Primary Infection Site as a Predictor of Sepsis Development in Emergency Department Patients.

BACKGROUND: Sepsis is a life-threatening condition but predicting its development and progression remains a challenge. OBJECTIVE: This study aimed to assess the impact of infection site on sepsis development among emergency department (ED) patients. METHODS: Data were collected from a single-center ED between January 2016 and December 2019. Patient encounters with documented infections, as defined by the Systematized Nomenclature of Medicine-Clinical Terms for upper respiratory tract (URI), lower respiratory tract (LRI), urinary tract (UTI), or skin or soft-tissue infections were included. Primary outcome was the development of sepsis or septic shock, as defined by Sepsis-1/2 criteria. Secondary outcomes included hospital disposition and length of stay, blood and urine culture positivity, antibiotic administration, vasopressor use, in-hospital mortality, and 30-day mortality. Analysis of variance and various different logistic regression approaches were used for analysis with URI used as the reference variable. RESULTS: LRI was most associated with sepsis (relative risk ratio [RRR] 5.63; 95% CI 5.07-6.24) and septic shock (RRR 21.2; 95% CI 17.99-24.98) development, as well as hospital admission rates (odds ratio [OR] 8.23; 95% CI 7.41-9.14), intensive care unit admission (OR 4.27; 95% CI 3.84-4.74), in-hospital mortality (OR 6.93; 95% CI 5.60-8.57), and 30-day mortality (OR 7.34; 95% CI 5.86-9.19). UTIs were also associated with sepsis and septic shock development, but to a lesser degree than LRI. CONCLUSIONS: Primary infection sites including LRI and UTI were significantly associated with sepsis development, hospitalization, length of stay, and mortality among patients presenting with infections in the ED.

IgE-mediated regulation of IL-10 and type I IFN enhances rhinovirus-induced Th2 differentiation by primary human monocytes.

Rhinovirus (RV) infections are linked to the development and exacerbation of allergic diseases including allergic asthma. IgE, another contributor to atopic disease pathogenesis, has been shown to regulate DC antiviral functions and influence T cell priming by monocytes. We previously demonstrated that IgE-mediated stimulation of monocytes alters multiple cellular functions including cytokine secretion, phagocytosis, and influenza-induced Th1 development. In this study, we investigate the effects of IgE-mediated stimulation on monocyte-driven, RV-induced T cell development utilizing primary human monocyte-T cell co-cultures. We demonstrate that IgE crosslinking of RV-exposed monocytes enhances monocyte-driven Th2 differentiation. This increase in RV-induced Th2 development was regulated by IgE-mediated inhibition of virus-induced type I IFN and induction of IL-10. These findings suggest an additional mechanism by which two clinically significant risk factors for allergic disease exacerbations-IgE-mediated stimulation and rhinovirus infection-may synergistically promote Th2 differentiation and allergic inflammation.

The ß2-adrenergic receptor controls inflammation by driving rapid IL-10 secretion.

The mammalian nervous system communicates important information about the environment to the immune system, but the underlying mechanisms are largely unknown. Secondary lymphoid organs are highly innervated by sympathetic neurons that secrete norepinephrine (NE) as the primary neurotransmitter. Immune cells express adrenergic receptors, enabling the sympathetic nervous system to directly control immune function. NE is a potent immunosuppressive factor and markedly inhibits TNF-α secretion from innate cells in response to lipopolysaccharide (LPS). In this study, we demonstrate that NE blocks the secretion of a variety of proinflammatory cytokines by rapidly inducing IL-10 secretion from innate cells in response to multiple Toll-like receptor (TLR) signals. NE mediated these effects exclusively through the ß2-adrenergic receptor (ADRB2). Consequently, Adrb2-/- animals were more susceptible to L. monocytogenes infection and to intestinal inflammation in a dextran sodium sulfate (DSS) model of colitis. Further, Adrb2-/- animals rapidly succumbed to endotoxemia in response to a sub-lethal LPS challenge and exhibited elevated serum levels of TNF-α and reduced IL-10. LPS-mediated lethality in WT animals was rescued by administering a ß 2-specific agonist and in Adrb2-/- animals by exogenous IL-10. These findings reveal a critical role for ADRB2 signaling in controlling inflammation through the rapid induction of IL-10. Our findings provide a fundamental insight into how the sympathetic nervous system controls a critical facet of immune function through ADRB2 signaling.

Cutting Edge: Critical Role of Glycolysis in Human Plasmacytoid Dendritic Cell Antiviral Responses.

Plasmacytoid dendritic cells (pDCs) are vital to antiviral defense, directing immune responses via secretion of huge concentrations of IFN-α. These cells are critical in protecting the lung against clinically relevant respiratory viruses, particularly influenza (Flu), a virus responsible for substantial worldwide morbidity and mortality. How pDC responses to such viral pathogens are regulated, however, is poorly understood in humans. Using an unbiased approach of gene chip analysis, we discovered that Flu significantly affects metabolism in primary human pDCs. We demonstrate that Flu and RV, another common respiratory virus, induce glycolysis in pDCs and that this metabolic pathway regulates pDC antiviral functions, including IFN-α production and phenotypic maturation. Intranasal vaccination of human volunteers with live influenza virus also increases glycolysis in circulating pDCs, highlighting a previously unrecognized potential role for metabolism in regulating pDC immune responses to viral infections in humans.

Sympathetic neural signaling via the ß2-adrenergic receptor suppresses T-cell receptor-mediated human and mouse CD8(+) T-cell effector function.

Postganglionic sympathetic neurons innervate secondary lymphoid organs and secrete norepinephrine (NE) as the primary neurotransmitter. NE binds and signals through five distinct members of the adrenergic receptor family. In this study, we show elevated expression of the ß2-adrenergic receptor (ADRB2) on primary human CD8(+) effector memory T cells. Treatment of both human and murine CD8(+) T cells with NE decreased IFN-γ and TNF-α secretion and suppressed their cytolytic capacity in response to T-cell receptor (TCR) activation. The effects of NE were specifically reversed by ß2-specific antagonists. Adrb2(-/-) CD8(+) T cells were completely resistant to the effects of NE. Further, the ADRB2-specific pharmacological ligand, albuterol, significantly suppressed effector functions in both human and mouse CD8(+) T cells. While both TCR activation and stimulation with IL-12 + IL-18 were able to induce inflammatory cytokine secretion, NE failed to suppress IFN-γ secretion in response to IL-12 + IL18. Finally, the long-acting ADRB2-specific agonist, salmeterol, markedly reduced the cytokine secretion capacity of CD8(+) T cells in response to infection with vesicular stomatitis virus. This study reveals a novel intrinsic role for ADRB2 signaling in CD8(+) T-cell function and underscores the novel role this pathway plays in adaptive T-cell responses to infection.

STAT4-mediated transcriptional repression of the IL5 gene in human memory Th2 cells.

Type I interferon (IFN-α/ß) plays a critical role in suppressing viral replication by driving the transcription of hundreds of interferon-sensitive genes (ISGs). While many ISGs are transcriptionally activated by the ISGF3 complex, the significance of other signaling intermediates in IFN-α/ß-mediated gene regulation remains elusive, particularly in rare cases of gene silencing. In human Th2 cells, IFN-α/ß signaling suppressed IL5 and IL13 mRNA expression during recall responses to T-cell receptor (TCR) activation. This suppression occurred through a rapid reduction in the rate of nascent transcription, independent of de novo expression of ISGs. Further, IFN-α/ß-mediated STAT4 activation was required for repressing the human IL5 gene, and disrupting STAT4 dimerization reversed this effect. This is the first demonstration of STAT4 acting as a transcriptional repressor in response to IFN-α/ß signaling and highlights the unique activity of this cytokine to acutely block the expression of an inflammatory cytokine in human T cells.

Housing Temperature-Induced Stress Is Suppressing Murine Graft-versus-Host Disease through ß2-Adrenergic Receptor Signaling.

Graft-versus-host disease (GVHD) is the major complication of allogeneic hematopoietic cell transplantation, a potentially curative therapy for hematologic diseases. It has long been thought that murine bone marrow-derived T cells do not mediate severe GVHD because of their quantity and/or phenotype. During the course of experiments testing the impact of housing temperatures on GVHD, we discovered that this apparent resistance is a function of the relatively cool ambient housing temperature. Murine bone marrow-derived T cells have the ability to mediate severe GVHD in mice housed at a thermoneutral temperature. Specifically, mice housed at Institutional Animal Care and Use Committee-mandated, cool standard temperatures (∼ 22°C) are more resistant to developing GVHD than are mice housed at thermoneutral temperatures (∼ 30°C). We learned that the mechanism underlying this housing-dependent immunosuppression is associated with increased norepinephrine production and excessive signaling through ß-adrenergic receptor signaling, which is increased when mice are cold stressed. Treatment of mice housed at 22°C with a ß2-adrenergic antagonist reverses the norepinephrine-driven suppression of GVHD and yields similar disease to mice housed at 30°C. Conversely, administering a ß2-adrenergic agonist decreases GVHD in mice housed at 30°C. In further mechanistic studies using ß2-adrenergic receptor-deficient (ß2-AR(-/-)) mice, we found that it is host cell ß2-AR signaling that is essential for decreasing GVHD. These data reveal how baseline levels of ß-adrenergic receptor signaling can influence murine GVHD and point to the feasibility of manipulation of ß2-AR signaling to ameliorate GVHD in the clinical setting.

IFN-α suppresses GATA3 transcription from a distal exon and promotes H3K27 trimethylation of the CNS-1 enhancer in human Th2 cells.

CD4(+) Th2 development is regulated by the zinc finger transcription factor GATA3. Once induced by acute priming signals, such as IL-4, GATA3 poises the Th2 cytokine locus for rapid activation and establishes a positive-feedback loop that maintains elevated GATA3 expression. Type I IFN (IFN-α/ß) inhibits Th2 cells by blocking the expression of GATA3 during Th2 development and in fully committed Th2 cells. In this study, we uncovered a unique mechanism by which IFN-α/ß signaling represses the GATA3 gene in human Th2 cells. IFN-α/ß suppressed expression of GATA3 mRNA that was transcribed from an alternative distal upstream exon (1A). This suppression was not mediated through DNA methylation, but rather by histone modifications localized to a conserved noncoding sequence (CNS-1) upstream of exon 1A. IFN-α/ß treatment led to a closed conformation of CNS-1, as assessed by DNase I hypersensitivity, along with enhanced accumulation of H3K27me3 mark at this CNS region, which correlated with increased density of total nucleosomes at this putative enhancer. Consequently, accessibility of CNS-1 to GATA3 DNA binding activity was reduced in response to IFN-α/ß signaling, even in the presence of IL-4. Thus, IFN-α/ß disrupts the GATA3-autoactivation loop and promotes epigenetic silencing of a Th2-specific regulatory region within the GATA3 gene.

IgE cross-linking critically impairs human monocyte function by blocking phagocytosis.

BACKGROUND: IgE cross-linking triggers many cellular processes that drive allergic disease. While the role of IgE in mediating allergic responses is best described on basophils and mast cells, expression of the high-affinity IgE receptor on other innate immune cells, including monocytes, suggests that it may affect the function of these cells in allergic environments. OBJECTIVE: To determine the effect of IgE cross-linking on the function of human monocytes. METHODS: Monocytes purified from healthy donor blood samples were cultured for 4 to 96 hours with media alone, a cross-linking anti-IgE antibody or control IgG. Surface CD14 and CD64 expression and secreted cytokine concentrations were determined. Monocyte function was determined by assessing (1) phagocytosis of Escherichia coli or apoptotic HEp2 cells and (2) killing of intracellular E coli. Select experiments were performed on monocytes obtained from participants with elevated versus normal serum IgE concentrations. RESULTS: IgE cross-linking on monocytes increased CD14 expression and induced secretion of TNF-α, IL-6, and autoregulatory IL-10. These effects were greatest in individuals with elevated serum IgE concentrations. In contrast, IgE cross-linking reduced CD64 expression and significantly impaired phagocytic function without disrupting the capacity of monocytes to kill bacteria. CONCLUSIONS: IgE cross-linking drives monocyte proinflammatory processes and autoregulatory IL-10 in a serum IgE-dependent manner. In contrast, monocyte phagocytic function is critically impaired by IgE cross-linking. Our findings suggest that IgE cross-linking on monocytes may contribute to allergic disease by both enhancing detrimental inflammatory responses and concomitantly crippling phagocytosis, a primary mechanism used by these cells to resolve inflammation.

The ß2-adrenergic receptor (ADRB2) entrains circadian gene oscillation and diurnal responses to virus infection in CD8 + T cells.

Adaptive immune cells are regulated by circadian rhythms (CR) under both steady state conditions and during responses to infection. Cytolytic CD8 + T cells display variable responses to infection depending upon the time of day of exposure. However, the neuronal signals that entrain these cyclic behaviors remain unknown. Immune cells express a variety of neurotransmitter receptors including nicotinic, glucocorticoid, and adrenergic receptors. Here, we demonstrate that the ß2-adrenergic receptor (ADRB2) regulates the periodic oscillation of select core clock genes, such as Per2 and Bmal1 , and selective loss of the Adrb2 gene dramatically perturbs the normal diurnal oscillation of clock gene expression in CD8 + T cells. Consequently, their circadian-regulated anti-viral response is dysregulated, and the diurnal development of CD8 + T cells into variegated populations of cytolytic T cell (CTL) effectors is dramatically altered in the absence of ADRB2 signaling. Thus, the Adrb2 directly entrains core clock gene oscillation and regulates CR-dependent T cell responses to virus infection as a function of time-of-day of pathogen exposure. One Sentence Summary: The ß2-adrenergic receptor regulates circadian gene oscillation and downstream daily timing of cytolytic T cell responses to virus infection.

Differences in systemic immune parameters in individuals with lung cancer according to race.

Introduction: Racial and ethnic disparities in the presentation and outcomes of lung cancer are widely known. To evaluate potential factors contributing to these observations, we measured systemic immune parameters in Black and White patients with lung cancer. Methods: Patients scheduled to receive cancer immunotherapy were enrolled in a multi-institutional prospective biospecimen collection registry. Clinical and demographic information were obtained from electronic medical records. Pre-treatment peripheral blood samples were collected and analyzed for cytokines using a multiplex panel and for immune cell populations using mass cytometry. Differences between Black and White patients were determined and corrected for multiple comparisons. Results: A total of 187 patients with non-small cell lung cancer (Black, 19; White, 168) were included in the analysis. There were no significant differences in baseline characteristics between Black and White patients. Compared to White patients, Black patients had significantly lower levels of CCL23 and CCL27, and significantly higher levels of CCL8, CXCL1, CCL26, CCL25, CCL1, IL-1 b, CXCL16, and IFN-γ (all P <0.05, FDR<0.1). Black patients also exhibited greater populations of non-classical CD16+ monocytes, NKT-like cells, CD4+ cells, CD38+ monocytes, and CD57+ gamma delta T cells (all P <0.05). Conclusions: Black and White patients with lung cancer exhibit several differences in immune parameters, with Black patients exhibiting greater levels of numerous pro-inflammatory cytokines and cell populations. The etiology and clinical significance of these differences warrant further evaluation.

IL-12 selectively programs effector pathways that are stably expressed in human CD8+ effector memory T cells in vivo.

CD8(+) cytotoxic T lymphocytes play a major role in defense against intracellular pathogens, and their functions are specified by antigen recognition and innate cytokines. IL-12 and IFN-α/ß are potent "signal 3" cytokines that are involved in both effector and memory cell development. Although the majority of effector cells are eliminated as inflammation resolves, some survive within the pool of memory cells and retain immediate effector function. In this study, we demonstrate that IL-12 instructs a unique program of effector cell differentiation that is distinct from IFN-α/ß. Moreover, effector memory (T(EM)) cells within peripheral blood display many common attributes of cells differentiated in vitro in response to IL-12, including proinflammatory cytokine secretion and lytic activity. A pattern of IL-12-induced genes was identified that demarcate T(EM) from central memory cells, and the ontologies of these genes correlated precisely with their effector functions. Further, we uncovered a unique program of gene expression that was acutely regulated by IL-12 and reflected in stable gene expression patterns within T(EM), but not T central memory cells in vivo. Thus, this study directly links a selective set of IL-12-induced genes to the programming of effector functions within the stable population of human CD8(+) T(EM) cells in vivo.

Impact of norepinephrine on immunity and oxidative metabolism in sepsis.

Sepsis is a major health problem in the United States (US), constituting a leading contributor to mortality among critically ill patients. Despite advances in treatment the underlying pathophysiology of sepsis remains elusive. Reactive oxygen species (ROS) have a significant role in antimicrobial host defense and inflammation and its dysregulation leads to maladaptive responses because of excessive inflammation. There is growing evidence for crosstalk between the central nervous system and the immune system in response to infection. The hypothalamic-pituitary and adrenal axis and the sympathetic nervous system are the two major pathways that mediate this interaction. Epinephrine (Epi) and norepinephrine (NE), respectively are the effectors of these interactions. Upon stimulation, NE is released from sympathetic nerve terminals locally within lymphoid organs and activate adrenoreceptors expressed on immune cells. Similarly, epinephrine secreted from the adrenal gland which is released systemically also exerts influence on immune cells. However, understanding the specific impact of neuroimmunity is still in its infancy. In this review, we focus on the sympathetic nervous system, specifically the role the neurotransmitter norepinephrine has on immune cells. Norepinephrine has been shown to modulate immune cell responses leading to increased anti-inflammatory and blunting of pro-inflammatory effects. Furthermore, there is evidence to suggest that norepinephrine is involved in regulating oxidative metabolism in immune cells. This review attempts to summarize the known effects of norepinephrine on immune cell response and oxidative metabolism in response to infection.

Characterizing inflammatory profiles of suicidal behavior in adolescents: Rationale and design.

BACKGROUND: The suicide rate in youth and young adults continues to climb - we do not understand why this increase is occurring, nor do we have adequate tools to predict or prevent it. Increased efforts to treat underlying depression and other disorders that are highly associated with suicide have had limited impact, despite considerable financial investments in developing and disseminating available methods. Thus, there is a tremendous need to identify potential markers of suicide behavior for youth during this high-risk period. METHODS: Funded by the American Foundation for Suicide Prevention (AFSP), this study aims to map immune dysfunction to suicidal behavior and establish a reliable immune signature of suicide risk that can 1) guide future research into fundamental pathophysiology and 2) identify targets for drug development. The study design is an observational study where blood samples and a comprehensive array of clinical measures are collected from three groups of adolescents (n = 75 each) (1) with suicidal behavior [recent (within 3 months) suicide attempt or suicidal ideation warranting urgent evaluation,] (2) at risk for mood disorders, and (3) who are healthy (no psychiatric history). Participants will complete self-report and clinical assessments, along with a blood draw, at baseline, 3 months, 6 months and 12 months, and online self-report assessments once a month. RESULTS: The recruitment for this study is ongoing. LIMITATIONS: Observational, variability in treatment regimens. CONCLUSIONS: This study will help elucidate immune mechanisms that may play a causal role in suicide and serve as targets for future therapeutic development.

Cutting edge: Type I IFN reverses human Th2 commitment and stability by suppressing GATA3.

T helper 2 cells regulate inflammatory responses to helminth infections while also mediating pathological processes of asthma and allergy. IL-4 promotes Th2 development by inducing the expression of the GATA3 transcription factor, and the Th2 phenotype is stabilized by a GATA3-dependent autoregulatory loop. In this study, we found that type I IFN (IFN-alpha/beta) blocked human Th2 development and inhibited cytokine secretion from committed Th2 cells. This negative regulatory pathway was operative in human but not mouse CD4(+) T cells and was selective to type I IFN, as neither IFN-gamma nor IL-12 mediated such inhibition. IFN-alpha/beta blocked Th2 cytokine secretion through the inhibition of GATA3 during Th2 development and in fully committed Th2 cells. Ectopic expression of GATA3 via retrovirus did not overcome IFN-alpha/beta-mediated inhibition of Th2 commitment. Thus, we demonstrate a novel role for IFN-alpha/beta in blocking Th2 cells, suggesting its potential as a promising therapy for atopy and asthma.

Adrenergic signaling controls early transcriptional programs during CD8+ T cell responses to viral infection.

Norepinephrine is a key sympathetic neurotransmitter, which acts to suppress CD8 + T cell cytokine secretion and lytic activity by signaling through the ß2-adrenergic receptor (ADRB2). Although ADRB2 signaling is considered generally immunosuppressive, its role in regulating the differentiation of effector T cells in response to infection has not been investigated. Using an adoptive transfer approach, we compared the expansion and differentiation of wild type (WT) to Adrb2-/- CD8 + T cells throughout the primary response to vesicular stomatitis virus (VSV) infection in vivo. We measured the dynamic changes in transcriptome profiles of antigen-specific CD8 + T cells as they responded to VSV. Within the first 7 days of infection, WT cells out-paced the expansion of Adrb2-/- cells, which correlated with reduced expression of IL-2 and the IL-2Rα in the absence of ADRB2. RNASeq analysis identified over 300 differentially expressed genes that were both temporally regulated following infection and selectively regulated in WT vs Adrb2-/- cells. These genes contributed to major transcriptional pathways including cytokine receptor activation, signaling in cancer, immune deficiency, and neurotransmitter pathways. By parsing genes within groups that were either induced or repressed over time in response to infection, we identified three main branches of genes that were differentially regulated by the ADRB2. These gene sets were predicted to be regulated by specific transcription factors involved in effector T cell development, such as Tbx21 and Eomes. Collectively, these data demonstrate a significant role for ADRB2 signaling in regulating key transcriptional pathways during CD8 + T cells responses to infection that may dramatically impact their functional capabilities and downstream memory cell development.

Immune characterization of suicidal behavior in female adolescents.

Background: To address the need to identify potential markers of suicide behavior for adolescents (ages 12-18 years), mass cytometry was used to explore the cellular mechanisms that may underpin immune dysregulation in adolescents with recent suicidal behavior. Methods: Peripheral blood mononuclear cell (PBMC) samples from 10 female adolescents with a recent suicide attempt and 4 healthy female adolescents were used. A panel of 30 antibodies was analyzed using mass cytometry. We used two complementary approaches to 1) identify the cell types that significantly differed between the two groups, and 2) explore differences in the expression profile of markers on the surface of these cells. Mass cytometry data were investigated using (Center for Disease Control, 2021) Opt-SNE for dimension reduced (Curtin and Heron, 2019), FlowSOM for clustering, and (Bridge et al., 2006) EgdeR and SAM for statistical analyses. Results: Opt-SNE (a data driven clustering analysis) identified 15 clusters of distinct cell types. From these 15 clusters, cluster 5 (classical monocytes) had statistically lower abundance in suicidal adolescents as compared to healthy controls, whereas cluster 7 (gamma-delta T cells) had statistically higher abundance in suicidal adolescents compared to healthy control. Furthermore, across the 15 cell types, chemokine receptors, CXCR3 (cluster 5) and CXCR5 (clusters 4, 5, 7, and 9), had an elevated expression profile in those with a recent suicide attempt versus healthy controls. Conclusion: This report demonstrates the utility of high dimensional cell phenotyping in psychiatric disorders and provides preliminary evidence for distinct immune dysfunctions in adolescents with recent suicide attempts as compared to healthy controls.

Regulation of effector and memory T-cell functions by type I interferon.

Type I interferon (IFN-α/ß) is comprised of a family of highly related molecules that exert potent antiviral activity by interfering with virus replication and spread. IFN-α/ß secretion is tightly regulated through pathogen sensing pathways that are operative in most somatic cells. However, specialized antigen-presenting plasmacytoid dendritic cells are uniquely equipped with the capacity to secrete extremely high levels of IFN-α/ß, suggesting a key role for this cytokine in priming adaptive T-cell responses. Recent studies in both mice and humans have demonstrated a role for IFN-α/ß in directly influencing the fate of both CD4(+) and CD8(+) T cells during the initial phases of antigen recognition. As such, IFN-α/ß, among other innate cytokines, is considered an important 'third signal' that shapes the effector and memory T-cell pool. Moreover, IFN-α/ß also serves as a counter-regulator of T helper type 2 and type 17 responses, which may be important in the treatment of atopy and autoimmunity, and in the development of novel vaccine adjuvants.

Reciprocal responsiveness to interleukin-12 and interferon-alpha specifies human CD8+ effector versus central memory T-cell fates.

Multiple innate signals regulate the genesis of effector and memory CD8+ T cells. In this study, we demonstrate that the innate cytokines interleukin (IL)-12 and interferon (IFN)-alpha/beta regulate distinct aspects of effector and memory human CD8+ T-cell differentiation. IL-12 exclusively promoted the development of IFN-gamma- and tumor necrosis factor (TNF)-alpha-secreting T effector memory (T(EM)) cells, whereas IFN-alpha drove the development of T central memory (T(CM)) cells. The development of T(EM) and T(CM) was linked to cell division. In rapidly dividing cells, IL-12 programmed T(EM) through induction of the IL-12 receptor beta2. In contrast, IFN-alpha regulated T(CM) development by slowing the progression of cell division in a subpopulation of cells that selectively expressed elevated IFN-alpha/beta receptor-2. The strength of signal delivered through T-cell receptor (TCR) engagement regulated the responsiveness of cells to IL-12 and IFN-alpha. In the presence of both IL-12 and IFN-alpha, these cytokine signals were amplified as the strength of the TCR signal was increased, promoting the simultaneous development of both T(CM) and T(EM). Together, our results support a novel model in which IL-12 and IFN-alpha act in a nonredundant manner to regulate the colinear generation of both effector and memory cells.