Identification and Growth Characteristics of a Gluten-Degrading Bacterium from Wheat Grains for Gluten-Degrading Enzyme Production.

Immunogenic peptides from wheat gluten can be produced during digestion, which are difficult to digest by gastrointestinal proteases and negatively affect immune responses in humans. Gluten intolerance is a problem in countries where wheat is a staple food, and a gluten-free diet is commonly recommended for its treatment and prevention. Enzyme approaches for degradation of the peptides can be considered as a strategy for its prevention. Here, we isolated a gluten-degrading bacterium, Bacillus amyloliquefaciens subsp. plantarum, from wheat grains. The culture conditions for enzyme production or microbial use were considered based on gluten decomposition patterns. Additionally, the pH range for the activity of the crude enzyme was investigated. The bacterium production of gluten-degrading enzymes was temperature-dependent within 25 °C to 45 °C, and the production time decreased with increasing culture temperature. However, it was markedly decreased with increasing biofilm formation. The bacterium decomposed high-molecular-weight glutenin proteins first, followed by gliadin proteins, regardless of the culture temperature. Western blotting with an anti-gliadin antibody revealed that the bacterium decomposed immunogenic proteins related to α/ß-gliadins. The crude enzyme was active in the pH ranges of 5 to 8, and enzyme production was increased by adding gliadin into the culture medium. In this study, the potential of the B. amyloliquefaciens subsp. plantarum for gluten-degrading enzyme production was demonstrated. If further studies for purification of the enzyme specific to the immunogenic peptides and its characteristics are conducted, it may contribute as a strategy for prevention of gluten intolerance.

Effects of Sodium Alginate on the Physical Properties and Storage Stability of Freeze-Dried Tofu Coagulated with Crab Shell Extract.

The amount of processed by-products such as crab shells is increasing, but industrial utilization is insufficient. In our previous study, crab shell extract (CSE) acted as a coagulant for tofu manufacturing. This study aimed to reduce freeze-dried (FD) tofu breakdown by improving its physical properties through adding sodium alginate (SA). FD state in tofu helps increase storage and availability, but FD tofu frequently fractures during processing, which is a concern for manufacturers. Tofu samples were prepared with either crab shell extract (CSE) or MgCl2, and SA, and freeze-dried. In the yields of FD tofu samples, there were no significant differences (p < 0.05). The brokenness of FD tofu samples was lower in CSE than in MgCl2 and was significantly reduced by SA in both tofu samples, which was affected by hardness. The water-holding capacity decreased after freeze-drying, and CSE reduced this decrease, regardless of SA addition. The microstructures differed depending on the coagulant and were dense upon SA addition. The FD tofu was packed into a multilayer film and stored at 25 °C or 45 °C for 6 months to investigate storage stability. During the storage, brokenness was unchanged in all tofu samples, indicating that they maintained their original structure. There were no significant differences in the volatile base nitrogen and thiobarbituric acid values according to the coagulant type and SA addition (p < 0.05). In conclusion, SA reduced FD tofu breakdown by improving the network structure, which may help increase FD tofu quality and decrease economic loss.

Preparation and Taste Profiling of the Enzymatic Protein Hydrolysate from a by-Product of Red Snow Crab Processing as a Natural Seasoning Compound.

The red snow crab (Chionoecetes japonicus) is the most industrially processed in the Republic of Korea, and the meat is very popular, owing to its savory taste and flavor. Its body meat production comprises a two-step separation to increase meat yield. However, during the secondary separation, broken shell debris is occasionally entrained in the meat products, which is a concern for manufacturers. As the residues from first separation contain 39.9% protein, it can be utilized as an enzymatic protein hydrolysate (FPH) rich in free amino acids (FAAs). A combination of flavourzyme and alcalase (1:1) superiorly hydrolyzed the protein of the residues, and the best hydrolysis condition was suggested at 60 °C for 15 h with fourfold water and 2% enzyme addition, achieving a 57.4% degree of hydrolysis. The EPH was mostly composed of FAAs containing most essential amino acids; however, bitter-tasting amino acids accounted for 46.4% of the FAAs. To reduce the bitter taste, different nonvolatile organic acids were considered as masking agents, and citric and malic acids were effective, though the umami taste is slightly decreased. In conclusion, the crab processing residues can be utilized as an FAA-based natural seasoning compound through enzymatic hydrolysis and organic acid treatment.

Lactococcus lactis-fermented spinach juice suppresses LPS-induced expression of adhesion molecules and inflammatory cytokines through the NF-κB pathway in HUVECs.

Spinach (Spinacia oleracea L.), a green leafy vegetable, is widely regarded as a functional food due to its biological activities; however, to the best of our knowledge, there are no previous studies that have investigated the protective effects of fermented spinach against endothelial dysfunction and its underlying mechanisms. Therefore, this study investigated the effects and possible mechanisms of action of fresh spinach juice (S.juice) and fermented S.juice on lipopolysaccharide (LPS)-induced inflammatory responses in human umbilical vein endothelial cells (HUVECs). The HUVECs were treated with S.juice and fermented S.juice for 18 h before LPS exposure, and the levels of cytokines and chemokines, such as monocyte chemoattractant protein-1 (MCP-1) and interleukin-6 (IL-6), were detected using enzyme-linked immunosorbent assays (ELISA). Furthermore, to examine the changes in inflammatory responses to the two treatments, immunofluorescence analysis was used to visualize the nuclear translocation of nuclear factor-κB (NF-κB). Western blot analysis was also performed to detect the differences in the expression of endothelial cell adhesion molecules, specifically vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1). Both S.juice and fermented S.juice inhibited the LPS-induced expression of MCP-1 and IL-6, and suppressed VCAM-1 and ICAM-1. Additionally, fermented S.juice inhibited the LPS-induced activation of NF-κB and degradation of the inhibitor of NF-κB (IκBα) in an LPS dose-dependent manner. These results suggest that the anti-inflammatory effect of vitamin K2-enriched fermented S.juice is mediated by the suppression of the NF-κB pathway, suggesting its potential as a novel therapeutic candidate for inflammatory cardiovascular disease.

Association of FUT2 and ABO with Crohn's disease in Koreans.

BACKGROUND AND AIM: Fucosyltransferase 2 (FUT2) at 19q13 is a well-established susceptibility locus for Crohn's disease (CD) in Caucasians. FUT2 encodes α-1,2-fucosyltransferase that regulates the secretion of the α-1-2-N-acetylgalactosaminyltransferase and α-1-3-galactosyltransferase (ABO) antigens in both the gastrointestinal mucosa and secretory glands. Given that CD is thought to arise from dysregulated mucosal immune responses to the gut flora and both the ABO blood group and the FUT2 secretor status affect the composition of the gut microbiota, the goal of this study was to evaluate the associations of variants of FUT2 and ABO with CD in Koreans. METHODS: Three single-nucleotide polymorphisms from the FUT2 and ABO genes were genotyped in 1735 patients with CD and 8074 healthy controls. RESULTS: The FUT2 non-secretor allele showed genome-wide significant association with CD in Koreans (rs1047781, odds ratio [OR] = 1.30, Pcombined  = 3.52 × 10-12 ). The ABO locus showed genome-wide significant association with CD in Asians (Pmeta  = 2.35 × 10-8 ). A moderate association was observed with the A and B groups (OR = 1.40, P = 2.26 × 10-6 ; and OR = 1.32, P = 1.92 × 10-4 , respectively) compared with the O group. Following stratification on the basis of FUT2 genotype, carriers of the secretor O blood group were significantly protective against CD than were those of the secretor non-O blood group (OR = 0.63, 95% confidence interval = 0.54-0.73, P = 2.86 × 10-9 ). CONCLUSIONS: These are the first results indicating that the O blood group and FUT2 secretor status are protective factors against CD in Asians.

Effects of smoking on the association of human leukocyte antigen with ulcerative colitis.

BACKGROUND AND AIM: Tobacco smoking is a risk factor for gastrointestinal disorders, causing mucosal damage and impairing immune responses. However, smoking has been found to be protective against ulcerative colitis (UC). Human leukocyte antigen (HLA) is a major susceptibility locus for UC, and HLA-DRB1*15:02 has the strongest effect in Asians. This study investigated the effects of smoking on the association between HLA and UC. METHODS: The study enrolled 882 patients with UC, including 526 never, 151 current, and 205 former smokers, and 3091 healthy controls, including 2124 never, 502 current, and 465 former smokers. Smoking-stratified analyses of HLA data were performed using a case-control approach. RESULTS: In a case-control approach, HLA-DRB1*15:02 was associated with UC in never smokers (ORnever smokers  = 3.20, Pnever smokers  = 7.88 × 10-23 ) but not in current or former smokers (Pcurrent smokers  = 0.72 and Pformer smokers  = 0.33, respectively). In current smokers, HLA-DQB1*06 was associated with UC (ORcurrent smokers  = 2.59, Pcurrent smokers  = 6.39 × 10-12 ). No variants reached genome-wide significance in former smokers. CONCLUSIONS: An association between UC and HLA-DRB1*15:02 was limited to never smokers. Our findings highlight that tobacco smoking modifies the effects of HLA on the risk of UC.

Association of TRAP1 with infliximab-induced mucosal healing in Crohn's disease.

BACKGROUND AND AIM: Anti-tumor necrosis factor (TNF) agents, such as infliximab (IFX), have been increasingly used to induce and maintain disease remission in patients with Crohn's disease (CD). Despite a considerable non-response rate, little is known about the genetic predictors of response to anti-TNF therapy in CD. Our aim in this study was to investigate the genetic factors associated with response to anti-TNF therapy in patients with CD. METHODS: We performed a two-stage genome-wide association study (GWAS) to identify loci influencing the response to IFX among Korean patients with CD, comprising 42 good responders with mucosal healing and 70 non-responders. The achievement of mucosal healing was assessed by endoscopy and imaging. The functional significance of TRAP1 (TNF receptor associated protein 1) was examined using dextran sodium sulfate-induced colitis model in TRAP1 transgenic mice. RESULTS: The GWAS identified rs2158962, an intronic single nucleotide polymorphism (SNP) of TRAP1, significantly associated with mucosal healing (odds ratio = 4.94; Pcombined  = 1.35 × 10-7 ). In the dextran sodium sulfate-induced acute colitis, TRAP1 transgenic mice showed a better response to IFX than the wild-type mice. CONCLUSIONS: The TRAP1 gene is associated with mucosal healing in CD patients following IFX therapy. Identifying the genetic predictors of mucosal healing to anti-TNF therapy can prevent patients from exposure to ineffective therapies.

Effects of crab shell extract as a coagulant on the textural and sensorial properties of tofu (soybean curd).

To find an economic use of red snow crab (Chionoecetes japonicus) shell waste, we focused on its high mineral content. To evaluate its usability as a coagulant for tofu making, the effects of the crab shell extracts on the textural and sensorial properties of the tofu samples were investigated. The crab shell powder (CSP) and ash (CSA) were used for their extract preparation, and 1%-5% acetic acid treatment led to an abundance of calcium in the resulting extracts. The tofu yields of all the acetic acid extracts were comparable with those of the commercial coagulants MgCl2 and glucono-δ-lactone (GDL). Furthermore, the results for the textural attributes and sensorial acceptability demonstrated that either the extract from CSP prepared with 3% acetic acid or the extracts from CSA prepared with 1% or 3% acetic acid could be used as coagulants, because all the values of the extracts were statistically equivalent to those of the MgCl2 and GDL (p < 0.05).

Antimicrobial and Antibiofilm Activities of Sulfated Polysaccharides from Marine Algae against Dental Plaque Bacteria.

Dental plaque biofilms cause various dental diseases; therefore, inhibiting the growths of the dental plaque bacteria which produce biofilms can be a strategy for preventing dental disease. Certain sulfated polysaccharides from marine algae exert antimicrobial activities against human bacterial pathogens in addition to their physiological benefits. On the basis of these observations, the antimicrobial and antibiofilm activities of sulfated polysaccharides from different marine algae were evaluated against dental plaque bacteria. Among the sulfated polysaccharides, a fucoidan from Fucus vesiculosus showed notable antimicrobial activities against the selected dental plaque bacteria, including some foodborne pathogenic bacteria. The minimum inhibitory concentrations were of 125 to 1000 µg mL-1. Regarding the antibiofilm activity, the fucoidan at the concentrations of above 250 µg mL-1 completely suppressed the biofilm formations and planktonic cell growths of Streptococcus mutans and S. sobrinus. However, no eliminative effect on the completed biofilm was observed. The fucoidan consisted of almost fucose base polysaccharide containing approximately 14.0% sulfate content. The average molecular weight of the fucoidan was changed by heat treatment (121 °C for 15 min) and it affected the antimicrobial activity.

An Intergenic Variant rs9268877 Between HLA-DRA and HLA-DRB Contributes to the Clinical Course and Long-term Outcome of Ulcerative Colitis.

BACKGROUND AND AIMS: The genetic contribution to the prognosis of ulcerative colitis [UC] is poorly understood, and most currently known susceptibility loci are not associated with prognosis. To identify genetic variants influencing the prognosis of UC, we performed an Immunochip-based study using an extreme phenotype approach. METHODS: Based on the finding that the only association, Pdiscovery-meta <1 × 10-4, was located in the human leukocyte antigen [HLA], we focused our analyses on the HLA region. We performed the analysis using HLA imputation data from three independent discovery cohorts of 607 UC patients [243 poor-prognosis and 364 good-prognosis], followed by replication in 274 UC patients [145 poor-prognosis and 129 good-prognosis]. RESULTS: We found that rs9268877, located between HLA-DRA and HLA-DRB, was associated with poor-prognosis of UC at genome-wide significance (odds ratio [ORdiscovery] = 1.82; ORreplication = 1.55; ORcombined-meta = 1.72, pcombined-meta = 1.04 × 10-8), with effect size [OR] increasing incrementally according to worsening of prognosis in each of the three independent discovery cohorts and the replication cohort. However, rs9268877 showed no association with UC susceptibility [ORcombined-meta = 1.07, pcombined-meta = 0.135]; rs9268877 influenced 30-year clinical outcomes, and the presence of the rs9268877 risk allele had a sensitivity of 80.0% and specificity of 38.1% for colectomy. CONCLUSIONS: Our results provide new insights into prognosis-associated genetic variation in UC, which appears to be distinct from the genetic contribution to disease susceptibility. These findings could be useful in identifying poor-prognosis patients who might benefit from early aggressive therapy.

Immunochip Meta-Analysis of Inflammatory Bowel Disease Identifies Three Novel Loci and Four Novel Associations in Previously Reported Loci.

BACKGROUND AND AIMS: Recent meta-analysis of genome-wide association studies have identified over 241 inflammatory bowel disease susceptibility loci. However, the known variants only account for a fraction of inflammatory bowel disease heritability. To identify additional susceptibility loci, we performed a trans-ethnic meta-analysis as well as an Asian-specific meta-analysis, using all published Immunochip association results of inflammatory bowel disease. METHODS: An inverse-variance fixed-effects meta-analysis was carried out across Korean and East Asian Immunochip datasets of 4156 cases and 4904 controls [Asian ancestry]. A trans-ethnic meta-analysis of inflammatory bowel disease was performed together with the European datasets of 38 155 cases and 48 485 controls genotyped on the immunochip using a Bayesian approach, Meta-Analysis of Trans-ethnic Association studies [MANTRA]. RESULTS: We identified seven novel associations, including three novel susceptibility loci at MYO10-BASP1, PPP2R3C/KIAA0391/PSMA6/NFKB1A and LRRK1 as well as four novel secondary associations within previously known loci at NCF4, TSPAN32, CIITA and VANGL2. The new loci further implicate alterations in B cell biology in Crohn's disease pathogenesis. The effects of five loci were universal across European and Asian ancestries, whereas the NCF4 and CIITA loci showed significant heterogeneity between European and East Asian populations. In addition, 103 previously known IBD loci showed supporting evidence of association with nominal significance [p < 0.05] in Asians. CONCLUSIONS: Our findings of new loci not previously associated with IBD support the importance of studying inflammatory bowel disease genetics in diverse populations.

Association of CDKN2A/CDKN2B with inflammatory bowel disease in Koreans.

BACKGROUND AND AIM: CDKN2A/CDKN2B locus on 9p21 is reported to be associated with various diseases, including cancer and cardiovascular and inflammatory diseases. Significant downregulation of CDKN2B-AS1 in inflamed colon tissue of inflammatory bowel disease (IBD) cases was reported in Europeans. This study aimed to confirm the suggestive association of CDKN2A/CDKN2B with IBD identified in our recent genome-wide association study (GWAS). METHODS: We examined the association of CDKN2A/CDKN2B locus with IBD in an additional sample of 574 IBD cases and 542 controls, totaling 4068 cases and 8074 controls. In silico study was performed at various levels for functional annotation of the causal variant. Co-localization of the GWAS association signals and the corresponding expression quantitative trait loci in IBD-related tissues was evaluated using eCAVIAR. RESULTS: An expanded GWAS showed genome-wide significant association of rs3731257 at 9p21 with IBD (odds ratio = 1.17, 95% confidence interval = 1.12-1.22, Pcombined  = 5.68 × 10-9 ) and Crohn's disease (odds ratio = 1.22, 95% confidence interval = 1.15-1.28, Pcombined  = 8.85 × 10-9 ) in the Korean population. Co-localization study suggested that both CDKN2B-AS1 and CDKN2A might be functionally associated with the locus in the small intestine. CONCLUSIONS: rs3731257 in CDKN2A/CDKN2B is an IBD-susceptible locus in Koreans, with a suggestive role for small intestine-specific gene regulation. Our findings suggested that alterations of the CDKN2A/CDKN2B locus could affect the pathophysiology of IBD.

Effects of the Polysaccharide from the Sporophyll of Brown Alga Undaria Pinnatifida on Serum Lipid Profile and Fat Tissue Accumulation in Rats Fed a High-Fat Diet.

We investigated the effects of the polysaccharide from the sporophyll of a selected brown alga Undaria pinnatifida on serum lipid profile, fat tissue accumulation, and gastrointestinal transit time in rats fed a high-fat diet. The algal polysaccharide (AP) was prepared by the treatment of multiple cellulase-producing fungi Trichoderma reesei and obtained from the sporophyll with a yield of 38.7% (dry basis). The AP was mostly composed of alginate and fucoidan (up to 89%) in a ratio of 3.75:1. The AP was added to the high-fat diet in concentrations of 0.6% and 1.7% and was given to male Sprague-Dawley rats (5-wk-old) for 5 wk. The 1.7% AP addition notably reduced body weight gain and fat tissue accumulation, and it improved the serum lipid profile, including triglycerides, total cholesterol, and very low-density lipoprotein-cholesterol. The effects were associated with increased feces weight and shortened gastrointestinal transit time. In addition, the lipid peroxidation of the liver was decreased in both groups.

Influence of squid liver powder on accumulation of cadmium in serum, kidney and liver of mice.

In this study, the effect of squid liver powder intake on accumulation of cadmium in mice was investigated. Subjects were divided into 4 groups including the control group (CON), squid liver powder group with lipids not removed (SLP100), and squid liver powder groups with lipids removed (LFSLP50 and LFSLP100). Feed intake and food efficiency ratio of squid liver powder groups was significantly higher than the CON. As a result of investigating cadmium content in hair, serum, liver, and kidney during intake of squid liver powder, all groups showed increase in cadmium accumulation through consistent, long-term intake. Especially, cadmium content in liver and kidney of LFSLP100 was significantly higher than the content of SLP100 and CON. As a result of pathological observation on liver and kidney tissues according to squid liver powder diet, LFSLP100 showed most serious pathological symptoms. In case of kidney tissues, degeneration was significantly more severe in LFSLP100 compared to other groups. Such results suggest that cadmium concentration in human body can be increased by ingestion of whole squid including internal organs and that tissues can be damaged by increased cadmium concentration. More specific and systematic studies are deemed necessary.

Effects of the Cynanchum wilfordii Ethanol Extract on the Serum Lipid Profile in Hypercholesterolemic Rats.

The purpose of this study was to investigate the effects of the ethanol extract of Cynanchum wilfordii (ECW) on the blood lipid profile of hypercholesterolemic rats. Thirty 7-week-old male Sprague-Dawley rats were allowed free access to either a normal diet (AIN-93 diet), or 1% high-cholesterol diet with or without 0.5% or 1% ECW for 5 weeks. After sacrifice, the rat serum lipid profile was analyzed. The diets containing ECW decreased body weight gains compared to the normal diet. Serum HDL-cholesterol levels of ECW-fed groups were significantly increased in the hypercholesterolemic groups and normal groups (P<0.05). When 1% ECW was fed to the normal group, total cholesterol level was increased. Moreover, treatment of ECW in hypercholesterolemic groups yielded a dose-dependent and highly significant decrease in the atherogenic index as compared to the control. These results suggest that intake of Cynanchum wilfordii may help reduce the risks of hypercholesterolemia by increasing blood HDL-cholesterol and lowering the atherogenic index.