Abietane Diterpenoids Isolated from Torreya nucifera Disrupt Replication of Influenza Virus by Blocking the Phosphatidylinositol-3-Kinase (PI3K)-Akt and ERK Signaling Pathway.

Although vaccines and antiviral drugs are available, influenza viruses continue to pose a significant threat to vulnerable populations globally. With the emergence of drug-resistant strains, there is a growing need for novel antiviral therapeutic approaches. We found that 18-hydroxyferruginol (1) and 18-oxoferruginol (2) isolated from Torreya nucifera exhibited strong anti-influenza activity, with 50% inhibitory concentration values of 13.6 and 18.3 µM against H1N1, 12.8 and 10.8 µM against H9N2, and 29.2 µM (only compound 2) against H3N2 in the post-treatment assay, respectively. During the viral replication stages, the two compounds demonstrated stronger inhibition of viral RNA and protein in the late stages (12-18 h) than in the early stages (3-6 h). Moreover, both compounds inhibited PI3K-Akt signaling, which participates in viral replication during the later stages of infection. The ERK signaling pathway is also related to viral replication and was substantially inhibited by the two compounds. In particular, the inhibition of PI3K-Akt signaling by these compounds inhibited viral replication by sabotaging influenza ribonucleoprotein nucleus-to-cytoplasm export. These data indicate that compounds 1 and 2 could potentially reduce viral RNA and viral protein levels by inhibiting the PI3K-Akt signaling pathway. Our results suggest that abietane diterpenoids isolated from T. nucifera may be potent antiviral candidates for new influenza therapies.

Deinococcus radiodurans-derived membrane vesicles protect HaCaT cells against H2O2-induced oxidative stress via modulation of MAPK and Nrf2/ARE pathways.

BACKGROUND: Deinococcus radiodurans is a robust bacterium that can withstand harsh environments that cause oxidative stress to macromolecules due to its cellular structure and physiological functions. Cells release extracellular vesicles for intercellular communication and the transfer of biological information; their payload reflects the status of the source cells. Yet, the biological role and mechanism of Deinococcus radiodurans-derived extracellular vesicles remain unclear. AIM: This study investigated the protective effects of membrane vesicles derived from D. radiodurans (R1-MVs) against H2O2-induced oxidative stress in HaCaT cells. RESULTS: R1-MVs were identified as 322 nm spherical molecules. Pretreatment with R1-MVs inhibited H2O2-mediated apoptosis in HaCaT cells by suppressing the loss of mitochondrial membrane potential and reactive oxygen species (ROS) production. R1-MVs increased the superoxide dismutase (SOD) and catalase (CAT) activities, restored glutathione (GSH) homeostasis, and reduced malondialdehyde (MDA) production in H2O2-exposed HaCaT cells. Moreover, the protective effect of R1-MVs against H2O2-induced oxidative stress in HaCaT cells was dependent on the downregulation of mitogen-activated protein kinase (MAPK) phosphorylation and the upregulation of the nuclear factor E2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway. Furthermore, the weaker protective capabilities of R1-MVs derived from ΔDR2577 mutant than that of the wild-type R1-MVs confirmed our inferences and indicated that SlpA protein plays a crucial role in R1-MVs against H2O2-induced oxidative stress. CONCLUSION: Taken together, R1-MVs exert significant protective effects against H2O2-induced oxidative stress in keratinocytes and have the potential to be applied in radiation-induced oxidative stress models.

Highly Efficient and Stable Inverted Perovskite Solar Cell Using Pure δ-FAPbI3 Single Crystals.

Pure δ-formamidinium lead triiodide (δ-FAPbI3 ) single crystal for highly efficient perovskite solar cell (PCS) with long-term stability is prepared by a new method consisting of liquid phase reaction of FAI and PbI2 in N,N-dimethyl formamide and antisolvent crystallization using acetonitrile. In this method, the incorporation of any impurity into the crystal is excluded by the molecular recognition of the crystal growth site. This pure crystal is used to fabricate α-FAPbI3 inverted PSCs which showed excellent power conversion efficiency (PCE) due to much-reduced trap-states. The champion device exhibited a high PCE of 23.48% under the 1-Sun condition. Surface-treated devices with 3-(aminomethyl)pyridine showed a significantly improved PCE of 25.07%. In addition, the unencapsulated device maintained 97.22% of its initial efficiency under continuous 1-Sun illumination for 1,000 h at 85 °C in an N2 atmosphere ensuring long-term thermal and photo stabilities of PSCs, whereas the control device kept only 89.93%.

Myristica fragrans Extract Inhibits Platelet Desialylation and Activation to Ameliorate Sepsis-Associated Thrombocytopenia in a Murine CLP-Induced Sepsis Model.

Sepsis, characterized by an uncontrolled host inflammatory response to infections, remains a leading cause of death in critically ill patients worldwide. Sepsis-associated thrombocytopenia (SAT), a common disease in patients with sepsis, is an indicator of disease severity. Therefore, alleviating SAT is an important aspect of sepsis treatment; however, platelet transfusion is the only available treatment strategy for SAT. The pathogenesis of SAT involves increased platelet desialylation and activation. In this study, we investigated the effects of Myristica fragrans ethanol extract (MF) on sepsis and SAT. Desialylation and activation of platelets treated with sialidase and adenosine diphosphate (platelet agonist) were assessed using flow cytometry. The extract inhibited platelet desialylation and activation via inhibiting bacterial sialidase activity in washed platelets. Moreover, MF improved survival and reduced organ damage and inflammation in a mouse model of cecal ligation and puncture (CLP)-induced sepsis. It also prevented platelet desialylation and activation via inhibiting circulating sialidase activity, while maintaining platelet count. Inhibition of platelet desialylation reduces hepatic Ashwell-Morell receptor-mediated platelet clearance, thereby reducing hepatic JAK2/STAT3 phosphorylation and thrombopoietin mRNA expression. This study lays a foundation for the development of plant-derived therapeutics for sepsis and SAT and provides insights into sialidase-inhibition-based sepsis treatment strategies.

Inhibition of melanogenesis by Aster yomena callus pellet extract in melanoma cells and patients with skin pigmentation.

Plant tissue culture holds immense potential for the production of secondary metabolites with various physiological functions. We recently established a plant tissue culture system capable of producing secondary metabolites from Aster yomena. This study aimed to uncover the mechanisms underlying the potential therapeutic effects of Aster yomena callus pellet extract (AYC-P-E) on photoaging-induced skin pigmentation. Excessive melanogenesis was induced in B16F10 melanoma cells using α-melanocyte stimulating hormone (α-MSH). The effects of AYC-P-E treatment on melanin biosynthesis inducers and melanin synthesis inhibition were assessed. Based on the results, a clinical study was conducted in subjects with skin pigmentation. AYC-P-E inhibited melanogenesis in α-MSH-treated B16F10 cells, accompanied by decreased mRNA and protein expression of melanin biosynthesis inducers, including cyclic AMP response element-binding protein (CREB), tyrosinase, microphthalmia-associated transcription factor (MITF), tyrosinase related protein-1 (TRP-1), and TRP-2. This anti-melanogenic effect was mediated by mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK) and protein kinase B (AKT) phosphorylation. Treatment of subjects with skin pigmentation with AYC-P-E-containing cream formulations resulted in 3.33%, 7.06%, and 8.68% improvement in the melanin levels at 2, 4, and 8 weeks, respectively. Our findings suggest that AYC-P-E inhibits excessive melanogenesis by activating MEK/ERK and AKT signaling, potentiating its cosmetic applications in hyperpigmentation treatment.

Protective Effect of Polysaccharides Extracted from Cudrania tricuspidata Fruit against Cisplatin-Induced Cytotoxicity in Macrophages and a Mouse Model.

Although cisplatin is one of most effective chemotherapeutic drugs that is widely used to treat various types of cancer, it can cause undesirable damage in immune cells and normal tissue because of its strong cytotoxicity and non-selectivity. This study was conducted to investigate the cytoprotective effects of Cudrania tricuspidata fruit-derived polysaccharides (CTPS) against cisplatin-induced cytotoxicity in macrophages, lung cancer cell lines, and a mouse model, and to explore the possibility of application of CTPS as a supplement for anticancer therapy. Both cisplatin alone and cisplatin with CTPS induced a significant cytotoxicity in A549 and H460 lung cancer cells, whereas cytotoxicity was suppressed by CTPS in cisplatin-treated RAW264.7 cells. CTPS significantly attenuated the apoptotic and necrotic population, as well as cell penetration in cisplatin-treated RAW264.7 cells, which ultimately inhibited the upregulation of Bcl-2-associated X protein (Bax), cytosolic cytochrome c, poly (adenosine diphosphateribose) polymerase (PARP) cleavage, and caspases-3, -8, and -9, and the downregulation of B cell lymphoma-2 (Bcl-2). The CTPS-induced cytoprotective action was mediated with a reduction in reactive oxygen species production and mitochondrial transmembrane potential loss in cisplatin-treated RAW264.7 cells. In agreement with the results obtained above, CTPS induced the attenuation of cell damage in cisplatin-treated bone marrow-derived macrophages (primary cells). In in vivo studies, CTPS significantly inhibited metastatic colonies and bodyweight loss as well as immunotoxicity in splenic T cells compared to the cisplatin-treated group in lung metastasis-induced mice. Furthermore, CTPS decreased the level of CRE and BUN in serum. In summation, these results suggest that CTPS-induced cytoprotective action may play a role in alleviating the side effects induced by chemotherapeutic drugs.

Bombyx batryticatus Protein-Rich Extract Induces Maturation of Dendritic Cells and Th1 Polarization: A Potential Immunological Adjuvant for Cancer Vaccine.

Bombyx batryticatus, a protein-rich edible insect, is widely used as a traditional medicine in China. Several pharmacological studies have reported the anticancer activity of B. batryticatus extracts; however, the capacity of B. batryticatus extracts as immune potentiators for increasing the efficacy of cancer immunotherapy is still unverified. In the present study, we investigated the immunomodulatory role of B. batryticatus protein-rich extract (BBPE) in bone marrow-derived dendritic cells (BMDCs) and DC vaccine-immunized mice. BBPE-treated BMDCs displayed characteristics of mature immune status, including high expression of surface molecules (CD80, CD86, major histocompatibility complex (MHC)-I, and MHC-II), increased production of proinflammatory cytokines (tumor necrosis factor-α and interleukin-12p70), enhanced antigen-presenting ability, and reduced endocytosis. BBPE-treated BMDCs promoted naive CD4+ and CD8+ T-cell proliferation and activation. Furthermore, BBPE/ovalbumin (OVA)-pulsed DC-immunized mice showed a stronger OVA-specific multifunctional T-cell response in CD4+ and CD8+ T cells and a stronger Th1 antibody response than mice receiving differently treated DCs, which showed the enhanced protective effect against tumor growth in E.G7 tumor-bearing mice. Our data demonstrate that BBPE can be a novel immune potentiator for a DC-based vaccine in anticancer therapy.

Chrysin Derivative CM1 and Exhibited Anti-Inflammatory Action by Upregulating Toll-Interacting Protein Expression in Lipopolysaccharide-Stimulated RAW264.7 Macrophage Cells.

Although our previous study revealed that gamma-irradiated chrysin enhanced anti-inflammatory activity compared to intact chrysin, it remains unclear whether the chrysin derivative, CM1, produced by gamma irradiation, negatively regulates toll-like receptor (TLR) signaling. In this study, we investigated the molecular basis for the downregulation of TLR4 signal transduction by CM1 in macrophages. We initially determined the appropriate concentration of CM1 and found no cellular toxicity below 2 µg/mL. Upon stimulation with lipopolysaccharide (LPS), CM1 modulated LPS-stimulated inflammatory action by suppressing the release of proinflammatory mediators (cytokines TNF-α and IL-6) and nitric oxide (NO) and downregulated the mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) signaling pathways. Furthermore, CM1 markedly elevated the expression of the TLR negative regulator toll-interacting protein (Tollip) in dose- and time-dependent manners. LPS-induced expression of cell surface molecules (CD80, CD86, and MHC class I/II), proinflammatory cytokines (TNF-α and IL-6), COX-2, and iNOS-mediated NO were inhibited by CM1; these effects were prevented by the knockdown of Tollip expression. Additionally, CM1 did not affect the downregulation of LPS-induced expression of MAPKs and NF-κB signaling in Tollip-downregulated cells. These findings provide insight into effective therapeutic intervention of inflammatory disease by increasing the understanding of the negative regulation of TLR signaling induced by CM1.

Vaccine efficacy of a Mycobacterium tuberculosis Beijing-specific proline-glutamic acid (PE) antigen against highly virulent outbreak isolates.

Bacillus Calmette-Guerin vaccine confers insufficient pulmonary protection against tuberculosis (TB), particularly the Mycobacterium tuberculosis (Mtb) Beijing strain infection. Identification of vaccine antigens (Ags) by considering Mtb genetic diversity is crucial for the development of improved TB vaccine. MTBK_20640, a new Beijing genotype-specific proline-glutamic acid-family Ag, was identified by comparative genomic analysis. Its immunologic features were characterized by evaluating interactions with dendritic cells (DCs), and immunogenicity and vaccine efficacy were determined against highly virulent Mtb Beijing outbreak Korean Beijing (K) strain and HN878 strain in murine infection model. MTBK_20640 induced DCs via TLR2 and downstream MAPK and NF-κB signaling pathways, effectively promoting naive CD4-positive (CD4+) T-cell proliferation and IFN-γ production. Different IFN-γ response was observed in mice infected with Mtb K or reference H37Rv strain. Significant induction of T helper type 1 cell-polarized Ag-specific multifunctional CD4+ T cells and a marked Ag-specific IgG2c response were observed in mice immunized with MTBK_20640/glucopyranosyl lipid adjuvant-stable emulsion. The immunization conferred long-term protection against 2 Mtb Beijing outbreak strains, as evidenced by a significant reduction in colony-forming units in the lung and spleen and reduced lung inflammation. MTBK_20640 vaccination conferred long-term protection against highly virulent Mtb Beijing strains. MTBK_20640 may be developed into a novel Ag component in multisubunit TB vaccines in the future.-Kwon, K. W., Choi, H.-H., Han, S. J., Kim, J.-S., Kim, W. S., Kim, H., Kim, L.-H., Kang, S. M., Park, J., Shin, S. J. Vaccine efficacy of a Mycobacterium tuberculosis Beijing-specific proline-glutamic acid (PE) antigen against highly virulent outbreak isolates.

Neuroprotective effect of Annona muricata-derived polysaccharides in neuronal HT22 cell damage induced by hydrogen peroxide.

Crude extracts and phytochemical compounds derived from Annona muricata leaves have been demonstrated to exert neuroprotective effects. However, the neuroprotective effects of Annona muricata leaves-derived polysaccharide extracts (ALPs) have not been investigated. ALP treatment was shown to induce concentration-dependent antioxidant activity in HT22 cells, and to increase cell viability in H2O2-treated HT22 cells. These effects were correlated with a decrease in major components of oxidation, including: Ca2+, ROS, and malondialdehyde (MDA). Mediators of the intracellular response to oxidation, including Bax, cytochrome c, and cleaved caspases-3, -8, -9, MAPKs, and NF-κB, were positively influenced by ALP treatment under conditions of H2O2-mediated oxidative stress. In addition, ALP restored the expression of superoxide dismutase (SOD) and associated signaling pathways (PARP, PI3K/AKT and Nrf2-mediated HO-1/NQO-1) following H2O2 treatment. These results provide new pharmacological evidence that ALP facilitates neuroprotection via prevention of neuronal oxidative stress and promotion of cell survival signaling pathways.Abbreviations: ABTS: 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonicacid); AD: Alzheimer's disease; ALP: polysaccharide extracts isolated from Annona muricata leaves; ARE: antioxidant response element; DPPH: 1,1-diphenyl-picrylhydrazyl; DCFH-DA: 2',7'-dichlorofluorescin diacetate; ECL: electrochemiluminescence; ERK: extracellular regulated kinase; FBS: Fetal bovine serum; FITC: fluorescein isothiocyanate; FRAP: ferric reducing antioxidant power; HO-1: Heme oxygenase-1; JNK: c-jun N-terminal kinase; MAPKs: mitogen-activated protein kinases; MDA: malondialdehyde; MMP: mitochondrial membrane potential; MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide; NQO1: NAD(P)H:quinine oxidoreductase 1, Nrf2: nuclear factor-E2-related factor 2; PD: parkinson's disease; PI3K: phosphatidylinositol-3kinase; PVDF: polyvinylidene difluoride; ROS: reactive oxygen species; SOD: Superoxidedismutase; TPTZ: tripydyltriazine.

4-Hydroxybenzaldehyde Restricts the Intracellular Growth of Toxoplasma gondii by Inducing SIRT1-Mediated Autophagy in Macrophages.

Toxoplasma gondii is an intracellular protozoan parasite that infects approximately one third of the human popu- lation worldwide. Considering the toxicity and side effects of anti-toxoplasma medications, it is important to develop effec- tive drug alternatives with fewer and less severe off-target effects. In this study, we found that 4-hydroxybenzaldehyde (4- HBA) induced autophagy and the expression of NAD-dependent protein deacetylase sirtuin-1 (SIRT1) in primary murine bone marrow-derived macrophages (BMDMs). Interestingly, treatment of BMDMs with 4-HBA significantly reduced the number of macrophages infected with T. gondii and the proliferation of T. gondii in infected cells. This effect was impaired by pretreating the macrophages with 3-methyladenine or wortmannin (selective autophagy inhibitors) or with sirtinol or EX527 (SIRT1 inhibitors). Moreover, we found that pharmacological inhibition of SIRT1 prevented 4-HBA-mediated expres- sion of LC3-phosphatidylethanolamine conjugate (LC3-II) and the colocalization of T. gondii parasitophorous vacuoles with autophagosomes in BMDMs. These data suggest that 4-HBA promotes antiparasitic host responses by activating SIRT1- mediated autophagy, and 4-HBA might be a promising therapeutic alternative for the treatment of toxoplasmosis.

Therapeutic Potential of Gamma-Irradiated Resveratrol in Ulcerative Colitis via the Anti-Inflammatory Activity and Differentiation of Tolerogenic Dendritic Cells.

BACKGROUND/AIMS: New therapeutic strategies and the development of treatments against inflammatory bowel disease (IBD) require the initiation of immune tolerance and inhibition of excessive inflammation. Resveratrol, a polyphenolic compound, is a powerful immunosuppressor, but it can lead to apoptotic death of normal cells at high concentrations. When we induced a structural modification of resveratrol by gamma irradiation, we were able to investigate the potential tolerogenic and anti-inflammatory effect of a new radiolysis product (named γ-Res) during dendritic cell (DC) activation/differentiation. METHODS: The potential tolerogenic and anti-inflammatory effect of γ-Res were investigated by cytokine secretion, surface molecule expression, antigen uptake ability, antigen presenting ability, signaling pathway, and mixed lymphocyte reaction (MLR) assay using enzyme-linked immunosorbent assay (ELISA), western blot and flow cytometry. RESULTS: LPS-activated DCs treated with γ-Res exhibited alterations in their mature and functional statuses including a strongly inhibited cytokine production, surface molecule expression, antigen-presenting ability, and activated DC-induced T cell proliferation/activation. In addition, the DCs generated by the γ-Res treatment during DC differentiation induced a decreased surface molecule expression and increased IL-10 production without altering the levels of TNF-α and IL-12p70, thereby promoting the inhibition of T cell proliferation/activation and the induction of regulatory T cells via interaction with DCs in vitro. Furthermore, in the in vivo DSS-induced colitis model, γ-Res treatment conferred protective immunity with a decrease in IFN-γ+CD4+ and IL-17A+CD4+ T cells and imparted protection by reducing the disease activity and histological disease score and increasing the survival rate in dextran sulfate sodium (DSS)-induced colitis in mice. CONCLUSION: Thus, our results suggest that γ-Res may be an excellent candidate for use in IBD treatment.

GLM, a novel luteolin derivative, attenuates inflammatory responses in dendritic cells: Therapeutic potential against ulcerative colitis.

GLM, a luteolin derivative, shows anti-melanogenic effect via regulation of various signal molecules; however, it is unclear whether it also exerts anti-inflammatory effect. This study investigated the mechanisms of the anti-inflammatory effect of GLM on activated dendritic cells (DCs) to elucidate its therapeutic potential for ulcerative colitis. The anti-inflammatory effect of GLM was firstly investigated based on its effect on DCs maturation and T cells proliferation/activation. GLM treatment downregulated pro-inflammatory cytokine productions, surface molecule expression, and antigen-presenting ability for MHC-II complex in LPS-activated DCs. Importantly, anti-inflammatory effect induced by GLM treatment were independent of MAPK/NF-κB signaling pathways. Furthermore, DCs that were co-treated with LPS and GLM impaired the proliferation and activation of naïve CD4+ T cells. Interestingly, GLM exerted in vivo protective effect in DSS-induced colitis models by decreasing Th1, Th2, and Th17 cells and myeloperoxidase (MPO) levels, as well as restoring body weight, disease activity, and DSS-induced pathology. Based on these results, GLM was shown to be a potential candidate treatment for ulcerative colitis.

The Synergistic Effect of a Bimetallic Catalyst for the Synthesis of Carbon Nanotube Aerogels and their Predominant Chirality.

Synthesis of continuous spinnable carbon nanotube (CNT) fibers is the most promising method for producing CNT fibers for commercial applications. The floating-catalyst chemical vapor deposition (FC-CVD) method is a rapid process that achieves catalyst formation, CNT nucleation and growth, and aerogel-like sock formation within a few seconds. However, the formation mechanism is unknown. Herein, the progress of CNT fiber formation with bimetallic catalysts was studied, and the effect of catalyst composition to CNT fiber synthesis and their structural properties was investigated. In the case of bimetallic catalysts, the carbon source rapidly decomposes and generates various secondary hydrocarbon species, such as CH4 , C2 H4 , C2 H2 , C3 H6 , and C4 H10 whereas monometallic catalysts generate only CH4 and C2 H4 on decomposition. CNT fiber formation with Fe1 Ni0 begins about 400 mm from the reactor entrance, whereas CNT formation with Fe0.8 Ni0.2 and Fe0.5 Ni0.5 begins at about 500 and 300 mm, respectively. The formed CNT bundles and individual CNTs are oriented along the gas flow at these locations. The enhanced rate of fiber formation and lowering of growth temperature associated with bimetallic catalysts is explained by the synergistic effects between the two metals. The synthesized CNTs become predominantly semiconducting with increasing Ni contents.

Identification of BR101549 as a lead candidate of non-TZD PPARγ agonist for the treatment of type 2 diabetes: Proof-of-concept evaluation and SAR.

The new class of PPARgamma non-TZD agonist originally derived from the backbone of anti-hypertensive Fimasartan, BR101549, was identified as a potential lead for anti-diabetic drug development. The X-ray crystallography of BR101549 with PPARgamma ligand binding domain (LBD) revealed unique binding characteristics versus traditional TZD full agonists. The lead candidate, BR101549, has been found activating PPARgamma to the level of Pioglitazone in vitro and indeed has demonstrated its effects on blood glucose control in mouse proof-of-concept evaluation. The attempts to improve its metabolic stability profile through follow-up SAR including deuterium incorporation have been also described.

Discovery of BR102375, a new class of non-TZD PPARγ full agonist for the treatment of type 2 diabetes.

As a potential treatment of type 2 diabetes, a novel PPARγ non-TZD full agonist, compound 18 (BR102375) was identified from the original lead BR101549 by the SAR efforts of the labile metabolite control through bioisosteres approach. In vitro assessments of BR102375 demonstrated its activating potential of PPARγ comparable to Pioglitazone as well as the induction of related gene expressions. Further in vivo evaluation of BR102375 in diabetic rodent models successfully proved its glucose lowering effect as a potential antidiabetic agent, but the anticipated suppression of weight gain was not evident. The X-ray co-crystal analysis of BR102375-PPARγ LBD unexpectedly revealed binding modes totally different from those of BR101549, which was found, instead, closely resembled to those of TZD full agonists.

Epigallocatechin-3-Gallate Regulates Anti-Inflammatory Action Through 67-kDa Laminin Receptor-Mediated Tollip Signaling Induction in Lipopolysaccharide-Stimulated Human Intestinal Epithelial Cells.

BACKGROUND/AIMS: Inflammatory bowel disease (IBD) is a condition that involves chronic inflammation in all or part of the digestive tract. Often painful and debilitating, IBD can lead to life-threatening complications and increase the risk for colon cancer. In this study, we investigated the epigallocatechin-3-gallate (EGCG) mediated anti-inflammation response in lipopolysaccharide (LPS)-stimulated human colorectal cells through the negative regulator of Toll-like receptor (TLR) signaling. METHODS: human intestinal epithelial cells (HT-29) were used in all experiments. Cell cytotoxicity and nitric oxide (NO) were evaluated by WST-1 and the Griess reagent. Western blot analysis and ELISA were used to determine inflammatory mediators and 67-kDa laminin receptor (67LR)-mediated Tollip signaling pathways. RESULTS: Treatment of EGCG and LPS did not affect the cytotoxicity in HT-29 cells. LPS treatment dose-dependently increased the pro-inflammatory cytokine, such as interleukin (IL)-8, whereas EGCG significantly reduced the LPS-stimulated IL-8 production. Additionally, EGCG treatment markedly increased the Toll-interacting protein (Tollip) expression, which negatively regulates the TLR signaling in a dose and time-dependent manner. In particular, in the result from an RNA interference-mediated assay, our finding showed that silencing of Tollip resulted in abrogation of the inhibitory action of EGCG on LPS-induced production of pro-inflammatory mediators (inducible nitric oxide synthase-mediated NO/COX2, and IL-8) and activation of MAPKs and NF-κB signaling pathways. Interestingly, we also found that Tollip expression induced by EGCG could be modulated through 67LR expressed on the surface of HT-29 cells. CONCLUSIONS: Our novel finding indicates that 67LR and Tollip signaling activated by EGCG treatment is essential for inhibition of inflammation in human intestinal epithelial cells.

A novel Th1-type T-cell immunity-biasing effect of malate dehydrogenase derived from Mycobacterium avium subspecies paratuberculosis via the activation of dendritic cells.

Mycobacterium avium subspecies paratuberculosis (MAP) is the causative pathogen of Johne's disease in ruminants, characterized by chronic granulomatous enteritis; it also has zoonotic potential and is associated with Crohn's disease in humans. A better understanding of the mycobacterial antigens and their roles in the host immune response may facilitate the rational design of control strategies, including the development of effective vaccines and diagnostic tools. However, the functional roles of a large proportion of MAP antigens involved in modulating the host immune response remain unknown. In this study, an immunological role of MAP malate dehydrogenase (MDH, MAP2541c), an antigen that is upregulated in stress culture conditions, such as nutrient starvation and hypoxia, in polarizing naïve CD4+/CD8+ T cells toward Th1-biased T-cell immunity via the activation of dendritic cells (DCs) was identified. DCs treated with MAP MDH displayed characteristics of the activated and mature immune status, with augmented expression of cell surface molecules and pro-inflammatory cytokines, including TNF-α, IL-1ß, IL-6, and IL-12p70, but not IL-10, along with a dose-dependent decrease in the antigen uptake capacity. A mechanistic investigation revealed that the observed DC maturation is mediated by the activation of JNK, ERK, and p38 MAP kinases, and the NF-κB signaling pathway. Notably, DCs activated by MAP MDH treatment promoted naïve CD4+/CD8+ T cell proliferation; in particular, they effectively polarized naïve CD4+ T cells to secrete IFN-γ and IL-2 and activate T-bet, but, unlike the LPS control, did not influence IL-5 and GATA-3. These results indicated that MAP MDH has the potential to induce the Th1 cell response via DC activation. Collectively, our data demonstrated that MAP MDH is a novel immunostimulatory antigen that drives Th1-biased T cell polarization via interactions with DCs, suggesting that MDP MDH has the potential to be an effective MAP vaccine antigen target and diagnostic marker.

Neuroprotective effect of Capsicum annuum var. abbreviatum against hydrogen peroxide-induced oxidative stress in HT22 hippocampus cells.

Phenolic compounds isolated from pepper (Capsicum annum) have been demonstrated to have neuroprotective effects, whereas the physiological properties of Capsicum annuum var. abbreviatum (CAA) have not been studied. Thus, we investigate the chemical composition and neuroprotective activity of CAA extract (CAAE) in HT22 hippocampus cells against H2O2-induced neurotoxicity. CAAE treatment resulted in a significant protection of H2O2-exposed HT22, this protection ultimately occurred through an inhibition of MDA and ROS levels and an induction of SOD activity. Furthermore, CAAE treatment reduced H202-induced apoptosis though decreasing the expression of pro-apoptotic factors (Bax, cytochrome c, and cleaved caspases-3) while increasing the expression of the anti-apoptotic factors (Bcl-2), as well as the accumulation of nucleus-Nrf2-mediated HO-1 signaling. Interestingly, CAAE has a high concentration of unique phenolic compositions (chlrogenic acid, tangeretin, etc.) than other capsicum annum extracts. Altogether, these findings suggest that CAAE can be a useful natural resource for alleviating neurodegenerative diseases.

Neuroprotective effect of polysaccharide separated from Perilla frutescens Britton var. acuta Kudo against H2O2-induced oxidative stress in HT22 hippocampus cells.

This study was carried out to evaluate the neuroprotective activity of polysaccharide extracts isolated from Perilla frutescens (PEPF) in H2O2-treated HT22 hippocampus cells. The PEPF treatment was found to increase the anti-oxidant activities of HT22 hippocampus cells. PEPF treatment resulted in a significant protection of HT22 hippocampus cells against H2O2-induced neurotoxicity, this protection ultimately occurred through an inhibition of ROS-mediated intracellular Ca2+ levels leading to MAPKs and NF-κB, as well as the accumulation of PI3K/AKT and Nrf2-mediated HO-1/NQO1 pathways. Furthermore, PEPF not only decreased the expression of Bax, cytochrome c, and cleaved caspases-3, -8, and -9, but also increased the expression of PARP and Bcl-2 in the H2O2-treated HT22 hippocampus cells, which overall contributed to the neuroprotective action. PEPF retains its mitochondrial membrane potential and reduces the elevated levels of sub-G1 phase and apoptotic morphological features induced by H2O2. It also reduces the malondialdehyde levels and enhances the intracellular SOD activity.