Therapeutic role of interferon-γ in experimental autoimmune encephalomyelitis is mediated through a tolerogenic subset of splenic CD11b+ myeloid cells.

Cumulative evidence has established that Interferon (IFN)-γ has both pathogenic and protective roles in Multiple Sclerosis and the animal model, Experimental Autoimmune Encephalomyelitis (EAE). However, the underlying mechanisms to the beneficial effects of IFN-γ are not well understood. In this study, we found that IFN-γ exerts therapeutic effects on chronic, relapsing-remitting, and chronic progressive EAE models. The frequency of regulatory T (Treg) cells in spinal cords from chronic EAE mice treated with IFN-γ was significantly increased with no effect on Th1 and Th17 cells. Consistently, depletion of FOXP3-expressing cells blocked the protective effects of IFN-γ, indicating that the therapeutic effect of IFN-γ depends on the presence of Treg cells. However, IFN-γ did not trigger direct in vitro differentiation of Treg cells. In vivo administration of blocking antibodies against either interleukin (IL)-10, transforming growth factor (TGF)-ß or program death (PD)-1, revealed that the protective effects of IFN-γ in EAE were also dependent on TGF-ß and PD-1, but not on IL-10, suggesting that IFN-γ might have an indirect role on Treg cells acting through antigen-presenting cells. Indeed, IFN-γ treatment increased the frequency of a subset of splenic CD11b+ myeloid cells expressing TGF-ß-Latency Associated Peptide (LAP) and program death ligand 1 (PD-L1) in a signal transducer and activator of transcription (STAT)-1-dependent manner. Furthermore, splenic CD11b+ cells from EAE mice preconditioned in vitro with IFN-γ and myelin oligodendrocyte glycoprotein (MOG) peptide exhibited a tolerogenic phenotype with the capability to induce conversion of naïve CD4+ T cells mediated by secretion of TGF-ß. Remarkably, adoptive transfer of splenic CD11b+ cells from IFN-γ-treated EAE mice into untreated recipient mice ameliorated clinical symptoms of EAE and limited central nervous system infiltration of mononuclear cells and effector helper T cells. These results reveal a novel cellular and molecular mechanism whereby IFN-γ promotes beneficial effects in EAE by endowing splenic CD11b+ myeloid cells with tolerogenic and therapeutic activities.

Single nucleotide polymorphisms associated with wine fermentation and adaptation to nitrogen limitation in wild and domesticated yeast strains.

For more than 20 years, Saccharomyces cerevisiae has served as a model organism for genetic studies and molecular biology, as well as a platform for biotechnology (e.g., wine production). One of the important ecological niches of this yeast that has been extensively studied is wine fermentation, a complex microbiological process in which S. cerevisiae faces various stresses such as limited availability of nitrogen. Nitrogen deficiencies in grape juice impair fermentation rate and yeast biomass production, leading to sluggish or stuck fermentations, resulting in considerable economic losses for the wine industry. In the present work, we took advantage of the "1002 Yeast Genomes Project" population, the most complete catalogue of the genetic variation in the species and a powerful resource for genotype-phenotype correlations, to study the adaptation to nitrogen limitation in wild and domesticated yeast strains in the context of wine fermentation. We found that wild and domesticated yeast strains have different adaptations to nitrogen limitation, corroborating their different evolutionary trajectories. Using a combination of state-of-the-art bioinformatic (GWAS) and molecular biology (CRISPR-Cas9) methodologies, we validated that PNP1, RRT5 and PDR12 are implicated in wine fermentation, where RRT5 and PDR12 are also involved in yeast adaptation to nitrogen limitation. In addition, we validated SNPs in these genes leading to differences in fermentative capacities and adaptation to nitrogen limitation. Altogether, the mapped genetic variants have potential applications for the genetic improvement of industrial yeast strains.

In-office bleaching with a commercial 40% hydrogen peroxide gel modified to have different pHs: Color change, surface morphology, and penetration of hydrogen peroxide into the pulp chamber.

OBJECTIVE: In-office bleaching gels are usually marketed in different pHs. This study is aimed at evaluating the efficacy, enamel surface morphology and concentration of hydrogen peroxide (HP) in the pulp chamber of teeth bleached with 40% HP with different pHs. MATERIALS AND METHODS: Forty premolars were randomly divided according to bleaching gel pH: 5.1, 6.3, 7.0, and control (no bleaching). Teeth were prepared, an acetate buffer was placed in the pulp chamber and teeth were bleached with two 20-minutes applications. The amount of HP was determined on a UV-VIS spectrophotometer. Color change was assessed by using a digital spectrophotometer before and 1 week after bleaching treatment. Five additional premolars were divided into four parts, assigned to the same groups above for analysis under scanning electron microscope. Data were subjected to anova and Tukey's tests (alpha = 0.05). RESULTS: The group pH 5.1 showed the highest HP diffusion in the pulp chamber (P < .001). No significant difference was detected in color change (P = .51). All groups presented the same pattern of enamel demineralization. CONCLUSIONS: The bleaching agent with pH 5.1 presented the highest HP amounts in the pulp chamber, but color change and enamel morphology were similar among groups. CLINICAL SIGNIFICANCE: Regardless of the pH, the bleaching effect can be observed in teeth submitted to high concentrations of HP, but a higher permeability of HP was found in the pulp chamber of teeth bleached with more acidic bleaching agents. Based on that, we suggest the use of alkaline gels for in-office bleaching to minimize damage to the pulpal tissue.

Interferon-gamma ameliorates experimental autoimmune encephalomyelitis by inducing homeostatic adaptation of microglia.

Compelling evidence has shown that interferon (IFN)-γ has dual effects in multiple sclerosis and in its animal model of experimental autoimmune encephalomyelitis (EAE), with results supporting both a pathogenic and beneficial function. However, the mechanisms whereby IFN-γ may promote neuroprotection in EAE and its effects on central nervous system (CNS)-resident cells have remained an enigma for more than 30 years. In this study, the impact of IFN-γ at the peak of EAE, its effects on CNS infiltrating myeloid cells (MC) and microglia (MG), and the underlying cellular and molecular mechanisms were investigated. IFN-γ administration resulted in disease amelioration and attenuation of neuroinflammation associated with significantly lower frequencies of CNS CD11b+ myeloid cells and less infiltration of inflammatory cells and demyelination. A significant reduction in activated MG and enhanced resting MG was determined by flow cytometry and immunohistrochemistry. Primary MC/MG cultures obtained from the spinal cord of IFN-γ-treated EAE mice that were ex vivo re-stimulated with a low dose (1 ng/ml) of IFN-γ and neuroantigen, promoted a significantly higher induction of CD4+ regulatory T (Treg) cells associated with increased transforming growth factor (TGF)-ß secretion. Additionally, IFN-γ-treated primary MC/MG cultures produced significantly lower nitrite in response to LPS challenge than control MC/MG. IFN-γ-treated EAE mice had a significantly higher frequency of CX3CR1high MC/MG and expressed lower levels of program death ligand 1 (PD-L1) than PBS-treated mice. Most CX3CR1highPD-L1lowCD11b+Ly6G- cells expressed MG markers (Tmem119, Sall2, and P2ry12), indicating that they represented an enriched MG subset (CX3CR1highPD-L1low MG). Amelioration of clinical symptoms and induction of CX3CR1highPD-L1low MG by IFN-γ were dependent on STAT-1. RNA-seq analyses revealed that in vivo treatment with IFN-γ promoted the induction of homeostatic CX3CR1highPD-L1low MG, upregulating the expression of genes associated with tolerogenic and anti-inflammatory roles and down-regulating pro-inflammatory genes. These analyses highlight the master role that IFN-γ plays in regulating microglial activity and provide new insights into the cellular and molecular mechanisms involved in the therapeutic activity of IFN-γ in EAE.

Intranasal delivery of interferon-ß-loaded nanoparticles induces control of neuroinflammation in a preclinical model of multiple sclerosis: A promising simple, effective, non-invasive, and low-cost therapy.

Multiple sclerosis (MS) is an autoimmune disease affecting the central nervous system (CNS). Interferon (IFN)-ß constitutes one of the first-line therapies to treat MS, but has limited efficacy due to the injectable systemic administration, short half-life, and limited CNS access. To address these limitations, we developed IFN-ß-loaded chitosan/sulfobutylether-ß-cyclodextrin nanoparticles (IFN-ß-NPs) for delivery of IFN-ß into the CNS via the intranasal (i.n.) route. The nanoparticles (NPs) (≈200 nm, polydispersity ≈0.1, and zeta potential ≈20 mV) were prepared by mixing two aqueous solutions and associated human or murine IFN-ß with high efficiency (90%). Functional in vitro assays showed that IFN-ß-NPs were safe and that IFN-ß was steadily released while retaining biological activity. Biodistribution analysis showed an early and high fluorescence in the brain after nasal administration of fluorescent probe-loaded NPs. Remarkably, mice developing experimental autoimmune encephalomyelitis (EAE), an experimental model of MS, exhibited a significant improvement of clinical symptoms in response to intranasal IFN-ß-NPs (inIFN-ß-NPs), whereas a similar dose of intranasal or systemic free IFN-ß had no effect. Importantly, inIFN-ß-NPs treatment was equally effective despite a reduction of 78% in the total amount of weekly administered IFN-ß. Spinal cords obtained from inIFN-ß-NPs-treated EAE mice showed fewer inflammatory foci and demyelination, lower expression of antigen-presenting and costimulatory proteins on CD11b+ cells, and lower astrocyte and microglia activation than control mice. Therefore, IFN-ß treatment at tested doses was effective in promoting clinical recovery and control of neuroinflammation in EAE only when associated with NPs. Overall, inIFN-ß-NPs represent a potential, effective, non-invasive, and low-cost therapy for MS.

Increases in norepinephrine release and ovarian cyst formation during ageing in the rat.

BACKGROUND: Depletion of ovarian follicles is associated with the end of reproductive function in ageing females. Recently, it has been described that this process parallels increases in the concentration of norepinephrine (NE) in the rat ovary. In sexually mature rats, experimentally-induced increases in the sympathetic tone of the ovary is causally related to ovarian cyst formation and deranged follicular development. Thus, there is a possibility that increased ovarian NE concentrations represent changes in the activity of sympathetic nerves, which consequently participate in the process of ovarian cyst formation observed during ageing in the human and experimental animal models. METHODS: Sprague-Dawley rats between 6 and 14 months old were used to analyse the capacity of the ovary to release 3H-NE recently incorporated under transmural depolarisation in relation to changes in the ovarian follicular population. Morphometric analysis of ovarian follicles and real time PCR for Bcl2 and Bax mRNA were used to assess follicular atresia. RESULTS: From 8 months old, the induced release of recently incorporated 3H-norepinephrine (3H-NE) from the ovary and ovarian NE concentrations increased, reaching their peak values at 12 months old and remained elevated up to 14 months old. Increases in sympathetic nerve activity paralleled changes in the follicular population, as well as disappearance of the corpus luteum. In contrast, luteinised follicles, precystic follicles, and cystic follicles increased. During this period, the relationship between Bax and Bcl2 mRNAs (the proapoptotic/antiapoptotic signals) increased, suggesting atresia as the principal mechanism contributing to the decreased follicular population. When NE tone was increased, the mRNA ratio favoured Bcl2 to Bax and antiapoptotic signals dominated this period of development. Thus, these changing ratios could be responsible for the increase in luteinised follicles, as well as precystic and cystic follicles. CONCLUSION: These data suggest that the ageing process in the ovary of the Sprague-Dawley rat is accompanied by an increased sympathetic tone of the ovary. Consequently, this sympathetic change could be related to a neuroendocrine-driven formation of a polycystic condition similar to that observed in the sympathetic-activated adult ovary.

Does the Use of Reservoirs Have Any Impact on the Efficacy of At-Home Bleaching? A Systematic Review.

To answer the following focused question through a systematic review: "Are the risk and intensity of tooth sensitivity (TS) and bleaching efficacy different between adult patients who undergo at-home bleaching using trays with reservoirs and those who use trays without reservoirs?". A comprehensive search was performed in the MEDLINE via PubMed, Scopus, Web of Science, Latin American and Caribbean Health Sciences Literature database, Brazilian Library in Dentistry, Cochrane Library, and grey literature without restrictions. Abstracts from conferences; unpublished and ongoing trial registries, dissertations and theses (ProQuest Dissertations and Periódicos Capes Theses databases) were searched. Only randomized clinical trials (RCTs) were included. We used the Risk of Bias tool (RoB) from the Cochrane Collaboration for quality assessment. After the removal of duplicates, title and abstract screening and full-text examination, nine RCTs remained for qualitative analyses. The great majority of the studies did not report the method of randomization, allocation concealment, and examiner blinding during color assessment. From the nine studies, eight were at unclear risk of bias. In regard to color change, four studies reported no change and two reported improved color change with reservoirs. Only four studies recorded tooth sensitivity and they reported no significant differences. Only one study reported greater gingival irritation with reservoirs. Lack of data reporting prevented us from running a meta-analysis. Further well-designed RCT should be conducted to answer this research question. So far there is not evidence to support that reservoirs in bleaching trays improve color change. PROSPERO - CRD42016037628.

Th1 and Th17 Cells and Associated Cytokines Discriminate among Clinically Isolated Syndrome and Multiple Sclerosis Phenotypes.

Multiple sclerosis (MS) is a chronic, inflammatory, and demyelinating disease of the central nervous system. It is a heterogeneous pathology that can follow different clinical courses, and the mechanisms that underlie the progression of the immune response across MS subtypes remain incompletely understood. Here, we aimed to determine differences in the immunological status among different MS clinical subtypes. Blood samples from untreated patients diagnosed with clinically isolated syndrome (CIS) (n = 21), different clinical forms of MS (n = 62) [relapsing-remitting (RRMS), secondary progressive, and primary progressive], and healthy controls (HCs) (n = 17) were tested for plasma levels of interferon (IFN)-γ, IL-10, TGF-ß, IL-17A, and IL-17F by immunoanalysis. Th1 and Th17 lymphocyte frequencies were determined by flow cytometry. Our results showed that IFN-γ levels and the IFN-γ/IL-10 ratio were higher in CIS patients than in RRMS patients and HC. Th1 cell frequencies were higher in CIS and RRMS than in progressive MS, and RRMS had a higher Th17 frequency than CIS. The Th1/Th17 cell ratio was skewed toward Th1 in CIS compared to MS phenotypes and HC. Receiver operating characteristic statistical analysis determined that IFN-γ, the IFN-γ/IL-10 ratio, Th1 cell frequency, and the Th1/Th17 cell ratio discriminated among CIS and MS subtypes. A subanalysis among patients expressing high IL-17F levels showed that IL-17F and the IFN-γ/IL-17F ratio discriminated between disease subtypes. Overall, our data showed that CIS and MS phenotypes displayed distinct Th1- and Th17-related cytokines and cell profiles and that these immune parameters discriminated between clinical forms. Upon validation, these parameters might be useful as biomarkers to predict disease progression.

Galectin-8 as an immunosuppressor in experimental autoimmune encephalomyelitis and a target of human early prognostic antibodies in multiple sclerosis.

Galectin-8 (Gal-8) is a member of a glycan-binding protein family that regulates the immune system, among other functions, and is a target of antibodies in autoimmune disorders. However, its role in multiple sclerosis (MS), an autoimmune inflammatory disease of the central nervous system (CNS), remains unknown. We study the consequences of Gal-8 silencing on lymphocyte subpopulations and the development of experimental autoimmune encephalitis (EAE), to then assess the presence and clinical meaning of anti-Gal-8 antibodies in MS patients. Lgals8/Lac-Z knock-in mice lacking Gal-8 expression have higher polarization toward Th17 cells accompanied with decreased CCR6+ and higher CXCR3+ regulatory T cells (Tregs) frequency. These conditions result in exacerbated MOG35-55 peptide-induced EAE. Gal-8 eliminates activated Th17 but not Th1 cells by apoptosis and ameliorates EAE in C57BL/6 wild-type mice. ß-gal histochemistry reflecting the activity of the Gal-8 promoter revealed Gal-8 expression in a wide range of CNS regions, including high expression in the choroid-plexus. Accordingly, we detected Gal-8 in human cerebrospinal fluid, suggesting a role in the CNS immune-surveillance circuit. In addition, we show that MS patients generate function-blocking anti-Gal-8 antibodies with pathogenic potential. Such antibodies block cell adhesion and Gal-8-induced Th17 apoptosis. Furthermore, circulating anti-Gal-8 antibodies associate with relapsing-remitting MS (RRMS), and not with progressive MS phenotypes, predicting clinical disability at diagnosis within the first year of follow-up. Our results reveal that Gal-8 has an immunosuppressive protective role against autoimmune CNS inflammation, modulating the balance of Th17 and Th1 polarization and their respective Tregs. Such a role can be counteracted during RRMS by anti-Gal-8 antibodies, worsening disease prognosis. Even though anti-Gal-8 antibodies are not specific for MS, our results suggest that they could be a potential early severity biomarker in RRMS.

Effect of the Purple Corn Beverage "Chicha Morada" in Composite Resin during Dental Bleaching.

During dental bleaching the staining potential of the surface would increase. This study aims to evaluate the staining susceptibility of one bleached composite resin after the exposure to three different beverages: Peruvian purple corn based beverage (chicha morada), green tea, and distilled water. Thirty disk-shaped specimens of one nanofill composite resin were prepared. The specimens were then divided into six groups (n = 5): purple corn (P), purple corn + bleaching (PB), green tea (T), green tea + bleaching (TB), distilled water (W), and distilled water + bleaching (WB). In groups that received bleaching, two sessions of bleaching with 35% hydrogen peroxide were done. Following bleaching, specimens were exposed to each liquid thirty minutes daily. Color was measured with a digital spectrophotometer. For statistical analysis, color measurement differences between the obtained results were used: during bleaching, after bleaching, and during + after bleaching. Two-way ANOVA was used to compare the color changes in the resins of all groups (p < 0.05). We conclude that all the evaluated beverages produced changes of color in the composite resin regardless of the bleaching procedure. However, purple corn was the only beverage that caused a perceptible color change (ΔE > 3.3).

Does the use of reservoirs have any impact on the efficacy of at-home bleaching? a systematic review

Abstract To answer the following focused question through a systematic review: "Are the risk and intensity of tooth sensitivity (TS) and bleaching efficacy different between adult patients who undergo at-home bleaching using trays with reservoirs and those who use trays without reservoirs?". A comprehensive search was performed in the MEDLINE via PubMed, Scopus, Web of Science, Latin American and Caribbean Health Sciences Literature database, Brazilian Library in Dentistry, Cochrane Library, and grey literature without restrictions. Abstracts from conferences; unpublished and ongoing trial registries, dissertations and theses (ProQuest Dissertations and Periódicos Capes Theses databases) were searched. Only randomized clinical trials (RCTs) were included. We used the Risk of Bias tool (RoB) from the Cochrane Collaboration for quality assessment. After the removal of duplicates, title and abstract screening and full-text examination, nine RCTs remained for qualitative analyses. The great majority of the studies did not report the method of randomization, allocation concealment, and examiner blinding during color assessment. From the nine studies, eight were at unclear risk of bias. In regard to color change, four studies reported no change and two reported improved color change with reservoirs. Only four studies recorded tooth sensitivity and they reported no significant differences. Only one study reported greater gingival irritation with reservoirs. Lack of data reporting prevented us from running a meta-analysis. Further well-designed RCT should be conducted to answer this research question. So far there is not evidence to support that reservoirs in bleaching trays improve color change. PROSPERO - CRD42016037628

Resumo Para responder a seguinte questão de pesquisa através de uma revisão sistemática: "O risco e a intensidade de sensibilidade dentária (SD) e eficácia de clareamento são diferentes entre pacientes adultos que realizam clareamento caseiro usando moldeiras com reservatórios e aqueles que usam moldeiras sem reservatórios?". Uma pesquisa abrangente foi realizada no MEDLINE via PubMed, Scopus, Web of Science, Literatura Latino-Americana e do Caribe em Ciências da Saúde (LILACS), Biblioteca Cochrane e literatura cinzenta, sem restrições. Os resumos da conferência anual da Associação Internacional para Pesquisa Dental além de estudos registrados ou em andamento também foram pesquisados. Dissertações e Teses foram pesquisados utilizando o Capes Journal Dissertações e Teses ProQuest. Apenas ensaios clínicos randomizados (ECR) foram incluídos. Usamos a ferramenta Risk of Bias (RoB) da Cochrane para avaliação de qualidade. Após a remoção de duplicatas, triagem de título e resumo e exame de texto completo, nove ECRs permaneceram para análises qualitativas. A grande maioria dos estudos não relatou o método de randomização, ocultação de alocação e cegamento do examinador durante a avaliação de cores. Dos nove estudos, oito estavam sob risco claro de viés. Em relação à mudança de cor, quatro estudos não relataram nenhuma mudança e dois relataram melhora na mudança de cor com reservatórios. Apenas quatro estudos registraram a sensibilidade dentária e não relataram diferenças significativas. Apenas um estudo relatou maior irritação gengival com reservatórios. A falta de relatórios de dados nos impediu de executar uma meta-análise. Outros ECR bem desenhados devem ser conduzidos para responder a esta questão de pesquisa. Até agora não há evidências que sustentem que reservatórios em moldeiras de clareamento melhorem a mudança de cor. PROSPERO - CRD42016037628