Stimulate Cytokine Production Research Articles

Unique Interplay Between Antinuclear Antibodies and Nuclear Molecules in the Pathogenesis of Systemic Lupus Erythematosus.

Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease that primarily affects young women and causes a wide range of inflammatory manifestations. The hallmark of SLE is the production of antibodies to components of the cell nucleus (antinuclear antibodies [ANAs]). These antibodies can bind to DNA, RNA, and protein complexes with nucleic acids. Among ANAs, antibodies to DNA (anti-DNA) are markers for classification and disease activity, waxing and waning disease activity in many patients. In the blood, anti-DNA antibodies can bind to DNA to form immune complexes with two distinct roles in pathogenesis: (1) renal deposition to provoke nephritis and (2) stimulation of cytokine production following uptake into innate immune cells and interaction with internal nucleic acid sensors. These sensors are part of an internal host defense system in the cell cytoplasm that can respond to DNA from infecting organisms; during cell stress, DNA from nuclear and mitochondrial sources can also trigger these sensors. The formation of immune complexes requires a source of extracellular DNA in an immunologically accessible form. As shown in in vivo and in vitro systems, extracellular DNA can emerge from dead and dying cells in both a free and a particulate form. Neutrophils undergoing the process of NETosis can release DNA in mesh-like structures called neutrophil extracellular traps. In SLE, therefore, the combination of ANAs and immunologically active DNA can create new structures that can promote inflammation throughout the body as well as drive organ inflammation and damage.

Deletion of codY impairs Staphylococcus epidermidis biofilm formation, generation of viable but non-culturable cells and stimulates cytokine production in human macrophages.

Introduction. Staphylococcus epidermidis biofilms are one of the major causes of bloodstream infections related to the use of medical devices. The diagnosis of these infections is challenging, delaying their treatment and resulting in increased morbidity and mortality rates. As such, it is urgent to characterize the mechanisms employed by this bacterium to endure antibiotic treatments and the response of the host immune system, to develop more effective therapeutic strategies. In several bacterial species, the gene codY was shown to encode a protein that regulates the expression of genes involved in biofilm formation and immune evasion. Additionally, in a previous study, our group generated evidence indicating that codY is involved in the emergence of viable but non-culturable (VBNC) cells in S. epidermidis.Gap statement/Hypothesis. As such, we hypothesized that the gene codY has have an important role in this bacterium virulence.Aim. This study aimed to assess, for the first time, the impact of the deletion of the gene codY in S. epidermidis virulence, namely, in antibiotic susceptibility, biofilm formation, VBNC state emergence and in vitro host immune system response.Methodology. Using an allelic replacement strategy, we constructed and then characterized an S. epidermidis strain lacking codY, in regards to biofilm and VBNC cell formation, susceptibility to antibiotics as well as their role in the interaction with human blood and plasma. Additionally, we investigate whether the codY gene can impact the activation of innate immune cells by evaluating the production of both pro- and anti-inflammatory cytokines by THP-1 macrophages.Results. We demonstrated that the deletion of the gene codY resulted in biofilms with less c.f.u. counts and fewer VBNC cells. Furthermore, we show that although WT and mutant cells were similarly internalized in vitro by human macrophages, a stronger cytokine response was elicited by the mutant in a toll-like receptor 4-dependent manner.Conclusion. Our results indicate that codY contributes to S. epidermidis virulence, which in turn may have an impact on our ability to manage the biofilm-associated infections caused by this bacterium.

Immunometabolic adaptation in monocytes underpins functional changes during pregnancy

Metabolic heterogeneity is a determinant of immune cell function. The normal physiological metabolic reprogramming of pregnancy that ensures the fuel requirements of mother and baby are met, might also underpin changes in immunity that occur with pregnancy and manifest as altered responses to pathogens and changes to autoimmune disease symptoms. Using peripheral blood from pregnant women at term, we reveal that monocytes lose M2-like and gain M1-like properties accompanied by reductions in mitochondrial mass, maximal respiration, and cardiolipin content in pregnancy; glycolysis is unperturbed. We establish that muramyl dipeptide (MDP)-stimulated cytokine production relies on oxidative metabolism, then show in pregnancy reduced cytokine production in response to MDP but not LPS. Overall, mitochondrially centered metabolic capabilities of late gestation monocytes are down-regulated revealing natural plasticity in monocyte phenotype and function that could reveal targets for improving pregnancy outcomes but also yield alternative therapeutic approaches to diverse metabolic and/or immune-mediated diseases beyond pregnancy.

Anti-PD-1 therapy triggers Tfh cell-dependent IL-4 release to boost CD8 T cell responses in tumor-draining lymph nodes.

Anti-PD-1 therapy targets intratumoral CD8+ T cells to promote clinical responses in cancer patients. Recent evidence suggests an additional activity in the periphery, but the underlying mechanism is unclear. Here, we show that anti-PD-1 mAb enhances CD8+ T cell responses in tumor-draining lymph nodes by stimulating cytokine production in follicular helper T cells (Tfh). In two different models, anti-PD-1 mAb increased the activation and proliferation of tumor-specific T cells in lymph nodes. Surprisingly, anti-PD-1 mAb did not primarily target CD8+ T cells but instead stimulated IL-4 production by Tfh cells, the major population bound by anti-PD-1 mAb. Blocking IL-4 or inhibiting the Tfh master transcription factor BCL6 abrogated anti-PD-1 mAb activity in lymph nodes while injection of IL-4 complexes was sufficient to recapitulate anti-PD-1 mAb activity. A similar mechanism was observed in a vaccine model. Finally, nivolumab also boosted human Tfh cells in humanized mice. We propose that Tfh cells and IL-4 play a key role in the peripheral activity of anti-PD-1 mAb.

Sciadonic acid ameliorates cyclophosphamide-induced immunosuppression by modulating the immune response and altering the gut microbiota.

Cyclophosphamide (Cy) is a frequently used chemotherapeutic drug, but long-term Cy treatment can cause immunosuppression and intestinal mucosal damage. The intestinal mucosal barrier and gut flora play important roles in regulating host metabolism, maintaining physiological functions and protecting immune homeostasis. Dysbiosis of the intestinal flora affects the development of the intestinal microenvironment, as well as the development of various external systemic diseases and metabolic syndrome. The present study investigated the influence of sciadonic acid (SA) on Cy-induced immunosuppression in mice. The results showed that SA gavage significantly alleviated Cy-induced immune damage by improving the immune system organ index, immune response and oxidative stress. Moreover, SA restored intestinal morphology, improved villus integrity and activated the nuclear factor κB signaling pathway, stimulated cytokine production, and reduced serum lipopolysaccharide (LPS) levels. Furthermore, gut microbiota analysis indicated that SA increased t beneficial bacteria (Alistipes, Lachnospiraceae_NK4A136_group, Rikenella and Odoribacter) and decreased pathogenic bacteria (norank-f-Oscillospiraceae, Ruminococcus and Desulfovibrio) to maintain intestinal homeostasis. The present study provided new insights into the SA regulation of intestinal flora to enhance immune responses. © 2024 Society of Chemical Industry.

Intermediate monocytes are associated with the first major adverse cardiovascular event in patients with stable coronary artery disease

BackgroundTraditional risk stratification modestly predicts adverse cardiovascular events in patients with coronary artery disease (CAD). Our aim was to investigate the association between monocyte subsets numbers and function, and the first major adverse cardiovascular event (MACE) in patients with symptomatic stable CAD and angiographically documented coronary atherosclerosis. MethodsPatients with stable CAD were screened for inclusion. Using flow cytometry, we identified classical, intermediate, and non-classical monocyte subsets and we assessed cytokine production capacity after ex-vivo stimulation of peripheral blood mononuclear cells. Clinical follow-up was performed after four years. The endpoint was the composite of cardiovascular death, acute myocardial infarction, and ischemic stroke. ResultsA cohort of 229 patients was recruited. The percentage of intermediate monocytes was positively associated with adverse cardiovascular events at follow-up (HR 1.09; 95%CI 1.02–1.16; p = 0.006), while the percentage of classical monocytes was identified as a protective factor for adverse outcomes (HR 0.96; 95%CI 0.94–0.99; p = 0.02). The percentage of intermediate monocytes remained independently associated with outcomes after adjusting for age, systolic blood pressure, and left ventricular ejection fraction (HR 1.07; 95% CI 1.01–1.14; p = 0.04). Several correlations were identified between monocyte subsets and stimulated cytokine production, but cytokine production capacity was not associated with adverse outcomes. ConclusionsIn patients with stable CAD, intermediate monocytes were associated with MACE at follow-up. The association was not due to an increased cytokine production capacity. Novel biomarkers could improve risk stratification in patients with stable CAD and could represent new pharmacological targets against atherosclerosis progression.

Saturated fatty acids stimulate cytokine production in tanycytes via the PP2Ac-dependent signaling pathway

The hypothalamic tanycytes are crucial for free fatty acids (FFAs) detection, storage, and transport within the central nervous system. They have been shown to effectively respond to fluctuations in circulating FFAs, thereby regulating energy homeostasis. However, the precise molecular mechanisms by which tanycytes modulate lipid utilization remain unclear. Here, we report that the catalytic subunit of protein phosphatase 2 A (PP2Ac), a serine/threonine phosphatase, is expressed in tanycytes and its accumulation and activation occur in response to high-fat diet consumption. In vitro, tanycytic PP2Ac responds to palmitic acid (PA) exposure and accumulates and is activated at an early stage in an AMPK-dependent manner. Furthermore, activated PP2Ac boosts hypoxia-inducible factor-1α (HIF-1α) accumulation, resulting in upregulation of an array of cytokines. Pretreatment with a PP2Ac inhibitor, LB100, prevented the PA-induced elevation of vascular endothelial growth factor (VEGF), fibroblast growth factor 1 (FGF1), hepatocyte growth factor (HGF), and dipeptidyl peptidase IV (DPPIV or CD26). Our results disclose a mechanism of lipid metabolism in tanycytes that involves the activation of PP2Ac and highlight the physiological significance of PP2Ac in hypothalamic tanycytes in response to overnutrition and efficacious treatment of obesity.

Hemalin vaccination modulates the host immune response and reproductive cycle of Haemaphysalis longicornis

Haemaphysalis longicornis can transmit high varieties of tick-borne pathogens (TBPs), and a primary strategy for preventing the transmission of those TBPs is to control ticks. Hemalin, a thrombin inhibitor of the Kunitz-type family and a crucial component in H. longicornis feeding process has been isolated from parthenogentic ticks. This study aimed to evaluate the validity of a recombinant Hemalin (rHlHemalin) vaccination as an anti-tick vaccine against H. longicornis in rabbits to find a new candidate for an effective tick control. In this study, mouse splenocytes were isolated and used to investigate immune responses after rHlHemalin stimulation. The rabbits were vaccinated with the rHlHemalin protein. After tick challenges, body weight at engorgement, egg mass, and the reproductive cycle of H. longicornis were evaluated. To confirm the vaccination, the passive immunization tests of α-rHlHemalin sera were performed. The results showed that the rHlHemalin protein could stimulate cytokine production in mouse splenocytes. Vaccination assay revealed that the periods from tick infestations to egg-hatch in the vaccination group were significantly longer than those in the phosphate buffer saline (PBS) group (P = 0.0003). In addition, the tick body weight at engorgement (P = 0.0019) and egg mass at 10 days after oviposition (P = 0.0232) were higher than those in the PBS group. These findings were consistent with the current passive immunization results and suggest rHlHemalin vaccination extended the reproductive cycle in H. longicornis but did not decrease the body weight at engorgement or weight of egg mass. Therefore, it is debatable whether Hemalin vaccination is highly-effective anti-tick vaccine or not. However, due to the importance of thrombin inhibitors in tick blood feeding and blood digestion, additional inhibitor-based vaccines should be developed aiming to find an effective and environmentally friendly biological strategy to combat ticks.

Immune-Boosting Potentiating Properties of Brassica nigra Hydroalcoholic Extract in Cyclophosphamide-Induced Immunosuppression in Rats.

The antioxidative and immune-boosting properties of the hydroalcoholic extract of Brassica nigra sprouts in cyclophosphamide-induced immunosuppression in rats were investigated in this study. B. nigra sprouts were prepared in the lab to monitor the bio-changes in bioactive compounds during the sprouting period up to 7 days at 17 ± 1 °C and 90% relative humidity. The total phenolic content (TPC), antioxidant activity (AOA), total flavonoids (TFs), total flavonols (TFLs), and total carotenoids (TCs) were evaluated. Consequently, the identification and quantification of phenolic acids, their derivatives, and flavonoids were carried out using HPLC. Subsequently, the selected BN sprout (6-day-old sprout) was biologically examined, and oxidative stress biomarkers, hematological parameters, immunoglobulins (Igs), and pro-inflammatory and anti-inflammatory cytokines were investigated. An increase in TPC, AOA, TFs, TFLs, and TCs was observed by increasing the sprouting time. The HPLC analysis indicated that the B. nigra seeds contained 10 phenolic acids and 4 flavonoids, predominantly syringic acid and quercetin, respectively. After 3 days, the number of phenolic acids increased to 16, predominantly syringic acid, and the number of flavonoids increased to 7, predominantly quercetin. On the 6th day, 13 phenolic acids were estimated, with the highest being benzoic acid, and 6 flavonoids were estimated, with the highest being quercetin. The greatest rise in phenols was seen on the sixth day of sprouting. These included caffeic acid, chlorogenic acid, cinnamic acid, ferulic acid, coumaric acid, benzoic acid, and rosmarinic acid. Flavonoids such as kaempferol and myricetin increased. The sprouts on day 6 were recorded as having the highest bioactive compounds and AOA content. The selected B. nigra sprouts were examined for antioxidative and immunomodulatory properties in a rat model. Dosing 250 and 500 mg kg-1, the rats exhibited significant improvements in terms of antioxidative stress and the number of white blood cells (WBCs), lymphocytes, and neutrophils in the blood, indicating stimulation of the immune response in a dose-dependent manner. In addition, the production of immune proteins, such as IgG, IgM, and IgA, was enhanced in the blood. Moreover, the 500 mg kg-1 concentration of BN extract stimulated cytokine production in a stronger manner than the 250 mg kg-1 concentration, indicating that the extract significantly increased immune activity. In conclusion, the results indicate that mustard seed extracts have immunosuppressive properties against cyclophosphamide-induced immunosuppression in rats.

Neutrophil-Mediated Progression of Mild Cognitive Impairment to Dementia.

Cognitive impairment is a serious condition that begins with amnesia and progresses to cognitive decline, behavioral dysfunction, and neuropsychiatric impairment. In the final stage, dysphagia and incontinence occur. There are numerous studies and developed drugs for cognitive dysfunction in neurodegenerative diseases, such as Alzheimer's disease (AD); however, their clinical effectiveness remains equivocal. To date, attempts have been made to overcome cognitive dysfunction and understand and delay the aging processes that lead to degenerative and chronic diseases. Cognitive dysfunction is involved in aging and the disruption of inflammation and innate immunity. Recent reports have indicated that the innate immune system is prevalent in patients with AD, and that peripheral neutrophil markers can predict a decline in executive function in patients with mild cognitive impairment (MCI). Furthermore, altered levels of pro-inflammatory interleukins have been reported in MCI, which have been suggested to play a role in the peripheral immune system during the process from early MCI to dementia. Neutrophils are the first responders of the innate immune system. Neutrophils eliminate harmful cellular debris via phagocytosis, secrete inflammatory factors to activate host defense systems, stimulate cytokine production, kill pathogens, and regulate extracellular proteases and inhibitors. This review investigated and summarized the regulation of neutrophil function during cognitive impairment caused by various degenerative diseases. In addition, this work elucidates the cellular mechanism of neutrophils in cognitive impairment and what is currently known about the effects of activated neutrophils on cognitive decline.

Bacterial lipopolysaccharide modulates immune response in the colorectal tumor microenvironment

Immune responses can have opposing effects in colorectal cancer (CRC), the balance of which may determine whether a cancer regresses, progresses, or potentially metastasizes. These effects are evident in CRC consensus molecular subtypes (CMS) where both CMS1 and CMS4 contain immune infiltrates yet have opposing prognoses. The microbiome has previously been associated with CRC and immune response in CRC but has largely been ignored in the CRC subtype discussion. We used CMS subtyping on surgical resections from patients and aimed to determine the contributions of the microbiome to the pleiotropic effects evident in immune-infiltrated subtypes. We integrated host gene-expression and meta-transcriptomic data to determine the link between immune characteristics and microbiome contributions in these subtypes and identified lipopolysaccharide (LPS) binding as a potential functional mechanism. We identified candidate bacteria with LPS properties that could affect immune response, and tested the effects of their LPS on cytokine production of peripheral blood mononuclear cells (PBMCs). We focused on Fusobacterium periodonticum and Bacteroides fragilis in CMS1, and Porphyromonas asaccharolytica in CMS4. Treatment of PBMCs with LPS isolated from these bacteria showed that F. periodonticum stimulates cytokine production in PBMCs while both B. fragilis and P. asaccharolytica had an inhibitory effect. Furthermore, LPS from the latter two species can inhibit the immunogenic properties of F. periodonticum LPS when co-incubated with PBMCs. We propose that different microbes in the CRC tumor microenvironment can alter the local immune activity, with important implications for prognosis and treatment response.

9-Hydroxyoctadecadienoic acid plays a crucial role in human skin photoaging

Human skin is regularly exposed to ultraviolet (UV) rays from sunlight, leading to photoaging, which differs from intrinsic aging. Although the acute effects of UV exposure have been extensively studied, limited research has addressed the long-term consequences of chronic UV exposure. This study aimed to investigate the underlying causes of chronic photoaging. A questionnaire-based assessment of sunlight exposure was conducted among volunteers in their 20s and 50s, and the stratum corneum of their skin was analyzed for bioactive lipid content. Volunteers were categorized into low and high UV exposure groups based on the questionnaire scores. The analysis results revealed a significant increase in 9-hydroxyoctadecadienoic acid (9-HODE) levels in the skin of individuals in their 50s with high UV exposure. However, UV exposure did not affect 9-HODE levels in the skin of individuals in their 20s. In vitro experiments further indicated that 9-HODE contributes to chronic inflammation, pigmentary changes, and extracellular matrix alterations during photoaging. Specifically, 9-HODE stimulated cytokine production [interleukin-6 (IL6), IL8, and granulocyte-macrophage colony-stimulating factor (GM-CSF)] and reduced dickkopf-1 (DKK1) production in keratinocytes. In fibroblasts, 9-HODE stimulated matrix metalloproteinase-1 (MMP1) and MMP3 production while reducing collagen I (COL1) production. The expression of G2A, the receptor for 9-HODE, was also confirmed in fibroblasts, suggesting that 9-HODE exerts its effects via G2A, as observed in keratinocytes. Overall, these findings indicate that 9-HODE is a mediator of chronic photoaging and highlight its potential significance in photoaging prevention.

Exploring redox imbalance and inflammation for asthma therapy.

Asthma is a prolonged inflammatory disorder of the airways, that affects an estimated 300 million people worldwide. Asthma is triggered by numerous endogenous and exogenous stimuli with symptoms like wheezing, cough, short of breath, chest tightening, airway obstruction, and hyperreactivity observed in patients. The review seeks to identify targets of redox imbalance and inflammation that could be explored to create effective treatments for asthma. Themethodology involved a search and review of literature relating to asthma pathogenesis, redox homeostasis, and inflammation. Eosinophils and neutrophils are involved in asthma pathogenesis. These inflammatory cells generate high levels of endogenous oxidants such as hydrogen peroxide and superoxide, which could result in redox imbalance in the airways of asthmatics. Redox imbalance occurs when the antioxidant systems becomes overwhelmed resulting in oxidative stress. Oxidative stress and inflammation have been linked with asthma inflammation and severity. Reactive oxygen species (ROS)/reactive nitrogen species (RNS) cause lung inflammation by activating nuclear factor kappa-B (NF-κB), mitogen-activated protein kinase (MAPK), activator protein-1, as well as additional transcription factors. These factors stimulate cytokine production which ultimately activates inflammatory cells in the bronchi, causing lung cellular injury and destruction. ROS/RNS is also produced by these inflammatory cells to eradicate invading bacteria. Antioxidant treatments for asthma have not yet been fully explored. Redox and inflammatory processes are viable targets that could be explored to create better therapy for asthma.

Effect of mevalonate, zoledronate and BCG on monocyte/macrophage phenotype

Cells of innate immunity, mainly monocytes/macrophages, form a long-term nonspecific immunological memory during the initial encounter with the pathogen, the so-called trained immunity. Mevalonate pathway metabolites play an important role in the formation of trained immunity. The aim of this investigation was to study the effect of modulators of mevalonate pathway, mevalonate and zoledronate, on the formation of trained immunity in human and animal monocytes/ macrophages.Material and methods. Human monocyte-like cell lines THP-1 and U-937, peritoneal macrophages of BALB/c mice were used. Trained immunity was induced in vitro by incubation of THP-1 and U-937 monocyte-like cell lines for 24 and 72 hours with inactivated Mycobacteria of BCG vaccine strain, and in vivo by intraperitoneal administration of BCG to BALB/c mice with isolation of peritoneal macrophages on day 7 after infection (lag phase). Cell hyperreactivity was assessed by response to a second stimulus with bacterial lipopolysaccharide (LPS) and mevalonate, zoledranate in the presence or absence of LPS. Lactate, cytokine (IL-1β, TNF-α, IL-10), nitric oxide and glucose level was measured in conditioned media from cells.Results and discussion. The study showed that monocyte-like cell lines THP-1 and U-937 responded differently by cytokine production, lactate, and glucose consumption to BCG stimulus in the presence or absence of lag phase. Mevalonate and zoledronate alone or in combination with LPS also stimulated cytokine production in different ways. The presence of lag phase for human monocyte-like cells is essential for the level of cytokine production and glucose consumption. Peritoneal macrophages have been shown to enhance pro-inflammatory cytokine production in response to LPS, mevalonate, and zoledronate.Conclusions. Mevalonate and zoledronate induce trained immunity in monocytes/macrophages.

Identification of Salmonella Pullorum Factors Affecting Immune Reaction in Macrophages from the Avian Host.

The host-specific Salmonella serovar S. Pullorum (SP) modulates the chicken immune response to a Th2-biased response associated with persistent infection. This is different from the Th1-biased immune response induced by the genetically close serovar, S. Enteritidis (SE). Based on core genome differences between SP and SE, we used three complementary bioinformatics approaches to identify SP genes, which may be important for stimulation of the immune response. Defined mutants were constructed in selected genes, and the infection potential and ability of mutants to stimulate cytokine production in avian derived HD11 macrophages were determined. Deletion of large genomic regions unique to SP did not change infection potential nor immune stimulation significantly. Mutants in genes with conserved single nucleotide polymorphisms (SNPs) between the two serovars in the region 100 bp upstream of the start codon (conserved upstream SNPs [CuSNPs]) such as sseE, osmB, tolQ, a putative immune antigen, and a putative persistent infection factor, exhibited differences in induction of inflammatory cytokines compared to wild-type SP, suggesting a possible role of these CuSNPs in immune regulation. Single nucleotide SP mutants correcting for the CuSNP difference were constructed in the upstream region of sifA and pipA. The SNP corrected pipA mutant expressed pipA at a higher level than the wild-type SP strain, and the mutant differentially caused upregulation of proinflammatory cytokines. It suggests that this CuSNP is important for the suppression of proinflammatory responses. In conclusion, this study has identified putative immune stimulating factors of relevance to the difference in infection dynamics between SP and SE in avian macrophages. IMPORTANCE Salmonella Pullorum is host specific to avian species, where it causes life-threatening infection in young birds. It is unknown why it is host restricted and causes systemic disease, rather than gastroenteritis normally seen with Salmonella. In the present study, we identified genes and single nucleotide polymorphisms (SNPs; relative to the broad-host-range type Salmonella Enteritidis), which affected survival and immune induction in macrophages from hens suggesting a role in development of the host specific infection. Further studies of such genes may enable understanding of which genetic factors determine the development of host specific infection by S. Pullorum. In this study, we developed an in silico approach to predict candidate genes and SNPs for development of the host-specific infection and the specific induction of immunity associated with this infection. This study flow can be used in similar studies in other clades of bacteria.

Antioxidant, Immunostimulatory, and Anticancer Properties of Hydrolyzed Wheat Bran Mediated through Macrophages Stimulation.

Previous studies demonstrated that enzymatic hydrolysis enhances wheat bran (WB) biological properties. This study evaluated the immunostimulatory effect of a WB hydrolysate (HYD) and a mousse enriched with HYD (MH) before and after in vitro digestion on murine and human macrophages. The antiproliferative activity of the harvested macrophage supernatant on colorectal cancer (CRC) cells was also analyzed. MH showed significantly higher content than control mousse (M) in soluble poly- and oligosaccharides (OLSC), as well as total soluble phenolic compounds (TSPC). Although in vitro gastrointestinal digestion slightly reduced the TSPC bioaccessibility of MH, ferulic acid (FA) levels remained stable. HYD showed the highest antioxidant activity followed by MH, which demonstrated a greater antioxidant activity before and after digestion as compared with M. RAW264.7 and THP-1 cells released the highest amounts of pro-inflammatory cytokines after being treated with 0.5 mg/mL of digested WB samples. Treatment with digested HYD-stimulated RAW264.7 supernatant for 96 h showed the most anticancer effect, and spent medium reduced cancer cell colonies more than direct WB sample treatments. Although a lack of inner mitochondrial membrane potential alteration was found, increased Bax:Bcl-2 ratio and caspase-3 expression suggested activation of the mitochondrial apoptotic pathway when CRC cells were treated with macrophage supernatants. Intracellular reactive oxygen species (ROS) were positively correlated with the cell viability in CRC cells exposed to RAW264.7 supernatants (r = 0.78, p < 0.05) but was not correlated in CRC cells treated with THP-1 conditioned media. Supernatant from WB-stimulated THP-1 cells may be able to stimulate ROS production in HT-29 cells, leading to a decrease of viable cells in a time-dependent manner. Therefore, our present study revealed a novel anti-tumour mechanism of HYD through the stimulation of cytokine production in macrophages and the indirect inhibition of cell proliferation, colony formation, and activation of pro-apoptotic proteins expression in CRC cells.

Elevated Interleukin-37 Associated with Dengue Viral Load in Patients with Dengue Fever

Dengue remains a public health issue worldwide. Similar to chronic infectious diseases, stimulation of cytokine production is not enough to drive immune effector cells for effective virus clearance. One possible mechanism is the virus induces a large number of negative stimulatory cytokines inhibiting immune response. Interleukin 37 (IL-37) plays a crucial regulatory role in infection and immunity, inhibits innate and adaptive immunity as an anti-inflammatory cytokine by inhibiting proinflammatory mediators and pathways. To date, there are few studies reporting correlations between dengue fever (DF) and IL-37. In this study we found that the serum IL-37b and IL-37b-producing monocytes in patients were significantly increased in DF patients. A majority of the IL-37b produced by DF patients was produced by monocytes, not lymphocytes. Increased levels of IL-6, IL-10, and IFN-α were also found in DF patients. However, we failed to detect IL-1β, IL-17A and TNF-α in plasma, because of off-target. In our study, there was no relation between IL-6, IL-10, and IFN-α expressions and IL-37b in serum (P > 0.05). The IL-37b-producing monocytes were negatively correlated with the level of IFN-α in serum and platelet count, and positively correlated with lymphocytes percentage (P < 0.05, respectively). Additionally, serum DENV nonstructural protein 1 levels were positively correlated with monocytes percentages (P < 0.05). Our data represents findings for IL-37b expression and its potential mechanisms in DF patients’ immune response.

Abstract 1624: eIF4E allosteric regulators cause rapid commitment to apoptosis in cancer cells while sparing immune cells

Abstract Advanced or metastatic breast cancers represents a large patient population. Targeted therapies following hormonal or chemotherapeutic approaches offer some benefit. Subsequently, molecular profiling has identified over 100 mutations that render inhibitor resistance, creating challenges for physicians, patients, and the healthcare system broadly. While advances in therapeutic options have improved treatment outcomes by prolonging PFS with better QOL, long-term solutions remain a challenge for the majority of patients. The aim of our work is to drive strong mechanistic responses in this large patient population by developing targeted therapies to a key fundamental mechanism at the convergence of many oncogenic signaling pathways through which resistance arises. Our work in established cell lines and primary patient derived organoids demonstrated potent and complete responses in breast cancers that represent patient populations where unmet needs remain high including ER positive, TNBC, PTEN and RB null molecular profiles. The regulation of CAP dependent protein translation is a central element in oncogenesis. One of a family of CAP binding proteins, eIF4E serves to anchor the m7GTP CAP of RNA into several protein complexes and regulates multiple aspects of mRNA processing such as nuclear export and ribosomal translation. Previous communications revealed our eIF4E regulators potently cause cell death in primary breast cancer organoid models differentially to models derived from healthy tissue. We have expanded our efforts following our observations of both rapid apoptosis and differential sensitivity of healthy cells. In vitro, in breast cancer cells we observed stable responses in mechanistic markers of pharmacodynamic response, including phospho-Rb, with short term drug incubation. In vivo, in a ZR-75-1 breast CDX model, intermittent dosing (QW) of our lead compound resulted in strong tumor growth inhibition (&amp;gt;95%). Consistent with growth impacts, we observed stable pharmacodynamic marker (p-Rb) changes in tumor tissue at 24 and 48 hours. To understand the impact on immune cells, we evaluated our regulators against primary cells including PBMCs, CD4+ T cells, and bone marrow mononuclear cells. Our eIF4E regulators did not impair survival of these cells up to a maximum tested dose of 25 micromolar. Primary CD4+ T cells were also functional in the presence of eIF4E regulators up to 5 micromolar, which included assessments of CD3/CD28 stimulated cytokine production and proliferative responses. Our findings suggest that allosteric modulation of eIF4E provides a unique and efficient way to address multiple resistance mechanisms while sparing normal immune cellular function. Thus, our allosteric eIF4E regulators represent a potential beneficial therapeutic approach to address multiple resistant cancer patient populations and fulfill the promise of this elusive target. Citation Format: Lisa-Marie Sturla, Xuemei Zhang, Katherine Bowdish, Susan E. Alters, Christopher VanDeusen. eIF4E allosteric regulators cause rapid commitment to apoptosis in cancer cells while sparing immune cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1624.

High Bacillary Burden and the ESX-1 Type VII Secretion System Promote MHC Class I Presentation by Mycobacterium tuberculosis-Infected Macrophages to CD8 T Cells.

We used a mouse model to study how Mycobacterium tuberculosis subverts host defenses to persist in macrophages despite immune pressure. CD4 T cells can recognize macrophages infected with a single bacillus invitro. Under identical conditions, CD8 T cells inefficiently recognize infected macrophages and fail to restrict M. tuberculosis growth, although they can inhibit M. tuberculosis growth during high-burden intracellular infection. We show that high intracellular M. tuberculosis numbers cause macrophage death, leading other macrophages to scavenge cellular debris and cross-present the TB10.4 Ag to CD8 T cells. Presentation by infected macrophages requires M. tuberculosis to have a functional ESX-1 type VII secretion system. These data indicate that phagosomal membrane damage and cell death promote MHC class I presentation of the immunodominant Ag TB10.4 by macrophages. Although this mode of Ag presentation stimulates cytokine production that we presume would be host beneficial, killing of uninfected cells could worsen immunopathology. We suggest that shifting the focus of CD8 T cell recognition to uninfected macrophages would limit the interaction of CD8 T cells with infected macrophages and impair CD8 T cell-mediated resolution of tuberculosis.

CCL26 in primary biliary cholangitis - Is it a novel disease mediator?

To verify the role of CX3C chemokine ligand 1 - CX3C chemokine receptor 1 (CX3CL1-CX3CR1) pathway in the pathogenesis of primary biliary cholangitis (PBC). To explore whether CCL26, a novel functional ligand to CX3CR1, participates in the immunological mechanism of PBC. Fifty-nine PBC patients and 54 healthy controls were recruited. Enzyme-linked immunosorbent assay and flow cytometry were used to measure CX3CL1 and CCL26 concentrations in plasma and CX3CR1 expression on peripheral lymphocytes, respectively. Chemotactic effects of CX3CL1 and CCL26 toward lymphocytes were detected by Transwell cell migration assays. CX3CL1 and CCL26 expressions in liver were assessed by immunohistochemical staining. Effects of CX3CL1 and CCL26 on stimulating cytokine production from lymphocytes were evaluated using intracellular flow cytometry. Significantly elevated CX3CL1 and CCL26 plasma concentration and CX3CR1 expression on CD4+ and CD8+ T cells were noted in PBC patients. CX3CL1 exhibited chemotactic activity toward CD8+ T, natural killer (NK) and NKT cells in a dose-dependent manner while such chemotactic effects were not detected for CCL26. In PBC patients, CX3CL1 and CCL26 were both increasingly expressed in biliary tracts and a concentration gradient of CCL26 in hepatocytes around portal areas was observed. Immobilized CX3CL1 could enhance interferon-γ production from T and NK cells while such effect was not exhibited by soluble CX3CL1 or CCL26. CCL26 expression is significantly elevated in plasma and biliary duct of PBC patients, yet does not appear to attract CX3CR1-expressing immune cells. CX3CL1-CX3CR1 pathway promotes the infiltration of T, NK and NKT cells into bile ducts and forms a positive feedback loop with T-helper 1 type cytokines in PBC.