Keunwook Lee

Direct role of NF-κB activation in Toll-like receptor-triggered HLA-DRA expression

Microbial components, such as DNA containing immunostimulatory CpG motifs (CpG‐DNA) and lipopolysaccharides (LPS), elicit the cell surface expression of MHC class II (MHC‐II) through Toll‐like receptor (TLR)/IL‐1R. Here, we show that CpG‐DNA and LPS induce expression of the HLA‐DRA in the human B cell line, RPMI 8226. Ectopic expression of the dominant negative mutant of CIITA and RNA interference targeting the CIITA gene indicate that CIITA activation is not enough for the maximal MHC‐II expression induced by CpG‐DNA and LPS. Additionally, nuclear factor (NF)‐κB activation is required for the CpG‐DNA‐activated and LPS‐activated HLA‐DRA expression, whereas IFN‐γ‐induced MHC‐II expression depends on CIITA rather than on NF‐κB. Comprehensive mutant analyses, electrophoretic mobility shift assays and chromatin immunoprecipitation assays, reveal that the functional interaction of NF‐κB with the promoter element is necessary for the TLR‐mediated HLA‐DRA induction by CpG‐DNA and LPS. This novel mechanism provides the regulation of MHC‐II gene expression with complexity and functional diversity.

Immune response and the tumor microenvironment: how they communicate to regulate gastric cancer.

Gastric cancer is the second most common cause of cancer-related death in the world. A growing body of evidence indicates that inflammation is closely associated with the initiation, progression, and metastasis of many tumors, including those of gastric cancer. In addition, approximately 60% of the worlds population is colonized by Helicobacter pylori, which accounts for more than 50% of gastric cancers. While the role of inflammation in intestinal and colonic cancers is relatively well defined, its role in stomach neoplasia is still unclear because of the limited access of pathogens to the acidic environment and the technical difficulties isolating and characterizing immune cells in the stomach, especially in animal models. In this review, we will provide recent updates addressing how inflammation is involved in gastric malignancies, and what immune characteristics regulate the pathogenesis of stomach cancer. Also, we will discuss potential therapeutics that target the immune system for the efficient treatment of gastric cancer.

Intrathecally injected nicotine enhances the antinociception induced by morphine but not β-endorphin, d-Pen2,5-enkephalin and U50,488H administered intrathecally in the mouse

The effect of nicotine injected intrathecally (i.t.) on the inhibition of the tail-flick response induced by morphine, beta-endorphin, D-Pen2,5-enkephalin (DPDPE), or [(trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl] benzeocetamide)] (U50,488H) administered i.t. was studied in ICR mice. The i.t. injection of nicotine alone at doses from 1 to 12 microg produced only a minimal inhibition of the tail-flick response. Morphine (0.2 microg), beta-endorphin (0.1 microg), DPDPE (0.5 microg) or U50,488H (6 microg) caused only slight inhibition of the tail-flick response. Nicotine injected i.t. dose dependently enhanced the inhibition of the tail-flick response induced by i.t. administered morphine (0.2 microg). However, i.t. injected nicotine at the same doses was not effective in enhancing the inhibition of the tail-flick response induced by beta-endorphin, DPDPE, or U50,488H administered i.t. Our results suggest that stimulating nicotinic receptors located in the spinal cord may enhance the antinociception induced by morphine administered spinally. However, the activation of nicotinic receptors at the spinal level may not be involved in modulating the antinociception induced by beta-endorphin, DPDPE, and U50,488H administered spinally.

CD11b regulates antibody class switching via induction of AID

HighlightsCD11b is expressed on the surface of conventional B2 B cells upon activation.CD11b is important for class switching and affinity maturation of antibodies.CD11b promotes induction of AID through the regulation of NF‐&kgr;B signaling pathway. Abstract The integrin CD11b, which is encoded by the integrin subunit alpha M (ITGAM), is primarily expressed on the surface of innate immune cells. Genetic variations in ITGAM are among the strongest risk factors for systemic lupus erythematosus, an autoimmune disease characterized by the presence of autoantibodies. However, the regulatory function of CD11b in the antibody responses remains unclear. Here, we report the induction of CD11b in activated B2 B cells and define its unexpected role in immunoglobulin heavy chain class switch recombination (CSR). LPS‐activated B cells lacking CD11b yielded fewer IgG subtypes such as IgG1 and IgG2a in vitro, and immunization‐dependent CSR and affinity maturation of antibodies were severely impaired in CD11b‐deficient mice. Notably, we observed the reduced expression of activation‐induced cytidine deaminase (AID), an enzyme that initiates CSR and somatic hypermutation, and ectopic expression of AID was sufficient to rescue the defective CSR of CD11b‐deficient B cells. LPS‐induced phosphorylation of NF‐&kgr;B p65 and I&kgr;B&agr; was attenuated in CD11b‐deficient B cells, and hyperactivation of I&kgr;B kinase 2 restored the defective AID expression and CSR, which implied that CD11b regulates the NF‐&kgr;B‐dependent induction of AID. Overall, our experimental evidence emphasized the function of CD11b in antibody responses and the role of CD11b as a vital regulator of CSR.

Extracellular Release of CD11b by TLR9 Stimulation in Macrophages

CpG-DNA upregulates the expression of pro-inflammatory cytokines, chemokines and cell surface markers. Investigators have shown that CD11b (integrin αM) regulates TLR-triggered inflammatory responses in the macrophages and dendritic cells. Therefore, we aimed to identify the effects of CpG-DNA on the expression of CD11b in macrophages. There was no significant change in surface expression of CD11b after CpG-DNA stimulation. However, CD11b was released into culture supernatants after stimulation with phosphorothioate-backbone modified CpG-DNA such as PS-ODN CpG-DNA 1826(S). In contrast, MB-ODN 4531 and non-CpG-DNA control (regardless of backbone type and liposome-encapsulation) failed to induce release of CD11b. Therefore, the context of the CpG-DNA sequence and phosphorothioate backbone modification may regulate the effects of CpG-DNA on CD11b release. Based on inhibitor studies, CD11b release is mediated by p38 MAP kinase activation, but not by the PI3K and NF-κB activation. CD11b release is mediated by lysosomal degradation and by vacuolar acidification in response to CpG-DNA stimulation. The amount of CD11b in the exosome precipitant was significantly increased by CpG-DNA stimulation in vivo and in vitro depending on TLR9. Our observations perhaps give more insight into understanding of the mechanisms involved in CpG-DNA-induced immunomodulation in the innate immunity.

Tat-PRAS40 prevent hippocampal HT-22 cell death and oxidative stress induced animal brain ischemic insults.

Proline rich Akt substrate (PRAS40) is a component of mammalian target of rapamycin complex 1 (mTORC1) and is known to play an important role against reactive oxygen species-induced cell death. However, the precise function of PRAS40 in ischemia remains unclear. Thus, we investigated whether Tat-PRAS40, a cell-permeable fusion protein, has a protective function against oxidative stress-induced hippocampal neuronal (HT-22) cell death in an animal model of ischemia. We showed that Tat-PRAS40 transduced into HT-22 cells, and significantly protected against cell death by reducing the levels of H2O2 and derived reactive species, and DNA fragmentation as well as via the regulation of Bcl-2, Bax, and caspase 3 expression levels in H2O2 treated cells. Also, we showed that transduced Tat-PARS40 protein markedly increased phosphorylated RRAS40 expression levels and 14-3-3σ complex via the Akt signaling pathway. In an animal ischemia model, Tat-PRAS40 effectively transduced into the hippocampus in animal brain and significantly protected against neuronal cell death in the CA1 region. We showed that Tat-PRAS40 protein effectively transduced into hippocampal neuronal cells and markedly protected against neuronal cell damage. Therefore, we suggest that Tat-PRAS40 protein may be used as a therapeutic protein for ischemia and oxidative stress-induced brain disorders.

Gomisin G Inhibits the Growth of Triple-Negative Breast Cancer Cells by Suppressing AKT Phosphorylation and Decreasing Cyclin D1

A type of breast cancer with a defect in three molecular markers such as the estrogen receptor, progesterone receptor, and human epidermal growth factor receptor is called triple-negative breast cancer (TNBC). Many patients with TNBC have a lower survival rate than patients with other types due to a poor prognosis. In this study, we confirmed the anti-cancer effect of a natural compound, Gomisin G, in TNBC cancer cells. Treatment with Gomisin G suppressed the viability of two TNBC cell lines, MDA-MB-231 and MDA-MB-468 but not non-TNBC cell lines such as MCF-7, T47D, and ZR75-1. To investigate the molecular mechanism of this activity, we examined the signal transduction pathways after treatment with Gomisin G in MDA-MB-231 cells. Gomisin G did not induce apoptosis but drastically inhibited AKT phosphorylation and reduced the amount of retinoblastoma tumor suppressor protein (Rb) and phosphorylated Rb. Gomisin G induced in a proteasome-dependent manner a decrease in Cyclin D1. Consequently, Gomisin G causes cell cycle arrest in the G1 phase. In contrast, there was no significant change in T47D cells except for a mild decrease in AKT phosphorylation. These results show that Gomisin G has an anti-cancer activity by suppressing proliferation rather than inducing apoptosis in TNBC cells. Our study suggests that Gomisin G could be used as a therapeutic agent in the treatment of TNBC patients.

mTOR kinase leads to PTEN-loss-induced cellular senescence by phosphorylating p53

Loss of PTEN, the major negative regulator of the PI3K/AKT pathway induces a cellular senescence as a failsafe mechanism to defend against tumorigenesis, which is called PTEN-loss-induced cellular senescence (PICS). Although many studies have indicated that the mTOR pathway plays a critical role in cellular senescence, the exact functions of mTORC1 and mTORC2 in PICS are not well understood. In this study, we show that mTOR acts as a critical relay molecule downstream of PI3K/AKT and upstream of p53 in PICS. We found that PTEN depletion induces cellular senescence via p53-p21 signaling without triggering DNA damage response. mTOR kinase, a major component of mTORC1 and mTORC2, directly binds p53 and phosphorylates it at serine 15. mTORC1 and mTORC2 compete with MDM2 and increase the stability of p53 to induce cellular senescence via accumulation of the cell cycle inhibitor, p21. In embryonic fibroblasts of PTEN-knockout mice, PTEN deficiency also induces mTORC1 and mTORC2 to bind to p53 instead of MDM2, leading to cellular senescence. These results collectively demonstrate for the first time that mTOR plays a critical role in switching cells from proliferation signaling to senescence signaling via a direct link between the growth-promoting activity of AKT and the growth-suppressing activity of p53.

Erratum to "Gomisin G Inhibits the Growth of Triple-Negative Breast Cancer Cells by Suppressing AKT Phosphorylation and Decreasing Cyclin D1" [Biomol.Ther. 26 (2018) 322-327]

The authors request to correct the author name from Yoonho Lim to Yoongho Lim page 322.

Gomisin G Suppresses the Growth of Colon Cancer Cells by Attenuation of AKT Phosphorylation and Arrest of Cell Cycle Progression

Colorectal cancer is one of the leading causes of cancer related death due to a poor prognosis. In this study, we investigated the effect of Gomisin G on colon cancer growth and examined the underlying mechanism of action. We found that Gomisin G significantly suppressed the viability and colony formation of LoVo cells. Gomisin G reduced the phosphorylation level of AKT implying that Gomisin G suppressed the PI3K-AKT signaling pathway. Gomisin G also induced apoptosis shown by Annexin V staining and an increased level of cleaved poly-ADP ribose polymerase (PARP) and Caspase-3 proteins. Furthermore, Gomisin G remarkably triggered the accumulation of cells at the sub-G1 phase which represents apoptotic cells. In addition, the level of cyclin D1 and phosphorylated retinoblastoma tumor suppressor protein (Rb) was also reduced by the treatment with Gomisin G thus curtailing cell cycle progression. These findings show the suppressive effect of Gomisin G by inhibiting proliferation and inducing apoptosis in LoVo cells. Taken together, these results suggest Gomisin G could be developed as a potential therapeutic compound against colon cancer.