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Background Vaccination with tumor peptide epitopes associated with major histocompatibility complex class I molecules is an attractive approach directed at inducing tumor-specific cytotoxic T lymphocytes (CTLs). However, challenges remain in improving the therapeutic efficacy of peptide epitope vaccines, including the low immunogenicity of peptide epitopes and insufficient stimulation of innate immune components in vivo. To overcome this, we aimed to develop and test an innovative strategy that elicits potent CTL responses against tumor epitopes. The essential feature of this strategy is vaccination using tumor epitope-loaded nanoparticles (NPs) in combination with polyinosinic-polycytidylic acid (poly-IC) and anti-PD1 monoclonal antibody (mAb). Methods Carboxylated NPs were prepared using poly(lactic-co-glycolic acid) and poly(ethylene/maleic anhydride), covalently conjugated with anti-H-2KbmAbs, and then attached to H-2Kb molecules isolated from the tumor mass (H-2b). Native peptides associated with the H-2Kbmolecules of H-2Kb-attached NPs were exchanged with tumor peptide epitopes. The tumor-specific CTL-inducing and anti-tumor activities of the tumor epitope-loaded NPs were examined in mice bearing EG7-OVA thymoma or B16-F10 melanoma. In addition, the anti-tumor therapeutic efficacy of the NPs was examined in combination with poly-IC, anti-PD1 mAb, or both. Results Tumor peptide epitope-loaded NPs efficiently induced tumor-specific CTLs when used to immunize tumor-bearing mice as well as normal mice. This activity of the NPs significantly was increased when co-administered with poly-IC. Accordingly, the NPs exerted significant anti-tumor effects in mice implanted with EG7-OVA thymoma or B16-F10 melanoma, and the anti-tumor activity of the NPs was significantly increased when applied in combination with poly-IC. The most potent anti-tumor activity was observed when the NPs were co-administered with both poly-IC and anti-PD1 mAb. Conclusions Immunization with tumor epitope-loaded NPs in combination with poly-IC and anti-PD1 mAb in tumor-bearing mice can be a powerful means to induce tumor-specific CTLs with therapeutic anti-tumor activity.

Human breast milk is a potential source of bacteria for the development of the intestinal microbiota of infants. Several species within the genera Lactobacillus and Bifidobacterium were demonstrated to shape the gut microbiota of infants. In this study, the bacterial diversity was investigated in the breast milk and feces of a mother-infant pair, and probiotic candidates were identified. Importantly, the novel L. gasseri EJL and B. breve JTL strains were isolated from breast milk and infant feces samples, respectively; their completed genome was resolved using de novo sequencing. In addition, the bacterial composition in the infant’s feces at 1 week revealed the prevalence of Bifidobacterium and Streptococcus; a higher diversity was observed after 3 weeks. In particular, the abundance of Akkermansia was sharply increased at 7 weeks, further increasing thereafter, up to 15 weeks. Our results suggest that human breast milk and infant’s feces are a source of probiotic candidates.

Knowledge of the impact of the gut microbiota on human health has increased, and modulation of the bacterial community is now considered a therapeutic target for various diseases. Certain novel bacterial species have probiotic properties associated with improvement in obesity and related metabolic disorders. The relative abundance of

This study investigated the retreatability of EndoSeal MTA (Maruch, Wonju, Korea) according to the presence or absence of a canal isthmus and the additional use of passive ultrasonic irrigation (PUI) through microcomputed tomography (micro-CT) imaging. An epoxy resin sealer (AH Plus (Dentsply DeTrey, Konstanz, Germany)) was used as a reference for comparison. Forty-five artificial mandibular molars (TRUETOOTH #19, DELABS, Santa Barbara, CA) with a mesial canal with an isthmus and a distal canal without an isthmus were obturated using gutta-percha and one of the following sealers ( n = 15 each): AH Plus, EndoSeal MTA, and EndoSeal MTA + PUI. Micro-CT scanning was performed to assess the void volume (as a percentage) at three root levels. After the root fillings were removed, second micro-CT scanning was conducted to evaluate the amount of remaining root filling material. The Kruskal-Wallis H test and post hoc analysis were used for between-group comparisons. The Mann–Whitney U test was used for comparisons between canals with and without an isthmus ( p < 0.05 ). In the EndoSeal MTA group, the void volume and remaining filling materials were higher irrespective of the presence or absence of an isthmus. In apical lesions in the EndoSeal MTA group, the void ratio was significantly lower, and there was a significantly higher amount of remaining filling material. Regardless of the presence of an isthmus, the amount of remaining filling material of the EndoSeal + PUI group was reduced to a similar degree as the AH plus group. When performing retreatment for root canals filled with EndoSeal MTA, removal of the filling material can be more difficult in the apical region. The additional use of PUI can improve the efficacy of removal.

Gut microbes are closely associated with disease onset and improvement. However, the effects of gut microbes on the occurrence, prevention, and treatment of benign prostatic hyperplasia (BPH) are still unclear. We investigated the alteration of gut microbiota with implications for the diagnosis, prevention, and treatment of BPH and identified correlations among various indicators, including hormone indicators, apoptosis markers in BPH, and finasteride treatment models. BPH induction altered the abundance of Lactobacillus, Flavonifractor, Acetatifactor, Oscillibacter, Pseudoflavonifractor, Intestinimonas, and Butyricimonas genera, which are related to BPH indicators. Among these, the altered abundance of Lactobacillus and Acetatifactor was associated with the promotion and inhibition of prostate apoptosis, respectively. Finasteride treatment altered the abundance of Barnesiella, Acetatifactor, Butyricimonas, Desulfovibrio, Anaerobacterium, and Robinsoniella genera, which are related to BPH indicators. Among these, altered abundances of Desulfovibrio and Acetatifactor were associated with the promotion and inhibition of prostate apoptosis, respectively. In addition, the abundances of Lactobacillus and Acetatifactor were normalized after finasteride treatment. In conclusion, the association between apoptosis and altered abundances of Lactobacillus and Acetatifactor, among other gut microbes, suggests their potential utility in the diagnosis, prevention, and treatment of BPH.

Increasing evidence suggests that gut microbiota play an important role in metabolic disorders. Rubus occidentalis (RO) (black raspberry) has antioxidant and anti-inflammatory effects and improves metabolic disorders such as hypertension and insulin resistance. This study investigated the composition of gut microbiota in RO-treated mice using an aged mouse model of high-fat diet-induced obesity. RO significantly improved mouse glucose profiles, including serum glucose levels and intraperitoneal glucose tolerance test results. RO treatment caused significant increases in the relative abundance of various genera including Bacteroides, Butyricimonas, Mucispirillum, and Ruminococcus. Moreover, these increased genera correlated with the glucose profiles. Our findings suggest that the effect of RO on glucose control is associated with an increase in short-chain fatty acids producing bacteria.

Abnormal inflammatory responses are closely associated with intestinal microbial dysbiosis. Oral administration of Qmatrix-diabetes-mellitus complex (QDMC), an Aloe gel-based formula, has been reported to improve inflammation in type 2 diabetic mice; however, the role of the gut microbiota in ameliorating efficacy of QDMC remains unclear. We investigated the effect of QDMC on the gut microbiota in a type 2 diabetic aged mouse model that was administered a high-fat diet. Proinflammatory (TNF-α and IL-6) and anti-inflammatory (IL-4 and IL-10) cytokine levels in the fat were normalized via oral administration of QDMC, and relative abundances of Bacteroides, Butyricimonas, Ruminococcus, and Mucispirillum were simultaneously significantly increased. The abundance of these bacteria was correlated to the expression levels of cytokines. Our findings suggest that the immunomodulatory activity of QDMC is partly mediated by the altered gut microbiota composition.

Natural catchments have formed efficient river networks for a long time. similarly, urban drainage networks have been developed with the purpose of efficiently draining rainfall from catchments to flood mitigation. In this study, we analyzes and compares the characteristics between the naturally formed river networks for a long time and the artificially formed drainage networks using Gibbs’ Model. Gibbs’ Model is a stochastic stream network model, which can generate multiple realizations of stochastic networks based on a single parameter value of . Gibbs‘ Model was applied to a total number of 239 urban catchments in Seoul, South Korea and 70 natural catchments in the Midwestern areas of US. Topographic characteristics of catchments are analyzed along with the efficiency of drainage networks, which are presented by for both natural and urban catchments. The result of this study demonstrates the difference between natural and artificial drainage network characteristics and suggests a new alternative measures to mitigate flood risks in urban catchments facing extreme hydrologic events with climate change.

Bolts tend to show different fatigue behavior with respect to the size or shape of a bolt and the level of preload though it is the same material. This makes it difficult to predict the fatigue life of bolts. In this study, a novel method to construct a unified stress-life (S-N) curve was proposed for the three different bolts with different sizes and lengths subjected to various external axial loading conditions and preloads, ranging from 44 % to 88% of the ultimate tensile stress. A total of 133 experiments for 46 different fatigue conditions were conducted and finite element analysis was also performed to calculate multiaxial stress components at the root of the thread. The fatigue characteristics of the bolts were investigated depending on various loading conditions, and the life prediction performance with fitted S-N curves was compared with respect to four mean stress correction models. The unified S-N curve showed high accuracy in life prediction; 15 % mean absolute error without exceeding ± 50 % for finite life loading conditions.

We developed spontaneous diet-induced metabolic disease in mice by feeding them a high-fat diet for 23 weeks and administered Aloe QDM complex for 16 weeks to examine its restorative effect on immune disorders and metabolic syndrome. A series of immune functional assays indicated Aloe QDM complex enhanced lymphocyte proliferation and antigen-specific immunity as determined by the restored functions of cytotoxic T lymphocytes (CTL) and IgG production. The elevated serum TNF-α level was also regulated by Aloe QDM complex treatment, which suggested its complex therapeutic potential. As for metabolic phenotypes, oral administration of Aloe QDM complex significantly improved diabetic symptoms, including high fasting glucose levels and glucose tolerance, and distinctly alleviated lipid accumulation in adipose and hepatic tissue. The simultaneous restoration of Aloe QDM complex on metabolic syndrome and host immune dysfunction, especially on the specific CTL killing was first elucidated in our study.

Hyperlipidemia, which is closely associated with a fatty diet and aging, is commonly observed in the western and aged society. Therefore, a novel therapeutic approach for this disease is critical, and an immunological view has been suggested as a novel strategy, because hyperlipidemia is closely associated with inflammation and immune dysfunction. In this study, the effects of an aqueous extract of Rubus occidentalis (RO) in obese mice were investigated using immunological indexes. The mice were fed a high-fat diet (HFD) to induce hyperlipidemia, which was confirmed by biochemical analysis and examination of the mouse physiology. Two different doses of RO and rosuvastatin, a cholesterol synthesis inhibitor used as a control, were orally administered. Disturbances in immune cellularity as well as lymphocyte proliferation and cytokine production were significantly normalized by oral administration of RO, which also decreased the elevated serum tumor necrosis factor (TNF)-α level and total cholesterol. The specific immune-related actions of RO comprised considerable improvement in cytotoxic T cell killing functions and regulation of antibody production to within the normal range. The immunological evidence confirms the significant cholesterol-lowering effect of RO, suggesting its potential as a novel therapeutic agent for hyperlipidemia and associated immune decline.

Dysbiosis of the gut microbiota is a contributing factor for obesity-related metabolic diseases such as hyperglycemia and hyperlipidemia. Pharmacotherapy for metabolic diseases involves the modulation of gut microbiota, which is suggested to be a potential therapeutic target. In this study, the modulation of gut microbiota by statins (cholesterol-lowering drugs: atorvastatin and rosuvastatin) was investigated in an aged mouse model of high-fat diet-induced obesity, and the association between gut microbiota and immune responses was described. Atorvastatin and rosuvastatin significantly increased the abundance of the genera Bacteroides, Butyricimonas, and Mucispirillum. Moreover, the abundance of these genera was correlated with the inflammatory response, including levels of IL-1β and TGFβ1 in the ileum. In addition, oral fecal microbiota transplantation with fecal material collected from rosuvastatin-treated mouse groups improved hyperglycemia. From these results, the effect of statins on metabolic improvements could be explained by altered gut microbiota. Our findings suggest that the modulation of gut microbiota by statins has an important role in the therapeutic actions of these drugs.

Objective This study evaluated the presence of residual root canal filling material after retreatment using micro-computed tomography (micro-CT). Materials and Methods Extracted human teeth (single- and double-rooted, n = 21/each; C-shaped, n = 15) were prepared with ProFile and randomly assigned to three subgroups for obturation with gutta-percha and three different sealers (EndoSeal MTA, EndoSequence BC sealer, and AH Plus). After 10 days, the filling material was removed and the root canals were instrumented one size up from the previous master apical file size. The teeth were scanned using micro-CT before and after retreatment. The percentage of remaining filling material after retreatment was calculated at the coronal, middle, and apical thirds. Data were analyzed using the Kruskal-Wallis test and Mann-Whitney U test with Bonferroni post hoc correction. Results The tested sealers showed no significant differences in the percentage of remaining filling material in single- and double-rooted teeth, although EndoSeal MTA showed the highest value in C-shaped roots (p < 0.05). The percentage of remaining filling material of AH Plus and EndoSeal MTA was significantly higher in C-shaped roots than in single- or double-roots (p < 0.05), while that of BC sealer was similar across all root types. EndoSeal MTA showed the highest values at the apical thirds of single- and double-roots (p < 0.05); otherwise, no significant differences were observed among the coronal, middle, and apical thirds. Conclusions Within the limitations of this study, a large amount of EndoSeal MTA remained after retreatment, especially in C-shaped root canals.

IL-18 is a crucial pro-inflammatory cytokine that mediates chronic intestinal inflammation. Metformin, an anti-diabetic drug, was reported to have ameliorative effects on inflammatory bowel disease. Recently, the mechanism of action of metformin was explained as a modulation of gut microbiota. In this study, fecal microbiota transplantation (FMT) using fecal material from metformin-treated mice was found to upregulate the expression of GLP-1 and pattern-recognition receptors TLR1 and TLR4 for the improvement in hyperglycemia caused by a high-fat diet. Further, FMT downregulated the expression of the inflammatory cytokine IL-18. Within the genera Akkermansia, Bacteroides, and Butyricimonas, which were promoted by metformin therapy, Butyricimonas was found to be consistently abundant following FMT. Our findings suggest that modulation of gut microbiota is a key factor for the anti-inflammatory effects of metformin which is used for the treatment of hyperglycemia.

This study was designed to characterize the potential therapeutic effects of two statin drugs commonly used to treat dyslipidemia in inflammation-linked metabolic disorders related to type 2 diabetes. Atorvastatin (10 mg/kg/day) and rosuvastatin (3 mg/kg/day) were administered to mice with diet-induced obesity (DIO). The statins lowered serum total and LDL cholesterol levels, and improved the atherogenic index and cardiac risk index. Furthermore, the drugs decreased fasting glucose levels, improved glucose tolerance, and decreased fat tissue weight and adipocyte size; this was accompanied by an overall body weight loss tendency. The statins also improved antigen-specific immunity. The killing activity of cytotoxic T cells and exacerbation of IgG secretion levels were considerably normalized. Most importantly, serum tumor necrosis factor-α and interleukin 6 levels decreased, while their RNA expression levels in fat tissue were regulated by the statins as well. This study is the first to indicate that low doses of atorvastatin and rosuvastatin, the dosing regimen for which has been controversial, could significantly improve diabetes-related metabolic disorders, and could modulate pro-inflammatory cytokines, alleviating inflammation and simultaneously restoring overall humoral and cell-mediated immunity.

Polymeric nanoparticles containing both antigens and drugs have remarkable advantages because they can specifically deliver the antigens and the drugs to phagocytes such as dendritic cells and macrophages, thereby reducing the potential systemic side effects of the drugs. In the present study, we fabricated polymeric nanoparticles (NP) containing both ovalbumin (OVA) and dexamethasone (Dex) (NP[OVA+Dex]) using biodegradable and biocompatible polymer poly(lactic-co-glycolic acid) (PLGA), after which their immunomodulatory effects were examined. Immature dendritic cells treated with NP[OVA+Dex] were profoundly suppressed in both MHC class II- and class I-restricted OVA peptide presentation. In addition, mice injected with NP[OVA+Dex] were profoundly suppressed in generating OVA-specific cytotoxic T cells and also in producing OVA-specific IgG with concomitant increase of regulatory T cells. Furthermore, feeding of mice with NP[OVA+Dex] induced OVA-specific immune tolerance, as demonstrated by significant decrease in generating OVA-specific immune responses upon subsequent intravenous challenge with the nanoparticles containing OVA only. The present study demonstrates that oral feeding as well as intravenous injection of PLGA nanoparticles encapsulated with both antigen and Dex is a useful means for inducing antigen-specific immune tolerance.

The objective of this study was to evaluate the association between overweight, obesity and the incidence of advanced dental caries in South Korean adults, using alternate measures. The participants included 376,077 people aged 20 years and older who had health examination at least one time between 2005 and 2008. This evaluation is based on a change of body mass index (BMI) category, for 10 years, using a nationally representative data resource available from the National Health Insurance System. Instead of using decayed, missing, and filled teeth (DMFT), the diagnostic codes which indicate dental caries, pulpal disease and visiting frequency at dental health professionals were used in this case. A multivariate adjusted Cox regression analysis was performed to examine the association between advanced dental caries and BMI. In addition to the BMI, a multivariate analysis of gender, age, lifestyle behaviors and systemic disease information was included. To this end, the hazard ratio (HR) and 95% confidence interval (CI) were calculated. Chiefly, it is noted that the overweight and obese people were more likely to develop advanced dental caries independent of the noted variables. The positive association between high BMI and incidence of advanced dental caries was more prominent in the population’s characteristic of people who were in a classification of the elderly and women. Among the health and lifecycle behaviors, smoking or not was found to be one of the factors affecting the results. The alternate method used in this study showed that being overweight and obesity had a direct association with the incidence of advanced dental caries in Korean adults.

Metabolic syndrome is characterized by a combination of several metabolic disorders, including obesity, hyperglycemia, and hyperlipidemia. A simultaneous occurrence is one of the most crucial features of metabolic syndrome, therefore, we selected an animal model in which this would be reflected. We fed C57BL/6N mice a high-fat diet for 23 weeks to develop metabolic syndrome and examined the efficacy of Rubus occidentalis (RO) for hyperglycemia and hypercholesterolemia. Oral administration of RO for 16 weeks improved hyperglycemia as indicated by significantly decreased fasting glucose levels and a glucose tolerance test. Improvements were also observed in hypercholesterolemia, in which significant decreases in serum total cholesterol, non-high-density lipoprotein (non-HDL) cholesterol, apolipoprotein A-1, and apolipoprotein B levels were observed. The time comparison of major biomarkers, observed at the initiation and termination of the experimental period, consistently supported the beneficial effects of RO on each metabolic phenotype. In addition, RO treatment attenuated the excessive fat accumulation in hepatic and adipose tissue by decreasing the size and number of lipid droplets. These results suggested that RO simultaneously exerted antihyperglycemic and antihyperlipidemic effects in mice with diet-induced metabolic syndrome.

Probiotics have been used for the treatment of various disorders or as alternative therapies. The stability of dual-coated probiotics is increased in the gastrointestinal environment. The aim of the present study was to evaluate the hepatoprotective effects of dual-coated and uncoated probiotic supplements, following liver injury. Albino Wistar rats were orally treated with probiotics daily and carbon tetrachloride (CCl4) was administered on the seventh and eighth days to induce acute liver damage. Hepatoprotective effects were determined by assessment of serum glutamic–oxaloacetic transaminase (SGOT) and serum glutamic–pyruvic transaminase (SGPT) activities, as well as by histopathological examination. The CCl4-treated control group showed increased SGOT and SGPT activities as compared with the normal control group. However, treatment with probiotics reduced SGOT and SGPT activities, following CCl4 administration. Animals treated with probiotics showed reduced liver weight than that in the standard CCl4 ...

Summary The purpose of this study was to analyze the sealing ability of different root canal filling materials over a 6-week period using a glucose penetration model. Forty-six recently extracted human premolars were used in this study. The root canals were enlarged to 40/0.06. Prepared canals were randomly assigned into four groups (n = 10) as follows: Group 1, Gutta-Percha (GP)/AH Plus with cold lateral compaction; Group 2, GP/AH Plus with continuous wave compaction; Group 3, RealSeal SE obturation system; and Group 4, OrthoMTA. The remaining specimens were used as positive and negative controls, and all specimens underwent thermocycling (10,000; 5–55 °C). The sealing ability of all samples was evaluated at 24 h, 1, 2, 4, and 6 weeks using a quantitative glucose leakage model, and scanning electron microscope (SEM) images were taken. A mixed effect analysis using R statistical language was performed. Groups 1, 2, and 4 showed low leakage levels during experimental periods. Group 3 showed low leakage levels for the first 2 weeks; however, the leakage level was significantly increased after 2 weeks compared to negative control group (p

The gut microbiota is a contributing factor in obesity-related metabolic disorders. The effect of metformin on the gut microbiota has been reported; however, the relationship between the gut microbiota and the mechanism of action of metformin in elderly individuals is unclear. In this study, the effect of metformin on the gut microbiota was investigated in aged obese mice. The abundance of the genera Akkermansia, Bacteroides, Butyricimonas, and Parabacteroides was significantly increased by metformin in mice fed a high-fat diet. Metformin treatment decreased the expression of IL-1β and IL-6 in epididymal fat, which was correlated with the abundance of various bacterial genera. In addition, both fecal microbiota transplantation from metformin-treated mice and extracellular vesicles of Akkermansia muciniphila improved the body weight and lipid profiles of the mice. Our findings suggest that modulation of the gut microbiota by metformin results in metabolic improvements in aged mice, and that these effects are associated with inflammatory immune responses.

Tolerogenic dendritic cells (tDCs) represent a promising tool for cellular therapy against autoimmune diseases, allergies, and transplantation rejection. Numerous pharmacological agents are known to induce tDC generation. Minocycline, which has long been used as a broad-spectrum antibiotic, was recently shown to significantly increase the generation of DCs with regulatory properties. Here, we examined the effect of the combination of minocycline with dexamethasone, rapamycin, vitamin D3, and interleukin (IL)-10, which are all known inducers of tDC generation. The highest number of tDCs was generated when minocycline and dexamethasone were used together with granulocyte colony-stimulating factor (GM-SCF) and IL-4. The tolerogenicity of the minocycline/dexamethasone-conditioned tDCs was much better than or at least equal to those of the tDCs generated with either one of these agents, as assessed through in vitro phenotypic and functional assays. In addition, pretreatment with MOG35-55 peptide-pulsed minocycline/dexamethasone-conditioned tDCs significantly ameliorated the clinical signs of experimental autoimmune encephalitis induced by MOG peptide injection in a murine model. These results confirmed that tDCs with potent tolerogenic properties could be efficiently generated by the combined use of minocycline and dexamethasone, along with GM-CSF and IL-4. Our results would help in the development of ex vivo tDC-based immunotherapies.

Myeloid-derived suppressor cells (MDSCs) mediate tumor-associated immune suppression in both cancer patients and tumor-bearing animals. Reduction or elimination of MDSCs reduces the rate of tumor progression and improves cancer therapies that employ mechanisms of immunity. Here we show that baccatin III, which is the precursor for the semisynthesis of paclitaxel, exerts anti-tumor immunomodulatory activity in very low doses (0.05-0.5mg/kg), although it is regarded as an inactive derivative of paclitaxel. Oral administration of baccatin III significantly reduced the growth of tumors induced by engrafting BALB/c mice with either 4 T1 mammary carcinoma or CT26 colon cancer cells. Baccatin III (0.5mg/kg) did not exert anti-tumor activity in athymic nude mice. Baccatin III decreased the accumulation of MDSCs in the spleens of the tumor-bearing mice. Furthermore, MDSCs isolated from baccatin III-treated mice, compared with those isolated from vehicle-treated mice, had a significantly reduced suppressive effect on T cells treated with the anti-CD3 and anti-CD28 monoclonal antibodies. Moreover, these cells produced significantly reduced amounts of reactive oxygen species and nitric oxide. These results suggest that baccatin III reduced tumor progression by inhibiting the accumulation and suppressive function of MDSCs.

Background Little information is available regarding the precise swine leukocyte antigen (SLA)-derived immunogenic peptides that are presented in the context of human HLA molecules. Here, we identified SLA-derived immunogenic peptides that are presented in association with human HLA-A2 molecule. Methods The SLA-derived peptides that bind to HLA-A*0201, a representative of the A2 supertype, were predicted using a computer-assisted algorithm. The candidate peptides were synthesized, and the stabilities of complexes formed between peptides and HLA-A*0201 were compared using major histocompatibility complex (MHC) stabilization assays. The cytotoxic T lymphocyte (CTL)-inducing activity of the selected peptides was examined in HLA-A*0201-transgenic mice. Results Among 15 candidate peptides synthesized, two peptides, peptide-35 (YLGPDGLLL) and peptide-43 (TLICHVDSI), were selected to have high affinity and stability with HLA-A*0201. Examination of the CTL-inducing activity of the two peptides in HLA-A*0201-transgenic mice showed that immunization with peptide-35, but not peptide-43, elicited potent CD8-specific CTL responses. The Peptide-35 is present in non-polymorphic α2 domains of 34 SLA-1 alleles, 18 SLA-2 alleles, and 1 SLA-3 allele. Conclusion This study identifies an immunogenic HLA-A*0201-restricted epitope derived from the SLA, which may be valuable for the development of epitope-specific immunoregulation strategies.

Biodegradable poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles (NPs) encapsulated with antigens can efficiently deliver the antigens to antigen-presenting cells such as dendritic cells (DCs). Active form of vitamin D, 1α,25-dihydroxyvitamin D3 (VD3), is known to have various immunosuppressive effects. Encapsulation of VD3 within NPs would enhance the delivery of VD3 to APCs, while preventing the potential systemic side effects of VD3. In this study, we examined the effects of PLGA NPs encapsulated with ovalbumin (OVA) and VD3. PLGA NPs containing OVA and VD3 (NP[OVA+VD3]) were prepared by W1/O/W2 double emulsion solvent evaporation method. Bone marrow-derived DCs treated with NP[OVA+VD3] were decreased in the expression of co-stimulatory molecules, and produced higher levels of immunosuppressive cytokines, while producing lower levels of pro-inflammatory cytokines, compared with DCs treated with NP[OVA]. Addition of NP[OVA+VD3] to allogeneic MLR reduced the proliferation of allogenic T cells. In addition, NP[OVA+VD3]-treated DCs efficiently generated CD4+CD25+Foxp3+Treg cells from CD4+CD25− T cells. Furthermore, mice injected i.v. with NP[OVA+VD3] were suppressed in inducing OVA-specific cytotoxic T cells. These results show that phagocytic delivery of antigen and VD3 to APCs using biodegradable NPs efficiently suppresses antigen-specific immune responses.

Adiponectin is an adipocyte hormone involved in glucose and lipid metabolism. The aim of this study was to develop a human adiponectin expression system in transgenic silkworm using a human adiponectin expression vector. The silk gland of the silkworm is a highly specialized organ that has the wonderful ability to synthesize and secrete silk protein. To express human adiponectin in the silk gland of transgenic silkworm, targeting vectors pB-A3-adiponectin-IRES-RFP and pB-Ser1-adiponectin-IRES-RFP were constructed and then introduced into the silkworm pupa. The transgenic silkworms were verified by PCR and then generated. The level of adiponectin in the transgenic silkworm was 6-10 ng/50 mg of freeze-dried powder, and western blotting using an antibody against human adiponectin demonstrated a specific band with a molecular weight of 30 kDa in the silkworm. These results showed that human adiponectin introduced into the silkworm genome was expressed successfully on a large-scale.

Chronic stress generally experienced in our daily lives; is known to augment disease vulnerability by suppressing the host immune system. In the present study; the effect of modified Aloe polysaccharide (MAP) on chronic stress-induced immunosuppression was studied; this Aloe compound was characterized in our earlier study. Mice were orally administered with MAP for 24 days and exposed to electric foot shock (EFS; duration; 3 min; interval; 10 s; intensity; 2 mA) for 17 days. The stress-related immunosuppression and restorative effect of MAP were then analyzed by measuring various immunological parameters. MAP treatment alleviated lymphoid atrophy and body weight loss. The numbers of lymphocyte subsets were significantly normalized in MAP-treated mice. Oral administration of MAP also restored the proliferative activities of lymphocytes; ovalbumin (OVA)-specific T cell proliferation; antibody production; and the cell killing activity of cytotoxic T lymphocytes. In summary; oral administration of MAP ameliorated chronic EFS stress-induced immunosuppression.

Dioscoreae Rhizome (DR) has been used in traditional medicine to treat numerous diseases and is reported to have anti-diabetes and anti-tumor activities. To identify a bioactive traditional medicine with anti-inflammatory activity of a water extract of DR (EDR), we determined the mRNA and protein levels of proinflammatory cytokines in macrophages through RT-PCR and western blot analysis and performed a FACS analysis for measuring surface molecules. EDR dose-dependently decreased the production of NO and pro-inflammatory cytokines such as IL-1β, IL-6, TNF-α, and PGE(2), as well as mRNA levels of iNOS, COX-2, and pro-inflammatory cytokines, as determined by western blot and RT-PCR analysis, respectively. The expression of co-stimulatory molecules such as B7-1 and B7-2 was also reduced by EDR. Furthermore, activation of the nuclear transcription factor, NF-κB, but not that of IL-4 and IL-10, in macrophages was inhibited by EDR. These results show that EDR decreased pro-inflammatory cytokines via inhibition of NF-κB-dependent inflammatory protein level, suggesting that EDR could be a useful immunomodulatory agent for treating immunological diseases.

Background Insulin resistance is an integral feature of metabolic syndromes, including obesity, hyperglycemia, and hyperlipidemia. In this study, we evaluated whether the aloe component could reduce obesity-induced inflammation and the occurrence of metabolic disorders such as blood glucose and insulin resistance. Methods Male C57BL/6 obese mice fed a high-fat diet for 54 days received a supplement of aloe formula (PAG, ALS, Aloe QDM, and Aloe QDM complex) or pioglitazone (PGZ) and were compared with unsupplemented controls (high-fat diet; HFD) or mice fed a regular diet (RD). RT-PCR and western blot analysis were used to quantify the expression of obesity-induced inflammation. Results Aloe QDM lowered fasting blood glucose and plasma insulin compared with HFD. Obesity-induced inflammatory cytokine (IL-1β, -6, -12, TNF-α) and chemokine (CX3CL1, CCL5) mRNA and protein were decreased markedly, as was macrophage infiltration and hepatic triglycerides by Aloe QDM. At the same time, Aloe QDM decreased the mRNA and protein of PPARγ/LXRα and 11β-HSD1 both in the liver and WAT. Conclusion Dietary aloe formula reduces obesity-induced glucose tolerance not only by suppressing inflammatory responses but also by inducing anti-inflammatory cytokines in the WAT and liver, both of which are important peripheral tissues affecting insulin resistance. The effect of Aloe QDM complex in the WAT and liver are related to its dual action on PPARγ and 11β-HSD1 expression and its use as a nutritional intervention against T2D and obesity-related inflammation is suggested.

Sulforaphane is an isothiocyanate found in cruciferous vegetables that has been reported to be an effective cancer preventive agent inducing growth arrest and/or cell death in cancer cells of various organs. This paper reports that sulforaphane exerts immunomodulatory activity on the MHC-restricted antigen presenting function. Sulforaphane efficiently increased the class II-restricted presentation of an exogenous antigen, ovalbumin (OVA), in both dendritic cells (DCs) and peritoneal macrophages in vitro. The class II-restricted OVA presentation-enhancing activity of sulforaphane was also confirmed using mice that had been injected with sulforaphane followed by soluble OVA. On the other hand, sulforaphane did not affect the class I-restricted presentation of exogenous OVA at concentrations that increase the class II-restricted antigen presentation. At a high concentration (), sulforaphane inhibited the class I-restricted presentation of exogenous OVA. Sulforaphane did not affect the phagocytic activity of the DCs, and the cell surface expression of total H-, B7-1, B7-2 and CD54 molecules, even though it increased the expression of I- molecules to a barely discernable level. These results show that sulforaphane increases the class II-restricted antigen presenting function preferentially, and might provide a novel insight into the mechanisms of the anti-cancer effects of sulforaphane.

The effects of intraphagosomal toll-like receptor (TLR) activation on the MHC-restricted presentation of exogenous antigen were examined in dendritic cells (DCs). For phagosomal targeting, nanoparticles containing both a TLR agonist and a model antigen, ovalbumin (OVA), were prepared using biodegradable polymer poly(D,L-lactic acid-co-glycolic acid) and were then opsonized with mouse IgG. After incubating mouse DCs with the nanoparticles, the efficacy of OVA peptide presentation was evaluated using OVA-specific CD8 and CD4 T cells. Inclusion of either the TLR3 agonist poly(I:C) or the TLR9 agonist CpG oligodeoxynucleotides (ODN) significantly increased and prolonged both MHC class I- and class II-restricted OVA presentation. Accordingly, the DCs that phagocytosed the nanoparticles containing poly(I:C) or CpG ODN together with OVA efficiently induced the proliferation of OVA-specific CD8 and CD4 T cells. The potency levels of poly(I:C) and CpG ODN in increasing the MHC-restricted presentation of the exogenous antigen appeared to be similar. A combination of the 2 TLR agonists was synergistic in increasing the MHC class I-restricted, but not the class II-restricted, presentation of exogenous antigen. These results show that IgG-opsonized biodegradable nanoparticles containing both intraphagosomal TLR agonists and antigens can be efficient carrier materials in inducing antigen-specific T cell responses.

－ Obesity-induced disorders contribute to the development of metabolic diseases such as insulin resistance, fatty liver diseases, and type 2 diabetes (T2D). In this study, we evaluated the hypoglycemic and hypolipidemic effects of aloe formula in high fat diet (HFD)-fed C57BL/6N mice. Male mice fed HFD for 28 weeks received a supplement of aloe formula, PAG, ALS, Aloe QDM, and an Aloe QDM complex for a further 8 weeks and were then compared with regular diet fed mice. After the experimental period, the blood glucose levels of the Aloe QDM complex-and PGZ-supplemented mice were significantly lower than those of the HFD-fed mice. Aloe formula, especially the Aloe QDM complex, and the PGZ treatment group profoundly affected the IPGTT and HOMA-IR. Immunochemistry was done for the morphological observation and the resulting sizes of adipocytes around the epididymis were significantly decreased when comparing the aloe formula-treated and HFD-fed groups. Further, aloe formula decreased mRNA expression of fatty acid synthesis enzymes and led to reduced hepatic steatosis in both liver and WAT. These results suggest that supplementation of Aloe QDM complex in the HFD-fed mice improved insulin resistance by lowering blood glucose levels and reducing adipocytes. Our data suggest that dietary aloe formula reduces obesity-induced glucose tolerance by suppressing fatty acid synthesis in the WAT and liver, both of which are important peripheral tissues affecting insulin resistance. The Aloe QDM complex could be used as a nutritional inter-vention against T2D.Keywords: High-fat diet, Type 2 diabetes, Diet-induced obesity, Insulin sensitivity

One hundred ninety-three frozen food samples collected in Korea various public bazaars from October 2006 to September 2007. Staphylococci were detected in 21.8% of frozen food samples. Staphylococcus aureus was isolated from 17 (8.8%) samples. Other staphylococci isolates were identified as S. warneri (7.8%), S. epidermidis (2.1%), S. xylosus (1.6%), S. eguorum (1%), and S. vitulinus (0.5%). Additionally, the antimicrobial susceptibility of 42 staphylococcal isolates to ten different antimicrobial agents was determined. The staphylococcal isolates demonstrated antimicrobial resistance to mupirocin (31%) oxacillin (14.3%), gentamicin (9.5%), teicoplanin (7.1%) and ciprofloxacin (7.1%). Most of the staphylococcal isolates showed high-level resistance to mupirocin (MIC(90), >128 microg/mL). Fortunately, most of the isolates were susceptible to vancomycin. The total bacteria and Escherichia coli count were tested to investigate the microbiological quality of frozen foods. From 193 frozen food samples, 43 (22.3%), 34 (17.6%) and 19 (9.8%) samples were shown to be of unacceptable quality due to total bacteria, coliform and E. coli counts, respectively.

The effects of processed Aloe vera gel (PAG) on the course of established diet-induced non-insulin-dependent diabetes mellitus (NIDDM) were studied in C57BL/6J mice. NIDDM was induced in C57BL/6J mice by feeding them a high-fat diet. Mice exhibiting diet-induced obesity (DIO) with blood glucose levels above 180mg/dl were selected to examine the antidiabetic effects of PAG. Oral administration of PAG for 8 weeks reduced circulating blood glucose concentrations to a normal level in these DIO mice. In addition, the administration of PAG significantly decreased plasma insulin. The antidiabetic effects of PAG were also confirmed by intraperitoneal glucose tolerance testing. PAG appeared to lower blood glucose levels by decreasing insulin resistance. The administration of PAG also lowered triacylglyceride levels in liver and plasma. Histological examinations of periepididymal fat pad showed that PAG reduced the average size of adipocytes. These results demonstrate that the oral administration of PAG prevents the progression of NIDDM-related symptoms in high-fat diet-fed mice, and suggest that PAG could be useful for treating NIDDM.

Background It has been recently noticed that type 2 diabetes (T2D), one of the most common metabolic diseases, causes a chronic low-grade inflammation and activation of the innate immune system that are closely involved in the pathogenesis of T2D. Cordyceps militaris, a traditional medicinal mushroom, produces a component compound, cordycepin (3'-deoxyadenosine). Cordycepin has been known to have many pharmacological activities including immunological stimulating, anti-cancer, and anti-infection activities. The molecular mechanisms of cordycepin in T2D are not clear. In the present study, we tested the role of cordycepin on the anti-diabetic effect and anti-inflammatory cascades in LPS-stimulated RAW 264.7 cells. Methods We confirmed the levels of diabetes regulating genes mRNA and protein of cytokines through RT-PCR and western blot analysis and followed by FACS analysis for the surface molecules. Results Cordycepin inhibited the production of NO and pro-inflammatory cytokines such as IL-1β, IL-6, and TNF-α in LPS-activated macrophages via suppressing protein expression of pro-inflammatory mediators. T2D regulating genes such as 11β-HSD1 and PPARγ were decreased as well as expression of co-stimulatory molecules such as ICAM-1 and B7-1/-2 were also decreased with the increment of its concentration. In accordance with suppressed pro-inflammatory cytokine production lead to inhibition of diabetic regulating genes in activated macrophages. Cordycepin suppressed NF-κB activation in LPS-activated macrophages. Conclusion Based on these observations, cordycepin suppressed T2D regulating genes through the inactivation of NF-κB dependent inflammatory responses and suggesting that cordycepin will provide potential use as an immunomodulatory agent for treating immunological diseases.

Background Macrophages generated in vitro using macrophage-colony stimulating factor (M-CSF) and interleukin (IL)-6 from bone marrow cells (BM-Mp) are defective in antigen presenting cell (APC) function as shown by their ability to induce the proliferation of anti-CD3 mAb-primed syngeneic T cells. However, they do express major histocompatibility (MHC) class I and II molecules, accessory molecules and intracellular adhesion molecules. Here we demonstrate that the defective APC function of macrophages is mainly due to production of TGF-β1 by BM-Mp. Methods Microarray analysis showed that TGF-β1 was highly expressed in BM-Mp, compared to a macrophage cell line, B6D, which exerted efficient APC function. Production of TGF-β1 by BM-Mp was confirmed by neutralization experiments of TGF-β1 as well as by real time-polymerase chain reaction (PCR). Results Addition of anti-TGF-β1 monoclonal antibody to cultures of BM-Mp and anti-CD3 mAb-primed syngeneic T cells efficiently induced the proliferation of syngeneic T cells. Conversely, the APC function of B6D cells was almost completely suppressed by addition of TGF-β1. Quantitative real time-PCR analysis also confirmed the enhanced expression of TGF-β1 in BM-Mp. Conclusion The defective APC function of macrophages generated in vitro with M-CSF and IL-6 was mainly due to the production of TGF-β1 by macrophages.

It is a hot clinical issue whether newly approved antimicrobial agents such as daptomycin, linezolid, quinupristin/dalfopristin (synercid) and tigecycline are active enough to be used for infections caused by vancomycin resistant bacteria. We performed susceptibility tests for mupirocin, which is in widespread clinical use in Korea, and four new antimicrobials, daptomycin, linezolid, quinupristin/dalfopristin and tigecycline, against vancomycin-resistant Enterococcus faecalis and Enterococcus faecium isolated from Korean patients in 1998 and 2005 to evaluate and compare the in vitro activity of these antimicrobials. Among these agents, quinupristin/dalfopristin, which is rarely used in hospitals in Korea, showed relatively high resistance to several vancomycin-resistant enterococci (VRE) isolated in 2005. Likewise, daptomycin, linezolid and tigecycline have not yet been in clinical use in Korea. However, our results showed that most of the 2005 VRE isolates were already resistant to linezolid and daptomycin (highest minimum inhibitory concentration (MIC) value >100 microg/ml). Compared with the other four antimicrobial agents tested in this study, tigecycline generally showed the greatest activity against VRE. However, four strains of 2005 isolates exhibited resistance against tigecycline (MIC >12.5 microg/ml). Almost all VRE were resistant to mupirocin, whereas all E. faecium isolated in 1998 were inhibited at concentrations between 0.8 to approximately 1.6 microg/ml. In conclusion, resistances to these new antimicrobial agents were exhibited in most of VRE strains even though these new antibiotics have been rarely used in Korean hospitals.

Biodegradable nanospheres generated from a biocompatible polymer, poly(D,L-lactide-co-gly-colide) (PLGA), have been studied extensively as implantable reservoirs for sustained-release drug delivery. PLGA-nanospheres have also been studied as vehicles to deliver antigens to phagocytes. The intracellular processing pathway of antigens delivered to phagocytes by PLGA particles was studied in the present study. Ovalbumin (OVA) encapsulated with PLGA (OVA-nanosphere) was efficiently captured, processed and presented on class I major histocompatibility complex (MHC-I) by dendritic cells (DCs). The MHC-I processing of OVA-nano-spheres was resistant to lactacystin, a proteosome inhibitor, and brefeldin A, which blocks anterograde transport from the endoplasmic reticulum (ER) through the Golgi apparatus. Chloroquine, which inhibits phagolysosomal enzymes by increasing phagolysosomal pH, inhibited MHC-I processing of OVA-nanospheres. In addition, DCs generated from TAP-/- mice were markedly suppressed in MHC-I processing of OVA-nanospheres. These results demonstrate that DCs process phagocytosed OVA-nanospheres via a vacuolar alternate MHC-I pathway for presentation of OVA peptides to T lymphocytes.

Enterococcus faecalis and Enterococcus faecium isolated from Korea patients in 1998 and 2005 were tested for susceptibility to nine different antimicrobial agents, including vancomycin and teicoplanin. Polymerase chain reaction (PCR) was used for the detection of several vancomycin resistance genes such as vanA ('high level'), vanB ('moderate high level'), vanC1 and vanC2 ('low level'). Both E. faecalis and E. faecium exhibited a resistance of 80% and more than 60% to synercid and mupirocin, respectively. Moreover, an average of 76% of all isolates was resistant to ciprofloxacin, gentamicin, lincomycin, cefotaxime, and meropenem, confirming the multiple drug resistance of most of the isolates. No resistance to vancomycin or teicoplanin was observed in the 1998 E. faecalis and E. faecium isolates. However, the 2005 E. faecalis and E. faecium isolates exhibited resistance of 16% and 12% to vancomycin and teicoplanin, respectively. In addition, vanA gene was detected in 4 strains of 2005 E. faecium isolates, thus showing a high resistance to vancomycin. No other vancomycin resistance genes, including vanB, vanC1, and vanC2, were found in our isolates. In this study, we compared the occurrence of antimicrobial resistance and the presence of specific resistance genes in E. faecalis and E. faecium. The results showed that the 2005 isolates were usually more resistant than the 1998 isolates.

Previous studies have shown that dendritic cells (DCs) can phagocytize, process and present a microencapsulated form of ovalbumin (OVA) in the context of class I MHC as well as class II MHC. In the present study, we examined the effects of recombinant human macrophage-colony stimulating factor (M-CSF) on the MHC-restricted presentation of microencapsulated OVA by DCs. Two types of DCs were generated from mouse bone marrow (BM) cells, one type with granulocyte/macrophage-colony stimulating factor (GM-CSF) alone, the other type with GM-CSF and interleukin (IL)-4. Pretreatment with M-CSF significantly enhanced both class I MHC and class II MHC-restricted presentation of exogenous OVA by both types of DCs. The enhancing activity of M-CSF on antigen presentation was more potent in DCs generated with GM-CSF alone compared to DCs generated with both GM-CSF and IL-4. Pretreatment of the DCs with M-CSF did not increase phagocytic activity or total level of expression of class I MHC (H-2K b ) molecules, but increased expression of OVA peptide-H-2K b complexes upon phagocytosis of microencapsulated OVA. These results demonstrate that M-CSF increases intracellular processing events of phagocytized antigen in DCs.

Leukotactin-1 (Lkn-1), a human CC chemokine that binds to both CC chemokine receptor (CCR)1 and CCR3, is distinct from other human CC chemokines in that it has long amino acid residues preceding the first cysteine at the NH 2 -terminus. Serial deletion studies showed that at least three amino acid residues, alanine–alanine–aspartic acid (A–A–D), preceding the first cysteine at the NH 2 -terminus are essential for the biological activity of Lkn-1. Point mutation and deletion studies for the three amino acids were performed in the present study. Substitutions of the first alanine residue with other amino acids did not cause significant loss of biological activities. Deletion of the third amino acid, aspartic acid, resulted in more than 100-fold loss of the activity. Deletion of two amino acids, alanine–alanine (A–A) or alanine–aspartic acid (A–D), resulted in almost complete loss of the activity. Loss of agonistic activity by deletion of two amino acids was due to impaired binding to CCR1. These results identify that alanine–aspartic acid residues preceding the first cysteine at the NH 2 -terminus are essential for the binding and biological activity of Lkn-1.

The main targets for the immunosuppressive calcineurin inhibitors, cyclosporin A (CsA) and tacrolimus, have been considered to be activated T cells, but not antigen-presenting cells. Here we demonstrate that CsA and tacrolimus, but not rapamycin, inhibit major histocompatibility complex (MHC)–restricted antigen presentation in dendritic cells (DCs). Microencapsulated ovalbumin (OVA) was efficiently captured, processed, and presented on both class I MHC molecules (cross-presentation) as well as on class II MHC molecules. Addition of CsA and tacrolimus, but not rapamycin, to cultures of DCs inhibited both the class I processing pathway and the class II processing pathway of exogenous OVA. In addition, CsA and tacrolimus, but not rapamycin, also inhibited the classic class I processing pathway of endogenous OVA. CsA and tacrolimus did not inhibit presentation of exogenously added OVA peptide, SIINFEKL, phagocytic activity of DCs, or the total level of expression of class I MHC (H-2Kb) molecules. CsA and tacrolimus, however, inhibited profoundly the expression of SIINFEKL-H-2Kb complexes in OVA-phagocytized DCs. These results demonstrate clearly that CsA and tacrolimus inhibit intracellular processing events of antigens, and further suggest that the immunosuppressive activity of CsA and tacrolimus is at least in part due to inhibition of antigen processing pathways.