SARS-CoV-2 vaccination induces immunological T cell memory able to cross-recognize variants from Alpha to Omicron.

We address whether T cell responses induced by different vaccine platforms (mRNA-1273, BNT162b2, Ad26.COV2.S, and NVX-CoV2373) cross-recognize early SARS-CoV-2 variants. T cell responses to early variants were preserved across vaccine platforms. By contrast, significant overall decreases were observed for memory B cells and neutralizing antibodies. In subjects ∼6 months post-vaccination, 90% (CD4+) and 87% (CD8+) of memory T cell responses were preserved against variants on average by AIM assay, and 84% (CD4+) and 85% (CD8+) preserved against Omicron. Omicron RBD memory B cell recognition was substantially reduced to 42% compared with other variants. T cell epitope repertoire analysis revealed a median of 11 and 10 spike epitopes recognized by CD4+ and CD8+ T cells, with average preservation > 80% for Omicron. Functional preservation of the majority of T cell responses may play an important role as a second-level defense against diverse variants.

Prior vaccination promotes early activation of memory T cells and enhances immune responses during SARS-CoV-2 breakthrough infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection of vaccinated individuals is increasingly common but rarely results in severe disease, likely due to the enhanced potency and accelerated kinetics of memory immune responses. However, there have been few opportunities to rigorously study early recall responses during human viral infection. To better understand human immune memory and identify potential mediators of lasting vaccine efficacy, we used high-dimensional flow cytometry and SARS-CoV-2 antigen probes to examine immune responses in longitudinal samples from vaccinated individuals infected during the Omicron wave. These studies revealed heightened spike-specific responses during infection of vaccinated compared to unvaccinated individuals. Spike-specific cluster of differentiation (CD)4 T cells and plasmablasts expanded and CD8 T cells were robustly activated during the first week. In contrast, memory B cell activation, neutralizing antibody production and primary responses to nonspike antigens occurred during the second week. Collectively, these data demonstrate the functionality of vaccine-primed immune memory and highlight memory T cells as rapid responders during SARS-CoV-2 infection.

Evidence from sperm whale clans of symbolic marking in non-human cultures.

Culture, a pillar of the remarkable ecological success of humans, is increasingly recognized as a powerful force structuring nonhuman animal populations. A key gap between these two types of culture is quantitative evidence of symbolic markers-seemingly arbitrary traits that function as reliable indicators of cultural group membership to conspecifics. Using acoustic data collected from 23 Pacific Ocean locations, we provide quantitative evidence that certain sperm whale acoustic signals exhibit spatial patterns consistent with a symbolic marker function. Culture segments sperm whale populations into behaviorally distinct clans, which are defined based on dialects of stereotyped click patterns (codas). We classified 23,429 codas into types using contaminated mixture models and hierarchically clustered coda repertoires into seven clans based on similarities in coda usage; then we evaluated whether coda usage varied with geographic distance within clans or with spatial overlap between clans. Similarities in within-clan usage of both "identity codas" (coda types diagnostic of clan identity) and "nonidentity codas" (coda types used by multiple clans) decrease as space between repertoire recording locations increases. However, between-clan similarity in identity, but not nonidentity, coda usage decreases as clan spatial overlap increases. This matches expectations if sympatry is related to a measurable pressure to diversify to make cultural divisions sharper, thereby providing evidence that identity codas function as symbolic markers of clan identity. Our study provides quantitative evidence of arbitrary traits, resembling human ethnic markers, conveying cultural identity outside of humans, and highlights remarkable similarities in the distributions of human ethnolinguistic groups and sperm whale clans.

A pediatric randomized, controlled trial of German cockroach subcutaneous immunotherapy.

BACKGROUND: Cockroach allergy contributes to morbidity among urban children with asthma. Few trials address the effect of subcutaneous immunotherapy (SCIT) with cockroach allergen among these at-risk children. OBJECTIVES: We sought to determine whether nasal allergen challenge (NAC) responses to cockroach allergen would improve following 1 year of SCIT. METHODS: Urban children with asthma, who were cockroach-sensitized and reactive on NAC, participated in a year-long randomized double-blind placebo-controlled SCIT trial using German cockroach extract. The primary endpoint was the change in mean Total Nasal Symptom Score (TNSS) during NAC after 12 months of SCIT. Changes in nasal transcriptomic responses during NAC, skin prick test wheal size, serum allergen-specific antibody production, and T-cell responses to cockroach allergen were assessed. RESULTS: Changes in mean NAC TNSS did not differ between SCIT-assigned (n = 28) versus placebo-assigned (n = 29) participants (P = .63). Nasal transcriptomic responses correlated with TNSS, but a treatment effect was not observed. Cockroach serum-specific IgE decreased to a similar extent in both groups, while decreased cockroach skin prick test wheal size was greater among SCIT participants (P = .04). A 200-fold increase in cockroach serum-specific IgG4 was observed among subjects receiving SCIT (P < .001) but was unchanged in the placebo group. T-cell IL-4 responses following cockroach allergen stimulation decreased to a greater extent among SCIT versus placebo (P = .002), while no effect was observed for IL-10 or IFN-γ. CONCLUSIONS: A year of SCIT failed to alter NAC TNSS and nasal transcriptome responses to cockroach allergen challenge despite systemic effects on allergen-specific skin tests, induction of serum-specific IgG4 serum production and down-modulation of allergen-stimulated T-cell responses.

An update on studies characterizing adaptive immune responses in SARS-CoV-2 infection and COVID-19 vaccination.

In this brief opinion piece, we highlight our studies characterizing adaptive SARS-CoV-2 immune responses in infection and vaccination, and the ability of SARS-CoV-2-specific T cells to recognize emerging variants of concern, and the role of pre-existing cross-reactive T cells. In the context of the debate on correlates of protection, the pandemic's progression in the past 3 years underlined the need to consider how different adaptive immune responses might differentially contribute to protection from SARS-CoV-2 infection versus COVID-19 disease. Lastly, we discuss how cross-reactive T cell responses may be useful in generating a broad adaptive immunity, recognizing different variants and viral families. Considering vaccines with broadly conserved antigens could improve preparedness for future infectious disease outbreaks.

Lymphotoxin beta receptor signaling directly controls airway smooth muscle deregulation and asthmatic lung dysfunction.

BACKGROUND: Dysregulation of airway smooth muscle cells (ASM) is central to the severity of asthma. Which molecules dominantly control ASM in asthma is unclear. High levels of the cytokine LIGHT (aka TNFSF14) have been linked to asthma severity and lower baseline predicted FEV1 percentage, implying that signals through its receptors might directly control ASM dysfunction. OBJECTIVE: Our study sought to determine whether signaling via lymphotoxin beta receptor (LTßR) or herpesvirus entry mediator from LIGHT dominantly drives ASM hyperreactivity induced by allergen. METHODS: Conditional knockout mice deficient for LTßR or herpesvirus entry mediator in smooth muscle cells were used to determine their role in ASM deregulation and airway hyperresponsiveness (AHR) in vivo. Human ASM were used to study signals induced by LTßR. RESULTS: LTßR was strongly expressed in ASM from normal and asthmatic subjects compared to several other receptors implicated in smooth muscle deregulation. Correspondingly, conditional deletion of LTßR only in smooth muscle cells in smMHCCreLTßRfl/fl mice minimized changes in their numbers and mass as well as AHR induced by house dust mite allergen in a model of severe asthma. Intratracheal LIGHT administration independently induced ASM hypertrophy and AHR in vivo dependent on direct LTßR signals to ASM. LIGHT promoted contractility, hypertrophy, and hyperplasia of human ASM in vitro. Distinguishing LTßR from the receptors for IL-13, TNF, and IL-17, which have also been implicated in smooth muscle dysregulation, LIGHT promoted NF-κB-inducing kinase-dependent noncanonical nuclear factor kappa-light-chain enhancer of activated B cells in ASM in vitro, leading to sustained accumulation of F-actin, phosphorylation of myosin light chain kinase, and contractile activity. CONCLUSIONS: LTßR signals directly and dominantly drive airway smooth muscle hyperresponsiveness relevant for pathogenesis of airway remodeling in severe asthma.

T Cell Responses in Pregnant Women Who Received mRNA-Based Vaccination to Prevent COVID-19 Revealed Unknown Exposure to the Natural Infection and Numerous SARS-CoV-2-Specific CD4- CD8- Double Negative T Cells and Regulatory T Cells.

We studied T-cell responses to SARS-CoV-2 in 19 pregnant subjects at different gestational weeks who received three doses of mRNA-based vaccination to prevent COVID-19. SARS-CoV-2 peptide pools were used for T-cell recognition studies: peptides were 15 amino acids long and had previously been defined in COVID-19-convalescent subjects. T-cell activation was evaluated with the AIM assay. Most subjects showed coordinated, spike-specific CD4+ and CD8+ T-cell responses and the development of T cell memory. Non-spike-specific T cells in subjects who were not aware of previous COVID-19 infection suggested a prior undetected, asymptomatic infection. CD4- CD8- double negative (DN) T cells were numerous, of which a percentage was specific for SARS-CoV-2 spike peptides. Regulatory T cells (Treg), both spike- and non-spike-specific, were also greatly expanded. Two Treg populations were defined: a population differentiated from naïve T cells, and pTreg, reverting from pro-inflammatory T cells. The Treg cells expressed CCR6, suggesting homing to the endometrium and vaginal epithelial cells. The pregnant women responded to SARS-CoV-2 vaccination. Asymptomatic COVID-19 was revealed by the T cell response to the non-spike peptides. The numerous DN T cells and Treg pointed our attention to new aspects of the adaptive immune response in vaccine recipients.

Antigen Discovery for Next-Generation Pertussis Vaccines Using Immunoproteomics and Transposon-Directed Insertion Sequencing.

BACKGROUND: Despite high vaccination rates, the United States has experienced a resurgence in reported cases of pertussis after switching to the acellular pertussis vaccine, indicating a need for improved vaccines that enhance infection control. METHODS: Bordetella pertussis antigens recognized by convalescent-baboon serum and nasopharyngeal wash were identified by immunoproteomics and their subcellular localization predicted. Genes essential or important for persistence in the baboon airway were identified by transposon-directed insertion-site sequencing (TraDIS) analysis. RESULTS: In total, 314 B. pertussis antigens were identified by convalescent baboon serum and 748 by nasopharyngeal wash. Thirteen antigens were identified as immunogenic in baboons, essential for persistence in the airway by TraDIS, and membrane-localized: BP0840 (OmpP), Pal, OmpA2, BP1485, BamA, Pcp, MlaA, YfgL, BP2197, BP1569, MlaD, ComL, and BP0183. CONCLUSIONS: The B. pertussis antigens identified as immunogenic, essential for persistence in the airway, and membrane-localized warrant further investigation for inclusion in vaccines designed to reduce or prevent carriage of bacteria in the airway of vaccinated individuals.

Anti-CD3 inhibits circulatory and tissue-resident memory CD4 T cells that drive asthma exacerbations in mice.

BACKGROUND: Exacerbations of asthma are thought to be strongly dependent on reactivation of allergen-induced lung tissue-resident and circulatory memory CD4 T cells. Strategies that broadly inhibit multiple T cell populations might then be useful to limit asthma. Accordingly, we tested whether targeting CD3 during exposure to inhaled allergen could prevent the accumulation of lung-localized effector memory CD4 T cells and block exacerbations of asthmatic inflammation. METHODS: House dust mite-sensitized and repetitively challenged BL/6 mice were transiently treated therapeutically with F(ab')2 anti-CD3ε and memory T cell responses and lung inflammation were assessed. PBMCs from HDM-allergic donors were examined for the effect of anti-CD3 on expansion of allergen-reactive T cells. RESULTS: Allergen-sensitized mice undergoing exacerbations of asthma were protected from lung inflammation by transient therapeutic treatment with F(ab')2 anti-CD3. Regardless of whether sensitized mice underwent a secondary or tertiary recall response to inhaled allergen, anti-CD3 inhibited all phenotypes of effector memory CD4 T cells in the lung tissue and lung vasculature by 80%-90%, including those derived from tissue-resident and circulatory memory T cells. This did not depend on Treg cells suggesting it was primarily a blocking effect on memory T cell signaling. Correspondingly, anti-CD3 also strongly inhibited proliferation of human allergen-reactive memory CD4 T cells from allergic individuals. In contrast, the number of surviving tissue-resident memory CD4 T cells that were maintained in the lungs at later times was not robustly reduced by anti-CD3. CONCLUSION: Anti-CD3 F(ab')2 administration at the time of allergen exposure represents a viable strategy for limiting the immediate activity of allergen-responding memory T cells and asthma exacerbations.

Selenized Chickpea Sprouts Hydrolysates as a Potential Anti-Aging Ingredient.

Skin aging represents a health and aesthetic problem that could result in infections and skin diseases. Bioactive peptides can potentially be used in skin aging regulation. Chickpea (Cicer arietinum L.) selenoproteins were obtained from germination with 2 mg Na2SeO3/100 g of seeds for 2 days. Alcalase, pepsin, and trypsin were used as hydrolyzers, and a membrane < 10 kDa was used to fractionate the hydrolysate. Se content, antioxidant capacity, elastase and collagen inhibition, functional stability, and preventative capacity were analyzed. Significant increases in Se content were found in germinated chickpea flour and protein related to the control. An increase of 38% in protein was observed in the selenized flour related to the control. A band (600-550 cm-1) observed in the selenized hydrolysates suggested the insertion of Se into the protein. Hydrolysates from pepsin and trypsin had the highest antioxidant potential. Se enhanced the stability of total protein and protein hydrolysates through time and increased their antioxidant capacity. Hydrolysates > 10 kDa had higher elastase and collagenase inhibition than the total protein and hydrolysates < 10 kDa. Protein hydrolysates < 10 kDa 6 h before UVA radiation had the highest inhibition of collagen degradation. Selenized protein hydrolysates showed promising antioxidant effects that could be related to skin anti-aging effects.

In vitro Evaluation of Anti-Inflammatory Activity of "Habanero" Chili Pepper (Capsicum chinense) Seeds Extracts Pretreated with Cellulase.

The aim of this study was to explore the effect of capsaicin and particular phenolic compounds profile from cellulase assisted extracts of Habanero (Capsicum chinense) chili pepper seeds (CPS) on the concentration of cytokines (IL-2, IL-6, TNF-α, IL-1ß) in murine macrophages (RAW 264.7) stimulated with lipopolysaccharides (LPS). Capsaicin was quantified by HPLC-DAD, and the phenolic profile was determined by UPLC-MS-QqQ. Anti-inflammatory activity was evaluated by Mouse Cytokine/Chemokine Magnetic Bead Panel 96-well plate assay. Among the 15 different phenolics found in CPS extracts obtained at 120 or 150 min of maceration with 2,500 UI/L at 30 ºC or 45 ºC in a 1:15 (w:v) proportion, the most abundant was vanillic acid (7.97-12.66 µg/g). The extract obtained at 30 ºC and 120 min, showed similar effects than the observed for synthetic anti-inflammatory drugs indomethacin and dexamethasone, and capsaicin standard. Beyond capsaicin, salicylic, protocatechuic and trans-cinnamic acids as well as vanillin in CPS extracts were correlated with the anti-inflammatory effect. On the other hand, capsaicin and chlorogenic acid contents were potential immunostimulants whose concentration varied depending on the cellulase treatment time.

Observations and Perspectives on Adaptive Immunity to Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2).

Since the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic began 2 years ago, the scientific community has swiftly worked to understand the transmission, pathogenesis, and immune response of this virus to implement public health policies and ultimately project an end to the pandemic. In this perspective, we present our work identifying SARS-CoV-2 epitopes to quantify T-cell responses and review how T cells may help protect against severe disease. We examine our prior studies which demonstrate durable humoral and cell-mediated memory in natural infection and vaccination. We discuss how SARS-CoV-2-specific T cells from either natural infection or vaccination can recognize emerging variants of concern, suggesting that the currently approved vaccines may be sufficient. We also discuss how pre-existing cross-reactive T cells promote rapid development of immune memory to SARS-CoV-2. We finally posit how identifying SARS-CoV-2 epitopes can help us develop a pan-coronavirus vaccine to prepare for future pandemics.

Biopolymer nanoparticles: a strategy to enhance stability, bioavailability, and biological effects of phenolic compounds as functional ingredients.

Phenolic compounds are abundant in nature and have multiple beneficial effects on human health due to their antioxidant, anti-inflammatory, antithrombotic, antiallergenic, anticancer, and antiatherosclerotic properties. For this reason, phenolics are becoming relevant functional ingredients for several industries, mainly the food industry, derived from food consumer exigencies and regulations. However, the use of their beneficial properties still presents some limitations, such as chemical instability under environmental and processing conditions, which leads to structural changes and compromises their biological activities. They also present poor water solubility and sensitivity to pH changes, decreasing their bioavailability in the organism. The technologies for extraction and stabilization of these compounds have evolved rapidly in the development of different delivery systems to encapsulate sensitive active molecules. Biopolymeric nanoparticles are biodegradable polymer-based colloidal systems with sizes ranging from 1 to 1000 nm, and different techniques can be carried out to develop them. These systems have emerged as a green and effective alternative to improve stability, bioavailability, and biological effects of phenolic compounds. This comprehensive review aims to present an overview of recent advances in encapsulation processes of phenolic compounds within biopolymer nanoparticles as delivery systems and the impact on their physicochemical properties and biological effects after encapsulation. © 2021 Society of Chemical Industry.

Differential T-Cell Reactivity to Endemic Coronaviruses and SARS-CoV-2 in Community and Health Care Workers.

Herein we measured CD4+ T-cell responses against common cold coronaviruses (CCC) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in high-risk health care workers (HCW) and community controls. We observed higher levels of CCC-reactive T cells in SARS-CoV-2-seronegative HCW compared to community donors, consistent with potential higher occupational exposure of HCW to CCC. We further show that SARS-CoV-2 T-cell reactivity of seronegative HCW was higher than community controls and correlation between CCC and SARS-CoV-2 responses is consistent with cross-reactivity and not associated with recent in vivo activation. Surprisingly, CCC T-cell reactivity was decreased in SARS-CoV-2-infected HCW, suggesting that exposure to SARS-CoV-2 might interfere with CCC responses, either directly or indirectly. This result was unexpected, but consistently detected in independent cohorts derived from Miami and San Diego. CD4+ T-cell responses against common cold coronaviruses (CCC) are elevated in SARS-CoV-2 seronegative high-risk health care workers (HCW) compared to COVID-19 convalescent HCW, suggesting that exposure to SARS-CoV-2 might interfere with CCC responses and/or cross-reactivity associated with a protective effect.

Lack of evidence supporting a role of IFN-ß and TGF-ß in differential polarization of Bordetella pertussis specific-T cell responses.

Bordetella Pertussis (BP) vaccine-induced immunity is waning worldwide despite excellent vaccine coverage. Replacement of the whole-cell inactivated vaccine (wP) by an acellular subunit vaccine (aP) is thought to play a major role and to be associated with the recurrence of whooping cough. Previously, we detected that the polarization towards a Th2 and Th1/Th17 response in aP and wP vaccinees, respectively, persists upon aP boosting in adolescents and adults. Additionally, IL-9 and TGF-ß were found to be up-regulated in aP-primed donors and network analysis further identified IFN-ß as a potential upstream regulator of IL-17 and IL-9. Based on these findings, we hypothesized that IFN-ß produced following aP vaccination may lead to increased IL-9 and decreased IL-17 production. Also, due to the well characterized role of TGF-ß in both Th17 and Th9 differentiation, we put forth that TGF-ß addition to BP-stimulated CD4 + T cells might modulate IL-17 and IL-9 production. To test this hypothesis, we stimulated in vitro cultures of PBMC or isolated naive CD4 + T cells from aP vs wP donors with a pool of BP epitopes and assessed the effect of IFN-ß or TGF-ß in proliferative responses as well as in the cytokine secretion of IL-4, IL-9, IL-17, and IFN-γ. IFN-ß reduced BP-specific proliferation in PBMC as well as cytokine production but increased IL-9, IL-4, and IFN-γ cytokines in naïve CD4 + T cells. These effects were independent of the childhood vaccination received by the donors. Similarly, TGF-ß reduced BP-specific proliferation in PBMC but induced proliferation in naïve CD4 + T cells. However, stimulation was associated with a generalized inhibition of cytokine production regardless of the original aP or wP vaccination received by the donors. Our study suggests that key T cell functions such as cytokine secretion are under the control of antigen stimulation and environmental cues but molecular pathways different than the ones investigated here might underlie the long-lasting differential cytokine production associated with aP- vs wP-priming in childhood vaccination.

Opuntia ficus-indica Extract and Isorhamnetin-3-O-Glucosyl-Rhamnoside Diminish Tumor Growth of Colon Cancer Cells Xenografted in Immune-Suppressed Mice through the Activation of Apoptosis Intrinsic Pathway.

This study aimed to evaluate the effects of Opuntia ficus-indica extract (OFI-E) and its glycoside isorhamnetin-3-O-glucosyl-rhamnoside (IGR) on the growth of human colorectal adenocarcinoma cells and in a xenografted-immunosuppressed mice model. The IC50 values of OFI-E and IGR on colon cancer cells (HT-29 RFP) were determinate, as well as their effects on the cell cycle and apoptosis induction. OFI-E and IGR produced an increased in apoptosis induction, ROS production and a G0/G1 cell cycle arrest. In xenografted-inmunosupressed mice, OFI-E and IGR reduced the tumor growth rate, myeloperoxidase activity and total cholesterol levels. OFI-E and IGR reduced the tumor growth through the overexpression of cleaved Caspase-9, Hdac11, and Bai1 proteins. OFI-E reduced the expression of bcl-2. Results demonstrated the chemopreventive effects of OFI-E, and its purified compound IGR, showing their potential as an alternative in the treatment of colorectal cancer.

Enhanced production and identification of antioxidants in in vitro cultures of the cacti Mammillaria candida and Turbinicarpus laui.

Cacti are an important source of metabolites but present limitations for their commercial exploitation, like slow growth and a decrease of wild populations. An alternative to obtain their biocompounds without affecting the natural environment are the in vitro culture techniques. We established in vitro cultures from Mammillaria candida Scheidweiler and Turbinicarpus laui Glass and Foster and used different stresses to increase metabolites and antioxidant activity. The cultures were exposed to 1.25% polyethylene glycol to induce a moderate drought stress, 50 g L-1 sucrose to generate an osmotic stress, chitosan (1.25 to 5 mg mL-1) to simulate a biotic attack, or to UV light. Chitosan was the best elicitor improving 1.5 times the concentration of phenolics, 9 to 10 times the content of flavonoids and betalains, and 16% the antioxidant activity in M. candida suspensions. In T. laui suspensions, this elicitor duplicates the flavonoids content and antioxidant activity. The antioxidant levels in elicited suspensions increased 5 to 10 times in relation to plant tubercles. Eleven compounds were identified in M. candida suspensions being digalloyl rhamnoside and epicatequin gallate the most abundant; in the T. laui suspensions, 16 compounds were detected and the most abundant were 17-decarboxi neobetanin and derivatives of luteolin. Thus, cacti in vitro culture is an efficient system to obtain high level of metabolites of biological interest.

The Proteins API: accessing key integrated protein and genome information.

The Proteins API provides searching and programmatic access to protein and associated genomics data such as curated protein sequence positional annotations from UniProtKB, as well as mapped variation and proteomics data from large scale data sources (LSS). Using the coordinates service, researchers are able to retrieve the genomic sequence coordinates for proteins in UniProtKB. This, the LSS genomics and proteomics data for UniProt proteins is programmatically only available through this service. A Swagger UI has been implemented to provide documentation, an interface for users, with little or no programming experience, to 'talk' to the services to quickly and easily formulate queries with the services and obtain dynamically generated source code for popular programming languages, such as Java, Perl, Python and Ruby. Search results are returned as standard JSON, XML or GFF data objects. The Proteins API is a scalable, reliable, fast, easy to use RESTful services that provides a broad protein information resource for users to ask questions based upon their field of expertise and allowing them to gain an integrated overview of protein annotations available to aid their knowledge gain on proteins in biological processes. The Proteins API is available at (http://www.ebi.ac.uk/proteins/api/doc).

A Cytokine-Independent Approach To Identify Antigen-Specific Human Germinal Center T Follicular Helper Cells and Rare Antigen-Specific CD4+ T Cells in Blood.

Detection of Ag-specific CD4(+) T cells is central to the study of many human infectious diseases, vaccines, and autoimmune diseases. However, such cells are generally rare and heterogeneous in their cytokine profiles. Identification of Ag-specific germinal center (GC) T follicular helper (Tfh) cells by cytokine production has been particularly problematic. The function of a GC Tfh cell is to selectively help adjacent GC B cells via cognate interaction; thus, GC Tfh cells may be stingy cytokine producers, fundamentally different from Th1 or Th17 cells in the quantities of cytokines produced. Conventional identification of Ag-specific cells by intracellular cytokine staining relies on the ability of the CD4(+) T cell to generate substantial amounts of cytokine. To address this problem, we have developed a cytokine-independent activation-induced marker (AIM) methodology to identify Ag-specific GC Tfh cells in human lymphoid tissue. Whereas Group A Streptococcus-specific GC Tfh cells produced minimal detectable cytokines by intracellular cytokine staining, the AIM method identified 85-fold more Ag-specific GC Tfh cells. Intriguingly, these GC Tfh cells consistently expressed programmed death ligand 1 upon activation. AIM also detected non-Tfh cells in lymphoid tissue. As such, we applied AIM for identification of rare Ag-specific CD4(+) T cells in human peripheral blood. Dengue, tuberculosis, and pertussis vaccine-specific CD4(+) T cells were readily detectable by AIM. In summary, cytokine assays missed 98% of Ag-specific human GC Tfh cells, reflecting the biology of these cells, which could instead be sensitively identified by coexpression of TCR-dependent activation markers.

Intestinal Permeability and Cellular Antioxidant Activity of Phenolic Compounds from Mango (Mangifera indica cv. Ataulfo) Peels.

Mango (Mangifera indica cv. Ataulfo) peel contains bound phenolics that may be released by alkaline or acid hydrolysis and may be converted into less complex molecules. Free phenolics from mango cv. Ataulfo peel were obtained using a methanolic extraction, and their cellular antioxidant activity (CAA) and permeability were compared to those obtained for bound phenolics released by alkaline or acid hydrolysis. Gallic acid was found as a simple phenolic acid after alkaline hydrolysis along with mangiferin isomers and quercetin as aglycone and glycosides. Only gallic acid, ethyl gallate, mangiferin, and quercetin were identified in the acid fraction. The acid and alkaline fractions showed the highest CAA (60.5% and 51.5%) when tested at 125 µg/mL. The value of the apparent permeability coefficient (Papp) across the Caco-2/HT-29 monolayer of gallic acid from the alkaline fraction was higher (2.61 × 10-6 cm/s) than in the other fractions and similar to that obtained when tested pure (2.48 × 10-6 cm/s). In conclusion, mango peels contain bound phenolic compounds that, after their release, have permeability similar to pure compounds and exert an important CAA. This finding can be applied in the development of nutraceuticals using this important by-product from the mango processing industry.