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Background: Eukaryotic initiation factor 5A hypusine (eIF5A Hyp ) stimulates the translation of proline repeat motifs. Salt inducible kinase 2 (SIK2) containing a proline repeat motif is overexpressed in ovarian cancers, in which it promotes cell proliferation, migration, and invasion., Methods and Results: Western blotting and dual luciferase analyses showed that depletion of eIF5A Hyp by GC7 or eIF5A-targeting siRNA downregulated SIK2 level and decreased luciferase activity in cells transfected with a luciferase-based reporter construct containing consecutive proline residues, whereas the activity of the mutant control reporter construct (replacing P825L, P828H, and P831Q) did not change. According to the MTT assay, GC7, which has a potential antiproliferative effect, reduced the viability of several ovarian cancer cell lines by 20-35% at high concentrations (ES2 > CAOV-3 > OVCAR-3 > TOV-112D) but not at low concentrations. In a pull-down assay, we identified eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) and 4E-BP1 (p4E-BP1) phosphorylated at Ser 65 as downstream binding partners of SIK2, and we validated that the level of p4E-BP1(Ser 65) was downregulated by SIK2-targeting siRNA. Conversely, in ES2 cells overexpressing SIK2, the p4E-BP1(Ser 65) level was increased but decreased in the presence of GC7 or eIF5A-targeting siRNA. Finally, the migration, clonogenicity, and viability of ES2 ovarian cancer cells were reduced by GC7 treatment as well as by siRNA for eIF5A gene silencing and siRNA for SIK2 and 4E-BP1 gene silencing. Conversely, those activities were increased in cells overexpressing SIK2 or 4E-BP1 and decreased again in the presence of GC7., Conclusion: The depletion of eIF5A Hyp by GC7 or eIF5A-targeting siRNA attenuated activation of the SIK2-p4EBP1 pathway. In that way, eIF5A Hyp depletion reduces the migration, clonogenicity, and viability of ES2 ovarian cancer cells., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

Host factors such as nutritional status and immune cell state are important for vaccine efficacy. Inflammasome activation may be important for triggering vaccine-induced humoral and cell-mediated immune responses. Formulations with alum as a typical adjuvant to overcome the effects of host factors have recently been shown to induce inflammasome activation, which augments vaccine efficacy. Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is one of the main components of inflammasomes, but it is not clear whether ASC affects the vaccine-induced immune response. Herein, we used two types of vaccines: inactivated influenza vaccine not formulated with alum, and HPV vaccine formulated with alum. We gave the vaccines to ASC knockout (ASC -/- ) mice to investigate the role of ASC in vaccine efficacy. Influenza vaccine-immunized ASC -/- mice did not show antibody titers in week 2 after the first vaccination. After boosting, the antibody titer in ASC -/- mice was about half that in wild type (WT) mice. Furthermore, a cytotoxic T-lymphocyte response against influenza vaccine was not induced in ASC -/- mice. Therefore, vaccinated ASC -/- mice did not show effective protection against viral challenge. ASC -/- mice immunized with alum-formulated HPV vaccine showed similar antibody titers and T-cell proliferation compared with immunized WT mice. However, the HPV vaccine without alum induced up to threefold lower titers of HPV-specific antibody titers in ASC -/- mice compared with those in WT mice. These findings suggest that alum in vaccine can overcome the ASC-deficient condition., (© 2018 The Societies and John Wiley & Sons Australia, Ltd.)

Mithramycin A (Mith) is a natural polyketide that has been used in multiple areas of research including apoptosis of various cancer cells. Here, we examined the critical role of Mith in apoptosis and its molecular mechanism in DU145 and PC3 prostate cancer cells and tumor xenografts. Mith decreased cell growth and induced apoptosis in DU145 and PC-3 cells. Myeloid cell leukemia-1 (Mcl-1) was over-expressed in both cell lines compared to RWPE1 cells. Mith inhibited Mcl-1 protein expression in both cells, but only altered Mcl-1 mRNA levels in PC-3 cells. We also found that Mith reduced Mcl-1 protein levels through both proteasome-dependent protein degradation and the inhibition of protein synthesis in DU145 cells. Studies using siRNA confirmed that the knockdown of Mcl-1 induced apoptosis. Mith significantly suppressed TPA-induced neoplastic cell transformation through the down-regulation of the Mcl-1 protein in JB6 cells, and suppressed the transforming activity of both cell types. Mith also inhibited tumor growth and Mcl-1 levels, in addition to inducing apoptosis, in athymic nude mice bearing DU145 cell xenografts without affecting five normal organs. Therefore, Mith inhibits cell growth and induces apoptosis by suppressing Mcl-1 in both prostate cancer cells and xenograft tumors, and thus is a potent anticancer drug candidate for prostate cancer., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Mitochondrial fusion and fission are dynamically regulated during apoptotic cell death, and mitofusin (Mfn) and related proteins have been shown to be involved in apoptosis-associated changes in mitochondrial morphology and function. Here, we investigated the involvement of Mfn proteins in the conformational activation and mitochondrial translocation of Bax, a key molecule responsible for apoptosis-associated mitochondrial changes. When ectopically expressed, Mfn1 inhibited the amino-terminal activation, but not the mitochondrial translocation, of Bax during staurosporine-induced apoptosis; overexpression of Mfn2 had no effect. Overexpression of Mfn1 mutants carrying point mutations in the GTPase domain (Mfn1-K88T and Mfn1-T109A) did not inhibit the amino-terminal activation of Bax. Furthermore, staurosporine-induced amino-terminal activation of Bax was significantly delayed in Mfn1-shRNA transfected (Mfn1-depleted) HeLa cells compared to cells transfected with control shRNA. These results collectively suggest a role for Mfn1 in regulating the activation of Bax on the outer mitochondrial membrane in a GTPase-dependent manner., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Background: 5'-Nitro-indirubinoxime (5'-NIO) is a new derivative of indirubin that exhibits anti-cancer activity in a variety of human cancer cells. However, its mechanism has not been fully clarified., Methods: Human salivary gland adenocarcinoma (SGT) cells were used in this study. Western blot and RT-PCR analyses were performed to determine cellular Notch levels. The cell cycle stage and level of apoptosis were analyzed using flow cytometry analysis., Results: 5'-NIO significantly inhibited the mRNA levels of Notch-1 and Notch-3 and their ligands (Delta1, 2, 3, and Jagged-2) in SGT cells. Immunocytochemistry analysis showed that 5'-NIO specifically decreased the level of Notch-1 in the nucleus. In addition, 5'-NIO induced G1 cell cycle arrest by reducing levels of CDK4 and CDK6 in SGT cells. Using flow cytometry and immunoblotting analysis, we found that 5'-NIO induces apoptosis following the secretion of cytochrome c and the activation of caspase-3 and caspase-7. Intracellular Notch-1 overexpression led to a decrease in G1 phase arrest and an inhibition of 5'-NIO-induced apoptosis., Conclusion: These observations suggest that 5'-NIO induces cell cycle arrest and apoptosis by down-regulating Notch-1 signaling., General Significance: This study identifies a new mechanism of 5'-NIO-mediated anti-tumor properties. Thus, 5'-NIO could be used as a candidate for salivary gland adenocarcinoma therapeutics., (Copyright (c) 2009 Elsevier B.V. All rights reserved.)

The antimicrobial effects of a lactic acid producing bacteria culture condensate mixture (LCCM) were assessed against Salmonella enteritidis. In the presence of LCCM, bacterial growth was assessed in vitro by the measurement of optical density (OD) and viable bacterial counting. At concentrations of 1.25 and 2.5% LCCM, OD values were significantly lower than that of the control broth, and at concentrations of 5 and 10% LCCM, OD values did not increase for the entire period of experiment. At 8 h after incubation, the viable bacterial numbers in 5% and 10% LCCM-containing broths were remarkably lower than that in the control broth. This antimicrobial ability of the LCCM was fundamentally attributed to causing cell death rather than inhibiting growth. Even when the pH of LCCM-containing broth was adjusted to 7.2, the number of viable bacteria was significantly lower in the broths containing LCCM over 2.5% than that in control broth at 8 h after incubation. However, the OD value of each culture in the presence of each concentration of the LCCM increased over 1.0 at the completion of the experiment. The in vivo antimicrobial effects of the LCCM against S. enteritidis were also assessed. In S. enteritidis-infected mice, the LCCM decreased both the viable bacteria found in the feces and the mortality rate of the mice. These findings showed that the LCCM might have an antimicrobial ability against S. enteritidis.

The adipose tissue NLRP3 inflammasome has recently emerged as a contributor to obesity-related metabolic inflammation. Recent studies have demonstrated that the activation of the NLRP3 inflammasome cleaves gasdermin D (GSDMD) and induces pyroptosis, a proinflammatory programmed cell death. However, whether GSDMD is involved in the regulation of adipose tissue function and the development of obesity-induced metabolic disease remains unknown. The aim of the present study was to investigate the role of GSDMD in adipose tissue inflammation as well as whole-body metabolism using GSDMD-deficient mice fed a high-fat diet (HFD) for 30 weeks. The effects of GSDMD deficiency on adipose tissue, liver, and isolated macrophages from wild-type (WT) and GSDMD knockout (KO) mice were examined. In addition, 3T3-L1 cells were used to examine the expression of GSDMD during adipogenesis. The results demonstrate that although HFD-induced inflammation was partly ameliorated in isolated macrophages and liver, adipose tissue remained unaffected by GSDMD deficiency. Compared with the WT HFD mice, GSDMD KO HFD mice exhibited a mild increase in HFD-induced glucose intolerance with increased systemic and adipose tissue IL-1β levels. Interestingly, GSDMD deficiency caused accumulation of fat mass when challenged with HFD, partly by suppressing the expression of peroxisome proliferator-activated receptor gamma (PPARγ). The expression of GSDMD mRNA and protein was dramatically suppressed during adipocyte differentiation and was inversely correlated with PPARγ expression. Together, these findings indicate that GSDMD is not a prerequisite for HFD-induced adipose tissue inflammation and suggest a noncanonical function of GSDMD in regulation of fat mass through PPARγ. [ABSTRACT FROM AUTHOR]

J Oral Pathol Med (2012) 41: 540-546 Background: Toll-like receptor (TLR) agonists have anticancer effect by inducing apoptosis or activating immune cells. In this study, we investigated whether imiquimod, TLR7 agonist, inhibits the proliferation of oral cancer cells. Methods: Toll-like receptor 7 expression and IL-6/8 production by imiquimod were examined using RT-PCR and Enzyme-linked immunosorbent assay, respectively. Cell viability was examined by MTT assay. To examine apoptotic cell death, Annexin V/PI staining for flow cytometry and Western blot analysis were performed. Necrotic cell death was determined by leakage of lactate dehydrogenase (LDH), HMGB1, and PI staining in imiquimod-treated oral squamous cell carcinoma (OSCC) cells. Results: Toll-like receptor7 mRNA was expressed in OSCC cells. Imiquimod induced IL-6 and IL-8 production in OSCC cells, suggesting the functional expression of TLR7. Imiquimod inhibited cells proliferation in a dose-dependent manner. The ratio of annexin V-positive cells and cleaved caspase-3/7 was increased by imiquimod treatment in OSCC cells, suggesting that imiquimod-induced cell death in OSCC cells may be owing to apoptosis. In addition, LDH secretion and PI staining were detected in OSCC cells treated with imiquimod, showing that imiquimod also induced necrotic cell death in the OSCC cells. Conclusions: Imiquimod inhibited effectively the growth of OSCC cells by inducing apoptosis and necrosis. [ABSTRACT FROM AUTHOR]

Objective. Toll-like receptors (TLR) signaling has dual effect of promoting tumor progression and anti-cancer property. This study was designed to determine the effect of polyinosinic-polycytidilic acid (poly I:C), a TLR3 agonist, on the proliferation of oral cancer cells. Materials and methods: Human oral squamous cell carcinoma cell lines, YD-10B and YD-8, were used. TLRs expression was examined by RT-PCR and IL-8 production by poly I:C was examined by ELISA. Cell proliferation was determined by MTT assay. Flow cytometry and Western blot analysis were performed to determine the molecular mechanism of poly I:C-induced cell death. Results. TLR3 was functionally expressed in YD-10B and YD-8 cells. Treatment of poly I:C inhibited the cell growth in a dose-dependent manner. Flow cytometry and Western blot analysis revealed that poly I:C induced apoptosis via a mitochondria-dependent pathway. In addition, combination treatment with poly I:C and paclitaxel more significantly inhibited cell proliferation compared with poly I:C or paclitaxel alone. Conclusions. Poly I:C effectively inhibits oral cancer cell proliferation and can be considered as a candidate to improve the inhibitory effect of anti-cancer drugs. [ABSTRACT FROM AUTHOR]

In this study, we investigated the effects of blue light exposure on nucleotide-binding oligomerization domain 2 (NOD2) expression on the mouse ocular surface and evaluated the role of NOD2 activation in light-induced cell death. Mice were divided into wild-type (WT), NOD2-knock out (KO), WT + blue light (WT + BL), and NOD2-KO + blue light (NOD2-KO + BL) groups, and the mice in the WT+BL and NOD2-KO + BL groups were exposed to blue light for 10 days. After 10 days of blue light exposure, increased reactive oxygen species and malondialdehyde were observed in the WT + BL and NOD2-KO + BL groups, and the WT + BL group showed a higher expression of NOD2 and autophagy related 16 like 1. Although both WT+BL and NOD2-KO + BL groups showed an increase in the expression of light chain 3-II, NOD2-KO + BL mice had a significantly lower p62 expression than WT + BL mice. In addition, NOD2-KO+BL mice had significantly lower corneal epithelial damage and apoptosis than WT + BL mice. In conclusion, blue light exposure can induce impaired autophagy by activation of NOD2 on the ocular surface. In addition, the reactive oxygen species (ROS)–NOD2–autophagy related 16 like 1 (ATG16L) signaling pathway may be involved in the blue-light-induced autophagy responses, resulting in corneal epithelial apoptosis. [ABSTRACT FROM AUTHOR]

Salicornia herbacea L. (Chenopodiaceae), an edible salt marsh plant with anti-inflammatory effects, was examined in macrophages and trophoblasts whether it modulates NLRP3 inflammasome activity. Pretreatment and delayed treatment of S. herbacea extract (SHE) in bone marrow-derived macrophages (BMDMs) reduced the activity of NLRP3 inflammasome induced by lipopolysaccharide (LPS) and adenosine triphosphate stimulation and downregulated interleukin (IL)-1β production. SHE also inhibited pyroptotic cell death, the adaptor molecule apoptosis-associated speck-like protein containing a CARD (ASC), oligomerization, and speck by NLRP3 inflammasome activity in BMDM. Similarly, SHE decreased the mRNA expression of NLRP3, ASC, IL-1β, and IL-6 in the LPS-stimulated human trophoblast cell line, Swan 71 cells. In addition, SHE inhibited the production of IL-6 and IL-1β and decreased the expression of cyclooxygenase-2 and prostaglandin E2 in stimulated Swan 71 cells. Finally, 3,5-dicaffeoylquinic acid (3,5-DCQA), one of the components of S. herbacea, inhibited IL-1β produced by NLRP3 inflammasome activity. In conclusion, SHE downregulated the activity of the NLRP3 inflammasome in macrophages and trophoblasts. [ABSTRACT FROM AUTHOR]