Cell-mediated Research Articles

Mitochondrial Hyperactivity and Reactive Oxygen Species Drive Innate Immunity to the Yellow Fever Virus-17D Live-Attenuated Vaccine.

The yellow fever virus 17D (YFV-17D) live attenuated vaccine is considered one of the successful vaccines ever generated associated with high antiviral immunity, yet the signaling mechanisms that drive the response in infected cells are not understood. Here, we provide a molecular understanding of how metabolic stress and innate immune responses are linked to drive type I IFN expression in response to YFV-17D infection. Comparison of YFV-17D replication with its parental virus, YFV-Asibi, and a related dengue virus revealed that IFN expression requires RIG-I-like Receptor signaling through MAVS, as expected. However, YFV-17D uniquely induces mitochondrial respiration and major metabolic perturbations, including hyperactivation of electron transport to fuel ATP synthase. Mitochondrial hyperactivity generates reactive oxygen species (mROS) and peroxynitrite, blocking of which abrogated IFN expression in non-immune cells without reducing YFV-17D replication. Scavenging ROS in YFV-17D-infected human dendritic cells increased cell viability yet globally prevented expression of IFN signaling pathways. Thus, adaptation of YFV-17D for high growth uniquely imparts mitochondrial hyperactivity generating mROS and peroxynitrite as the critical messengers that convert a blunted IFN response into maximal activation of innate immunity essential for vaccine effectiveness.

Abstract C042: Identification of a non-canonical pathway used by CAF to deliver gamma-aminobutyric acid in pancreatic cancer

Abstract Cancer-associated fibroblasts (CAFs) are cells responsible for deposition and remodeling of the extracellular matrix, and modification of the stromal microenvironment, which facilitates tumor growth. Moreover, CAFs are extremely immunosuppressive because they produce large amounts of inhibitory cytokines, growth factors and metabolites. Thus, understanding how CAFs impart severe immunosuppression in the tumor microenvironment is critical to make PDAC amenable to immunotherapies. The link between metabolism and immunosuppression in tumors has become clearer in recent years, and we postulate that one such metabolite, gamma-aminobutyric acid (GABA), a key inhibitory neurotransmitter, can act as a major immunosuppressive metabolite in PDAC. Our preliminary data shows that CAFs produce high levels of glutamate (Glu) and that metabolites derived from glutamate, such as GABA, are immunosuppressive to NK cells. Moreover, we found that CAFs could produce GABA, de novo, as determined by heavy isotope tracing. Thus, our goal is to uncover the mechanisms used by CAFs to produce GABA, and to evaluate the immunosuppressive mechanisms behind CAF- derived GABA. GABA production was measured in the conditioned media of CAFs grown in 3D culture, where it was observed that inhibition of glutaminase (GLS) and glutamine synthetase (GS) enzymes was not able to decrease the GABA levels. In addition, CAFs did not express glutamate decarboxylase (GAD), the canonical enzyme responsible for the conversion of Glu to GABA, which led us to explore the expression of a non-canonical GABA synthesis pathways in CAFs. Intriguingly, we found that CAFs derived from patient tumors expressed the enzymes of non-canonical pathway (ODC1, DAO, ALDH1A1 and ALDH9A1), and to a greater degree when compared to normal fibroblasts isolated from donor pancreata (Gift of Life Program). CRISPRi mediated knockdown (KD) of aldehyde dehydrogenase 1 (ALDH1a1) in CAFs decreased GABA production compared to control CAFs. On the other hand, CAFs knocked down for diamine oxidase (DAO), showed increased GABA production, suggesting a compensatory mechanism for GABA production upstream of ALDH1a1. Overall, our preliminary data demonstrate that CAFs produce GABA de novo, via a putative alternative GABA synthesis pathway. Our next steps are to identify the cytokine profile of KDs CAFs and perform functional assays utilizing our 3D co-culturing system, uncovering natural killer cell responses to tumor cells in the presence of CAFs with and without the enzymes necessary for GABA production. Translationally, we are analyzing the tumor interstitial fluid from PDAC patients to uncover metabolites/cytokines (secreted factors) that can be correlated with patient parameters (ex. overall survival) and immune cell activation and functionality. It is our hope that the insight provided by this study can lead to a novel set of prognostic or diagnostic factors to improve clinical outcomes in this devasting disease. Citation Format: Ariana Musa de Aquino, Myree Graves, Farrah Ali Ali, David A. Rangel, Langley Kyra, Julie Clark, David Kwon, Guillaume Cognet, Alex Muir, Débora Barbosa Vendramini Costa, Ralph Francescone. Identification of a non-canonical pathway used by CAF to deliver gamma-aminobutyric acid in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr C042.

Toxic effects of ZnO NPs on immune response and tissue pathology in Mytilus galloprovincialis

Nano-zinc oxide (ZnO NPs), as widely used nanomaterials, are inevitably released into aquatic environments, posing potential threats to aquatic organisms. Mytilus galloprovincialis is a bivalve species sensitive to changes in marine ecological environments, but there has been limited research on its toxicity response to ZnO NPs. Therefore, we selected M. galloprovincialis as the research subject and exposed them to 50 µg/L ZnO NPs for 96 h and 30 days to determine the dissolution of ZnO NPs in seawater and their distribution in M. galloprovincialis. The toxicity of ZnO NPs in M. galloprovincialis was then evaluated through gene expression, tissue pathology, and cellular immune response. The results showed that ZnO NPs could enrich Zn in various tissues of the mussel, in the order of gills > hepatopancreas > adductor muscle > mantle. Seven immune-related genes including four heat shock protein genes (HSPA12A, sHSP24.1, sHSP22, TCTP) and three apoptotic genes (Ras, p63 and Bcl-2) were altered to varying degrees. There was a downward trend in lysosomal membrane stability of M. galloprovincialis after exposure to ZnO NPs for 96 h and 30 days, while ROS and apoptosis rates increased significantly. Furthermore, the seven genes, apoptosis, LMS, and ROS were dependent on exposure time, treatment, and their interaction. Histopathological damage included disorganisation of hepatopancreas epithelial cells, gill filament swelling, and contraction of blood sinuses. These results indicated that ZnO NPs exerted toxicity in M. galloprovincialis, affecting the immune system, resulting in changes in the expression of immune-related genes and ultimately leading to histopathological changes. Our research findings could contribute to systematically understand the impact of ZnO NPs on bivalves in aquatic environments and provide a theoretical basis for marine pollution assessment.

Biodegradable oxygen-evolving metalloantibiotics for spatiotemporal sono-metalloimmunotherapy against orthopaedic biofilm infections

Pathogen-host competition for manganese and intricate immunostimulatory pathways severely attenuates the efficacy of antibacterial immunotherapy against biofilm infections associated with orthopaedic implants. Herein, we introduce a spatiotemporal sono-metalloimmunotherapy (SMIT) strategy aimed at efficient biofilm ablation by custom design of ingenious biomimetic metal-organic framework (PCN-224)-coated MnO2-hydrangea nanoparticles (MnPM) as a metalloantibiotic. Upon reaching the acidic H2O2-enriched biofilm microenvironment, MnPM can convert abundant H2O2 into oxygen, which is conducive to significantly enhancing the efficacy of ultrasound (US)-triggered sonodynamic therapy (SDT), thereby exposing bacteria-associated antigens (BAAs). Moreover, MnPM disrupts bacterial homeostasis, further killing more bacteria. Then, the Mn ions released from the degraded MnO2 can recharge immune cells to enhance the cGAS-STING signaling pathway sensing of BAAs, further boosting the immune response and suppressing biofilm growth via biofilm-specific T cell responses. Following US withdrawal, the sustained oxygenation promotes the survival and migration of fibroblasts, stimulates the expression of angiogenic growth factors and angiogenesis, and neutralizes excessive inflammation. Our findings highlight that MnPM may act as an immune costimulatory metalloantibiotic to regulate the cGAS-STING signaling pathway, presenting a promising alternative to antibiotics for orthopaedic biofilm infection treatment and pro-tissue repair.

Molecular characterization of the IgH locus and V(D)J recombination in large yellow croaker (Larimichthys crocea)

V(D)J recombination is crucial for generating a diverse repertoire of immunoglobulins. Although the V(D)J recombination process has been well characterized in mammals, this process remains largely unexplored in teleosts. In this study, we comprehensively analyzed the IgH locus of a marine fish species large yellow croaker (Larimichthys crocea), and identified 28 V, 19 D, and 8 J gene segments, following a pattern of V-Dζ-Jζ-Cζ-Dμ-Jμ-Cμ1-Cμ2. The V, D, and J gene segments are flanked by consensus recombination signal sequences, with spacer lengths similar to those observed in mammals. The V gene segments are categorized into three distinct families, and exhibited a higher sequence identity compared to those in mammals. Additionally, we designed a set of primers for the examination of the V(D)J recombination in large yellow croaker. RNA-seq analysis showed increased expression of genes related to immunoglobulin production and lymphocyte chemotaxis in IgM + B cells upon Pseudomonas plecoglossicida infection, accompanied by altered expression of V gene segments, suggesting their involvement in the response to P. plecoglossicida infection. Taken together, we identified the IgH locus and V(D)J recombination process of large yellow croaker, which contribute to the understanding of immunoglobulin production and B cell immunity in teleosts, and may provide insights into vaccine development in large yellow croaker.

New insights into mechanism of ellagic acid alleviating arsenic-induced oxidative stress through MAPK/keap1-Nrf2 signaling pathway response, molecular docking and metabolomics analysis in HepG2 cells

The increase of oxidative stress level is one of the vital mechanisms of liver toxicity induced by arsenic (As). Ellagic acid (EA) is widely known due to its excellent antioxidation. Nevertheless, whether EA could alleviate As-induced oxidative stress and the underlying mechanisms remain unknown. Herein, As (2 and 4 μM) and EA (25 and 50 μM) were selected for alone and combined exposure of HepG2 cells to investigate the effects of EA on As-induced oxidative stress. Results indicated that EA could alleviate the oxidative stress caused by As via decreasing intracellular ROS level and MDA content, as well as improving SOD, CAT and GSH-PX activities. qRT-PCR showed that EA might enhance the expression levels of antioxidant enzymes NQO1, CAT and GPX1 by activating MAPK (JNK, p38 and ERK)/keap1-Nrf2 signaling pathway. EA was found to promote dissociation from keap1 and nuclear translocation of Nrf2 by competing with Nrf2 at ARG-380 and ARG-415 sites on keap1 to exert antioxidation using molecular docking. Moreover, metabolomics revealed that EA might maintain the redox balance of HepG2 cells by modulating or reversing disorders of carbon, amino acid, lipid and other metabolisms caused by As. This study provides diversified new insights for the removal of liver toxicity of As and the application of EA.

Programming Bordetella pertussis lipid A to promote adjuvanticity

BackgroundBordetella pertussis is the causative agent of whooping cough or pertussis. Although both acellular (aP) and whole-cell pertussis (wP) vaccines protect against disease, the wP vaccine, which is highly reactogenic, is better at preventing colonization and transmission. Reactogenicity is mainly attributed to the lipid A moiety of B. pertussis lipooligosaccharide (LOS). Within LOS, lipid A acts as a hydrophobic anchor, engaging with TLR4-MD2 on host immune cells to initiate both MyD88-dependent and TRIF-dependent pathways, thereby influencing adaptive immune responses. Lipid A variants, such as monophosphoryl lipid A (MPLA) can also act as adjuvants. Adjuvants may overcome the shortcomings of aP vaccines.ResultsThis work used lipid A modifying enzymes from other bacteria to produce an MPLA-like adjuvant strain in B. pertussis. We created B. pertussis strains with distinct lipid A modifications, which were validated using MALDI-TOF. We engineered a hexa-acylated monophosphorylated lipid A that markedly decreased human TLR4 activation and activated the TRIF pathway. The modified lipooligosaccharide (LOS) promoted IRF3 phosphorylation and type I interferon production, similar to MPLA responses. We generated three other variants with increased adjuvanticity properties and reduced endotoxicity. Pyrogenicity studies using the Monocyte Activation Test (MAT) revealed that these four lipid A variants significantly decreased the IL-6, a marker for fever, response in peripheral blood mononuclear cells (PBMCs).ConclusionThese findings pave the way for developing wP vaccines that are possibly less reactogenic and designing adaptable adjuvants for current vaccine formulations, advancing more effective immunization strategies against pertussis.

Multitask machine learning-based tumor-associated collagen signatures predict peritoneal recurrence and disease-free survival in gastric cancer.

Accurate prediction of peritoneal recurrence for gastric cancer (GC) is crucial in clinic. The collagen alterations in tumor microenvironment affect the migration and treatment response of cancer cells. Herein, we proposed multitask machine learning-based tumor-associated collagen signatures (TACS), which are composed of quantitative collagen features derived from multiphoton imaging, to simultaneously predict peritoneal recurrence (TACSPR) and disease-free survival (TACSDFS). Among 713 consecutive patients, with 275 in training cohort, 222 patients in internal validation cohort, and 216 patients in external validation cohort, we developed and validated a multitask machine learning model for simultaneously predicting peritoneal recurrence (TACSPR) and disease-free survival (TACSDFS). The accuracy of the model for prediction of peritoneal recurrence and prognosis as well as its association with adjuvant chemotherapy were evaluated. The TACSPR and TACSDFS were independently associated with peritoneal recurrence and disease-free survival in three cohorts, respectively (all P < 0.001). The TACSPR demonstrated a favorable performance for peritoneal recurrence in all three cohorts. In addition, the TACSDFS also showed a satisfactory accuracy for disease-free survival among included patients. For stage II and III diseases, adjuvant chemotherapy improved the survival of patients with low TACSPR and low TACSDFS, or high TACSPR and low TACSDFS, or low TACSPR and high TACSDFS, but had no impact on patients with high TACSPR and high TACSDFS. The multitask machine learning model allows accurate prediction of peritoneal recurrence and survival for GC and could distinguish patients who might benefit from adjuvant chemotherapy.

Impact of HDAC6-mediated progesterone receptor expression on the response of breast cancer cells to hormonal therapy

Modulation of estrogen receptor (ER) and progesterone receptor (PR) expression, as well as their emerging functional crosstalk, remains a potential approach for enhancing the response to hormonal therapy in breast cancer. Aberrant epigenetic alterations induced by histone deacetylases (HDACs) were massively implicated in dysregulating the function of hormone receptors in breast cancer. Although much is known about the regulation of ER signaling by HDAC, the precise role of HDAC in modulating the expression of PR and its impact on the outcomes of hormonal therapy is poorly defined. Here, we demonstrate the involvement of HDAC6 in regulating PR expression in breast cancer cells. The correlation between HDAC6 and hormone receptors was investigated in patients’ tissues by immunohistochemistry (n = 80) and publicly available data (n = 3260) from breast cancer patients. We explored the effect of modulating the expression of HDAC6 as well as its catalytic inhibition on the level of hormone receptors by a variety of molecular analyses, including Western blot, immunofluorescence, Real-time PCR, RNA-seq analysis and chromatin immunoprecipitation. Based on our in-silico and immunohistochemistry analyses, HDAC6 levels were negatively correlated with PR status in breast cancer tissues. The downregulation of HDAC6 enhanced the expression of PR-B in hormone receptor-positive and triple-negative breast cancer (TNBC) cells. The selective targeting of HDAC6 by tubacin resulted in the enrichment of the H3K9 acetylation mark at the PGR-B gene promoter region and enhanced the expression of PR-B. Additionally, transcriptomic analysis of tubacin-treated cells revealed enhanced activity of acetyltransferase and growth factor signaling pathways, along with the enrichment of transcription factors involved in the transcriptional activity of ER, underscoring the crucial role of HDAC6 in regulating hormone receptors. Notably, the addition of HDAC6 inhibitor potentiated the effects of anti-ER and anti-PR drugs mainly in TNBC cells. Together, these data highlight the role of HDAC6 in regulating PR expression and provide a promising therapeutic approach for boosting breast cancer sensitivity to hormonal therapy.

A nanobody-guided multifunctional T cell engager promotes strong anti-tumor responses via synergistic immuno-photothermal effects

BackgroundT cell-based immunotherapies are facing great challenges in the recruitment and activation of tumor-specific T cells against solid tumors. Among which, utilizing nanobody (Nb) or nanobodies (Nbs) to construct T cell engager has emerged as a more practical potential for enhancing the anti-tumor effectiveness of T cells. Here, we designed a new Nb-guided multifunctional T cell engager (Nb-MuTE) that not only recruited effector T cells into the tumor tissues, but also efficiently activated T cells anti-tumor immunity when synergies with photothermal effect.ResultsThe Nb-MuTE, which was constructed based on an indocyanine green (ICG)-containing liposome with surface conjugation of CD105 and CD3 Nbs, and showed excellent targetability to both tumor and T cells, following enhancement of activation, proliferation and cytokine secretion of tumor-specific T cells. Notably, the immunological anti-tumor functions of Nb-MuTE-mediated T cells were further enhanced by the ICG-induced photothermal effect in vitro and in vivo.ConclusionsSuch a new platform Nb-MuTE provides a practical and “all-in-one” strategy to potentiate T cell responses for the treatment of solid tumor in clinic.

Comparison of The Effectiveness of Thymoquinone, St. John Wort Oil and Silver Sulfadiazine in Experimental Burn Wounds.

We aimed to compare the effectiveness of thymoquinone (TQ), most important bioactive component of black cumin, St. John wort (SJW) oil, a traditional medicinal plant used in burns, and silver sulfadiazine (AgSD), well-known anti-inflammatory agent used in modern medicine, in an experimental burn rat-model. 63 Wistar-Albino rats were randomly divided into 9 groups (n=7). TQ, SJW were administered topically and systemically but AgSD was applied topically. Epithelialization, inflammatory cell response, granulation tissue, vascularization, and fibrosis were evaluated. Malondialdehyde (MDA), total antioxidant status (TAS), total oxidant status (TOS), vitamin E, 8-hydroxy-deoxyguanosine (8-OHdG), coenzyme Q10 (CoQ10) were analyzed in serum. Topical TQ accelerated theepithelialization, enabled granulation, vascularization and fibrosis in wounds (P=0.001). Topical and systemic TQ increased Vitamin-E levels (P=0.003) but reduced TOS and 8-OHdG levels (P=0.001). Topical SJW reduced granulation and vascularization. Topical and systemic SJW decreased TOS, MDA and 8-OHdG levels (P=0.001), but increased TAS (P=0.001), and Vitamin-E levels (P=0.003). Topical AgSD reduced TOS, 8-OHdG, MDA levels (P=0.001). Topical and systemic TQ demonstrated significant advantages in accelerating wound healing process, while also enhancing antioxidant defenses and reducing oxidative damage. SJW oil, particularly in topical application, improved epithelialization and antioxidant status but showed less efficacy in systemic use. AgSD, while effective in reducing oxidative stress, was less successful in promoting wound healing and appeared to delay granulation and fibrosis. TQ offers superior protective and healing benefits, SJW is effective locally but less so systemically, and AgSD should be used cautiously, potentially combined with antioxidants to mitigate its negative impact on wound healing.

Regular cold shower exposure modulates humoral and cell-mediated immunity in healthy individuals

BackgroundCold hydrotherapy is an ancient practice that has recently gained scientific interest for its potential health benefits. This study explored the effects of regular cold shower exposure on immune cell function. MethodsSixty healthy Egyptian adults were randomized to take cold or hot showers daily for 90 days. Levels of immunoglobulins, cytokines, and interferon-gamma were measured in blood samples at baseline, 30, 60, and 90 days. ResultsThe cold shower group exhibited significant increases in immunoglobulin levels. Conversely, the hot shower group showed a significant decrease in IgM levels at 60 and 90 days compared to baseline, alongside nonsignificant decrease of IgG and IgA. the cold shower group demonstrated elevated levels of IL-2 and IL-4 at 90 days, indicating enhanced T-cell proliferation and humoral immunity, respectively. In contrast, the hot shower group did not exhibit significant changes in cytokine levels. There were no significant differences in IFN-γ and TNF-α levels between the groups. ConclusionsRegular cold shower exposure appears to enhance humoral and cell-mediated immunity through the upregulation of antibodies, interleukin-2, and interleukin-4. Brief cold stressors may induce physiological adaptations that prime the immune response. This accessible, sustainable lifestyle modification could potentially serve as an alternative therapy to boost immunity. Further research on larger populations is warranted to better understand the physiological effects of cold temperatures on immunity.

Proanthocyanidins-based adjuvant for enhanced immune responses of SARS-CoV-2 subunit vaccine

Aluminum-containing adjuvants have been widely used in human vaccines, such as anti-COVID-19 vaccine, to enhance antigen-specific immune responses. However, a major limitation of aluminum adjuvants in clinical application is that they typically enhance humoral immune responses but have weaker effects on cellular immune responses. Therefore, there has been a continuous and active pursuit of novel adjuvants that can address the shortcomings of traditional adjuvants and improve vaccine efficacy. In this paper, we proposed a novel vaccine-adjuvant system utilizing the convenient physical mixing of natural proanthocyanidins (PCs) and its derivative with a recombinant RBD subunit vaccine, which induced comparable humoral immune responses compared to aluminum adjuvant. In the context of cellular immunity, PCs modified with 4-bromomethyl phenylboronic acid not only enhanced the maturation and migration of dendritic cells but also improved the antigen uptake and presentation capacity of monocytes, leading to robust activation of immune cells, particularly involving CD4+ and CD8+ T cells, thereby demonstrating a clear Th1/CTL response bias. In general, PCs andtheir derivatives could be potential innovative natural adjuvants which effectively balance the Th1/Th2 immune response to recompense for the deficiency of aluminum adjuvant, and our study also offers an approach to discovering safe and efficient natural adjuvants.

The Complex Landscape of Immune Dysregulation in Multisystem Inflammatory Syndrome in Children with COVID-19

Abstract The immune responses following SARS-CoV-2 infection in children are still under investigation. While coronavirus disease 2019 (COVID-19) is usually mild in the pediatric population, some children develop severe clinical manifestations or multisystem inflammatory syndrome in children (MIS-C) after infection. MIS-C, typically emerging 2-6 weeks after SARS-CoV-2 exposure, is characterized by a hyperinflammatory response affecting multiple organs. This review aims to explore the complex landscape of immune dysregulation in MIS-C, focusing on innate, T cell-, and B cell-mediated immunity, and discusses the role of SARS-CoV-2 spike protein as a superantigen in MIS-C pathophysiology. Understanding these mechanisms is crucial for improving the management and outcomes for affected children.

Expansion of tumor-reactive CD8+ T cell clonotypes occurs in the spleen in response to immune checkpoint blockade.

Immune checkpoint blockade (ICB) enhances T cell responses against cancer, leading to long-term survival in a fraction of patients. CD8+ T cell differentiation in response to chronic antigen stimulation is highly complex, and it remains unclear precisely which T cell differentiation states at which anatomic sites are critical for the response to ICB. We identified an intermediate-exhausted population in the white pulp of the spleen that underwent substantial expansion in response to ICB and gave rise to tumor-infiltrating clonotypes. Increased systemic antigen redirected differentiation of this population toward a more circulatory exhausted KLR state, whereas a lack of cross-presented tumor antigen reduced its differentiation in the spleen. An analogous population of exhausted KLR CD8+ T cells in human blood samples exhibited diminished tumor-trafficking ability. Collectively, our data demonstrate the critical role of antigen density within the spleen for the differentiation and expansion of T cell clonotypes in response to ICB.

Oxidative Stress and Age-Related Tumors

Oxidative stress is the result of the imbalance between reactive oxygen and nitrogen species (RONS), which are produced by several endogenous and exogenous processes, and antioxidant defenses consisting of exogenous and endogenous molecules that protect biological systems from free radical toxicity. Oxidative stress is a major factor in the aging process, contributing to the accumulation of cellular damage over time. Oxidative damage to cellular biomolecules, leads to DNA alterations, lipid peroxidation, protein oxidation, and mitochondrial dysfunction resulting in cellular senescence, immune system and tissue dysfunctions, and increased susceptibility to age-related pathologies, such as inflammatory disorders, cardiovascular and neurodegenerative diseases, diabetes, and cancer. Oxidative stress-driven DNA damage and mutations, or methylation and histone modification, which alter gene expression, are key determinants of tumor initiation, angiogenesis, metastasis, and therapy resistance. Accumulation of genetic and epigenetic damage, to which oxidative stress contributes, eventually leads to unrestrained cell proliferation, the inhibition of cell differentiation, and the evasion of cell death, providing favorable conditions for tumorigenesis. Colorectal, breast, lung, prostate, and skin cancers are the most frequent aging-associated malignancies, and oxidative stress is implicated in their pathogenesis and biological behavior. Our aim is to shed light on the molecular and cellular mechanisms that link oxidative stress, aging, and cancers, highlighting the impact of both RONS and antioxidants, provided by diet and exercise, on cellular senescence, immunity, and development of an antitumor response. The dual role of ROS as physiological regulators of cell signaling responsible for cell damage and diseases, as well as its use for anti-tumor therapeutic purposes, will also be discussed. Managing oxidative stress is crucial for promoting healthy aging and reducing the risk of age-related tumors.

50 Awardee Talk: The role of non-traditional inheritance patterns among Bos indicus-Bos taurus crosses for global beef value chain considerations

Abstract Bos indicus-Bos taurus crosses have been effectively used in many production environments globally. Substantially higher amounts of hybrid vigor have been reported in these F1 crosses as compared with those involving only Bos taurus or only Bos indicus breeds. However, for many years, and in many environments, “unusual” aspects of calf growth have been documented among these crosses. Among F1 crosses, Bos indicus-sired calves (Bos indicus x Bos taurus) have birth weight 8 to 12 kg heavier than contemporary Bos taurus-sired calves (Bos taurus x Bos indicus); additionally, pronounced sexual dimorphism occurs where males may weigh 5 to 10 kg more than females from Bos indicus sires (Bos taurus dams) but 1 kg or less from Bos taurus sires (Bos indicus dams) in both natural service and embryo transfer calves. These reciprocal differences have been reported for fetal and placentome size mid gestation. Reciprocal differences in birth weight have also been reported among backcrosses and F2 calves, and also exist for longer gestation length from Bos indicus sires (Bos taurus dams). Increased birth weight and gestation length occur when more Bos indicus is present in the sire vs. the dam, and when Bos indicus influence is inherited through the sire vs. the dam. There is speculation that sex-specific genomic imprinting may be involved for reciprocal birth weight differences that are analogous to mouse interspecific crosses for litter weight. Reports of hybrid vigor retention among inter se matings after the F1 have been variable historically, and this variation may relate to how the foundational F1 parents were produced (Bos indicus- vs. Bos taurus-sired). Breed-of-origin haplotype on BTA5 has been shown to affect heifer pregnancy and rebreeding among F2 Nellore-Angus cows where Nellore-derived haplotypes had much lower performance compared with Angus-derived and hybrid haplotypes. Disproportionate calf sex has been reported among reciprocal F2 Nellore (Bos indicus)-Angus (Bos taurus) with 66.6% male calves from Nellore x Angus F1 sires. When retrospectively evaluating historical literature for reciprocal crosses, it appears that several instances occur with disproportionate calf sex percentage, and this phenomenon warrants further study. The extent that potential reciprocal differences may affect post-weaning growth and carcass traits has not been widely evaluated. Recent investigations have shown reciprocal F1 differences for some cell-mediated immune responses regarding T-lymphocyte proliferation, but no differences for humoral response. There needs to be additional research that completely characterizes the X chromosome of Bos indicus and Bos taurus origins and their roles for multiple production traits. There also needs to be more investigation of non-additive and parent-of-origin effects in genetic evaluations of Bos indicus-Bos taurus composites, particularly with desired first-generation breed percentage. Improved utilization of Bos indicus cattle and crosses will be instrumental in meeting global dietary protein demands.

201 Transcriptome analysis of bovine abomasal mucosa following experimental infection with Ostertagia ostertagi

Abstract Ostertagia ostertagi is a cattle stomach nematode that causes significant disease; however, little is known about host responses to infection. Due to the emergence of drug resistance in these parasites, there is a need for alternative treatments. To understand host responses to O. ostertagi infection in the gastric mucosa, gene expression in fundic and pyloric mucosa was compared between infected and uninfected calves for 21 d post-infection. The number of differentially expressed genes (DEGs) increased over time with more upregulated than downregulated DEGs at each time point. Fundic mucosa had more DEGs than pyloric mucosa across all time points. The largest changes occurred between d 7 to 9 and 10 post-infection. Most DEGs are associated with immunity, cellular reorganization, cell migration, and proliferation. Tuft/epithelial cell response to the infection was atypical, lacking key cytokines. Numerous genes associated with T cell exhaustion were upregulated. The data collectively indicate that O. ostertagi infection elicited host immune responses, yet also suggest parasite-induced immunosuppression is present. This may explain why cattle are slow in developing protective immunity to O. ostertagi. Understanding mechanisms of parasite evasion will facilitate the rational design of protective vaccines against complex nematode parasites.

N-Glycan Branching Regulates BTLA Opposite to PD-1 to Limit T Cell Hyperactivity Induced by Branching Deficiency.

N-glycan branching is a potent and multifaceted negative regulator of proinflammatory T cell and B cell function. By promoting multivalent galectin-glycoprotein lattice formation at the cell surface, branching regulates clustering and/or endocytosis of the TCR complex (TCR+CD4/CD8), CD45, CD25, BCR, TLR2 and TLR4 to inhibit T cell and B cell activation/proliferation and proinflammatory TH1 and TH17 over TH2 and induced T regulatory cell responses. In addition, branching promotes cell surface retention of the growth inhibitory receptor CTLA-4. However, the role of N-glycan branching in regulating cell surface levels of other checkpoint receptors such as BTLA (B and T lymphocyte attenuator) and PD-1 (programmed cell death protein 1) is unknown. In this study, we report that whereas branching significantly enhances PD-1 cell surface expression by reducing loss from endocytosis, the opposite occurs with BTLA in both T cells and B cells. T cell hyperactivity induced by branching deficiency was opposed by BTLA ligation proportional to increased BTLA expression. Other members of the BTLA/HVEM (herpesvirus entry mediator) signaling axis in T cells, including HVEM, LIGHT, and CD160, are largely unaltered by branching. Thus, branching-mediated endocytosis of BTLA is opposite of branching-induced inhibition of PD-1 endocytosis. In this manner, branching deficiency-induced upregulation of BTLA appears to serve as a checkpoint to limit extreme T cell hyperactivity and proinflammatory outcomes in T cells with low branching.

Autoregulated splicing of TRA2β programs T cell fate in response to antigen-receptor stimulation.

T cell receptor (TCR) sensitivity to peptide-major histocompatibility complex (MHC) dictates T cell fate. Canonical models of TCR sensitivity cannot be fully explained by transcriptional regulation. In this work, we identify a posttranscriptional regulatory mechanism of TCR sensitivity that guides alternative splicing of TCR signaling transcripts through an evolutionarily ultraconserved poison exon (PE) in the RNA-binding protein (RBP) TRA2β in mouse and human. TRA2β-PE splicing, seen during cancer and infection, was required for TCR-induced effector T cell expansion and function. Tra2β-PE skipping enhanced T cell response to antigen by increasing TCR sensitivity. As antigen levels decreased, Tra2β-PE reinclusion allowed T cell survival. Finally, we found that TRA2β-PE was first included in the genome of jawed vertebrates that were capable of TCR gene rearrangements. We propose that TRA2β-PE splicing acts as a gatekeeper of TCR sensitivity to shape T cell fate.