Evaluation and selection of potent fluorescent immunosensors by combining fluorescent peptide and nanobodies displayed on yeast surface.

Quenchbody (Q-body) is a quench-based fluorescent immunosensor labeled with fluorescent dye(s) near the antigen-binding site of an antibody. Q-bodies can detect a range of target molecules rapidly and directly. However, because Q-bodies show different antigen responses depending on the antibody used, time-consuming optimization of the Q-body structure is often necessary, and a high-throughput screening method for discriminating and selecting good Q-bodies is required. Here, we aimed to develop a molecular display method of nanobody-based "mini Q-bodies" by combining yeast surface display and coiled-coil forming E4/K4 peptide-based fluorescence labeling. As a result, the yeast-displayed mini Q-body recognizing the anti-cancer agent methotrexate (MTX) showed significant quenching and MTX-dependent dequenching on cells. To demonstrate the applicability of the developed method to select highly responsive mini Q-bodies, a small nanobody library consisting of 30 variants that recognize human serum albumin was used as a model. The best variant, showing a 2.4-fold signal increase, was obtained through selection by flow cytometry. Furthermore, the same nanobody prepared from Escherichia coli also worked as a mini Q-body after dye labeling. The described approach will be applied to quickly obtain well-behaved Q-bodies and other fluorescent biosensors for various targets through directed evolutionary approaches.

SPINK7 Recognizes Fungi and Initiates Hemocyte-Mediated Immune Defense Against Fungal Infections.

Serine protease inhibitors of Kazal-type (SPINKs) were widely identified in vertebrates and invertebrates, and played regulatory roles in digestion, coagulation, and fibrinolysis. In this study, we reported the important role of SPINK7 in regulating immune defense of silkworm, Bombyx mori. SPINK7 contains three Kazal domains and has 6 conserved cysteine residues in each domain. Quantitative real-time PCR analyses revealed that SPINK7 was exclusively expressed in hemocytes and was upregulated after infection with two fungi, Saccharomyces cerevisiae and Candida albicans. Enzyme activity inhibition test showed that SPINK7 significantly inhibited the activity of proteinase K from C. albicans. Additionally, SPINK7 inhibited the growth of three fungal spores, including S. cerevisiae, C. albicans, and Beauveria bassiana. The pathogen-associated molecular patterns (PAMP) binding assays suggested that SPINK7 could bind to ß-D-glucan and agglutinate B. bassiana and C. albicans. In vitro assays were performed using SPINK7-coated agarose beads, and indicated that SPINK7 promoted encapsulation and melanization of agarose beads by B. mori hemocytes. Furthermore, co-localization studies using immunofluorescence revealed that SPINK7 induced hemocytes to aggregate and entrap the fungi spores of B. bassiana and C. albicans. Our study revealed that SPINK7 could recognize fungal PAMP and induce the aggregation, melanization, and encapsulation of hemocytes, and provided valuable clues for understanding the innate immunity and cellular immunity in insects.

Enhanced immune response by vacuoles isolated from Saccharomyces cerevisiae in RAW 264.7 macrophages.

Vacuoles are membrane vesicles in eukaryotic cells, the digestive system of cells that break down substances absorbed outside the cell and digest the useless components of the cell itself. Researches on anticancer and intractable diseases using vacuoles are being actively conducted. The practical application of the present study to animals requires the determination of the biocompatibility of vacuole. In the present study, we evaluated the effects of vacuoles isolated from Saccharomyces cerevisiae in RAW 264.7 cells. This showed a significant increase in the production of nitric oxide (NO) produced by macrophage activity. Using Reactive Oxygen Species (ROS) assay, we identified that ROS is increased in a manner dependent on vacuole concentration. Western blot analysis showed that vacuole concentration-dependently increased protein levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2). Therefore, iNOS expression was stimulated to induce NO production. In addition, pro-inflammatory cytokines levels promoted, such as interleukin (IL) 6 (IL-6) and tumor necrosis factor (TNF) α (TNF-α). In summary, vacuoles activate the immune response of macrophages by promoting the production of immune-mediated transporters NO, ROS, and pro-inflammatory cytokines.

Serological markers facilitate the diagnosis of Crohn's disease.

Background and aim: The diagnosis of Crohn's disease (CD) is challenging. Ongoing search for biomarkers to facilitate the diagnosis is a worthwhile endeavor. The aim of this study was to explore the role of serological markers in the diagnosis of CD at an inflammatory bowel disease (IBD) referral center.Methods: This was a retrospective study including 196 suspected CD patients. The expression of ASCA-IgG, ASCA-IgA, AYMA-IgG, AYCA-IgA, FI2Y-IgG, and pANCA in the patient's serum was determined by enzyme-linked immunosorbent assay (ELISA) and indirect immunofluorescence (IF).Results: ASCA was a relatively specific marker for CD (p = 0.0005), but not AYMA-IgG, AYCA-IgA, F12Y-IgG (p = 0.5936, 0.7974, 0.1085, respectively). However, a high sensitivity of 96.77% (95% CI 90.19%-99.83%) was noted for ASCA+/FI2Y+ to identify CD patients among the suspected cases, albeit with low PPV. The more combinations of serological markers, the higher sensitivity, and NPV. No correlation was found between the age of onset or disease location and the expression of ASCA, AYMA, AYCA, FI2Y, or pANCA. There was no significant difference between the expression of ASCA and the disease behavior at diagnosis (p = 0.3307). However, a decreased proportion of AYMA+ CD patients was found in those who received surgery compared with their non-surgical counterparts (p = 0.0488).Conclusions: ASCA was found to be the most accurate serological marker for the differential diagnosis of CD. Combinations of ASCA, AYMA, AYCA, and FI2Y improved diagnostic accuracy of CD.

Saccharomyces cerevisiae as a skin physiology, pathology, and treatment model.

Saccharomyces cerevisiae serves as a useful model in experimental biology. Within dermatology research, several studies have examined this organism's role in skin physiology, pathology, and treatment. Saccharomyces cerevisiae has been used to explore the mechanisms of melanogenesis as its extract inhibits key enzymes involved in melanogenesis and melanosome transfer. Additionally, the lack of probiotic intestinal Saccharomyces cerevisiae has been associated with psoriasis, potentially related to the anti-inflammatory effects of the yeast. Furthermore, antibodies against Saccharomyces cerevisiae have been observed in skin conditions, including atopic dermatitis. Saccharomyces cerevisiae may even cause skin infections, such as septic emboli in a patient with acute myelogenous leukemia. Lastly, Saccharomyces cerevisiae has potential use in vaccine development against melanoma and is utilized to study various treatment modalities such as zinc pyrithione, an ingredient often used in anti-dandruff shampoo.

A Case Report of Sequential Use of a Yeast-CEA Therapeutic Cancer Vaccine and Anti-PD-L1 Inhibitor in Metastatic Medullary Thyroid Cancer.

Medullary thyroid cancer (MTC) accounts for ~4% of all thyroid malignancies. MTC derives from the neural crest and secretes calcitonin (CTN) and carcinoembryonic antigen (CEA). Unlike differentiated thyroid cancer, MTC does not uptake iodine and I-131 RAI (radioactive iodine) treatment is ineffective. Patients with metastatic disease are candidates for FDA-approved agents with either vandetanib or cabozantinib; however, adverse effects limit their use. There are ongoing trials exploring the role of less toxic immunotherapies in patients with MTC. We present a 61-year-old male with the diagnosis of MTC and persistent local recurrence despite multiple surgeries. He was started on sunitinib, but ultimately its use was limited by toxicity. He then presented to the National Cancer Institute (NCI) and was enrolled on a clinical trial with heat-killed yeast-CEA vaccine (NCT01856920) and his calcitonin doubling time improved in 3 months. He then came off vaccine for elective surgery. After surgery, his calcitonin was rising and he enrolled on a phase I trial of avelumab, a programmed death-ligand 1 (PD-L1) inhibitor (NCT01772004). Thereafter, his calcitonin decreased > 40% on 5 consecutive evaluations. His tumor was subsequently found to express PD-L1. CEA-specific T cells were increased following vaccination, and a number of potential immune-enhancing changes were noted in the peripheral immunome over the course of sequential immunotherapy treatment. Although calcitonin declines do not always directly correlate with clinical responses, this response is noteworthy and highlights the potential for immunotherapy or sequential immunotherapy in metastatic or unresectable MTC.

Pneumocystis carinii Major Surface Glycoprotein Dampens Macrophage Inflammatory Responses to Fungal ß-Glucan.

BACKGROUND: Pneumocystis major surface glycoprotein (Msg) is a 120-kD surface protein complex on the organism with importance in adhesion and immune recognition. In this study, we show that Msg significantly impairs tumor necrosis factor (TNF)-α secretion by macrophages induced by Saccharomyces cerevisiae and Pneumocystis carinii (Pc) ß-glucans. METHODS: Major surface glycoprotein was shown to greatly reduce ß-glucan-induced Dectin-1 immunoreceptor tyrosine-based activating motif (ITAM) phosphorylation. Major surface glycoprotein also down regulated Dectin-1 receptor messenger ribonucleic acid (mRNA) expression in the macrophages. It is interesting that Msg incubation with macrophages resulted in significant mRNA upregulation of both C-type lectin receptors (CLR) Mincle and MCL in Msg protein presence alone but to even greater amounts in the presence of Pc ß-glucan. RESULTS: The silencing of MCL and Mincle resulted in TNF-α secretions similar to that of macrophages treated with Pneumocystis ß-glucan alone, which is suggestive of an inhibitory role for these 2 CLRs in Msg-suppressive effects on host cell immune response. CONCLUSIONS: Taken together, these data indicate that the Pneumocystis Msg surface protein complex can act to suppress host macrophage inflammatory responses to the proinflammatory ß -glucan components of the organisms.

Cochlear supporting cells function as macrophage-like cells and protect audiosensory receptor hair cells from pathogens.

To protect the audiosensory organ from tissue damage from the immune system, the inner ear is separated from the circulating immune system by the blood-labyrinth barrier, which was previously considered an immune-privileged site. Recent studies have shown that macrophages are distributed in the cochlea, especially in the spiral ligament, spiral ganglion, and stria vascularis; however, the direct pathogen defence mechanism used by audiosensory receptor hair cells (HCs) has remained obscure. Here, we show that HCs are protected from pathogens by surrounding accessory supporting cells (SCs) and greater epithelial ridge (GER or Kölliker's organ) cells (GERCs). In isolated murine cochlear sensory epithelium, we established Theiler's murine encephalomyelitis virus, which infected the SCs and GERCs, but very few HCs. The virus-infected SCs produced interferon (IFN)-α/ß, and the viruses efficiently infected the HCs in the IFN-α/ß receptor-null sensory epithelium. Interestingly, the virus-infected SCs and GERCs expressed macrophage marker proteins and were eliminated from the cell layer by cell detachment. Moreover, lipopolysaccharide induced phagocytosis of the SCs without cell detachment, and the SCs phagocytosed the bacteria. These results reveal that SCs function as macrophage-like cells, protect adjacent HCs from pathogens, and provide a novel anti-infection inner ear immune system.

Serological antibodies and surgery in a population-based inception cohort of Crohn's disease patients - the IBSEN study.

Introduction: Serological antibodies have been associated with complicated disease course in Crohn's disease (CD), including the need for surgery.Aim: The aim of this study was to investigate if a panel of relevant antibodies could predict surgery in a prospective population-based cohort of patients with CD.Methods: The population-based IBSEN cohort has been followed prospectively for 20 years. At the 10- and 20-year follow-up, the following panel of serological antibodies was analysed: pANCA, ASCA IgA, ASCA IgG, anti-OmpC, anti-I2, and anti-CBir1. At the 20-year follow-up or until lost to follow-up, all CD-related surgeries were registered.Results: Serum was available from 159 patients at 10-year follow-up and 135 patients at 20-year follow-up. In 113 patients, serum was available at both time points. No significant change of antibody status (positive vs. negative) was found from 10-year to 20-year follow-up. Negative pANCA, positive ASCA IgA and positive ASCA IgG at 10-year follow-up were all individually associated with increased risk for CD-related surgery. There was no association between anti-OmpC, anti-I2 or anti-CBir1 and CD-related surgery. In a multiple regression model including disease location and behaviour, only stricturing or penetrating disease behaviour and negative pANCA remained significantly associated with higher odds for surgery.Conclusion: Positive ASCA IgA and IgG, and negative pANCA were associated with higher odds for CD-related surgery in univariate analysis. Since disease phenotype changes during the disease course, while serological antibodies are stable, our results support the use of pANCA, ASCA IgA and ASCA IgG as prognostic markers in CD.

Serum Biomarkers Identify Patients Who Will Develop Inflammatory Bowel Diseases Up to 5 Years Before Diagnosis.

BACKGROUND & AIMS: Biomarkers are needed to identify patients at risk for development of inflammatory bowel diseases. We aimed to identify serum biomarkers of Crohn's disease and ulcerative colitis that can be detected and quantified before diagnosis. METHODS: We obtained serum samples from patients archived before a diagnosis of Crohn's disease (n = 200) or ulcerative colitis (n = 199), as well as from 200 healthy individuals (controls), collected from 1998 through 2013 as part of the US Defense Medical Surveillance System. We measured levels of antibodies against microbes (anti-Saccharomyces cerevisiae IgA or IgG, anti-Escherichiacoli outer membrane porin C, anti-CBir1, anti-flagellin 2, anti-flagellin X, and perinuclear anti-neutrophil cytoplasmic antibodies) and 1129 proteins in each sample. We then used functional principal component analysis to derive the time-varying trajectory for each marker, which then was used in a multivariate model to predict disease status. Predictive performances at different prediagnosis timepoints were evaluated using area under the receiver operating characteristic curves (AUROCs). Biological pathways that were up-regulated in serum from patients with Crohn's disease were identified based on changes in protein abundance at different time periods preceding diagnosis. RESULTS: We identified a panel of 51 protein biomarkers that were predictive of Crohn's disease within 5 years with an AUROC of 0.76 and a diagnosis within 1 year with an AUROC of 0.87. Based on the proteins included in the panel, imminent development of CD was associated with changes in the complement cascade, lysosomes, innate immune response, and glycosaminoglycan metabolism. Serum antibodies and proteins identified patients who received a diagnosis of ulcerative colitis within 5 years with an AUROC of only 0.56 and within 1 year with an AUROC of 0.72. CONCLUSIONS: We identified a panel of serum antibodies and proteins that were predictive of patients who will receive a diagnosis of Crohn's disease within 5 years with high accuracy. By contrast we did not identify biomarkers associated with future diagnosis of ulcerative colitis.

Treatment of hidradenitis suppurativa: Surgery and yeast (Saccharomyces cerevisiae)-exclusion diet. Results after 6 years.

BACKGROUND: Hidradenitis suppurativa is a complex disorder, the pathogenesis of which is still unsolved. The known association between hidradenitis suppurativa and Crohn's disease, an autoimmune disease diagnosed with the presence of Anti-Saccharomyces cerevisiae antibodies of the IgG family, suggests that a much more complex mechanism than a simple infectious disorder is involved. The goal of this study is to report patients' characteristics and the outcome of 6 years of a yeast (Saccharomyces cerevisiae)-exclusion diet and surgery in the treatment of hidradenitis suppurativa. METHOD: We analyzed 185 patients with hidradenitis suppurativa with a self-evaluative questionnaire. Thirty-seven patients were treated in our center following our protocol. The other 148 were members of a support group for patients with hidradenitis suppurativa treated by other centers. RESULTS: In 80% of patients who had the onset of hidradenitis suppurativa before the age of 30, the female to male ratio was 3.34:1, 74% were active smokers, and 5% also had Crohn's disease. In the diet group, 70% had an improvement of hidradenitis suppurativa symptomatology, 81% of whom in less than 6 months. Also, 87% of patients demonstrated an immediate recurrence of skin lesions less than a week after consuming a food containing the yeast. Immunologic testing showed intolerance to yeast, wheat, and cow's milk in 20%, 29%, and 23% of patients, respectively. CONCLUSION: The analysis confirmed the stabilization and regression of hidradenitis suppurativa with our diet, presumably by decreasing the local and systemic inflammation, leading to a less invasive operative treatment. These new findings seem to link hidradenitis suppurativa to food intolerance and gut dysbiosis.

Pancreatic anti-GP2 and anti-Saccharomyces cerevisiae antibodies in ruminants with paratuberculosis: A better understanding of the immunopathogenesis of Crohn's disease.

INTRODUCTION: Ruminants (cattle and sheep) with Mycobacterium avium (MAP)-induced paratuberculosis (ptb), the ruminant model of Crohn's disease (CD), exhibit pancreatic specific autoantibodies (PAB) against GP2 but not against CUZD1. Since anti-Saccharomyces cerevisiae antibodies (ASCAs) is a CD marker, we tested MAP-infected ptb ruminants for ASCA, and compared them with ruminants lacking evidence of anti-MAP serology or with ruminants, which were positive for anti-GP2 antibodies. MATERIAL AND METHODS: A total of 98 samples from ruminants (48 cattle and 50 sheep) were studied. IgG anti-MAP antibodies, and CD-related ASCA and anti-GP2 antibodies were tested by modified ELISAs. RESULTS: Nine cattle (18.75%) and 20 sheep (40%) were suffered from ptb. ASCA antibodies were present in 21/48 (43.7%) cattle and 10/50 (20%) sheep while anti-GP2 antibodies were present in 14/48 (29.2%) cattle, and 8/50 (16%) sheep. ASCA antibodies were more prevalent in anti-MAP antibody positive (14/29, 48.3%) than in anti-MAP negative ruminants (17/69, 24.6%, P=0.022) and also in anti-GP2 antibody positive (13/23, 56.5%) than in anti-GP2 negative ruminants (18/75, 24%, P=0.003). No association between ASCA and anti-MAP antibody concentrations were found (r=0.159, P=0.117). A significant association between ASCA and anti-GP2 antibody concentration were observed (r=0.211 and P=0.037). CONCLUSION: ASCA are present in a significant proportion of ruminants with ptb and correlate with anti-GP2 antibody positivity, a finding further supporting the notion that Crohn's disease and ptb share common immunological mechanisms of antigen-driven loss of self-tolerance.

Antibodies to Crohn's disease peptide 353 as a diagnostic marker for pediatric Crohn's disease: a prospective multicenter study in Japan.

BACKGROUND: Various serologic markers such as anti-glycoprotein 2 antibodies and anti-Saccharomyces cerevisiae antibodies have been reported to be diagnostically useful in Crohn's disease. Mitsuyama et al. reported that antibodies to Crohn's disease peptide 353, a newly proposed serologic marker, were more useful in Japanese adults than anti-Saccharomyces. We addressed the same issue in Japanese children and adolescents. METHODS: Prospectively enrolled subjects under 17 years old assessed and treated at 12 pediatric centers in Japan included groups with Crohn's disease, ulcerative colitis, other intestinal diseases, or good health. The 3 serum markers were analyzed by enzyme-linked immunosorbent assays. RESULTS: Enrolled subjects, numbering 367, included 120 with Crohn's disease, 148 with ulcerative colitis, 56 with other intestinal diseases, and 43 healthy subjects. In Crohn's disease, anti-Crohn's disease peptide 353, anti-glycoprotein 2, and anti-Saccharomyces concentrations (median, 2.25, 3.0, and 8.9 U/mL) were significantly greater than in ulcerative colitis (1.1, 1.9, and 3.4; all P < 0.001), other intestinal diseases (1.1, 1.85, and 2.95; all P < 0.001), and healthy controls (1.1, 1.7, and 2.8; all P < 0.001), respectively. At 95% specificity, sensitivity of anti-Crohn's disease peptide (45.0%) was significantly higher than for anti-glycoprotein 2 (30.8%; P < 0.05) or anti-Saccharomyces (26.7%; P < 0.01). CONCLUSIONS: Anti-Crohn's disease peptide 353 proved more useful for diagnosis of Crohn's disease in Japanese children than the other 2 markers. To our knowledge, this is the first pediatric report to that effect.

Rapid Generation of Chicken Immune Libraries for Yeast Surface Display.

Fluorescence-activated cell sorting (FACS) in combination with yeast surface display has emerged as a vital tool for the isolation and engineering of antibodies and antibody-derived fragments from synthetic, naïve, and immune libraries. However, the generation of antibodies against certain human antigens from immunized animals, e.g., mice, can remain challenging due to the homology to the murine counterpart. Due to the phylogenetic distance from humans, avian immunization can be a powerful technique for the generation of antibodies with high specificity against human antigens. Additionally, the peculiar Ig gene diversification in chickens enables the amplification of heavy and light chain genes utilizing single primer pairs, resulting in a convenient library generation. Herein, we describe the protocol for the construction of a single chain fragment variable (scFv) library derived from chickens after immunization with epidermal growth factor receptor (EGFR) for subsequent yeast surface display as well as the screening process utilizing FACS for the isolation of high-affinity antibodies.

Sneaking Out for Happy Hour: Yeast-Based Approaches to Explore and Modulate Immune Response and Immune Evasion.

Many pathogens (virus, bacteria, fungi, or parasites) have developed a wide variety of mechanisms to evade their host immune system. The budding yeast Saccharomyces cerevisiae has successfully been used to decipher some of these immune evasion strategies. This includes the cis-acting mechanism that limits the expression of the oncogenic Epstein-Barr virus (EBV)-encoded EBNA1 and thus of antigenic peptides derived from this essential but highly antigenic viral protein. Studies based on budding yeast have also revealed the molecular bases of epigenetic switching or recombination underlying the silencing of all except one members of extended families of genes that encode closely related and highly antigenic surface proteins. This mechanism is exploited by several parasites (that include pathogens such as Plasmodium, Trypanosoma, Candida, or Pneumocystis) to alternate their surface antigens, thereby evading the immune system. Yeast can itself be a pathogen, and pathogenic fungi such as Candida albicans, which is phylogenetically very close to S. cerevisiae, have developed stealthiness strategies that include changes in their cell wall composition, or epitope-masking, to control production or exposure of highly antigenic but essential polysaccharides in their cell wall. Finally, due to the high antigenicity of its cell wall, yeast has been opportunistically exploited to create adjuvants and vectors for vaccination.

Antimicrobial properties and phenoloxidase activation of the lectin isolated from kadal shrimp (Metapenaeus dobsoni).

The present study reveals purification and characterization of the lectin from the haemolymph of Metapenaeus dobsoni. The Md-Lec was purified by affinity chromatography with mannose coupled sepharose CL-4B column and it exhibits single band with a molecular weight of 68 kDa in SDS-PAGE. Furthermore, the molecular mass was confirmed by MALDI-TOF and functional groups present were analysed by FTIR. The surface morphology of purified Md-Lec displays the homogeneous nature of protein. The X-ray diffraction (XRD) analysis expresses three peaks at 10.7716̊, 21.6258̊ and 31.7523̊which indicate the crystalline nature of the protein and the retention time of 3.068 min evident from HPLC reveals the purity of the sample. Functional analysis of purified Md-Lec exhibits yeast agglutination activity against Saccharomyces cerevisiae and has the ability to agglutinate the human erythrocytes, which was observed by light microscopy. It also exhibited phenoloxidase activation, encapsulation and phagocytic activities. In addition, purified Md-Lec showed the broad spectrum of bacterial agglutination activity against Gram negative Vibrio parahaemolyticus and Aeromonas hydrophila, important fish pathogens. Antiviral potential and anticancer activity of purified Md-Lec against CyHV-2 virus and MDA-MB-231 breast cancer cell lines were also evaluated in this study.

Reappraisal of antibodies against Saccharomyces cerevisiae (ASCA) as persistent biomarkers in quiescent Crohn's disease.

A clear correlation exists between microbiota and the dysregulation of the immune response in Inflammatory Bowel Diseases (IBD), which comprise Crohn's disease (CD) and ulcerative colitis (UC). These unbalanced reactions also involve humoral responses, with antibodies against Saccharomyces cerevisiae. Thus, here we aimed to quantify IgA and IgG specific to S. cerevisiae (ASCA) in quiescent CD and UC, to correlate the production of these antibodies with patient's inflammatory response and disease clinical presentation. Twenty-nine subjects (16 CD and 13 UC) and 45 healthy controls were enrolled in this study and had plasma samples tested for ASCA and cytokines (IL-2, IL-4, IL-6, IL-10, IFN-γ, TNF-α), besides clinical evaluation. IBD patients had increase IgA and IgG ASCA, especially those with colonic (L2) and fistulizing (B3) CD. Similarly, patients who dropped out the treatment had augmented ASCA, while IgG was reduced in those receiving sulfasalazine treatment. Furthermore, the quiescent CD patients had elevated IL-6 on plasma, especially in the absence of treatment, together with increased counter regulatory response of IL-10. There was a positive correlation between IgA and IgG on CD but not UC, as well as between IgA and TNF in total IBD patients. In addition, the levels of IgG x TNF, IgA x IL-10 and IgG x IL-10 were also correlated in CD, indicating that ASCA production may be influenced by the inflammatory response. Finally, we concluded that ASCA could be pointed as relevant biomarker of CD presentation and residual inflammation, even in clinical remission patients.

Anti-Saccharomyces cerevisiae Antibodies as a Prognostic Biomarker in Children With Crohn Disease.

OBJECTIVE: Although anti-Saccharomyces cerevisiae antibodies (ASCAs) could be a useful biomarker in differentiating Crohn disease (CD) from ulcerative colitis (UC), their role as prognostic markers in children with CD has been underinvestigated. This longitudinal prospective observational study aimed to assess the prognostic value of ASCA status among children with CD managed using biologics. METHODS: The study population comprised children with inflammatory bowel disease diagnosed with CD from 2012 to 2018. Cox regression model with adjustment for a priori covariates was used to examine the response to anti-tumor necrosis factor (TNF) biological therapy among ASCA-positive patients in comparison to ASCA-negative patients. RESULTS: There were 273 measurements available from the study cohort comprising children with CD, who were followed up for a median duration of 14 months (interquartile range 5-42). ASCA-positive patients had a higher risk for moderate to severe clinical disease (odds ratio 2.88; 95% confidence interval [CI] 1.2-7.55) and extensive endoscopic distribution (odds ratio 3.30; CI 1.12-9.74) at baseline in comparison to ASCA-negative patients, respectively. In comparison to ASCA immunoglobulin G (IgG)-negative patients, ASCA IgG-positive patients who were treated with biologics had a significantly lower relapse rate (adjusted hazard ratio 0.12; CI 0.02-0.93). Ten (14%) patients had an unstable ASCA value with either ASCA immunoglobulin A or ASCA IgG status changing from positive to negative or vice versa. CONCLUSIONS: ASCA-positive children with CD present with more extensive (endoscopic) and clinically severe disease. ASCA IgG is a useful prognostic marker among children with CD who receive biologics.

Yeast-based vaccines: New perspective in vaccine development and application.

In presently licensed vaccines, killed or attenuated organisms act as a source of immunogens except for peptide-based vaccines. These conventional vaccines required a mass culture of associated or related organisms and long incubation periods. Special requirements during storage and transportation further adds to the cost of vaccine preparations. Availability of complete genome sequence, well-established genetic, inherent natural adjuvant and non-pathogenic nature of yeast species viz. Saccharomyces cerevisiae, Pichia pastoris makes them an ideal model system for the development of vaccines both for public health and for on-farm consumption. In this review, we compile the work in this emerging field during last two decades with major emphases on S. cerevisiae and P. pastoris which are routinely used worldwide for expression of heterologous proteins with therapeutic value against infectious diseases along with possible use in cancer therapy. We also pointed towards the developments in use of whole recombinant yeast, yeast surface display and virus-like particles as a novel strategy in the fight against infectious diseases and cancer along with other aspects including suitability of yeast in vaccines preparations, yeast cell wall component as an immune stimulator or modulator and present status of yeast-based vaccines in clinical trials.

Altered uric acid metabolism in isolated colonic Crohn's disease but not ulcerative colitis.

BACKGROUND AND AIM: Patients with inflammatory bowel disease (IBD) have higher incidence of developing nephrolithiasis. Increased uric acid production induced by Saccharomyces cerevisiae exacerbates colitis in mice. We aimed to evaluate the association between serum uric acid level and disease activity in IBD population. METHODS: Four hundred and thirty-five patients enrolled in Jinling Hospital from January 1, 2015 to August 31, 2017 were included in the retrospective study. Clinical parameters were collected and compared with non-IBD matched controls (n = 51). Serum uric acid to creatinine ratio (UA/Cr) was used as a biomarker for uric acid metabolism. Sixty-five active IBD patients were longitudinally studied to investigate the UA/Cr before and after therapy. Linear mixed models were estimated for Crohn's disease (CD) group to explore the relationship between UA/Cr and other parameters. RESULTS: Uric acid to creatinine ratio was significantly correlated with Crohn's disease activity index (ρ = 0.184, P = 0.002) and Harvey Bradshaw index (ρ = 0.154, P = 0.010) and C-reactive protein (ρ = 0.591, P < 0.001) in CD group. Colonic CD and anti-Saccharomyces cerevisiae antibody (ASCA) positive CD had an increased UA/Cr compared with L1, L3, and ASCA negative CD (P = 0.027, P = 0.0013, and P = 0.043, respectively). A significant decrease in UA/Cr was observed after induction therapy in active CD (P = 0.0002) but not in ulcerative colitis (P = 0.076). CONCLUSION: Uric acid to creatinine ratio correlated with disease activity in CD. Colonic CD and ASCA positive CD had an increased UA/Cr. Effective treatment for CD patients lowered UA/Cr. Uric acid metabolism might be a novel aspect to investigate disease activity of IBD.