Additional use of α-IgM antibodies potentiates CpG ODN2006-induced B cell activation by targeting mainly naïve and marginal zone-like B cells.

CpG ODN2006 is widely used as a potent B cell stimulant in vitro and in vivo. However, it shows a deficit in targeting naïve B cells in vitro. In this study, we investigated whether α-IgM can support ODN2006-induced effects on B cells to obtain enhanced activation with focus on different B cell subsets. Our results delineated robust B cell activation, shown by increased activation marker expression and cytokine secretion by each agent alone, and further augmented when used in combination. Interestingly, α-IgM targeted mainly naïve and marginal zone-like B cells, thus complementing the pronounced effects of ODN2006 on memory B cells and achieving optimal activation for all B cell subsets. Taken together, combining ODN2006 and α-IgM is beneficial for in vitro activation including all B cell subsets. Furthermore, our results suggest that α-IgM could enhance efficacy of ODN2006 in vivo with further need of investigation.

SARS-CoV-2 infection induces adaptive NK cell responses by spike protein-mediated induction of HLA-E expression.

HLA-E expression plays a central role for modulation of NK cell function by interaction with inhibitory NKG2A and stimulatory NKG2C receptors on canonical and adaptive NK cells, respectively. Here, we demonstrate that infection of human primary lung tissue with SARS-CoV-2 leads to increased HLA-E expression and show that processing of the peptide YLQPRTFLL from the spike protein is primarily responsible for the strong, dose-dependent increase of HLA-E. Targeting the peptide site within the spike protein revealed that a single point mutation was sufficient to abrogate the increase in HLA-E expression. Spike-mediated induction of HLA-E differentially affected NK cell function: whereas degranulation, IFN-γ production, and target cell cytotoxicity were enhanced in NKG2C+ adaptive NK cells, effector functions were inhibited in NKG2A+ canonical NK cells. Analysis of a cohort of COVID-19 patients in the acute phase of infection revealed that adaptive NK cells were induced irrespective of the HCMV status, challenging the paradigm that adaptive NK cells are only generated during HCMV infection. During the first week of hospitalization, patients exhibited a selective increase of early NKG2C+CD57- adaptive NK cells whereas mature NKG2C+CD57+ cells remained unchanged. Further analysis of recovered patients suggested that the adaptive NK cell response is primarily driven by a wave of early adaptive NK cells during acute infection that wanes once the infection is cleared. Together, this study suggests that NK cell responses to SARS-CoV-2 infection are majorly influenced by the balance between canonical and adaptive NK cells via the HLA-E/NKG2A/C axis.

From Immunosenescence to Aging Types-Establishing Reference Intervals for Immune Age Biomarkers by Centile Estimation.

Immunological aging type definition requires establishing reference intervals from the distribution of immunosenescence biomarkers conditional on age. For 1605 individuals (18-97 years), we determined the comprehensive immune age index IMMAX from flow-cytometry-based blood cell sub-populations and identified age-specific centiles by fitting generalized additive models for location, scale, and shape. The centiles were uncorrelated with age and facilitated the categorization of individuals as immunologically slow or fast aging types. Using its 50th percentile as a reference, we rescaled the IMMAX to equivalent years of life (EYOL) and computed the immunological age gap as the difference between EYOL and chronological age. Applied to preliminary baseline and follow-up measurements from 53 participants of the Dortmund Vital Study (Clinical-Trials.gov Identifier: NCT05155397), the averaged changes in the IMMAX and EYOL conformed to the 5-year follow-up period, whereas no significant changes occurred concerning IMMAX centiles and age gap. This suggested that the participants immunologically adapted to aging and kept their relative positions within the cohort. Sex was non-significant. Methodical comparisons indicated that future confirmatory analyses with the completed follow-up examinations could rely on percentile curves estimated by simple linear quantile regression, while the selection of the immunosenescence biomarker will greatly influence the outcome, with IMMAX representing the preferable choice.

Early expansion of activated adaptive but also exhausted NK cells during acute severe SARS-CoV-2 infection.

The analysis of immunological parameters during the course of a SARS-CoV-2 infection is of great importance, both to identify diagnostic markers for the risk of a severe course of COVID-19, and to better understand the role of the immune system during the infection. By using multicolor flow cytometry we compared the phenotype of Natural Killer (NK) cells from hospitalized COVID-19 patients during early SARS-CoV-2 infection with samples from recovered and SARS-CoV-2 naïve subjects. Unsupervised high-dimensional analysis of 28-color flow cytometric data revealed a strong enrichment of NKG2C expressing NK cells in response to the acute viral infection. In addition, we found an overrepresentation of highly activated NK cell subsets with an exhausted phenotype. Moreover, our data show long-lasting phenotypic changes within the NK cell compartment that did not completely reverse up to 2 months after recovery. This demonstrates that NK cells are involved in the early innate immune response against SARS-CoV-2.

A Systematic Analysis of Biological, Sociodemographic, Psychosocial, and Lifestyle Factors Contributing to Work Ability Across the Working Life Span: Cross-sectional Study.

BACKGROUND: As employees age, their physical and mental abilities decline and work ability decreases, enhancing the risk for long-term sick leave or even premature retirement. However, the relative impact of biological and environmental determinants on work ability with increasing age is poorly understood in terms of their complexity. OBJECTIVE: Previous research has shown relationships between work ability and job and individual resources, as well as specific demographic and lifestyle-related variables. However, other potentially important predictors of work ability remain unexplored, such as personality traits and biological determinants, including cardiovascular, metabolic, immunological, and cognitive abilities or psychosocial factors. Our aim was to systematically evaluate a wide range of factors to extract the most crucial predictors of low and high work ability across the working life span. METHODS: As part of the Dortmund Vital Study, 494 participants from different occupational sectors, aged between 20 and 69 years, completed the Work Ability Index (WAI) assessing employee's mental and physical resources. A total of 30 sociodemographic variables were grouped into 4 categories (social relationships, nutrition and stimulants, education and lifestyle, and work related), and 80 biological and environmental variables were grouped into 8 domains (anthropometric, cardiovascular, metabolic, immunologic, personality, cognitive, stress related, and quality of life) and have been related to the WAI. RESULTS: Using the analyses, we extracted important sociodemographic factors influencing work ability, such as education, social activities, or sleep quality, and identified age-dependent and age-independent determinants of work ability. Regression models explained up to 52% of the WAI variance. Negative predictors of work ability were chronological and immunological age, immunological inefficiency, BMI, neuroticism, psychosocial stress, emotional exhaustion, demands from work, daily cognitive failures, subclinical depression, and burnout symptoms. Positive predictors were maximum heart rate during ergometry, normal blood pressure, hemoglobin and monocyte concentration, weekly physical activity, commitment to the company, pressure to succeed, and good quality of life. CONCLUSIONS: The identified biological and environmental risk factors allowed us to evaluate work ability in its complexity. Policy makers, employers, and occupational safety and health personnel should consider the modifiable risk factors we identified to promote healthy aging at work through focused physical, dietary, cognitive, and stress-reduced preventive programs, in addition to well-balanced working conditions. This may also increase the quality of life, commitment to the job, and motivation to succeed, which are important factors to maintain or even enhance work ability in the aging workforce and to prevent early retirement. TRIAL REGISTRATION: ClinicalTrials.gov NCT05155397; https://clinicaltrials.gov/ct2/show/NCT05155397. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): RR2-10.2196/32352.

Calibrating a Comprehensive Immune Age Metric to Analyze the Cross Sectional Age-Related Decline in Cardiorespiratory Fitness.

Cardiorespiratory fitness (CRF) is essential for sustained work ability in good health, but declines with aging, as does the functionality of the immune system, the latter process commonly referred to as immunosenescence. This study aimed to compare the capacity of immunosenescence biomarkers with chronological age for predicting low CRF in a cross-sectional sample recruited from the regional working population. CRF was determined by submaximal bicycle ergometer testing in a cross-sectional sample of 597 volunteers aged 20-70 years from the 'Dortmund Vital Study' (DVS, ClinicalTrials.gov Identifier: NCT05155397). Low CRF was scored if the ergometer test was not completed due to medical reasons or if the power output projected to a heart rate of 130 bpm divided by body mass was below sex-specific reference values of 1.25 W/kg for females and 1.5 W/kg for males, respectively. In addition to established biomarkers of immunosenescence, we calibrated a comprehensive metric of immune age to our data and compared its predictive capacity for low CRF to chronological age, while adjusting our analysis for the influence of sex, obesity, and the level of regular physical activity, by applying univariate and multiple logistic regression. While obesity, low physical activity, chronological and immune age were all associated with increased probability for low CRF in univariate analyses, multiple logistic regression revealed that obesity and physical activity together with immune age, but not chronological age, were statistically significant predictors of low CRF outcome. Sex was non-significant due to the applied sex-specific reference values. These results demonstrate that biological age assessed by our immunological metric can outperform chronological age as a predictor for CRF and indicate a potential role for immunosenescence in explaining the inter-individual variability of the age-related decline in cardiorespiratory fitness.

Human NK cells responses are enhanced by CD56 engagement.

Natural Killer (NK) cells are important innate lymphocytes for effective immune responses against intracellular pathogens and tumors. CD56 is a well-known marker for human NK cells, but there is very limited information about a functional role of this surface receptor. Here, we show that engagement of CD56 can induce NK cell activation resulting in degranulation, IFN-γ secretion and morphological changes, making CD56 a potential co-activating receptor in NK cells. Interestingly, this effect was only observed in cytokine pre-activated and not in freshly isolated human NK cells, demonstrating that NK cell reactivity upon CD56 engagement was dependent on cytokine stimulation. Inhibition of Syk, PI3K, Erk, and src-family-kinases impaired CD56-mediated NK cell stimulation. Finally, we used CRISPR/Cas9 to delete CD56 from primary human NK cells. While this abolished the stimulatory effect of CD56 on pre-activated NK cells, the cytotoxic activity of NK cells against several tumor target cells was not affected by the absence of CD56. This demonstrates that the stimulating effect of CD56 on pre-activated NK cells does not have a major impact on their cytotoxic activity, but it may contribute to the function of CD56 as a fungal recognition receptor and in the NK cell developmental synapse.

Neutralizing antibody responses 300 days after SARS-CoV-2 infection and induction of high antibody titers after vaccination.

Neutralizing antibodies against SARS-CoV-2 are important to protect against infection and/or disease. Using an assay to detect antibodies directed against the receptor binding domain (RBD) of SARS-CoV-2 Spike, we identified individuals with SARS-CoV-2 infection after an outbreak at a local health institution. All but one COVID-19 patient developed detectable anti-RBD antibodies and 77% had virus neutralizing antibody titers of >1:25. Antibody levels declined slightly over time. However, we still detected virus neutralizing antibody titers in 64% of the COVID-19 patients at >300 days after infection, demonstrating durability of neutralizing antibody levels after infection. Importantly, full COVID-19 vaccination of these individuals resulted in higher antibody titers compared to fully vaccinated individuals in the absence of prior infection. These data demonstrate long-lived antibody-mediated immunity after SARS-CoV-2 infection, and a clear benefit of two vaccine doses for recovered individuals.

Impact of Biological and Lifestyle Factors on Cognitive Aging and Work Ability in the Dortmund Vital Study: Protocol of an Interdisciplinary, Cross-sectional, and Longitudinal Study.

BACKGROUND: Previous research revealed several biological and environmental factors modulating cognitive functioning over a human's lifespan. However, the relationships and interactions between biological factors (eg, genetic polymorphisms, immunological parameters, metabolic products, or infectious diseases) and environmental factors (eg, lifestyle, physical activity, nutrition, and work type or stress at work) as well as their impact on cognitive functions across the lifespan are still poorly understood with respect to their complexity. OBJECTIVE: The goal of the Dortmund Vital Study is to validate previous hypotheses as well as generate and validate new hypotheses about the relationships among aging, working conditions, genetic makeup, stress, metabolic functions, the cardiovascular system, the immune system, and mental performance over the human lifespan with a focus on healthy working adults. The Dortmund Vital Study is a multidisciplinary study involving the Departments of Ergonomics, Immunology, Psychology and Neurosciences, and Toxicology at the Leibniz Research Centre for Working Environment and Human Factors at the Technical University of Dortmund (IfADo) in Germany, as well as several national and international partners. METHODS: The Dortmund Vital Study is designed as a combined cross-sectional and longitudinal study. Approximately 600 healthy subjects aged between 20 and 70 years will participate. A wide range of demographic, psychological, behavioral, sensory, cardiovascular, immunological, and biochemical data, a comprehensive electroencephalography (EEG)-based cognitive test battery as well as structural and functional magnetic resonance imaging (MRI) have been included in the study. RESULTS: The study was approved by the Ethics Committee of IfADo in October 2015. The baseline testing was conducted between 2016 and 2021 and will be repeated every 5 years (3 follow-up measures until 2035). As of March 2020 (until the outbreak of the COVID-19 pandemic), 593 participants have been enrolled. Some results from the cross-sectional part of the study were already published, further results will be published soon. Longitudinal data will be analyzed and published by 2025. CONCLUSIONS: We anticipate that the study will shed light on sources of interindividual differences in the alterations of cognitive functioning with increasing age and reveal biological and lifestyle markers contributing to work ability, longevity, and healthy aging on the one hand, and to risk factors for cognitive decline, mild cognitive impairment, or even dementia on the other hand. TRIAL REGISTRATION: ClinicalTrials.gov NCT05155397; https://clinicaltrials.gov/ct2/show/NCT05155397. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/32352.

Results of the Optimune trial: A randomized controlled trial evaluating a novel Internet intervention for breast cancer survivors.

INTRODUCTION: After the acute treatment phase, breast cancer patients often experience low quality of life and impaired mental health, which could potentially be improved by offering cognitive behavioural therapy (CBT) and addressing exercise and dietary habits. However, CBT and other behavioural interventions are rarely available beyond the acute treatment phase. Internet-based interventions could bridge such treatment gaps, given their flexibility and scalability. In this randomized controlled trial (RCT), we investigated the effects of such an intervention ("Optimune") over three months. METHODS: This RCT included 363 female breast cancer survivors (age range = 30-70), recruited from the community, who had completed the active treatment phase. Inclusion criteria were: breast cancer diagnosis less than 5 years ago and acute treatment completion at least 1 month ago. Participants were randomly assigned to (1) an intervention group (n = 181), in which they received care as usual (CAU) plus 12-month access to Optimune immediately after randomization, or (2) a control group (n = 182), in which they received CAU and Optimune after a delay of 3 months. Primary endpoints were quality of life (QoL), physical activity, and dietary habits at three months. We hypothesized that intervention group participants would report better QoL, more physical activity, and improved dietary habits after 3 months. RESULTS: Intention-to-treat (ITT) analyses revealed significant effects on QoL (d = 0.27, 95% CI: 0.07-0.48) and dietary habits (d = 0.36, 95% CI: 0.15-0.56), but the effect on physical exercise was not significant (d = 0.30; 95% CI: 0.10-0.51). DISCUSSION: These findings suggest the effectiveness of Optimune, a new CBT-based Internet intervention for breast cancer survivors, in facilitating improvements in quality of life and dietary habits. Efforts to disseminate this intervention more broadly may be warranted. TRIAL REGISTRATION: ClinicalTrials.gov, NCT03643640. Registered August 23rd 2018, https://clinicaltrials.gov/ct2/show/NCT03643640.

Low self-reported stress despite immune-physiological changes in paramedics during rescue operations.

Despite the high stress levels, paramedics seem to ignore or even negate the stress. This can be detrimental and lead to stress-related diseases. Therefore, we investigated the divergence between physiological and psychological stress responses of paramedics. Participants were 16 paramedics and 17 white-collar workers. We assessed psychological stress parameters, cortisol awakening response (CAR), and quantified immune parameters. In paramedics, electrocardiogram (ECG) was measured during one complete 24-hour shift. Our results revealed that CAR was higher in paramedics compared to controls. An alteration of immune parameters was observed even during days of free time. Also, ECG recordings showed acute stress in paramedics during rescue situations. Questionnaires revealed that rescue-service specific stressors affect psychological outcomes. However, paramedics reported significantly less mental stress and higher levels of depersonalization than controls. Taken together, our results suggest higher stress in paramedics compared to controls. However, paramedics negate their daily stress. Our findings underline therefore the importance to develop stress-management interventions for paramedics including sensitization for their stress reactions.

Regulation of natural killer cell activity by glucocorticoids, serotonin, dopamine, and epinephrine.

The immune system and the nervous system are highly complex organs composed of various different cells that must interact with each other for proper function of the system. This communication can be mediated by soluble factors. The factors released by the nervous system (neurotransmitters) differ from those released by the immune system (cytokines). Nevertheless, the nervous and immune systems can influence each other's activity because immune cells express neurotransmitter receptors, and neurons express cytokine receptors. Moreover, immune cells can synthesize and release neurotransmitters themselves, thus using neurotransmitter-mediated pathways via autocrine and paracrine mechanisms. Natural killer (NK) cells are innate lymphocytes that are important for early and effective immune reactions against infections and cancer. Many studies have shown the strong influence of stress and the nervous system on NK cell activity. This phenomenon may be one reason why chronic stress leads to a higher incidence of infections and cancer. Here, we review the effects of neuroendocrine factors on the different activities of NK cells. Understanding the effects of neuroendocrine factors on NK cell activities during physiological and pathophysiological conditions may result in novel therapeutic strategies to enhance NK cell functions against tumors.

Analysis of Natural Killer cell functions in patients with hereditary hemochromatosis.

Hereditary hemochromatosis (HH) is an autosomal-recessive disorder of the iron metabolism. Patients are typically affected by dysregulated iron levels, which can lead to iron accumulation within essential organs, such as liver, heart and pancreas. Furthermore, many HH patients are also afflicted by several immune defects and increased occurrence of autoimmune diseases that are linked to human homeostatic iron regulator protein (HFE) in the immune response. Here we examined immune cell phenotype and function in 21 HH patients compared to 21 healthy controls with a focus on Natural Killer (NK) cells. We observed increased basal and stimulated production of pro-inflammatory cytokines such as IL-1ß or IL-18 in HH patients compared to healthy controls. However, we did not find major changes in the phenotype, the amount or the cytotoxic function of NK cells in HH patients. Instead, our data show a general decrease in the total number of granulocytes in HH patients (2774 ± 958 per µl versus 3457 ± 1122 per µl in healthy controls). These data demonstrate that NK cells of HH patients are not significantly affected and that the patients' treatment by regular phlebotomy is sufficient to avoid systemic iron overload and its consequences to the immune system.

Activation of natural killer cells by rituximab in granulomatosis with polyangiitis.

OBJECTIVE: In the last few years, anti-CD20 antibody rituximab profoundly changed the therapeutic landscape of granulomatosis with polyangiitis (GPA). Here, we investigated whether natural killer (NK) cells may play a role in rituximab's mechanism of action in GPA. METHODS: B cell depletion, NK cell degranulation, and the expression of CD69 and CD16 on NK cells were measured in a series of in vitro experiments using peripheral blood mononuclear cells (PBMCs). In vivo activation of NK cells was investigated in patients receiving rituximab infusions. Cells were analyzed by seven-color flow cytometry. RESULTS: NK cells from GPA patients were activated by immobilized rituximab. Also soluble rituximab activated NK cells, provided that B cells were present. NK cells degranulated and expressed the activation marker CD69 while CD16 expression was decreased. This activation of NK cells by soluble rituximab was accompanied by a reduction of B cells. The next-generation anti-CD20 antibody obinutuzumab showed stronger effects compared to rituximab on both the reduction of B cells and the activation of NK cells. Finally, we found that rituximab led to the activation of NK cells in vivo, provided that B cells were not depleted due to prior rituximab infusions. CONCLUSION: B cell-bound rituximab activates NK cells in GPA. While NK cells therefore participate in rituximab's mechanism of action in humans, their potential may be more efficiently exploited, e.g., by Fc engineering of therapeutic antibodies.

NK cells switch from granzyme B to death receptor-mediated cytotoxicity during serial killing.

NK cells eliminate virus-infected and tumor cells by releasing cytotoxic granules containing granzyme B (GrzB) or by engaging death receptors that initiate caspase cascades. The orchestrated interplay between both cell death pathways remains poorly defined. Here we simultaneously measure the activities of GrzB and caspase-8 in tumor cells upon contact with human NK cells. We observed that NK cells switch from inducing a fast GrzB-mediated cell death in their first killing events to a slow death receptor-mediated killing during subsequent tumor cell encounters. Target cell contact reduced intracellular GrzB and perforin and increased surface-CD95L in NK cells over time, showing how the switch in cytotoxicity pathways is controlled. Without perforin, NK cells were unable to perform GrzB-mediated serial killing and only killed once via death receptors. In contrast, the absence of CD95 on tumor targets did not impair GrzB-mediated serial killing. This demonstrates that GrzB and death receptor-mediated cytotoxicity are differentially regulated during NK cell serial killing.

Circulating growth/differentiation factor 15 is associated with human CD56bright natural killer cell dysfunction and nosocomial infection in severe systemic inflammation.

BACKGROUND: Systemic inflammation induced by sterile or infectious insults is associated with an enhanced susceptibility to life-threatening opportunistic, mostly bacterial, infections due to unknown pathogenesis. Natural killer (NK) cells contribute to the defence against bacterial infections through the release of Interferon (IFN) γ in response to Interleukin (IL) 12. Considering the relevance of NK cells in the immune defence we investigated whether the function of NK cells is disturbed in patients suffering from serious systemic inflammation. METHODS: NK cells from severely injured patients were analysed from the first day after the initial inflammatory insult until the day of discharge in terms of IL-12 receptor signalling and IFN-γ synthesis. FINDINGS: During systemic inflammation, the expression of the IL-12 receptor ß2 chain, phosphorylation of signal transducer and activation 4, and IFN-γ production on/in NK cells was impaired upon exposure to Staphylococcus aureus. The profound suppression of NK cells developed within 24 h after the initial insult and persisted for several weeks. NK cells displayed signs of exhaustion. Extrinsic changes were mediated by the early and long-lasting presence of growth/differentiation factor (GDF) 15 in the circulation that signalled through the transforming growth factor ß receptor I and activated Smad1/5. Moreover, the concentration of GDF-15 in the serum inversely correlated with the IL-12 receptor ß2 expression on NK cells and was enhanced in patients who later acquired septic complications. INTERPRETATION: GDF-15 is associated with the development of NK cell dysfunction during systemic inflammation and might represent a novel target to prevent nosocomial infections. FUND: The study was supported by the Department of Orthopaedics and Trauma Surgery, University Hospital Essen.

LIPG-promoted lipid storage mediates adaptation to oxidative stress in breast cancer.

Endothelial lipase (LIPG) is a cell surface associated lipase that displays phospholipase A1 activity towards phosphatidylcholine present in high-density lipoproteins (HDL). LIPG was recently reported to be expressed in breast cancer and to support proliferation, tumourigenicity and metastasis. Here we show that severe oxidative stress leading to AMPK activation triggers LIPG upregulation, resulting in intracellular lipid droplet accumulation in breast cancer cells, which supports survival. Neutralizing oxidative stress abrogated LIPG upregulation and the concomitant lipid storage. In human breast cancer, high LIPG expression was observed in a limited subset of tumours and was significantly associated with shorter metastasis-free survival in node-negative, untreated patients. Moreover, expression of PLIN2 and TXNRD1 in these tumours indicated a link to lipid storage and oxidative stress. Altogether, our findings reveal a previously unrecognized role for LIPG in enabling oxidative stress-induced lipid droplet accumulation in tumour cells that protects against oxidative stress, and thus supports tumour progression.

SLAM family receptors in natural killer cells - Mediators of adhesion, activation and inhibition via cis and trans interactions.

SLAM family receptors are important for the fine-tuning of immune reactions. Their expression is restricted to cells of hematopoietic origin and most SLAM family receptors are their own ligand. Here we review how these receptors are involved in regulating the functions of Natural Killer (NK) cells. We discuss that promoting cellular adhesion may be a main function of SLAM family receptors in NK cells. The homophilic interactions of SLAM family receptors can not only occur in trans between different cells, but also in cis on the surface of the same cell. This cis interaction additionally modulates the function of the receptors and subsequently affects the activities of NK cells. Finally, SLAM-family receptors can also mediate inhibitory signals under certain conditions. These inhibitory signals can contribute to the functional maturation of NK cells during NK cell education. Therefore, SLAM family receptors are critically involved in many aspects of NK cell functionality.

Attention to Emotional Information Is Associated With Cytokine Responses to Psychological Stress.

This study aimed to investigate the impact of mental stress on salivary cytokines and attention to emotional stimuli, as well as associations between stress-induced changes of immune and cognitive parameters. In a randomized order a total of 60 young adults were assigned to one of two stress conditions with varying stress intensity. High stress was induced by a socially evaluated Paced Auditory Serial Addition Test (PASAT). As a low stress task a paper-and-pencil version of PASAT was administered. Salivary cytokines were measured before, 5 min after, and 45 min after completion of the stress task, and were assayed for pro- and anti-inflammatory cytokines. Three distinct types of attention - alerting, orienting, and executive control - were measured by the modified Emotional Attention Network Test Integration (E-ANTI). IL-1ß and IL-6 increased only in the high-stress group. Significant increases in IFN-α, IFN-γ, TNF-α, and IL-10 at 45 min after stress induction (all p's < 0.05) were observed in both the high-stress and the low-stress group. Alerting attention was positively related to more pronounced increases in IFN-α and TNF-α in both groups. Further, better orienting attention after presentation of negative cues was associated with higher increases in IFN-α, TNF-α, IL-2, IL-5, and IL-10 in both groups, and higher overall levels of IFN-α, IFN-γ, and IL-12p70 in the high-stress group. There were no systematic gender differences in cytokine responses. We conclude that attention processes modulate the increases of salivary cytokines after stress exposure, and that these effects depend on stress level, particular attention network, and stimulus valence.