Characterization and anti-tumor activities of polysaccharide isolated from Brassica rapa L. via activation of macrophages through TLR2-and TLR4-Dependent pathways.

We have previously shown the immunostimulatory effects by Nozawana (Brassica rapa L.). In this report, we determined the characteristics of Nozawana polysaccharide (NPS) and evaluated the immunomodulatory effects and anti-tumor activity of NPS mediated by macrophage activation. The molecular weight of NPS was determined by gel filtration chromatography with an average molecular weight of approximately 100.6 kDa. HPLC analysis showed that NPS contained glucose, galacturonic acid, galactose, and arabinose. NPS increased cytokine and nitric oxide (NO) production by macrophages in a Toll-like receptor (TLR)2 and TLR4-dependent manner. Furthermore, NPS induced apoptosis significantly against 4T1 murine breast cancer cells cultured in conditioned medium from NPS-treated macrophages through tumor necrosis factor-α. In tumor-bearing mouse model, tumor growth was significantly reduced in NPS-treated mice compared with control mice. These results support the potential use of NPS as an immunotherapeutic material found in health food products.

Antioxidant and Antimicrobial Activities of Thai Edible Plant Extracts Prepared Using Different Extraction Techniques.

This study investigated the antioxidant and antimicrobial activities of six Thai edible plant leaf extracts, including Cashew (CN), Chamuang (CM), Monpu (MP), Thurianthet (TT), Kradon (KD) and Pakliang (PL), extracted using ethanol extraction (EE), microwave-assisted extraction (MAE), and ultrasonic-assisted extraction (UAE). The leaf extracts were characterized for percentage yield, total phenolic content (TPC), total flavonoid content (TFC), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and-ferric reducing antioxidant power (FRAP) activity, and antimicrobial activity against spoilage. MAE produced the highest percentage yields, among which MAE-extracted MP exhibited the highest yield. Furthermore, the highest TPC and TFC were obtained for MAE, with MAE-extracted KD and CN showing the highest TPC and TFC, respectively, among the samples. The highest DPPH and FRAP values were seen in MAE-processed CN, KD, and MP extracts. The inhibition zone of pathogenic bacteria, minimum inhibitory concentration, and minimum bacterial concentration were determined in all samples except TT. These findings indicate that, compared to EE and UAE, MAE improved the antioxidant and antimicrobial efficacy of the leaf extracts. The aforementioned extracts could be employed as natural food additives to prevent chemical and microbial spoilage of foods.

Glochidion littorale Leaf Extract Exhibits Neuroprotective Effects in Caenorhabditis elegans via DAF-16 Activation.

A number of plants used in folk medicine in Thailand and Eastern Asia are attracting interest due to the high bioactivities of their extracts. The aim of this study was to screen the edible leaf extracts of 20 plants found in Thailand and investigate the potential neuroprotective effects of the most bioactive sample. The total phenol and flavonoid content and 2,2-diphenyl-1-picrylhydrazyl radical-scavenging activity were determined for all 20 leaf extracts. Based on these assays, Glochidion littorale leaf extract (GLE), which showed a high value in all tested parameters, was used in further experiments to evaluate its effects on neurodegeneration in Caenorhabditis elegans. GLE treatment ameliorated H2O2-induced oxidative stress by attenuating the accumulation of reactive oxygen species and protected the worms against 1-methyl-4-phenylpyridinium-induced neurodegeneration. The neuroprotective effects observed may be associated with the activation of the transcription factor DAF-16. The characterization of this extract by LC-MS identified several phenolic compounds, including myricetin, coumestrin, chlorogenic acid, and hesperidin, which may play a key role in neuroprotection. This study reports the novel neuroprotective activity of GLE, which may be used to develop treatments for neurodegenerative diseases such as Parkinson's syndrome.

Intracellular cAMP contents regulate NAMPT expression via induction of C/EBPß in adipocytes.

A decline in intracellular nicotinamide adenine mononucleotide (NAD+) causes adipose tissue dysfunction. Nicotinamide phosphoribosyltransferase (NAMPT) catalyzes the rate-limiting step in the NAD+ biosynthesis pathway. However, the molecular mechanism that mediates regulation of NAMPT expression in adipocytes is yet to be elucidated. This study found that intracellular cAMP regulates NAMPT expression and promoter activity in 3T3-L1 adipocytes. cAMP-mediated Nampt promoter activity was suppressed by protein kinase A inhibitor H89, whereas AMP-activated protein kinase inhibitor compound C did not affect cAMP-mediated Nampt promoter activity. Intracellular cAMP induced CCAAT/enhancer-binding protein ß (C/EBPß) expression. Knockdown of C/EBPß suppressed NAMPT expression and promoter activity. Furthermore, the Nampt promoter was activated by C/EBPß, while LIP activated the dominant-negative form of C/EBPß. Promoter sequence analysis revealed that the region from -96 to -76 on Nampt was required for C/EBPß-mediated promoter activity. Additionally, chromatin immunoprecipitation assay demonstrated that C/EBPß was bound to the promoter sequences of Nampt. Finally, NAMPT inhibitor FK866 suppressed adipogenesis in 3T3-L1 cells, and this suppressive effect was restored by nicotinamide mononucleotide treatment. These findings showed that intracellular cAMP increased NAMPT levels by induction of C/EBPß expression and indicated that the induction of NAMPT expression was important for adipogenesis.

Enhanced Anti-Allergic Activity of Milk Casein Phosphopeptide by Additional Phosphorylation in Ovalbumin-Sensitized Mice.

The proteolytic digest of milk casein, known as casein phosphopeptide (CPP-III), exhibits diverse biological activities, including calcium absorption and antioxidant activities. We hypothesized that the additional phosphorylation of this peptide can enhance its immunomodulatory activity such as suppression of allergy-associated cytokine and antigen-specific immune response. This study was conducted to assess whether oral intake of additionally phosphorylated CPP-III (P-CPP) attenuates ovalbumin (OVA)-induced IgE-mediated allergic reactions because of the additional phosphate groups. Female BALB/c mice were intraperitoneally sensitized with OVA twice at intervals of 14 days and then orally fed native CPP-III (N-CPP), P-CPP, and dephosphorylated CPP-III (D-CPP) for 6 weeks. Next, the mice were orally challenged with 50 mg of OVA. Oral administration of P-CPP suppressed total and specific IgE levels in the serum. Mice fed P-CPP exhibited low levels of OVA-specific IgG1 and increased OVA-specific IgG2a. P-CPP also suppressed IL-4 production, while D-CPP showed similar a level compared to that of the control. Further, P-CPP increased the population of the T follicular helper (Tfh) cell in the spleen. These results suggest that additional phosphorylation of CPP can enhance the attenuation of allergen-specific IgE-modulated allergic reactions in a murine food allergy model.

Theobromine suppresses adipogenesis through enhancement of CCAAT-enhancer-binding protein ß degradation by adenosine receptor A1.

Theobromine, a methylxanthine derived from cacao beans, reportedly has various health-promoting properties but molecular mechanism by which effects of theobromine on adipocyte differentiation and adipogenesis remains unclear. In this study, we aimed to clarify the molecular mechanisms of the anti-adipogenic effect of theobromine in vitro and in vivo. ICR mice (4week-old) were administered with theobromine (0.1g/kg) for 7days. Theobromine administration attenuated gains in body and epididymal adipose tissue weights in mice and suppressed expression of adipogenic-associated genes in mouse adipose tissue. In 3T3-L1 preadipocytes, theobromine caused degradation of C/EBPß protein by the ubiquitin-proteasome pathway. Pull down assay showed that theobromine selectively interacts with adenosine receptor A1 (AR1), and AR1 knockdown inhibited theobromine-induced C/EBPß degradation. Theobromine increased sumoylation of C/EBPß at Lys133. Expression of the small ubiquitin-like modifier (SUMO)-specific protease 2 (SENP2) gene, coding for a desumoylation enzyme, was suppressed by theobromine. In vivo knockdown studies showed that AR1 knockdown in mice attenuated the anti-adipogenic effects of theobromine in younger mice. Theobromine suppresses adipocyte differentiation and induced C/EBPß degradation by increasing its sumoylation. Furthermore, the inhibition of AR1 signaling is important for theobromine-induced C/EBPß degradation.

Theophylline suppresses interleukin-6 expression by inhibiting glucocorticoid receptor signaling in pre-adipocytes.

Adipose tissues in obese individuals are characterized by a state of chronic low-grade inflammation. Pre-adipocytes and adipocytes in this state secrete pro-inflammatory adipokines, such as interleukin 6 (IL-6), which induce insulin resistance and hyperglycemia. Theophylline (1,3-dimethylxanthine) exerts anti-inflammatory effects, but its effects on pro-inflammatory adipokine secretion by pre-adipocytes and adipocytes have not been examined. In this study, we found that theophylline decreased IL-6 secretion by 3T3-L1 pre-adipocytes and mouse-derived primary pre-adipocytes. The synthetic glucocorticoid dexamethasone (DEX) induced IL-6 expression in 3T3-L1 pre-adipocytes, and this effect was suppressed by theophylline at the mRNA level. Knockdown of CCAAT/enhancer binding protein (C/EBP) δ inhibited DEX-induced IL-6 expression, and theophylline suppressed C/EBPδ expression. Furthermore, theophylline suppressed transcriptional activity of the glucocorticoid receptor (GR) through suppression of nuclear localization of GR. In vivo, glucocorticoid corticosterone treatment (100 µg/mL) increased fasting blood glucose and plasma IL-6 levels in C57BL/6 N mice. Theophylline administration (0.1% diet) reduced corticosterone-increased fasting blood glucose, plasma IL-6 levels, and Il6 gene expression in adipose tissues. These results show that theophylline administration attenuated glucocorticoid-induced hyperglycemia and IL-6 production by inhibiting GR activity. The present findings indicate the potential of theophylline as a candidate therapeutic agent to treat insulin resistance and hyperglycemia.

Role of phosphate groups on antiviral activity of casein phosphopeptide against feline calicivirus as a surrogate for norovirus.

BACKGROUND: Current research on the gastrointestinal digestion of milk-casein strongly suggests the existence of novel bioactive peptides with antiviral activities that are attributable to their immunostimulatory effects. In the present study, we investigated the antiviral activity of casein peptides rich in phosphate groups, such as casein phosphopeptide (CPP-III). RESULTS: We prepared two types of CPP with different phosphorylation levels to clarify the role of the phosphate group. Further phosphorylation of CPP-III was conducted by dry heating with sodium pyrophosphate, whereas dephosphorylation was performed enzymatically using alkaline phosphatase and alkaline treatment. Feline calicivirus (FCV) strain F9, a typical norovirus surrogate, and Crandell Rees feline kidney cells were used as the target virus and host cells, respectively. Antiviral activity was determined based on the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and quantitative polymerase chain reaction quantification of antiviral cytokine mRNA expression. Higher cell viability was observed in the host cells treated with phosphorylated CPP-III, and a significant up-regulation of type 1 interferon expression was induced compared to that treated with native CPP-III. However, dephosphorylation of CPP-III resulted in a decrease in the anti-FCV effect. CONCLUSION: The CPP effect was enhanced by the introduction of additional phosphates and conversely weakened by their elimination. Therefore, CPP-III phosphorylation represents an emerging approach for the production of food-grade antiviral agents. © 2016 Society of Chemical Industry.

Functional expression of amyloidogenic human stefins A and B in Pichia pastoris using codon optimization.

Complementary DNAs encoding human stefins A (HSA) and B (HSB) were synthesized using Pichia-preferred codons by overlap extension PCR. The full-length genes were ligated downstream of the glyceraldehyde-3-phosphate dehydrogenase promoter in the Pichia expression vector pGAPZαC and successfully expressed in Pichia pastoris strain X-33. Functional recombinant HSA and HSB proteins were purified from culture medium at yields of 121.3 ± 13.5 (n = 3) and 95.4 ± 4.1 (n = 3) mg/L, respectively. Using this expression strategy, we demonstrated that high levels of bioactive recombinant HSA and HSB can be produced by fermentation in P. pastoris.

Orally administered low-molecular weight agaro-oligosaccharides are absorbed into the plasma of healthy humans.

Agaro-oligosaccharides (AOSs) are known to have biological activities, such as anti-inflammatory, anti-tumor, and anti-obesity effects. Although existing evidence suggests the presence of AOSs in peripheral tissues after oral administration, whether AOSs permeate into the blood circulation remains unknown. Thus, we hypothesized that AOSs with low-molecular weight can permeate the human gastrointestinal tract. To test this hypothesis, the time course of absorption was examined by analyzing plasma samples before and 1, 2, and 4 h after ingestion. Analysis was performed using liquid chromatography/mass spectrometry after labeling with p-aminobenzoic ethyl ester. Our results showed that the plasma concentration of agarobiose (Abi) was higher than that of agarotetraose (Ate); however, agarohexaose was not detected. Additionally, plasma levels of Abi and Ate were proportional to the dose. These results suggest that permeation efficiency is dependent on the molecular weight and that the systemic absorption of Abi via the gastrointestinal tract is better than that of Ate. These findings will contribute to a better understanding of the bioactivity of orally administered AOSs in peripheral tissues.

Fermented soybean foods (natto) ameliorate age-related cognitive decline by hippocampal TAAR1-mediated activation of the CaMKII/CREB/BDNF signaling pathway in senescence-accelerated mouse prone 8 (SAMP8).

Natto is a traditional fermented soybean-based food that has been an integral part of Japanese cuisine for several centuries. Although there have been extensive studies on the cognitive benefits of soybeans, only limited studies have examined the effects of natto on cognitive function. This study investigated the potential cognitive benefits of natto in senescence-accelerated mouse-prone 8 (SAMP8) mice. After 12 weeks of oral administering natto fermented for 18 h, the spatial learning and memory performance were improved compared with those in SAMP8 control mice. Furthermore, activation of the brain-derived neurotrophic factor (BDNF)/tropomyosin receptor kinase B (TrkB)/cAMP response element-binding protein (CREB) signaling and N-methyl-D-aspartate receptor (NMDAR)-calcium/calmodulin-dependent protein kinase II (CaMKII) cascade was observed in the hippocampus of SAMP8 mice that were fed natto. Additionally, natto administration upregulated trace amine-associated receptor 1 (TAAR1) as a modulator of NMDAR. These findings suggest that natto ameliorates cognitive decline by activating the TAAR1-mediated CaMKII/CREB/BDNF signaling pathway in the hippocampus of SAMP8 mice.

Genistein enhances NAD+ biosynthesis by upregulating nicotinamide phosphoribosyltransferase in adipocytes.

A decrease in the NAD+ level in adipocytes causes adipose-tissue dysfunction, leading to systemic glucose, and lipid metabolism failure. Therefore, it is necessary to develop small molecules and nutraceuticals that can increase NAD+ levels in adipocytes. Genistein, a nutraceutical derived from soybeans, has various physiological activities and improves glucose and lipid metabolism. In this study, we aimed to unravel the effects of genistein on the NAD+ level in adipocytes and the underlying molecular mechanisms. Genistein enhanced NAD+ biosynthesis by increasing the expression of nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in NAD+ biosynthesis. A pull-down assay using genistein-immobilized beads revealed prohibitin 1 (PHB1) as a target protein of genistein. The knockdown of Phb1 suppressed the genistein-induced increase in NAMPT expression and NAD+ level in adipocytes. Genistein-bound PHB1 contributed to the stabilization of the transcription factor CCAAT/enhancer-binding protein ß through the activation of extracellular signal-regulated kinase, resulting in increased NAMPT expression at the transcriptional level. Genistein induced the dephosphorylation of peroxisome proliferator-activated receptor at serine 273 and increased the level of the insulin-sensitizing adipokine adiponectin in adipocytes, whereas the knockdown of Nampt and Phb1 abolished these genistein-mediated effects. Our results proved the potential efficacy of genistein in increasing the NAD+ level and restoring metabolic function in adipocytes. Furthermore, we identified PHB1, localized to the plasma membrane, as a novel candidate target protein for increased expression of NAMPT in adipocytes. Overall, these findings will assist in developing NAD+-boosting nutraceuticals to alleviate metabolic dysfunctions in adipose tissues.

Lacticaseibacillus paracasei K71 Alleviates UVB-Induced Skin Barrier Dysfunction by Attenuating Inflammation via Increased IL-10 Production in Mice.

SCOPE: Ultraviolet B (UVB) radiation causes skin barrier dysfunction, leading to decreased water-holding capacity, impaired epidermal barrier function, and increased skin thickness. This study investigates the protective effects of oral administration of Lacticaseibacillus paracasei K71 against skin barrier dysfunction in UVB-irradiated mice. METHODS AND RESULTS: Mice are fed diets with or without K71 and irradiated with UVB three times a week for 12 weeks. Oral administration of K71 suppresses UVB-induced decrease in stratum corneum water content, mitigates the increase of transepidermal water loss, and decreases epidermal thickness of the dorsal skin. Treatment with K71 reverses the upregulation of inflammatory cytokines and the activation of nuclear factor-κB induced by UVB irradiation and upregulates the expression of anti-inflammatory IL-10 in the dorsal skin. Notable upregulation of IL-10 is observed in the spleens of K71-treated mice. K71 treatment enhances IL-10 production in J774.1 macrophages; however, this enhancement is diminished by inhibiting K71 phagocytosis and TLR3. Furthermore, transfection using K71 RNAs significantly increases IL-10 production. CONCLUSION: These results indicate that K71 may alleviate UVB-induced skin barrier dysfunction by attenuating inflammation via increasing IL-10 production and that K71 RNAs may induce IL-10 production in macrophages. Therefore, K71 may be beneficial for preventing skin barrier dysfunction.

Benign papillary mesothelioma of the peritoneum associated with gastric cancer: report of a case.

Benign papillary mesothelioma of the peritoneum is an uncommon lesion that is usually discovered by chance during a surgical procedure. This lesion resembles metastatic carcinoma in gross appearance; therefore, intraoperative diagnosis can be difficult. This report presents a case of benign papillary mesothelioma concurrent with gastric cancer. The tumor was located on the hepatogastric ligament and resembled a metastatic peritoneal implant. A pathological review confirmed the diagnosis to be benign papillary mesothelioma.

Immunomodulatory Properties of Highly Viscous Polysaccharide Extract from the Gagome Alga (Kjellmaniella crassifolia).

Marine brown algae are rich in sulfated polysaccharides, which have the ability to form gels and viscous solution. Sulfated polysaccharides exhibit many biological activities; however, little is known whether the viscoelastic property in the polysaccharide extract is correlated with biological activities. We examined the immunomodulatory properties of highly viscous polysaccharide extract (HVPE) from Gagome Kjellmaniella crassifolia in a murine model, and the effects were compared with those of a less viscous polysaccharide extract (LVPE). HVPE or LVPE (10, 30, and 100 mg/kg/day) were orally administered to C57BL/6 mice for 14 days. Secretions of cytokine and IgA in Con A-stimulated spleen and Peyer's patch (PP) cells and phagocytic activity of peritoneal macrophages was determined. IFN-γ, IL-12, IL-6, and IgA secretions showed high levels in spleen cell cultures from mice administered HVPE, whereas these effects were diminished in the LVPE-administered mice. The phagocytic activity of peritoneal macrophages was enhanced by the continuous oral administration of HVPE, and these effects were higher than those of LVPE. Furthermore, an increase in IgA secretion by administration of HVPE was observed in Con A-stimulated PP cells. These results suggest that the polysaccharide extract from K. crassifolia has immunomodulatory activities, which depend on the viscosity.

Current Trends in Food-Derived Peptidic Antioxidants.

Recently, peptidic antioxidants have attracted much attention due to their promising applications in the production of valuable functional food and nutraceuticals with health-promoting properties [...].

Intestinal permeability of agaro-oligosaccharides: Transport across Caco-2 cell monolayers and pharmacokinetics in rats.

Agaro-oligosaccharides (AOSs), even-numbered oligosaccharides prepared from agar, are applied to various food, including supplements, drinks, and jellies because of their biological activities. This study aimed to evaluate the AOS permeation in the gastrointestinal tract in vivo and in vitro. Agarobiose (Abi), agarotetraose (Ate), and agarohexaose (Ahe) were detected in rat plasma after oral administration of AOSs. The detection level of agarobiose in the plasma was higher than that of agarohexaose, which was consistent with the permeation study using Caco-2 cell monolayers. Further, the adenosine triphosphate inhibitor (sodium azide) or endocytosis inhibitor (colchicine) did not inhibit AOS permeation through Caco-2 cell monolayers. Conversely, AOS permeation enhanced upon treatment with cytochalasin B, a tight junction disrupter, suggesting that AOSs might have passed mainly through the tight junctions between the intestinal epithelial cells. These results indicate that AOSs, especially agarobiose, can be absorbed as an intact form via the gastrointestinal tract across the intestinal epithelium through the paracellular pathway.

Buckwheat Flour and Its Starch Prevent Age-Related Cognitive Decline by Increasing Hippocampal BDNF Production in Senescence-Accelerated Mouse Prone 8 Mice.

Buckwheat is an important pseudo-cereal crop worldwide. This study investigated whether long-term administration of buckwheat can suppress age-related cognitive decline in senescence-accelerated mouse prone 8 (SAMP8) mice. For 26 weeks, 18-week-old male SAMP8 mice were fed a standard diet containing 5% (w/w) buckwheat, Tartary buckwheat, wheat, or rice flour. In the Barnes maze and passive avoidance tests, mice fed buckwheat whole flour (BWF) showed improved cognitive performance compared to those fed a control diet, while no improvement was noticed in case of the other diets. Analysis of the gut microbiota showed that BWF and buckwheat outer flour administration increased the abundance of Lactococcus and Ruminiclostridium, respectively, at the genus level. The expression levels of brain-derived neurotrophic factor (BDNF), postsynaptic Arc and PSD95, and the mature neuronal marker NeuN in the hippocampus were increased after BWF administration, which was induced by the activation of the ERK/CREB signaling pathway and histone H3 acetylation. A similar increase in cognitive performance-related hippocampal BDNF expression in SAMP8 mice was observed after the oral administration of starch prepared from BWF. Therefore, the long-term administration of BWF suppresses cognitive decline by increasing hippocampal BDNF production in SAMP8 mice.

Antioxidative ability of chicken myofibrillar protein developed by glycosylation and changes in the solubility and thermal stability.

Myofibrillar protein prepared from chicken breast muscle was incubated with several concentrations of glucose or maltose for 6 h at 60 °C and 35% relative humidity in order to obtain glycosylated chicken protein. When the ratio of the weights of the myofibrillar protein and glucose or maltose had respectively reached 1:6 or 1:3-5, the solubility of each type of glycosylated chicken protein in a 0.1 M NaCl solution was exceeded by about 60%, although the myofibrillar protein was insoluble in a low ionic strength solution. Moreover, when the myofibril and maltose reaction (myofibril:maltose = 1:4) was extended to 36 h, the glycosylated protein did not undergo denaturation when held at 50 °C for 2 h, while it also exhibited an antioxidative function against superoxide anion radicals.

Potential of plant-derived peptides for the improvement of memory and cognitive function.

Recently, there has been an increased demand for functional foods, to reduce the risk of age-related cognitive impairment, dementia, and Alzheimer's disease. Among them, plant-derived bioactive compounds, such as phytochemicals and peptides, have notable potential in improving memory and cognitive functions. Many studies have provided potential data concerning the characteristics and structure-activity relationships of memory-enhancing peptides. When considering the proof of efficacy of these plant-based peptides in humans as neurological treatment options, it is necessary to accumulate evidence concerning their bioavailability and permeability through blood-brain barrier (BBB). This review focuses on the memory-enhancing effects of peptides derived from plant proteins and presents a current perspective on their structure-activity relationships and BBB permeability.