Wine fermentation process evaluation through NMR analysis: Polysaccharides, ethanol quantification and biological activity.

Winemaking production is old knowledge of the combination of saccharification and fermentation processes. During the fermentation process, ethanol concentration is one of the main key parameters that provides the quality of wine and is linked to the consumption of carbohydrates present in wine. In this work was determined the better fermentation time, where the wine retains its highest concentration of ethanol and a higher concentration of the polysaccharides of Bordo wine of Vitis labrusca by 1D and 2D NMR measurements. The study provides information on the polysaccharide content for improving features and quality control of winemaking. Moreover, following previous studies by our group (de Lacerda Bezerra et al., 2018, de Lacerda Bezerra, Caillot, de Oliveira, Santana-Filho, & Sassaki, 2019; Stipp et al., 2023) showed that the soluble polysaccharides also inhibited the production of inflammatory cytokines (TNF-α and IL-1ß) and mediator (NO) in macrophage cells stimulated with LPS, bringing some important health benefits of wine.

Recombinant human interferon-α14 for the treatment of canine allergic pruritic disease in eight dogs.

BACKGROUND: Allergic pruritic diseases are increasingly common in dogs. This group of conditions hampers life quality as pruritus progressively interferes with normal behaviours. Therefore, new treatment modalities for canine allergic pruritic diseases are necessary. While novel drugs have recently reached the market, there is still the need for other therapeutic approaches. Some dogs are refractory even to the newer compounds, and cost is also an important issue for these. Older therapeutic modalities are only moderately successful or have considerable secondary effects, as is the case with glucocorticoids. OBJECTIVES: Report on the use of recombinant human interferon-α14 (rhIFN-α14) for the treatment of canine allergic pruritus. Following the experience with a similar compound in the Japanese market, it was expected that rhIFN-α14 could alter the Th1/Th2 disbalance that drives these diseases. METHODS: Here, we present an uncontrolled trial in which eight dogs with clinical diagnosis of allergic pruritus were treated with rhIFN-α14, either orally or via subcutaneous injections. Skin condition, microbiota and anti-interferon antibody levels were assessed. RESULTS: The parenteral use of interferon induced hypersensitivity in two of the three dogs in which it was used. The oral administration was consistently safe and could reduce signs of the allergic condition in three of the five treated animals. Treatment also altered the skin microbiota, as verified by next-generation sequencing. CONCLUSION: The present results indicate that rhIFN-α14 is a viable candidate for the treatment of canine allergic pruritus. Future controlled studies are needed, and the oral route is indicated for further trials.

Relevance of antibodies against the Chicken Anaemia Virus.

Chicken Infectious Anaemia (CIA) Virus (CAV) inhibits the function of multiple immune compartments. Mortality due to clinical infection is controlled in broilers by passive immunization derived from vaccinated breeders. Therefore, serological tests are often used in chicks to determine maternally-derived antibodies (MDA). We used a vaccine overdose-induced model of CIA. The model replicated the most common features of the disease. This model was used to determine the role of MDA in the protection of chicks. Hatchlings were tested for anti-CAV titers by ELISA and were sorted into groups based on antibody levels. SPF chicks were used as a no-antibody control. Lower specific antibody levels seemed to facilitate viral entry into the thymus, but viral levels, CD4+ and CD8+ counts, thymus architecture, and haematocrit were preserved by MDA, regardless of its levels. Levels of MDA are not correlated with protection from CIA, but are important for the progression CAV infection.

Comparative genomics of a novel clade shed light on the evolution of the genus Erysipelothrix and characterise an emerging species.

Erysipelothrix sp. isolates obtained from a deadly outbreak in farmed turkeys were sequenced and compared to representatives of the genus. Phylogenetic trees-supported by digital DNA:DNA hybridization and Average Nucleotide Identity-revealed a novel monophyletic clade comprising isolates from pigs, turkeys, and fish, including isolates previously described as E. sp. Strain 2. Genes coding for the SpaC protein, typically found in E. sp. Strain 2, were detected in all isolates of the clade. Therefore, we confirm E. sp. Strain 2 represents a unique species that may be isolated from a broad host range, and the name "Erysipelothrix takahashiae" is suggested. Core genome analysis showed that the pathogenic species of this genus, E. rhusiopathiae and the clade E. sp. Strain 2, are enriched in core functionalities related to nutrient uptake and transport, but not necessarily homologous pathways. For instance, whereas the aerobic DctA transporter may uptake C4-dicarboxylates in both species, the anaerobic DcuC transporter is exclusive of the E. sp. Strain 2. Remarkably, the pan-genome analysis uncovered that genes related to transport and metabolism, recombination and repair, translation and transcription in the fish isolate, within the novel clade, have undergone a genomic reduction through pseudogenization. This reflects distinct selective pressures shaping the genome of species and strains within the genus Erysipelothrix while adapting to their respective niches.

Effects of aroA deleted E. coli vaccine on intestinal microbiota and mucosal immunity.

E. coli infection of broilers can result in systemic diseases and productivity losses. Use of antimicrobials against this condition is common but other approaches, such as vaccination, are gaining ground. Anecdotal field reports indicate that intestinal health is improved unspecifically following E. coli live vaccination. We hypothesized that the intestine may be an important site for the functionality of the vaccine. Vaccine effects on the intestine were assessed. Spray vaccination induced marked alterations of the caecum microbiota of broilers within 3 days, and this effect gradually waned. However, T cell activation occurred in the spleen, but not in caecal tonsils, and anti-E. coli IgA was concentrated in the respiratory mucosae. Accordingly, IL-6 mRNA was produced in the lungs following immunization. Overall, these data are an initial indication that any vaccine-induced effects on the intestine are greatly associated with the microbiota. However, immunity conferred by vaccination is not primarily induced in gut-associated lymphoid tissues.

Fish oil supplementation enhances colon recovery after experimental colitis.

INTRODUCTION: Fish oil (FO) has an anti-inflammatory and pro-resolution activity and it has been used to restore physiological disturbances on inflammatory conditions. Here, we investigate whether FO supplementation could, acutely, prevent or restore inflammatory damages on experimental colitis. METHODS: Wistar rats orally received 2 g.kg-1.day-1 of FO for 30 days before induction of experimental colitis. Specimens were collected on the 2nd and 7th days after colitis-induction and intestinal mucus, inflammatory activity and colon integrity were determined. RESULTS: Experimental colitis did cause colon disruption and FO, acutely, did not prevent the loss of intestinal and fecal mucus, neither the increase of inflammatory activity and intestinal permeability. On the 7th day of colitis, FO soften the perturbations of experimental colitis, increasing histological and fecal mucus and, also decreased inflammatory activity, but this was not accompanied by intestinal permeability. CONCLUSION: FO did not protect, acutely, intestinal damages from experimental colitis, but at long run promotes higher intestinal recovery.

A blocking antibody against canine CSF-1R maturated by limited CDR mutagenesis.

CSF-1R is a receptor mostly associated with the mononuclear phagocytic system. However, its expression within tumors has been linked with poor prognosis in both humans and dogs. Accordingly, several reports have demonstrated the beneficial effects of blocking CSF-1R in model systems of cancer. In this study, we generated a monoclonal antibody that could block CSF-1R in dogs as the first step to develop an anticancer drug for this species. Initially, an antibody was raised by the hybridoma methodology against the fragment responsible for receptor dimerization. mAb3.1, one of the resulting hybridoma clones, was able to bind macrophages in fixed tissues and was shown to inhibit cells of the mononuclear phagocytic line. Nevertheless, mAb 3.1 could not bind to some glycoforms of the receptor in its native form, while also demonstrating cross-reactivity with other proteins. To enhance binding properties of the mAb, five amino acids of the complementarity-determining region 2 of the variable heavy chain of mAb3.1 were mutated by PCR, and the variant scFv clones were screened by phage display. The selected scFv clones demonstrated improved binding to the native receptor as well as increased anti-macrophage activity. The resulting scFv antibody fragment presented here has the potential for use in cancer patients and in inflammatory diseases. Furthermore, this work provides insights into the use of such restricted mutations in antibody engineering.

Monoclonal Antibody DL11C8 Identifies ADAM23 as a Component of Lipid Raft Microdomains.

A disintegrin and metalloprotease protein 23 (ADAM23) is a transmembrane type I glycoprotein involved with the development and maintenance of the nervous system, including neurite outgrowth, neuronal adhesion and differentiation and regulation of synaptic transmission. In addition, ADAM23 seems to participate in immune response and tumor establishment through interaction with different members of integrin receptors. Here, we describe a novel monoclonal antibody (DL11C8) that specifically recognizes the cysteine-rich domain of both pre-protein (100 kDa) and mature (70 kDa) forms of ADAM23 from different species, including human, rodents and avian orthologs. Using this antibody, we detected both forms of ADAM23 on the cell surface of three neuronal cell lineages (Neuro-2a, SH-SY5Y and CHLA-20), with a higher relative content of ADAM23100 kDa. Furthermore, we demonstrate for the first time that a catalytically inactive member of the ADAM family is present in the membrane signaling platforms, namely lipid rafts. Indeed, the mature ADAM2370 kDa partitions between raft and non-raft membrane domains, while the pro-protein ADAM23100 kDa is mainly expressed in non-raft domains. These membranous distributions were observed in both different brain regions homogenates and primary cultured neurons lysates from mouse cortex and cerebellum. Taken together, these findings point out ADAM23 as a lipid raft molecular component.

Effect of an Enterococcus faecium probiotic on specific IgA following live Salmonella Enteritidis vaccination of layer chickens.

Probiotics and immunization are being widely adopted by the poultry industry with the goal of controlling Salmonella enterica. However, the interaction between these two management protocols has been sparsely studied. The present study aimed to understand the role of an Enterococcus faecium probiotic in the production of salmonella-specific IgA in layers immunized with a live vaccine. Four groups were used: "Control" (no vaccine or probiotic); "Probiotic" (which received an E. faecium product); "Vaccine" (immunized with two doses of a live attenuated S. Enteritidis vaccine); and "Vaccine + probiotic". Faecal salmonella-specific IgA was analysed 7 and 20 days post-vaccination (dpv) boost. At 7 dpv, the "Vaccine" and "Vaccine + probiotic" groups had similar IgA levels. However, at 20 dpv, IgA levels were two times higher in the "Vaccine + probiotic" group compared to the "Vaccine" group. To understand the role of the intestinal microbiota in this finding, bacterial diversity in faeces was analysed by 16S rRNA gene sequencing. The improvement in IgA production in probiotic-treated birds was accompanied by marked changes in the faecal microbiome. Some of the main differences between the "Vaccine" and "Vaccine + probiotic" groups included reduction of Escherichia-Shigella and increases in Blautia, Anaerotruncus and Lactobacillus in the latter group. Although no direct causal link can be established from this study design, it is possible that the E. faecium probiotic induces improved antibody production following vaccination via modulation of the intestinal microbiota.

Effect of spray Escherichia coli vaccine on the immunity of poultry.

Avian pathogenic Escherichia coli is a current problem in the poultry industry, causing mortality and economic losses. This paper evaluates the dynamics in immune response after the use of spray vaccination against E. coli and, thereby, seeks to understand how the vaccine can provide protection. During the early stages of response to vaccination the presence of antigen-presenting cells is predominant, but these diminish within the first 7 days after vaccination. The immune correlate of protection of vaccination using the E. coli vaccine Poulvac E. coli (aroA-deficient mutant strain) probably does not depend on the production of circulating antibodies (as assessed through the presence of B lymphocytes) and is linked to the presence of CD4+TCRVbeta1+. These cells act on mucosa tissue stimulating the production of immunoglobulin A. Vaccination stimulated a high state of immunocompetence, as assessed by measurement of several cellular subsets. This state of "immune alertness," however, may be associated with reduced weight gain. The high presence of naive and memory CD8 cells in the vaccinated group at 14 and 21 days postvaccination may indicate greater ability in the future to prevent tissue invasion by E. coli, based on the possibility that these cells will proliferate rapidly to a new stimulus. The simultaneous use of vaccine with the antibiotic ceftiofur sodium interferes with the immune response obtained through vaccination. In combination, the data obtained in this study indicate that the immune response produced by a spray vaccine against E. coli is mainly a cellular response, especially relevant to the sites in contact with the pathogen. It is suggested that there is a strong cell migration to the mucous membranes, where macrophages act first and then lymphocytes take part to protect the host. It is believed that recruited lymphocytes will act in the production of secreted IgA, which probably plays a greater role in the defense when compared with circulating immunoglobulins. The assessment of cellular dynamics by flow cytometry made it possible to elucidate the operation mechanism of the live E. coli vaccine.

Plasma cysteine/cystine reduction potential correlates with plasma creatinine levels in chronic kidney disease.

BACKGROUND/AIMS: Oxidative stress has been considered a nontraditional risk factor for cardiovascular disease in the chronic kidney disease (CKD) population, possibly triggered by uremic toxicity. METHODS: A chromatographic method with coulometric detection was adapted to directly and simultaneously determine cysteine (Cys) and cystine (Cyss) in plasma samples. Healthy subjects and CKD subjects in different stages were analyzed. The free Cys and free Cyss levels in their plasma were determined, and the reduction potential [Eh(Cyss/2Cys)] was calculated with the Nernst equation. RESULTS: Healthy plasma presented Eh(Cyss/2Cys) of -123 ± 7 mV. Plasma Eh(Cyss/2Cys) correlated significantly with creatinine levels (p < 0.0001, r = 0.62). CONCLUSION: Plasma Eh(Cyss/2Cys) correlated with increased levels of plasma creatinine, supporting the view that uremia triggers oxidative stress. In addition, it may be used as a quantitative oxidative stress biomarker in uremic conditions.

Assessing the use of 35 percent sodium ascorbate for removal of residual hydrogen peroxide after in-office tooth bleaching.

BACKGROUND: The authors conducted a study to quantify the amount of hydrogen peroxide remaining after in-office bleaching and to investigate the effectiveness of 35 percent sodium ascorbate (SA) on removal of the bleaching agent from dentin. METHODS: The authors sectioned 70 extracted human third molars to form 4×4×2-millimeter blocks of dentin and then categorized them into eight groups as follows: group 1, no bleaching; group 2, bleaching (35 percent hydrogen peroxide) and daily measurement; group 3, bleaching plus one application of 35 percent SA for 60 minutes; group 4, bleaching plus one application of SA for 10 minutes; group 5, bleaching plus two applications of SA for 10 minutes each; group 6, bleaching plus two applications of SA for five minutes each; group 7, bleaching plus three applications of SA for one minute each; and group 8, bleaching plus two applications of SA for one minute each. They quantified the amount of residual hydrogen peroxide by using a colorimetric assay with horseradish peroxidase and o-phenylenediamine dihydrochloride as the enzyme and substrate, respectively. RESULTS: The authors analyzed the data by using the Kruskal-Wallis test (α = .05). The results showed that no hydrogen peroxide remained after 120 hours in group 2 and at their respective measurement points in groups 5, 6, 7 and 8, whereas hydrogen peroxide had been removed only partially in groups 3 and 4. CONCLUSION: Two applications of 35 percent SA for one minute each produced the same result--complete removal of the bleaching agent--as that observed five days after tooth bleaching with 35 percent hydrogen peroxide without any application of SA. CLINICAL IMPLICATIONS: In an appropriate concentration, SA facilitates rapid elimination of residual peroxide up to two minutes after bleaching with 35 percent hydrogen peroxide. This finding suggests that clinicians can perform bonding procedures shortly after the bleaching procedure.

Assessment of MMP-9, TIMP-1, and COX-2 in normal tissue and in advanced symptomatic and asymptomatic carotid plaques.

BACKGROUND: Mature carotid plaques are complex structures, and their histological classification is challenging. The carotid plaques of asymptomatic and symptomatic patients could exhibit identical histological components. OBJECTIVES: To investigate whether matrix metalloproteinase 9 (MMP-9), tissue inhibitor of MMP (TIMP), and cyclooxygenase-2 (COX-2) have different expression levels in advanced symptomatic carotid plaques, asymptomatic carotid plaques, and normal tissue. METHODS: Thirty patients admitted for carotid endarterectomy were selected. Each patient was assigned preoperatively to one of two groups: group I consisted of symptomatic patients (n = 16, 12 males, mean age 66.7 ± 6.8 years), and group II consisted of asymptomatic patients (n = 14, 8 males, mean age 67.6 ± 6.81 years). Nine normal carotid arteries were used as control. Tissue specimens were analyzed for fibromuscular, lipid and calcium contents. The expressions of MMP-9, TIMP-1 and COX-2 in each plaque were quantified. RESULTS: Fifty-eight percent of all carotid plaques were classified as Type VI according to the American Heart Association Committee on Vascular Lesions. The control carotid arteries all were classified as Type III. The median percentage of fibromuscular tissue was significantly greater in group II compared to group I (p < 0.05). The median percentage of lipid tissue had a tendency to be greater in group I than in group II (p = 0.057). The percentages of calcification were similar among the two groups. MMP-9 protein expression levels were significantly higher in group II and in the control group when compared with group I (p < 0.001). TIMP-1 expression levels were significantly higher in the control group and in group II when compared to group I, with statistical difference between control group and group I (p = 0.010). COX-2 expression levels did not differ among groups. There was no statistical correlation between MMP-9, COX-2, and TIMP-1 levels and fibrous tissue. CONCLUSIONS: MMP-9 and TIMP-1 are present in all stages of atherosclerotic plaque progression, from normal tissue to advanced lesions. When sections of a plaque are analyzed without preselection, MMP-9 concentration is higher in normal tissues and asymptomatic surgical specimens than in symptomatic specimens, and TIMP-1 concentration is higher in normal tissue than in symptomatic specimens.

Transplantation of SNAP-treated adipose tissue-derived stem cells improves cardiac function and induces neovascularization after myocardium infarct in rats.

Stem cell therapy has been considered a promise for damaged myocardial tissue. We have previously shown that S-nitroso-N-acetyl-D,L-penicillamine (SNAP) increases the expression of several muscular markers and VEGF in mesenchymal stem cells, indicating that transplantation of SNAP-treated cells could provide better functional outcomes. Here, we transplanted SNAP-treated adipose tissue-derived stem cells (ADSCs) in rat infarcted myocardium. After 30days, we observed a significant improvement of the ejection fraction in rats that received SNAP-treated ADSCs, compared with those that received untreated cells (p=0.008). Immunohistochemical reactions showed an increased expression of troponin T-C and von Willebrand factor, and organized vascular units in the infarcted area of tissue transplanted with treated ADSCs. SNAP exposure induced intracellular S-nitrosation, a decreased GSH/GSSG ratio, but did not increase cGMP levels. Collectively, these results indicate that SNAP alters the redox environment of ADSCs, possibly associated with a pre-differentiation state, which may improve cardiac function after transplantation.