The NLRP3 inflammasome plays a critical role in mediating the innate immune defense against pathogenic infections, but aberrant activation of NLRP3 inflammasome has been linked to a variety of inflammatory diseases. Thus targeting the NLRP3 inflammasome represents a promising therapeutic for the treatment of such diseases. Scutellarin is a flavonoid isolated from Erigeron breviscapus (Vant.) Hand.-Mazz. and has been reported to exhibit potent anti-inflammatory activities, but the underlying mechanism is only partly understood. In this study, we aimed to investigate whether scutellarin could affect the activation of NLRP3 inflammasome in macrophages. The results showed that scutellarin dose-dependently reduced caspase-1 activation and decreased mature interleukin-1β (IL-1β) release in lipopolysaccharide (LPS)-primed macrophages upon ATP or nigericin stimulation, indicating that scutellarin inhibited NLRP3 inflammasome activation in macrophages. Consistent with this, scutellarin also suppressed pyroptotic cell death in LPS-primed macrophages treated with ATP or nigericin. ATP or nigericin-induced ASC speck formation and its oligomerization were blocked by scutellarin pre-treatment. Intriguingly, scutellarin augmented PKA-specific phosphorylation of NLRP3 in LPS-primed macrophages, which was completely blocked by selective PKA inhibitor H89, suggesting that PKA signaling had been involved in the action of scutellarin to suppress NLRP3 inflammasome activation. Supporting this, the inhibitory effect of scutellarin on NLRP3 inflammasome activation was completely counteracted by H89 or adenyl cyclase inhibitor MDL12330A. As NLRP3-dependent release of IL-1β has a critical role in sepsis, the in vivo activity of scutellarin was assayed in a mouse model of bacterial sepsis, which was established by intraperitoneally injection of a lethal dose of viable Escherichia coli. Oral administration of scutellarin significantly improved the survival of mice with bacterial sepsis. In line with this, scutellarin treatment significantly reduced serum IL-1β levels and attenuated the infiltration of inflammatory cells in the liver of E. coli-infected mice. These data indicated that scutellarin suppressed NLRP3 inflammasome activation in macrophages by augmenting PKA signaling, highlighting its potential therapeutic application for treating NLRP3-related inflammatory diseases.

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Scutellarin Suppresses NLRP3 Inflammasome Activation in Macrophages and Protects Mice against Bacterial Sepsis

Downregulation of TRIM27 expression inhibits the proliferation of ovarian cancer cells in vitro and in vivo

// Jen-Ho Tseng 1, 2 , Cheng-Yi Chen 3 , Pei-Chun Chen 3 , Sheng-Huang Hsiao 1, 4 , Chi-Chen Fan 5 , Yu-Chih Liang 6 , Chie-Pein Chen 3, 7 1 Department of Neurosurgery, Taipei City Hospital, Renai Branch, Taipei 106, Taiwan 2 Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei 110, Taiwan 3 Department of Medical Research, MacKay Memorial Hospital, New Taipei City 251, Taiwan 4 College of Science, National Chengchi University, Taipei 116, Taiwan 5 Department of Physiology, MacKay Memorial Hospital, Taipei 104, Taiwan 6 School of Medical Laboratory Science and Biotechnology, College of Medical Science and Technology, Taipei Medical University, Taipei 110, Taiwan 7 Department of Medicine, Taipei Medical University, Taipei 110, Taiwan Correspondence to: Chie-Pein Chen, email: cpchen@mmh.org.tw Keywords: cell growth, glioblastoma multiforme, histone deacetylase, paraoxonase 2, valproic acid Received: September 30, 2015      Accepted: January 10, 2017      Published: January 18, 2017 ABSTRACT We studied the potential mechanisms of valproic acid (VPA) in the treatment of glioblastoma multiforme (GBM). Using the human U87, GBM8401, and DBTRG-05MG GBM-derived cell lines, VPA at concentrations of 5 to 20 mM induced G2/M cell cycle arrest and increased the production of reactive oxygen species (ROS). Stress-related molecules such as paraoxonase 2 (PON2), cyclin B1, cdc2, and Bcl-xL were downregulated, but p27, p21 and Bim were upregulated by VPA treatment. VPA response element on the PON2 promoter was localized at position -400/-1. PON2 protein expression was increased in GBM cells compared with normal brain tissue and there was a negative correlation between the expression of PON2 and Bim. These findings were confirmed by the public Bredel GBM microarray (Gene Expression Omnibus accession: GSE2223) and the Cancer Genome Atlas GBM microarray datasets. Overexpression of PON2 in GBM cells significantly decreased intracellular ROS levels, and PON2 expression was decreased after VPA stimulation compared with controls. Bim expression was significantly induced by VPA in GBM cells with PON2 silencing. These observations were further shown in the subcutaneous GBM8401 cell xenograft of BALB/c nude mice. Our results suggest that VPA reduces PON2 expression in GBM cells, which in turn increases ROS production and induces Bim production that inhibits cancer progression via the PON2–Bim cascade.

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Valproic acid inhibits glioblastoma multiforme cell growth via paraoxonase 2 expression

// Chen-Guang Li 1,* , Liang Yan 1,* , Yan-Yun Jing 1,* , Li-Hui Xu 2 , Yi-Dan Liang 1 , Hong-Xia Wei 1 , Bo Hu 3 , Hao Pan 1 , Qing-Bing Zha 4 , Dong-Yun Ouyang 1 and Xian-Hui He 1 1 Department of Immunobiology, College of Life Science and Technology, Jinan University, Guangzhou, China 2 Department of Cell Biology, College of Life Science and Technology, Jinan University, Guangzhou, China 3 Department of Nephrology, the First Affiliated Hospital of Jinan University, Guangzhou, China 4 Department of Fetal Medicine, the First Affiliated Hospital of Jinan University, Guangzhou, China * These authors have contributed equally to this work Correspondence to: Xian-Hui He, email: // Keywords : antibacterial infection, berberine, AMP-activated protein kinase (AMPK), inflammasome, macrophages, Immunology and Microbiology Section, Immune response, Immunity Received : August 19, 2016 Accepted : December 01, 2016 Published : December 12, 2016 Abstract The isoquinoline alkaloid berberine possesses many pharmacological activities including antibacterial infection. Although the direct bactericidal effect of berberine has been documented, its influence on the antibacterial functions of macrophages is largely unknown. As inflammasome activation in macrophages is important for the defense against bacterial infection, we aimed to investigate the influence of berberine on inflammasome activation in murine macrophages. Our results showed that berberine significantly increased ATP-induced inflammasome activation as reflected by enhanced pyroptosis as well as increased release of caspase-1p10 and mature interleukin-1β (IL-1β) in macrophages. Such effects of berberine could be suppressed by AMP-activated protein kinase (AMPK) inhibitor compound C or by knockdown of AMPKα expression, indicating the involvement of AMPK signaling in this process. In line with increased IL-1β release, the ability of macrophages to kill engulfed bacteria was also intensified by berberine. This was corroborated by the in vivo finding that the peritoneal live bacterial load was decreased by berberine treatment. Moreover, berberine administration significantly improved survival of bacterial infected mice, concomitant with increased IL-1β levels and elevated neutrophil recruitment in the peritoneal cavity. Collectively, these data suggested that berberine could enhance bacterial killing by augmenting inflammasome activation in macrophages through AMPK signaling.

/pdf/berberine-augments-atp-induced-inflammasome-activation-in-20a9ky7w8s.pdf

Berberine augments ATP-induced inflammasome activation in macrophages by enhancing AMPK signaling

The flavonoid baicalin has been reported to possess potent anti-inflammatory activities by suppressing inflammatory signaling pathways. However, whether baicalin can suppress the activation of NOD-like receptor (NLR) family, pyrin containing domain 3 (NLRP3) inflammasome in macrophages is largely unknown. Here, we showed that baicalin treatment dose-dependently inhibited adenosine triphosphate (ATP) or nigericin-induced NLRP3 inflammasome activation, as revealed by the decreased release of mature interleukin (IL)-1β, active caspase-1p10, and high-mobility group box-1 protein from lipopolysaccharide (LPS)-primed bone marrow-derived macrophages. The formation of ASC specks, a critical marker of NLRP3 inflammasome assembly, was robustly inhibited by baicalin in the macrophages upon ATP or nigericin stimulation. All these inhibitory effects of baicalin could be partly reversed by MDL12330A or H89, both of which are inhibitors of the protein kinase A (PKA) signaling pathway. Consistent with this, baicalin strongly enhanced PKA-mediated phosphorylation of NLRP3, which has been suggested to prevent ASC recruitment into the inflammasome. Of note, the PKA inhibitor H89 could block baicalin-induced NLRP3 phosphorylation on PKA-specific sites, further supporting PKA's role in this process. In addition, we showed that when administered pre and post exposure to Escherichia coli infection baicalin treatment significantly improved mouse survival in bacterial sepsis. Baicalin administration also significantly reduced IL-1β levels in the sera of bacterial infected mice. Altogether, our results revealed that baicalin inhibited NLRP3 inflammasome activation at least partly through augmenting PKA signaling, highlighting its therapeutic potential for the treatment of NLRP3-related inflammatory diseases.

/pdf/baicalin-inhibits-nod-like-receptor-family-pyrin-containing-44l77pr4q7.pdf

Baicalin Inhibits NOD-Like Receptor Family, Pyrin Containing Domain 3 Inflammasome Activation in Murine Macrophages by Augmenting Protein Kinase A Signaling.

// Gao-Xiang Zhao 1, * , Li-Hui Xu 2, * , Hao Pan 1 , Qiu-Ru Lin 1 , Mei-Yun Huang 1 , Ji-Ye Cai 3 , Dong-Yun Ouyang 1 , Xian-Hui He 1 1 Department of Immunobiology, College of Life Science and Technology, Jinan University, Guangzhou, China 2 Department of Cell Biology, College of Life Science and Technology, Jinan University, Guangzhou, China 3 Department of Chemistry, College of Life Science and Technology, Jinan University, Guangzhou, China * These authors have contributed equally to this work Correspondence to: Xian-Hui He, e-mail: thexh@jnu.edu.cn Dong-Yun Ouyang, e-mail: dongyun1967@aliyun.com Keywords: gossypol, valproic acid, apoptosis, akt, FOXO3a Received: April 29, 2015      Accepted: October 06, 2015      Published: October 16, 2015 ABSTRACT Previously we reported that valproic acid (VPA) acts in synergy with GOS to enhance cell death in human DU145 cells. However, the underlying mechanism remains elusive. In this study, we observed that such synergistic cytotoxicity of GOS and VPA could be extended to human A375, HeLa, and PC-3 cancer cells. GOS and VPA co-treatment induced robust apoptosis as evidenced by caspase-8/-9/-3 activation, PARP cleavage, and nuclear fragmentation. GOS and VPA also markedly decreased cyclin A2 protein expression. Owing to the reduction of cyclin A2, Akt signaling was suppressed, leading to dephosphorylation of FOXO3a. Consequently, FOXO3a was activated and the expression of its target genes, including pro-apoptotic FasL and Bim , was upregulated. Supporting this, FOXO3a knockdown attenuated FasL and Bim upregulation and apoptosis induction in GOS+VPA-treated cells. Furthermore, blocking proteasome activity by MG132 prevented the downregulation of cyclin A2, dephosphorylation of Akt and FOXO3a, and induction of apoptosis in cells co-treated with GOS and VPA. In mouse model, GOS and VPA combination significantly inhibited the growth of A375 melanoma xenografts. Our findings indicate that GOS and VPA co-treatment induces apoptosis in human cancer cells by suppressing the cyclin-A2/Akt/FOXO3a pathway.

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Mei Yun Huang

Papers

The BH3-mimetic gossypol and noncytotoxic doses of valproic acid induce apoptosis by suppressing cyclin-A2/Akt/FOXO3a signaling