y.K.,Kim, W.S.,Kim, G.b.,Kwon, S.,Huh, Y.S. North-Holland Pub. Co 2017 Journal of crystal growth Vol.469 No.-

Alpha fetoprotein (AFP) is a cancer marker, particularly for hepatocellular carcinoma. Normal levels of AFP are less than 20ng/mL; however, its levels can reach more than 400ng/mL in patients with HCC. Enzyme linked immunosorbent assay (ELISA) and radioimmunoassay (RIA) have been employed for clinical diagnosis of AFP; however, these methods are time consuming and labor intensive. In this study, we developed a localized surface plasmon resonance (LSPR) based biosensor for simple and rapid detection of AFP. This biosensor consists of a UV-Vis spectrometer, a cuvette cell, and a biosensor chip nanopatterned with gold nanoparticles (AuNPs). In our LSPR biosensor, binding of AFP to the surface of the sensor chip led to an increasing magnitude of the LSPR signals, which was measured by an ultraviolet-visible (UV-Vis) spectrometer. Our LSPR biosensor showed sufficient detectability of AFP at concentrations of 1ng/mL to 1μg/mL. Moreover, the overall procedure for detection of AFP was completed within 20min. This biosensor could also be utilized for a point of care test (POCT) by employing a portable UV-Vis spectrometer. Owing to the simplicity and rapidity of the detection process, our LSPR biosensor is expected to replace traditional diagnostic methods for the early detection of diseases.

Structural and functional effects of manipulating the degree of methylesterification in a model homogalacturonan with a pseudo-random fungal pectin methylesterase followed by a processive methylesterase

Kim, Yang,Cameron, Randall G.,Williams, Martin A.K.,Luzio, Gary A. Elsevier 2018 Food hydrocolloids Vol.77 No.-

<P><B>Abstract</B></P> <P>We explored the possibility of controlling charge distribution in the homogalacturonan regions of pectin to produce a population of demethylesterified molecules with desirable functional properties by utilizing consecutive treatments with pectin methylesterases (PME) having different modes of action. A fungal PME from <I>Aspergillus aculeatus</I> (<I>Aa</I>-PME), with a pseudo-random mode of action, was used to demethylesterify a extremely high methylesterified HG (DM 94%, average degree of polymerization 246) by reducing the degree of methylesterification (DM) from 94% to either 70% or 80%. A second demethylesterification step, to 50% DM, was performed using a processive PME from <I>Carica papaya</I> (<I>CpL</I>-PME). Introduced demethylesterified blocks were released either by exhaustive or limited endo polygalacturonase (EPG) digestion. Degree of blockiness (DB), absolute degree of blockiness (DB<SUB>abs</SUB>), average demethylesterified block size ( B S ¯ ) and number of average sized demethylesterified blocks per molecule ( B N ¯ ) were estimated. B S ¯ and B N ¯ as well as DB/DB<SUB>abs</SUB> differed depending on the initial DM reduction by <I>Aa</I>PME, the number of activity units of <I>CpL</I>PME used and the reaction pH (<I>P</I> < 0.05). Consecutive demethylesterification of HG by <I>AaPME</I> to 80% DM and then by <I>CpL</I>PME to 50% DM at pH 4.5 produced significantly longer oligomer blocks compared to <I>Aa-PME</I> demethylesterification to 70% DM followed by <I>CpL</I>-PME to 50% DM at pH 7.0. Limited EPG digestion released nearly intact demethylesterified blocks and the released oligomers were coupled with <I>in silico</I> modeling. Resulting oligomer distribution corresponded to the <I>in silico</I> mode of action representing contiguous demethylesterification of 10 GalA residues rather than that of random or complete block-wise demethylesterification. Calcium-mediated gels of the modified HGs displayed G′ higher than G″ values and both moduli differed significantly according to the amount of <I>CpL</I>PME applied even though their final DMs were identical. These results suggest the possibility of controlling B S ¯ and engineering a population of demethylesterified pectin molecules with specified demethylesterified B S ¯ and functional properties.</P> <P><B>Highlights</B></P> <P> <UL> <LI> A fungal and a plant pectin methylesterase were sequentially applied to a model homogalacturonan. </LI> <LI> Average demethylesterified block size and number differed based on the treatment. </LI> <LI> A processive multiple attack mode of action best explained block distributions. </LI> <LI> Significant correlations between B S ¯ , B N ¯ , degree of blockiness and G′ were observed. </LI> <LI> Functional properties could be customized using PMEs of different mode of actions. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

A study of nerve agent model organophosphonate binding with manganese-A₂B-corrole and -A₂B₂-porphyrin systems

Kim, K.,Kim, I.,Maiti, N.,Kwon, S.J.,Bucella, D.,Egorova, O.A.,Lee, Y.S.,Kwak, J.,Churchill, D.G. Pergamon Press 2009 Polyhedron Vol.28 No.12

Herein the synthesis and binding studies of novel trans-A<SUB>2</SUB>B-corrole and trans-A<SUB>2</SUB>B<SUB>2</SUB>-porphyrin derivatives are presented in comparing manganese(III)-organophosphonate (OP) binding (e.g., M<SUP>n+</SUP>←O?PR(OR)<SUB>2</SUB>) capabilities. H<SUB>3</SUB>(PFP-VC) [PFP-VC=5,15-di(pentafluorophenyl)-10-(3-vinylphenyl)corrolate] was synthesized by way of literature procedures and was characterized by a variety of 2-D NMR spectroscopic techniques and single-crystal X-ray diffraction. These compounds represent the first example of 3-vinyl-phenyl-containing meso-substituted corroles or porphyrins. Mn(PFP-VC) (3) was treated separately with (CH<SUB>3</SUB>CH<SUB>2</SUB>O)<SUB>2</SUB>P?O(C<SUB>3</SUB>H<SUB>6</SUB>NMe<SUB>2</SUB>), (C<SUB>4</SUB>H<SUB>9</SUB>O)<SUB>2</SUB>P?O(Me), (C<SUB>2</SUB>H<SUB>5</SUB>O)<SUB>2</SUB>P?O(CH<SUB>2</SUB>COCH<SUB>3</SUB>), (CH<SUB>3</SUB>CH<SUB>2</SUB>O)<SUB>2</SUB>P?O(Me), to give 1:1 adducts, as determined by UV-Vis spectroscopy (Job Plot), giving a red shift; Ph<SUB>3</SUB>P?O, was also found to bind, but very weakly. The trans-A<SUB>2</SUB>B<SUB>2</SUB>-porphyrin analogue Mn(PFP-VP) (4) was also prepared by way of a literature procedure; related binding studies gave 1:1 organophosphonate-Mn(PFP-VP) adducts (Job Plot). A clean blue shift occurred for the Mn-porphyrins at higher organophosphonate loadings (K<SUB>a</SUB> values: 6.7 (0.9)-11.9 (0.4)M<SUP>-1</SUP>). DFT geometry optimizations of O?P(OMe)<SUB>2</SUB>Me binding and formal Mn-O or P-O cleavage products in the unsubstituted neutral Mn-corrolato and -porphyrinato systems with a range of metal-based spin states revealed greatest stability in formal phosphoryl oxygen binding (energies: 11-13kcal/mol) for the Mn-corrole (singlet); the Mn-porphyrin (sextet) was also quite stable.

Gut-Specific Delivery of T-Helper 17 Cells Reduces Obesity and Insulin Resistance in Mice

Hong, C.P.,Park, A.,Yang, B.G.,Yun, C.H.,Kwak, M.J.,Lee, G.W.,Kim, J.H.,Jang, M.S.,Lee, E.J.,Jeun, E.J.,You, G.,Kim, K.S.,Choi, Y.,Park, J.H.,Hwang, D.,Im, S.H.,Kim, J.F.,Kim, Y.K.,Seoh, J.Y.,Surh, C. Elsevier North Holland [etc.] 2017 Gastroenterology Vol.152 No.8

<P>BACKGROUND & AIMS: Obesity and metabolic syndrome have been associated with alterations to the intestinal microbiota. However, few studies examined the effects of obesity on the intestinal immune system. We investigated changes in subsets of intestinal CD4(+) T-helper (T-H) cells with obesity and the effects of gut-tropic T(H)17 cells in mice on a high-fat diet (HFD). METHODS: We isolated immune cells from small intestine and adipose tissue of C57BL/6 mice fed a normal chow diet or a HFD for 10 weeks and analyzed the cells by flow cytometry. Mice fed a vitamin A-deficient HFD were compared with mice fed a vitamin A-sufficient HFD. Obese RAG1-deficient mice were given injections of only regulatory T cells or a combination of regulatory T cells and T(H)17 cells (wild type or deficient in integrin beta 7 subunit or interleukin 17 [IL17]). Mice were examined for weight gain, fat mass, fatty liver, glucose tolerance, and insulin resistance. Fecal samples were collected before and after T cell transfer and analyzed for microbiota composition by metagenomic DNA sequencing and quantitative polymerase chain reaction. RESULTS: Mice placed on a HFD became obese, which affected the distribution of small intestinal CD4(+) T-H cells. Intestinal tissues from obese mice had significant reductions in the proportion of T(H)17 cells but increased proportion of T(H)1 cells, compared with intestinal tissues from nonobese mice. Depletion of vitamin A in obese mice further reduced the proportion of T(H)17 cells in small intestine; this reduction correlated with more weight gain and worsening of glucose intolerance and insulin resistance. Adoptive transfer of in vitro-differentiated gut-tropic T(H)17 cells to obese mice reduced these metabolic defects, which required the integrin beta 7 subunit and IL17. Delivery of T(H)17 cells to intestines of mice led to expansion of commensal microbes associated with leanness. CONCLUSIONS: In mice, intestinal T(H)17 cells contribute to development of a microbiota that maintains metabolic homeostasis, via IL17. Gut-homing T(H)17 cells might be used to reduce metabolic disorders in obese individuals.</P>

Novel anti-apoptotic mechanism of A20 through targeting ASK1 to suppress TNF-induced JNK activation

Won, M,Park, K A,Byun, H S,Sohn, K-C,Kim, Y-R,Jeon, J,Hong, J H,Park, J,Seok, J H,Kim, J M,Yoon, W-H,Jang, I-S,Shen, H M,Liu, Z G,Hur, G M Macmillan Publishers Limited 2010 CELL DEATH AND DIFFERENTIATION Vol.17 No.12

The zinc-finger protein A20 has crucial physiological functions as a dual inhibitor of nuclear factor-κB (NF-κB) activation and apoptosis in tumor necrosis factor (TNF) receptor 1 signaling pathway. Although the molecular basis for the anti-NF-κB function of A20 has been well elucidated, the anti-apoptotic function of A20 is largely unknown. Here, we report a novel mechanism underlying the anti-apoptotic function of A20: A20 blocks TNF-induced apoptosis through suppression of c-jun N-terminal kinase (JNK) by targeting apoptosis signal-regulating kinase1 (ASK1). First, the ectopic expression of A20 drastically inhibits TNF-induced JNK activation and apoptosis in multiple cell types including those deficient of NF-κB activation. Unexpectedly, the blunting effect of A20 on TNF-induced JNK activation is not mediated by affecting the TNFR1 signaling complex formation. Instead, A20 interacts with ASK1, an important MAPKK kinase in the JNK signaling cascade. More importantly, overexpression of wild-type A20, but not of mutant A20 (ZnF4; C624A, C627A), promotes degradation of the ASK1 through the ubiquitin-proteasome system. Taken together, the results from this study reveal a novel anti-apoptotic mechanism of A20 in TNF signaling pathway: A20 binds to ASK1 and mediates ASK1 degradation, leading to suppression of JNK activation and eventually blockage of apoptosis.

Simulation of single grid-based phase-contrast x-ray imaging (g-PCXI)

Lim, H.W.,Lee, H.W.,Cho, H.S.,Je, U.K.,Park, C.K.,Kim, K.S.,Kim, G.A.,Park, S.Y.,Lee, D.Y.,Park, Y.O.,Woo, T.H.,Lee, S.H.,Chung, W.H.,Kim, J.W.,Kim, J.G. Elsevier BV * North-Holland 2017 Nuclear Instruments & Methods in Physics Research. Vol. No.

<P><B>Abstract</B></P> <P>Single grid-based phase-contrast x-ray imaging (g-PCXI) technique, which was recently proposed by Wen et al. to retrieve absorption, scattering, and phase-gradient images from the raw image of the examined object, seems a practical method for phase-contrast imaging with great simplicity and minimal requirements on the setup alignment. In this work, we developed a useful simulation platform for g-PCXI and performed a simulation to demonstrate its viability. We also established a table-top setup for g-PCXI which consists of a focused-linear grid (200-lines/in strip density), an x-ray tube (100-μm focal spot size), and a flat-panel detector (48-μm pixel size) and performed a preliminary experiment with some samples to show the performance of the simulation platform. We successfully obtained phase-contrast x-ray images of much enhanced contrast from both the simulation and experiment and the simulated contract seemed similar to the experimental contrast, which shows the performance of the developed simulation platform. We expect that the simulation platform will be useful for designing an optimal g-PCXI system.</P> <P><B>Highlights</B></P> <P> <UL> <LI> It is proposed for the single grid-based phase-contrast x-ray imaging (g-PCXI) technique. </LI> <LI> We implemented for a numerical simulation code. </LI> <LI> The preliminary experiment with several samples to compare is performed. </LI> <LI> It is expected to be useful to design an optimal g-PCXI system. </LI> </UL> </P>

인플루엔자 유행시 국가방역 관리 능력 향상을 위한 기초연구

김지희,김우주,이주연,나병국,박종원,신구철,김선숙,김진아,고연아,박순희,김도근,임윤구 국립보건연구원 1999 국립보건원보 Vol.36 No.-

Aβ-Induced Drp1 phosphorylation through Akt activation promotes excessive mitochondrial fission leading to neuronal apoptosis

Kim, D.I.,Lee, K.H.,Gabr, A.A.,Choi, G.E.,Kim, J.S.,Ko, S.H.,Han, H.J. Elsevier Biomedical Press 2016 Biochimica et biophysica acta, Molecular cell rese Vol.1863 No.11

Mitochondrial dysfunction is known as one of causative factors in Alzheimer's disease (AD), inducing neuronal cell death. Mitochondria regulate their functions through changing their morphology. The present work was undertaken to investigate whether Amyloid β (Aβ) affects mitochondrial morphology in neuronal cells to induce apoptosis. Aβ treatment induced not only the fragmentation of mitochondria but also neuronal apoptosis in association with an increase in caspase-9 and -3 activity. Calcium influx induced by Aβ up-regulated the activation of Akt through CaMKII resulting in changes to the phosphorylation level of Drp1 in a time-dependent manner. Translocation of Drp1 from the cytosol to mitochondria was blocked by CB-124005 (an Akt inhibitor). Recruitment of Drp1 to mitochondria led to ROS generation and mitochondrial fission, accompanied by dysfunction of mitochondria such as loss of membrane potential and ATP production. ROS generation and mitochondrial dysfunction by Aβ were attenuated when treated with Mdivi-1, a selective Drp1 inhibitor. Furthermore, the sustained Akt activation induced not only the fragmentation of mitochondria but also the activation of mTOR, eventually suppressing autophagy. Inhibition of autophagic clearance of Aβ led to increased ROS levels and aggravating mitochondrial defects, which were blocked by Rapamycin (an mTOR inhibitor). In conclusion, sustained phosphorylation of Akt by Aβ directly activates Drp1 and inhibits autophagy through the mTOR pathway. Together, these changes elicit abundant mitochondrial fragmentation resulting in ROS-mediated neuronal apoptosis.

Pinoresinol-4,4'-di-O-β-d-glucoside from Valeriana officinalis root stimulates calcium mobilization and chemotactic migration of mouse embryo fibroblasts

Do, K.H.,Choi, Y.W.,Kim, E.K.,Yun, S.J.,Kim, M.S.,Lee, S.Y.,Ha, J.M.,Kim, J.H.,Kim, C.D.,Son, B.G.,Kang, J.S.,Khan, I.A.,Bae, S.S. G. Fischer 2009 Phytomedicine Vol.16 No.6

Lignans are major constituents of plant extracts and have important pharmacological effects on mammalian cells. Here we showed that pinoresinol-4,4'-di-O-β-d-glucoside (PDG) from Valeriana officinalis induced calcium mobilization and cell migration through the activation of lysophosphatidic acid (LPA) receptor subtypes. Stimulation of mouse embryo fibroblast (MEF) cells with 10μM PDG resulted in strong stimulation of MEF cell migration and the EC<SUB>50</SUB> was about 2μM. Pretreatment with pertussis toxin (PTX), an inhibitor of G<SUB>i</SUB> protein, completely blocked PDG-induced cell migration demonstrating that PDG evokes MEF cell migration through the activation of the G<SUB>i</SUB>-coupled receptor. Furthermore, pretreatment of MEF cells with Ki16425 (10μM), which is a selective antagonist for LPA<SUB>1</SUB> and LPA<SUB>3</SUB> receptors, completely blocked PDG-induced cell migration. Likewise, PDG strongly induced calcium mobilization, which was also blocked by Ki16425 in a dose-dependent manner. Prior occupation of the LPA receptor with LPA itself completely blocked PDG-induced calcium mobilization. Finally, PDG-induced MEF cell migration was attenuated by pretreatment with a phosphatidylinositol 3-kinase (PI3K) inhibitor such as LY294002. Cells lacking downstream mediator of PI3K such as Akt1 and Akt2 (DKO cells) showed loss of PDG-induced migration. Re-expression of Akt1 (but not Akt2) completely restored PDG-induced DKO cell migration. Given these results, we conclude that PDG is a strong inducer of cell migration. We suggest that the pharmacological action of PDG may occur through the activation of an LPA receptor whereby activation of PI3K/Akt signaling pathway mediates PDG-induced MEF cell migration.