The Peptidylarginine Deiminase Inhibitor Cl-Amidine Suppresses Inducible Nitric Oxide Synthase Expression in Dendritic Cells

Jang, Byungki,Ishigami, Akihito,Kim, Yong-Sun,Choi, Eun-Kyoung MDPI AG 2017 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.18 No.11

<P>The conversion of peptidylarginine into peptidylcitrulline by calcium-dependent peptidylarginine deiminases (PADs) has been implicated in the pathogenesis of a number of diseases, identifying PADs as therapeutic targets for various diseases. The PAD inhibitor Cl-amidine ameliorates the disease course, severity, and clinical manifestation in multiple disease models, and it also modulates dendritic cell (DC) functions such as cytokine production, antigen presentation, and T cell proliferation. The beneficial effects of Cl-amidine make it an attractive compound for PAD-targeting therapeutic strategies in inflammatory diseases. Here, we found that Cl-amidine inhibited nitric oxide (NO) generation in a time- and dose-dependent manner in maturing DCs activated by lipopolysaccharide (LPS). This suppression of NO generation was independent of changes in NO synthase (NOS) enzyme activity levels but was instead dependent on changes in inducible NO synthase (iNOS) transcription and expression levels. Several upstream signaling pathways for iNOS expression, including the mitogen-activated protein kinase, nuclear factor-κB p65 (NF-κB p65), and hypoxia-inducible factor 1 pathways, were not affected by Cl-amidine. By contrast, the LPS-induced signal transducer and the activator of transcription (STAT) phosphorylation and activator protein-1 (AP-1) transcriptional activities (c-Fos, JunD, and phosphorylated c-Jun) were decreased in Cl-amidine-treated DCs. Inhibition of Janus kinase/STAT signaling dramatically suppressed iNOS expression and NO production, whereas AP-1 inhibition had no effect. These results indicate that Cl-amidine-inhibited STAT activation may suppress iNOS expression. Additionally, we found mildly reduced cyclooxygenase-2 expression and prostaglandin E2 production in Cl-amidine-treated DCs. Our findings indicate that Cl-amidine acts as a novel suppressor of iNOS expression, suggesting that Cl-amidine has the potential to ameliorate the effects of excessive iNOS/NO-linked immune responses.</P>

The Redox-Sensitive DNA Binding Sites Responsible for Age-Related Downregulation of SMP30 by ERK Pathway and Reversal by Calorie Restriction

Jung, Kyung Jin,Maruyama, Naoki,Ishigami, Akihito,Yu, Byung Pal,Chung, Hae Young Mary Ann Liebert 2006 Antioxidants & redox signaling Vol.8 No.3

Vitamin C deficiency increases the binucleation of hepatocytes in SMP30 knock-out mice

Park, Jin-Kyu,Hong, Il-Hwa,Ki, Mi-Ran,Chung, Hae-Young,Ishigami, Akihito,Ji, Ae-Ri,Goo, Moon-Jung,Kim, Dong-Hwan,Kwak, Ji-Hoon,Min, Chang-Woo,Lee, Seung-Sook,Jeong, Kyu-Shik Blackwell Publishing Asia 2010 Journal of Gastroenterology and Hepatology Vol.25 No.11

<P>Abstract</P><P>Background and Aims: </P><P>The binucleation of hepatocytes, which was known as an important feature of liver growth and physiology, has been reported to be increased during the chronic oxidative injury stage and has been regarded as an age-related change of hepatic structures. Therefore, we investigated the binuclearity pattern in the livers of senescence marker proteins-30 (SMP30) knock-out (KO) mice compared with wild-type (WT) mice and vitamin C-treated KO (KO + VC) mice.</P><P>Methods: </P><P>The WT, KO and KO + VC mice were fed a vitamin C free diet and VC(+) group mice were given vitamin C water containing 1.5 g/L of vitamin C, whereas VC(−) group was given normal drinking water without vitamin C, for 16 weeks.</P><P>Results: </P><P>In microscopic examination, the livers of KO mice showed a significantly increased number of binuclear hepatocytes compared with that of WT mice and KO + VC mice. KO mice also showed the most increased expression level of CYP2E1 and PCNA determined by immunohistochemistry and immunoblot analysis. Moreover, KO mice indicated the highest level of serum alanine aminotransferase and aspartate aminotransferase level in serum biochemical analysis. Accordingly, significantly decreased levels of reactive oxygen species, MDA (malondialdehyde) and HAE (4-hydroxyalkenals) were detected in KO + VC mice compared with KO mice.</P><P>Conclusion: </P><P>Therefore, it is concluded that vitamin C deficiency induces an increase of CYP2E1 expression and elevated ROS production, which causes oxidative liver injury and the elevation of hepatocyte binucleation in SMP30 KO mice.</P>

Protective effects of KI against on the acute hepatic Liver injury of SMP30 knock-out mice

( Moon Jung Goo ),( Hye Rim Lee ),( Mi Ran Ki ),( Hae Young Chung ),( Akihito Ishigami ),( Kyu Shik Jeong ),( Jin Kyu Park ),( Da Hee Jeong ),( Ho Yong Park ),( Kwang Hee Son ),( Dong Ha Sin ),( Sun H 한국수의병리학회 2006 한국수의병리학회 학술발표논문집 Vol.10 No.-

Senescence marker protein 30 protects intestinal epithelial cells against inflammation-induced cell death by enhancing Nrf2 activity

Choo, Jieun,Heo, Gwangbeom,Kim, Su Jin,Lee, Yunna,Ishigami, Akihito,Maruyama, Naoki,Chung, Hae Young,Im, Eunok Elsevier 2018 Biochimica et biophysica acta. Molecular basis of Vol.1864 No.12

<P><B>Abstract</B></P> <P>Senescence marker protein 30 (SMP30) is a calcium-binding protein whose expression decreases during senescence. SMP30 deficiency increases susceptibility to cytokine-induced apoptosis in the liver and to radiation-induced apoptosis in the small intestine. Furthermore, colonic epithelial cell death is associated with the severity of colitis. Therefore, in the present study, we investigated the function of SMP30 during intestinal inflammation. In SMP30 deficient mice, colitis was significantly exacerbated as demonstrated by increased mortality (<I>p</I> = 0.001), body weight loss (<I>p</I> = 0.0105 at day 8), rectal bleeding (<I>p</I> = 0.0047 at day 8) and diarrhea (<I>p</I> = 0.0030 at day 8), histological scores (ulcers, <I>p</I> = 0.0002; edema, <I>p</I> = 0.0125; leukocyte infiltration, <I>p</I> = 0.0016) and productions of pro-inflammatory cytokines (IL-1α, <I>p</I> = 0.0452; IL-6, <I>p</I> = 0.0074; G-CSF, <I>p</I> = 0.0036). In addition, greater proportions of apoptotic cells and lower levels of anti-apoptotic marker proteins (total PARP-1 and Bcl-2) were observed in the inflamed intestines of SMP30 deficient mice than in wild type controls. In vitro experiments on colonic epithelial cells showed that stable SMP30 expression inhibited but that SMP30 siRNA expression increased TNF-α-induced apoptosis. SMP30 inhibition decreased Nrf2 mRNA expression levels (<I>p</I> < 0.0001), but SMP30 overexpression increased Nrf2 mRNA expression levels (<I>p</I> = 0.0495). The underlying mechanism by which SMP30 protected cells appeared to be by inhibiting Nrf2 ubiquitination and Keap1 expression, and thus enhancing Nrf2 activity. Moreover, SMP30 deficiency increased the incidence of colitis-associated colon cancer as determined by increased mortality (<I>p</I> = 0.0572) and average polyp number (<I>p</I> = 0.0277). Collectively, these findings suggest that SMP30 protects intestinal epithelial cells from apoptosis and this can contribute to amelioration of colitis and colitis-associated colon cancer.</P> <P><B>Highlights</B></P> <P> <UL> <LI> SMP30 deficiency exacerbates colitis and colitis-associated colon cancer. </LI> <LI> SMP30 prevents epithelial cells from cytokine-induced cell death. </LI> <LI> SMP30 increases Nrf2 activity by inhibiting Nrf2 ubiquitination and Keap1 expression. </LI> <LI> Protective effect of SMP30 is mediated by increased Nrf2 activity. </LI> </UL> </P>

Poster Session 2 : DNA Replication, Recombination, and Repair Transcription, RNA Processing and Translation Protein Structure and Function ; Transcriptional Regulation of Senescence Marker Protein 30 by Age and Calorie Restriction

( Kyung Jin Jung ),( Bo Kyung Sung ),( Chul Hong Kim ),( Dae Hyun Kim ),( Akihito Ishigami ),( Naoki Maruyama ),( Hae Young Chung ) 생화학분자생물학회 2002 생화학분자생물학회 춘계학술발표논문집 Vol.2002 No.-

Subcellular Localization of Peptidylarginine Deiminase 2 and Citrullinated Proteins in Brains of Scrapie-Infected Mice: Nuclear Localization of PAD2 and Membrane Fraction-Enriched Citrullinated Proteins.

Jang, Byungki,Shin, Hae-Young,Choi, Jin-Kyu,Nguyen, Du Phuong Thao,Jeong, Byung-Hoon,Ishigami, Akihito,Maruyama, Naoki,Carp, Richard I,Kim, Yong-Sun,Choi, Eun-Kyoung Association of the Journal of Neuropathology and E 2011 Journal of neuropathology and experimental neurolo Vol.70 No.2

<P>Peptidylarginine deiminase (PAD) and citrullinated proteins have emerged as key molecules in various human diseases, but detailed subcellular localizations of PAD2 and citrullinated proteins are poorly mapped in brain under normal and pathologic conditions. We performed subcellular fractionation and electron microscopic analysis using brains of normal and scrapie-infected mice. Peptidylarginine deiminase 2 was abundantly present in cytosol and weakly in microsomal and mitochondrial fractions and expression in these fractions was higher in brains of scrapie-infected mice. Despite relatively low PAD2 expression, in microsomal and mitochondrial fractions, citrullinated proteins were present at high levels in these fractions in scrapie-infected brains. Surprisingly, increased PAD2 expression and accumulated citrullinated proteins were also found in nuclear fractions in scrapie-infected brains. By electron microscopy, PAD2 and citrullinated proteins in scrapie-infected brains were widely distributed in most cellular compartments including mitochondria, endoplasmic reticulum, glial filaments, nuclei, and Golgi apparatus in astrocytes and hippocampal neurons. Taken together, we report for the first time the nuclear localization of PAD2 and the detailed subcellular localization of PAD2 and of citrullinated proteins in scrapie-infected brains. Our findings suggest that different subcellular compartmentalization of PAD2 and citrullinated proteins may have different physiological roles in normal and neurodegenerative conditions.</P>

<i>Smad3</i> Deficiency Ameliorates Hepatic Fibrogenesis through the Expression of Senescence Marker Protein-30, an Antioxidant-Related Protein

Jeong, Da-Hee,Hwang, Meeyul,Park, Jin-Kyu,Goo, Moon-Jung,Hong, Il-Hwa,Ki, Mi-Ran,Ishigami, Akihito,Kim, Ah-Young,Lee, Eun-Mi,Lee, Eun-Joo,Jeong, Kyu-Shik Molecular Diversity Preservation International (MD 2013 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.14 No.12

<P>Smad3 is a key mediator of the transforming growth factor (TGF)-β1 signaling pathway that plays central role in inflammation and fibrosis. In present study, we evaluated the effect of Smad3 deficiency in <I>Smad3</I><SUP>−/−</SUP> mice with carbon tetrachloride (CCl<SUB>4</SUB>)-induced liver fibrosis. The animals were received CCl<SUB>4</SUB> or olive oil three times a week for 4 weeks. Histopathological analyses were performed to evaluate the fibrosis development in the mice. Alteration of protein expression controlled by Smad3 was examined using a proteomic analysis. CCl<SUB>4</SUB>-induced liver fibrosis was rarely detected in <I>Smad3</I><SUP>−/−</SUP> mice compared to <I>Smad3</I><SUP>+/+</SUP>. Proteomic analysis revealed that proteins related to antioxidant activities such as senescence marker protein-30 (<I>SMP30</I>), selenium-binding proteins (SP56) and glutathione <I>S</I>-transferases (GSTs) were up-regulated in <I>Smad3</I><SUP>−/−</SUP> mice. Western blot analysis confirmed that <I>SMP30</I> protein expression was increased in <I>Smad3</I><SUP>−/−</SUP> mice. And <I>SMP30</I> levels were decreased in CCl<SUB>4</SUB>-treated <I>Smad3</I><SUP>+/+</SUP> and <I>Smad3</I><SUP>−/−</SUP> mice. These results indicate that Smad3 deficiency influences the proteins level related to antioxidant activities during early liver fibrosis. Thus, we suggest that Smad3 deteriorate hepatic injury by inhibitor of antioxidant proteins as well as mediator of TGF-β1 signaling.</P>

Developmental and age-related changes of peptidylarginine deiminase 2 in the mouse brain

Shimada, Nobuko,Handa, Setsuko,Uchida, Yoshiaki,Fukuda, Mitsugu,Maruyama, Naoki,Asaga, Hiroaki,Choi, Eun-Kyoung,Lee, Jaewon,Ishigami, Akihito Wiley Subscription Services, Inc., A Wiley Company 2010 Journal of neuroscience research Vol.88 No.4

<P>Peptidylarginine deiminases (PADs) are a group of posttranslational modification enzymes that citrullinate (deiminate) protein arginine residues in a Ca<SUP>2+</SUP>-dependent manner. Enzymatic citrullination abolishes positive charges of native protein molecules, inevitably causing significant alterations in their structure and functions. Among the five isoforms of PADs, PAD2 and PAD4 are proved occupants of the central nervous system (CNS), and especially PAD2 is a main PAD enzyme expressed in the CNS. We previously reported that abnormal protein citrullination by PAD2 has been closely associated with the pathogenesis of neurodegenerative disorders such as Alzheimer's disease and prion disease. Protein citrullination in these patients is thought to play a role during the initiation and/or progression of disease. However, the contribution of changes in PAD2 levels, and consequent citrullination, during developmental and aging processes remained unclear. Therefore, we used quantitative real-time RT-PCR, Western blot analysis, and immunohistochemical methods to measure PAD2 expression and localization in the brain during those processes. PAD2 mRNA expression was detected in the brains of mice as early as embryonic day 15, and its expression in cerebral cortex, hippocampus, and cerebellum increased significantly as the animals aged from 3 to 30 months old. No citrullinated proteins were detected during that period. Moreover, we found here, for the first time, that PAD2 localized specifically in the neuronal cells of the cerebral cortex and Purkinje cells of the cerebellum. These findings indicate that, despite PAD2's normally inactive status, it becomes active and citrullinates cellular proteins, but only when the intracellular Ca<SUP>2+</SUP> balance is upset during neurodegenerative changes. © 2009 Wiley-Liss, Inc.</P>

Senescence marker protein 30 is up-regulated in kainate-induced hippocampal damage through ERK-mediated astrocytosis

Son, Tae Gen,Park, Hee Ra,Kim, So Jung,Kim, Keunho,Kim, Min-Sun,Ishigami, Akihito,Handa, Setsuko,Maruyama, Naoki,Chung, Hae Young,Lee, Jaewon Wiley Subscription Services, Inc., A Wiley Company 2009 Journal of neuroscience research Vol.87 No.13

<P>Senescence maker protein 30 (SMP30) is decreased in an androgen-independent manner in kidney and liver with age. However, regulation of SMP30 expression in the brain has not been examined in aging and neurodegenerative diseases. To investigate SMP30 expression in the brain, we utilized aging and kainate (KA)-induced neurodegenerative disease models. Interestingly, expression of SMP30 was unlikely to decrease in the aged brain, but total levels of SMP30 protein were increased at 4 weeks after KA injury. Increased glial fibrillary acidic protein (GFAP) with elevated SMP30 expression was observed at the same time post-KA, indicating that regulation of SMP30 expression in the brain may be associated with astrocytosis. We confirmed that KA induced GFAP expression with increased SMP30 in rat astrocyte cells. Moreover, we found that ERK1/2 activation was involved in the up-regulation of SMP30 in astrocytes. Our results suggest that elevated SMP30 in activated astrocytes plays an important supportive role after brain damage. © 2009 Wiley-Liss, Inc.</P>