Transcription factor activation and protein phosphorylation patterns are distinct for CD28 and ICAM-1 co-stimulatory molecules.

We examined signaling differences between two co-stimulatory molecules, CD28 and ICAM-1 by analyzing transcription factors and proteins that are activated downstream of these co-stimulations. We observed that FAST-1, a crucial protein in the TGFß signaling pathway, was activated by only ICAM-1 co-stimulation, and not by CD28. We also observed that receptor tyrosine kinases Csk, Dtk, FGFR1 and ROR2 were phosphorylated upon CD28 co-stimulation and IGF-1R, HGFR, MuSK and EphA8 were phosphorylated upon ICAM-1 co-stimulation. Together, these findings suggest that these two co-stimulators induce the activation of different sets of proteins, suggesting that each co-stimulatory molecule has its unique signaling profile.

Engagement of CD45 alters early signaling events in human T cells co-stimulated through TCR + CD28.

Previously our lab has shown that co-stimulation of human T cells through different co-stimulatory molecules tune differentiation to different phenotypes. An open question is where in the signaling pathways induced by the co-stimulation do differences occur that contribute to outcome of differentiation. In this project, we investigate the early signaling process by comparing events that follow engagement of CD45 alone or in association with a co-stimulatory molecule: CD28. CD45 plays a crucial role to initiate T cell signaling by dephosphorylating a negatively regulatory tyrosine residue in Src family kinases such as Lck. First, we observed that engagement of CD45 alone induced signaling in T cells. We observed that TCR/CD3 stimulation with CD45 promoted prolonged Lck association with TCR/CD3 complex and Lck remained associated during TCR/CD3 + CD28 + CD45 stimulation as well. We concluded that Lck association is dependent on TCR/CD3 and CD45 engagement. Hence, CD45 altered early signaling events in T cells.

T cell activation and proliferation following acute exercise in human subjects is altered by storage conditions and mitogen selection.

Recent work investigating exercise induced changes in immunocompetence suggests that some of the ambiguity in the literature is resultant from different cell isolation protocols and mitogen selection. To understand this effect, we compared post-exercise measures of T cell activation and proliferation using two different stimulation methods (costimulation through CD28 or stimulation with phytohaemagglutinin [PHA]). Further, we investigated whether exercise induced changes are maintained when T cell isolation from whole blood is delayed overnight in either a room temperature or chilled (4°C) environment. As expected, an increased proliferation response was observed post-exercise in T cells isolated from whole blood of previously trained individuals immediately after blood collection. Also, cells stimulated with PHA after resting overnight in whole blood were not adversely impacted by the storage conditions. In contrast, allowing cells to rest overnight in whole blood prior to stimulation through CD28, lessened the proliferation observed by cells following exercise rendering both the room temperature and chilled samples closer to the results seen in the control condition. Changes in early markers of activation (CD25), followed a similar pattern, with activation in PHA stimulated cells remaining fairly robust after overnight storage; whereas cell activation following stimulation through CD3+CD28 was disproportionately decreased by the influence of overnight storage. These findings indicate that decisions regarding cell stimulation methods need to be paired with the timeline for T cell isolation from whole blood. These considerations will be especially important for field based studies of immunocompetence where there is a delay in getting whole blood samples to a lab for processing as well as clinical applications where a failure to isolate T cells in a timely manner may result in loss of the response of interest.

Acute bouts of exercise induce a suppressive effect on lymphocyte proliferation in human subjects: A meta-analysis.

OBJECTIVE: Lymphocyte proliferative responses are commonly used to assess immune function in clinical settings, yet it is unclear how proliferative capacity is altered by exercise. This analysis aims to quantitatively assess the proliferative response of lymphocytes following an acute bout of exercise. METHODS: Electronic databases were searched for articles containing the keywords "exercise" OR "acute" OR "aerobic" OR "resistance training" OR "immune function" AND "proliferation" AND "lymphocyte." Initial results yielded 517 articles of which 117 were reviewed in full. Twenty-four articles met the inclusion criteria. Calculated standardized mean difference (SMD) and corresponding standard errors (SE) were integrated using random-effect models. RESULTS: Analyses uncovered evidence for suppression of proliferative capacity following acute exercise in general (SMD=-0.18, 95% CI: -0.21, -0.16) with long duration, high intensity exercise exhibiting a moderate suppressive effect (SMD=-0.55, 95% CI: -0.86, -0.24). Discordant proliferative responses for long duration, high intensity exercise in competitive versus non-competitive settings were identified with enhanced proliferation (SMD=0.46, 95% CI: 0.03, 0.89) observed following competitive events and a large suppressive effect detected for similar activities outside of a competitive environment (SMD: -1.28, 95% CI: -1.61, -0.96) (p=0.02). CONCLUSION: Evidence suggests lymphocyte proliferation is suppressed following acute bouts of exercise, with exercise lasting longer than one hour having a greater magnitude of effect regardless of exercise intensity. Variations in observed effect sizes across intensity, duration, and competitive environment further highlight our need to acknowledge the impact of study designs in advancing our understanding of exercise immunology.

Immunoendocrine alterations following Marine Corps Martial Arts training are associated with changes in moral cognitive processes.

Combined physical and psychological stress events have been associated with exacerbated endocrine responses and increased alterations in immune cell trafficking when compared to exercise stress alone. Military training programs are rigorous in nature and often purposefully delivered in environments combining high levels of both physical and mental stress. The objective of this study was to assess physiological and cognitive changes following U.S. Marine Corps Martial Arts training. Seven active-duty, male Marines were observed during a typical Marine Corps Martial Arts training session. Immune parameters, including immunomodulatory cytokines, and hormone concentrations were determined from blood samples obtained at baseline, immediately post training (IP) and at 15min intervals post-training to 1h (R15, R30, R45, R60). Assessments of cognitive moral functioning (moral judgment and intent) were recorded at intervals during recovery. There were significant fluctuations in immunoendocrine parameters. Peak endocrine measures were observed within the IP-R15 time interval. Distributions of circulating immune cells were significantly altered with neutrophils and all lymphocyte subsets elevated at IP. IFN-γ and IL-17a exhibited small, non-significant, parallel increases over the recovery period. Moral functioning was informed by different social identities during the recovery resulting in changes in moral decision-making. These data demonstrate that the Marine Corps Martial Arts Program induces significant alterations in lymphocyte and leukocyte distributions, but does not shift the balance of Th1/Th2 cytokines or induce a systemic inflammatory response. The program does, however, induce alterations in moral decision-making ability associated with the observed endocrine responses, even suggesting a potential interaction between one's social identities and endocrine responses upon moral decision-making.

Timely blockade of ICAM-1.LFA-1 interaction prevents disease onset in a mouse model of emphysema.

AIM: It is becoming apparent that emphysema is partly driven by self-reactive T cells inducing inflammatory damage. Thus, T cells become targets for therapy similar to other autoimmune diseases. Costimulatory blockade therapy targets disease-specific T cells, rendering them ineffective by blocking a necessary costimulatory event on the T-cell surface. This therapy is tested here in mouse emphysema. MATERIALS & METHODS: Peptides representing contact domains of counter receptors LFA-1 and ICAM-1 were used as blockade therapy in elastase-induced emphysema. RESULTS: When administered during the first week after disease induction, blockade prevented lung destruction, reduced leukocyte infiltration and inhibited the decrease in T-cell CD4:CD8 ratio, also common in human emphysema. CONCLUSION: Costimulatory blockade therapy can affect the progress of emphysema.

Low density lipoprotein promotes human naive T cell differentiation to Th1 cells.

Oxidized LDL (oxLDL) in the arterial wall and its incorporation into foam cells leads to inflammation and nucleation of atherosclerotic plaque; this is opposed by HDL. OxLDL and HDL regulate activation of macrophages and endothelial cells, and study of T cell participation has been limited to mature, differentiated cells such as Th1 cells, which perpetuate atherogenesis by promoting cell-mediated responses and inflammation. Immature naïve T cells, just emerged from the thymus, have remained largely unstudied. We hypothesized that LDL and HDL provide selective modulation of immature naïve T cell differentiation and participation in plaque development. In our in vitro model, naïve cells become activated and differentiate to mature effector T cells that are Th1, Th2 or Treg cells. Addition of oxLDL favored differentiation to Th1 cells, reduced Th2 cell activity and prolonged cell survival. In contrast, HDL inhibited T cell proliferation and reduced cell survival. The data suggest a novel mechanism where oxLDL enhances differentiation of human naïve CD4+ T cells to Th1 cells capable of promoting inflammation and plaque progression, and this is opposed by HDL.

Variations in rodent models of type 1 diabetes: islet morphology.

Type 1 diabetes (T1D) is characterized by hyperglycemia due to lost or damaged islet insulin-producing ß -cells. Rodent models of T1D result in hyperglycemia, but with different forms of islet deterioration. This study focused on 1 toxin-induced and 2 autoimmune rodent models of T1D: BioBreeding Diabetes Resistant rats, nonobese diabetic mice, and Dark Agouti rats treated with streptozotocin. Immunochemistry was used to evaluate the insulin levels in the ß -cells, cell composition, and insulitis. T1D caused complete or significant loss of ß -cells in all animal models, while increasing numbers of α -cells. Lymphocytic infiltration was noted in and around islets early in the progression of autoimmune diabetes. The loss of lymphocytic infiltration coincided with the absence of ß -cells. In all models, the remaining α - and δ -cells regrouped by relocating to the islet center. The resulting islets were smaller in size and irregularly shaped. Insulin injections subsequent to induction of toxin-induced diabetes significantly preserved ß -cells and islet morphology. Diabetes in animal models is anatomically heterogeneous and involves important changes in numbers and location of the remaining α - and δ -cells. Comparisons with human pancreatic sections from healthy and diabetic donors showed similar morphological changes to the diabetic BBDR rat model.

Elimination of T cell reactivity to pancreatic ß cells and partial preservation of ß cell activity by peptide blockade of LFA-1:ICAM-1 interaction in the NOD mouse model.

In insulin dependent diabetes mellitus (T1D), self-reactive T cells infiltrate pancreatic islets and induce beta cell destruction and dysregulation of blood glucose. A goal is to control only the self-reactive T cells, leaving the remainder of the T cell population free to protect the host. One approach is blockade of the second signal for T cell activation while allowing the first (antigen-specific) signal to occur. This work proposes that small peptides that block interaction of second signals delivered through the counter receptors LFA-1:ICAM-1 will induce attacking T cells (receiving the antigen signal) to become anergic or undergo apoptosis. In NOD mice, the peptides eliminated T cell reactivity against pancreatic antigens and reduced cellular infiltration into islets, which retained stronger density of insulin staining at five weeks after cessation of therapy. In in vitro studies the peptides induced nonresponsiveness during activation of T cells from mice and from human peripheral blood.

Potential immunological consequences of pharmacological suppression of gastric acid production in patients with multiple sclerosis.

Corticosteroids are standard treatment for patients with multiple sclerosis experiencing acute relapse. Because dyspeptic pain is a common side effect of this intervention, patients can be given a histamine receptor-2 antagonist, proton pump inhibitor or antacid to prevent or ameliorate this disturbance. Additionally, patients with multiple sclerosis may be taking these medications independent of corticosteroid treatment. Interventions for gastric disturbances can influence the activation state of the immune system, a principal mediator of pathology in multiple sclerosis. Although histamine release promotes inflammation, activation of the histamine receptor-2 can suppress a proinflammatory immune response, and blocking histamine receptor-2 with an antagonist could shift the balance more towards immune stimulation. Studies utilizing an animal model of multiple sclerosis indicate that histamine receptor-2 antagonists potentially augment disease activity in patients with multiple sclerosis. In contrast, proton pump inhibitors appear to favor immune suppression, but have not been studied in models of multiple sclerosis. Antacids, histamine receptor-2 antagonists and proton pump inhibitors also could alter the intestinal microflora, which may indirectly lead to immune stimulation. Additionally, elevated gastric pH can promote the vitamin B12 deficiency that patients with multiple sclerosis are at risk of developing. Here, we review possible roles of gastric acid inhibitors on immunopathogenic mechanisms associated with multiple sclerosis.

The Salmonella type III secretion system inner rod protein PrgJ is partially folded.

The type III secretion system (T3SS) is essential in the pathogenesis of many bacteria. The inner rod is important in the assembly of the T3SS needle complex. However, the atomic structure of the inner rod protein is currently unknown. Based on computational methods, others have suggested that the Salmonella inner rod protein PrgJ is highly helical, forming a folded 3 helix structure. Here we show by CD and NMR spectroscopy that the monomeric form of PrgJ lacks a tertiary structure, and the only well-structured part of PrgJ is a short α-helix at the C-terminal region from residues 65-82. Disruption of this helix by glycine or proline mutation resulted in defective assembly of the needle complex, rendering bacteria incapable of secreting effector proteins. Likewise, CD and NMR data for the Shigella inner rod protein MxiI indicate this protein lacks a tertiary structure as well. Our results reveal that the monomeric forms of the T3SS inner rod proteins are partially folded.

Choice of resident costimulatory molecule can influence cell fate in human naïve CD4+ T cell differentiation.

With antigen stimulation, naïve CD4+ T cells differentiate to several effector or memory cell populations, and cytokines contribute to differentiation outcome. Several proteins on these cells receive costimulatory signals, but a systematic comparison of their differential effects on naïve T cell differentiation has not been conducted. Two costimulatory proteins, CD28 and ICAM-1, resident on human naïve CD4+ T cells were compared for participation in differentiation. Under controlled conditions, and with no added cytokines, costimulation through either CD3+CD28 or CD3+CAM-1 induced differentiation to T effector and T memory cells. In contrast, costimulation through CD3+ICAM-1 induced differentiation to Treg cells whereas costimulation through CD3+CD28 did not.

Deferiprone modulates in vitro responses by peripheral blood T cells from control and relapsing-remitting multiple sclerosis subjects.

T cells are important mediators of autoimmune inflammation in relapsing-remitting multiple sclerosis (RRMS). Previous studies found that deferiprone, an iron chelator, suppressed disease activity in a mouse model of multiple sclerosis, and inhibition of T cell proliferation was implicated as a putative mechanism. The objective of the present study was to examine the effects of deferiprone on suppressing in vitro responses of T cells from control and RRMS subjects. Peripheral blood T cells were co-stimulated with anti-CD3+anti-CD28 and cultured with or without interleukin 2 (IL-2). Proliferating CD4+ T cells from control and RRMS subjects, cultured with or without IL-2, decreased in response to 75 µM deferiprone, although the extent of decreased proliferation of CD4+ T cells from RRMS subjects was less than for control subjects. Proliferating CD8+ T cells from control subjects, cultured with or without IL-2, also decreased in response to 75 µM deferiprone, and this decrease was seen in proliferating CD8+ T cells from RRMS cultured with IL-2. CD4+CD25+ and CD8+CD25+ cells from control subjects, cultured with or without IL-2, declined in 75 µM deferiprone, but the decrease was smaller than for the CD4+ and CD8+ proliferative responses. CD4+CD25+ and CD8+CD25+ cells from RRMS subjects showed more variability than for control subjects, but CD4+CD25+ cultured with IL-2 and CD8+CD25+ cells cultured without IL-2 significantly declined in 75 µM deferiprone. CD4+FoxP3+ and CD4+CD25+FoxP3+ cells tended to remain constant or increase. In summary, deferiprone induced declines in proliferative responses at a dosage that is within peak serum pharmacological concentrations.

Two different modes of costimulation predispose human T lymphocytes to differential responses in the presence of HDL or oxidized LDL.

In addition to levels of high-density lipoprotein (HDL), oxidized (ox) low-density lipoprotein (LDL), the inflammatory process and certain genetic factors, T cells are crucial for expansion of atherosclerotic plaque. The interrelationships among these influences are still being defined. Here, we examined how HDL and oxLDL affect T cell function. T cells require two activation signals to achieve functional activity. The first signal is specific and is delivered by appropriately presented antigen. The second (costimulatory) signal can be received through any of several T cell surface proteins, the most widely studied of which are CD28 and LFA-1. We have identified ICAM-1, resident on T cells, as a costimulatory protein. Here, we describe differential effects when T cells were costimulated through either LFA-1 or ICAM-1 in the presence of HDL or oxLDL. In general, T cells costimulated through either LFA-1 or ICAM-1 in the presence of oxLDL were predisposed to a decrease in proliferation and increased apoptosis, although ICAM-1-costimulated cells were protected from apoptosis induced by lower levels of oxLDL. T cell subsets also were examined. In the presence of HDL, CD8(+) T cells increased proliferation when costimulated through LFA-1. HDL exerted no effect on proliferation of CD4(+) T cells whereas proliferation decreased in the presence of oxLDL. Naïve T cells proliferated better in response to costimulation through LFA-1 in the presence of HDL but proliferation of effector/memory cells was not altered in the presence of HDL. When T cells were costimulated through LFA-1, in the presence of either HDL or oxLDL synthesis of Th-1 but not Th-2 cytokines was increased. T cells costimulated through ICAM-1 increased Th-1 but not Th-2 cytokines but this was not altered in the presence of HDL or oxLDL. Thus, the nature of costimulation seems to influence T cell responses in the presence of the lipoproteins.

Costimulation of naive human CD4 T cells through intercellular adhesion molecule-1 promotes differentiation to a memory phenotype that is not strictly the result of multiple rounds of cell division.

The process by which naive T cells become activated, differentiate into effector cells and ultimately generate long-lived memory cells is dependent upon a number of factors, including the costimulatory signals received by the T cell. To best understand the multiple events involved, it is important to understand the potential contributions by individual signalling proteins using both in vitro and in vivo studies. Here, the potential for costimulation through intercellular adhesion molecule-1 (ICAM-1; CD54), resident on the surface of naive human T cells, to influence differentiation was investigated. Costimulation of naive T cells through ICAM-1 resulted in expansive cell division, high interleukin-2 production, and protection from apoptosis. Prolonged culture led to outgrowth of a subpopulation of cells with a highly differentiated CD45RA- CD11a(hi) CD27- phenotype. In this respect, costimulation through ICAM-1 was similar to costimulation through CD28 and different from costimulation through leucocyte function-associated antigen-1. The CD45RA- CD11a(hi) CD27- cells responded to suboptimal stimulation through the T-cell receptor alone with a more robust proliferative response compared with naive cells from the same subject. These cells also secreted higher levels of T helper type 1 cytokines in response to lower levels of stimulation than their naive counterparts. The surface phenotype and more sensitive response characteristics suggest the creation of a memory T-cell subpopulation as a result of costimulation through ICAM-1. Finally, generation of this memory population was the result of specific costimulatory signals, and not merely because of a high number of cell divisions. These data reveal a new role for resident ICAM-1 to influence the differentiation of naive T cells.

Inflammatory mediators and immune function are altered in home parenteral nutrition patients.

OBJECTIVES: Patients who used home parenteral nutrition (HPN) and healthy, volunteer control subjects were examined to assess relative immune potential and inflammatory marker expression and to investigate the association between HPN and immune parameters. METHODS: Subjective Global Assessments were performed on all subjects. The peripheral blood concentration of C-reactive protein was determined by enzyme-linked immunosorbent assay. The peripheral blood concentration of systemic inflammatory mediators that included tumor necrosis factor-alpha (TNF-alpha), soluble TNF-alpha receptors p55 and p75, and interleukin-6 were similarly determined. Peripheral blood lymphocytes were isolated and the percentage of circulating CD4+ and CD8+ lymphocytes was determined by flow cytometry. In addition, peripheral blood lymphocytes were cultured in the presence of the T-cell mitogen, phytohemagglutinin, and the proliferative response of the CD3+ population was assessed by flow cytometry. Results of these experiments were obtained for 10 clinically stable patients who had used HPN longer than 2 y and these results were compared by Student's t test with data obtained for 12 normal, volunteer control subjects. RESULTS: Of the 10 patients who used HPN and were examined, seven had short bowel syndrome, two had dysmotility, and one required HPN due to radiation enteritis. Based on Subjective Global Assessments, all patients were well nourished. No difference was observed in TNF-alpha level between groups and C-reactive protein levels were within normal limits (1.2 mg/L in patients, 0.99 mg/L in controls). Soluble TNF-alpha receptors p55 and p75 were significantly increased (P < 0.001), but serum interleukin-6 was not (P = 0.07). The percentage of CD8+ cells and the CD4+/CD8+ ratio were not statistically different between groups. In contrast to this result, the percentage of CD4+ cells and the proliferative T-cell response to phytohemagglutinin were significantly depressed in patients who used HPN versus control subjects. CONCLUSIONS: These data suggest the presence of an underlying inflammatory process and subsequent abnormal T-lymphocyte function in patients who use HPN.

MIP-1alpha induces differential MAP kinase activation and IkappaB gene expression in human B lymphocytes.

The chemokine macrophage inflammatory protein-1alpha (MIP-1alpha) stimulates migration of B cells and affects B cell immunoglobulin production. However, the molecular mechanisms by which MIP-1alpha modulates these biologic effects have not been completely defined. Previously, we demonstrated that treatment of B cells with MIP-1alpha induced the transcription factor, nuclear factor (NF)-kappaB, to bind to DNA, concomitant with the degradation of IkappaBalpha, a cytoplasmic inhibitor of NF-kappaB activation. Here, we report that MIP-1alpha treatment of tonsil B cells induced IkappaB gene expression that was dependent on MIP-1alpha-mediated activation of a pathway(s) involving NF-kappaB and phosphatidylinositol-3 kinase (PI3K). The NF-kappaB pathway is understood to be controlled in an autoregulatory fashion, so expression of IkappaB is thought to provide a means by which B cells modulate this pathway after stimulation with MIP-1alpha. Although the idea of NF-kappaB autoregulation is not novel, this is the first report to suggest the regulation of B cell gene expression by MIP-1alpha. In addition, we observed the activation of Jun N-terminal kinase (JNK) and p38 mitogenic-activated protein kinase (MAPK), but not extracellular signal-related kinase (ERK) in response to MIP-1alpha. Although p38 and NF-kappaB activity were both necessary for B cell migration, IkappaB gene expression was not affected by p38 inhibition, suggesting that p38 is involved in a separate MIP-1alpha-mediated signal transduction pathway.

The outcome of T-cell costimulation through intercellular adhesion molecule-1 differs from costimulation through leucocyte function-associated antigen-1.

Optimal T-cell activation requires both an antigen-specific and a costimulatory signal. The outcome of T-cell activation can be influenced by the nature of the costimulatory signal the T cell receives. We recently demonstrated the ability of stimulation through intercellular adhesion molecule-1 (ICAM-1), resident on the T-cell surface, to provide a second signal for T-cell activation, and have extended that work here to begin an examination of the functional outcome of this set of signals. Costimulation through ICAM-1 resulted in a greater percentage of cells having undergone more than three divisions when compared to costimulation through leucocyte function-associated antigen-1 (LFA-1). Costimulation through ICAM-1 also had an effect similar to costimulation through CD28 in its ability to down-regulate the cyclin dependent kinase inhibitor p27kip1. Costimulation through ICAM-1 provided greater protection from apoptosis than costimulation through LFA-1, especially in cells having divided more than three times. This was supported by the ability of costimulation through ICAM-1 to up-regulate the anti-apoptotic protein Bcl-2. Finally, costimulation through ICAM-1 or CD28 produced a greater number of T cells with a memory phenotype than costimulation through LFA-1.

Influence of carbohydrate ingestion on cytokine responses following acute resistance exercise.

The effect of carbohydrate supplementation (CHO) on interleukin 2 (IL-2) and interleukin 5 (IL-5) secretion following acute resistance exercise was examined in 9 resistance-trained males. Subjects completed a randomized, double-blind protocol with exercise separated by 14 days. The exercise consisted of a high-intensity, short rest interval squat workout. Subjects consumed 1.0 g x kg body mass(-1) CHO or an equal volume of placebo (PLC) 10 min prior to and 10 min following exercise. Blood was collected at rest (REST), immediately post exercise (POST), and at 1.5 h of recovery (1.5 h POST). Isolated peripheral blood mononuclear cells were stimulated with PHA and assayed for IL-2 and IL-5 secretion. IL-2 secretion was significantly decreased at POST for both the PLC and CHO groups. However, the degree of decrease was less in the CHO group (16%) than in the PLC group (48%), and this difference was statistically significant. These responses were transient, and the values returned to normal by 1.5 h POST. A mild and transient but significant decrease in IL-5 secretion by the PLC group was observed at POST (26%) compared to REST. No significant decrease was observed in IL-5 secretion for CHO from REST to POST (12%). These data support a possible effect of carbohydrate supplementation on IL-2 and IL-5 secretion following high-intensity resistance exercise.

Stimulation through intercellular adhesion molecule-1 provides a second signal for T cell activation.

Regulation of T cell activation requires two signals. First, appropriately presented Ag in the context of MHC interacts with the T cell Ag receptor-CD3 complex. The best-studied second signal is CD28, which resides on the T cell and responds to its counter receptor, B7. A second signal also can be delivered through LFA-1 residing on the T cell, responding to its counter receptor ICAM-1 residing on a different cell. Characterization of a second signal is tied to its ability to costimulate (along with stimulation through the TCR) proliferation, IL-2 secretion, and coactivation of phosphatidylinositol 3-kinase. We examined whether ICAM-1, residing on the T cell surface, could deliver a second signal into that T cell. Costimulation through CD3 plus ICAM-1 caused increased T cell proliferation, increased expression of the activation marker CD69, increased transcription through the IL-2 regulatory region, and increased secretion of selected Th1 but not Th2 cytokines. Costimulation through CD3 plus ICAM-1 caused synergistic activation of phosphatidylinositol 3-kinase. Finally, the combination of anti-CD3 plus anti-ICAM-1 (but not anti-CD3 alone) caused prolonged proliferation of naive T cells in a manner similar to costimulation through LFA-1 or CD28. Thus, we demonstrate for the first time that ICAM-1 resident on a T cell can deliver a costimulatory signal into that T cell.