Memory T lymphocytes are characterized by their ability to mount a rapid and robust response to the recall antigen. Norepinephrine (NE) is one of the primary catecholamines of the sympathetic nervous system released during a stress response and plays an important role in modulating immune function. NE binds to the adrenergic receptors on immune cells, including T cells, resulting in either suppressed or enhanced function depending on the type of cell, activation status of the cell, duration of NE exposure and concentration of NE. Here, we aim to analyze the effects of NE on the functionality of nave (Tn), central memory (Tcm) and effector memory (Tem) CD8 T cells. We isolated CD8 T cell subsets from healthy human adults and treated cells in vitro with NE (1x10-6 M) for 16 hours; we then stimulated NE treated and untreated CD8 T cell subsets with antibodies for CD3 and CD28 for 24 and 72 hours. We assessed the level of beta-2 adrenergic receptor (ADRB2) expression in these cells as well as global gene expression changes in NE treated Tcm cells by microarray analysis. Altered expressed genes after NE treatment were identified and further confirmed by RT-qPCR, and by ELISA for protein changes. We further determined whether the observed NE effects on memory CD8 T cells are mediated by ADRB2 using specific adrenergic receptor agonist and antagonists. Finally, we examined the levels of mRNA and protein of the NE-induced genes in healthy adults with high serum levels of NE (>150 pg/mL) compared to low levels (<150 pg/mL). We found that memory (Tcm and Tem) CD8 T cells expressed a significantly higher level of ADRB2 compared to nave cells. Consequently, memory CD8 T cells were significantly more sensitive than nave cells to NE induced changes in gene expressions in vitro. Global gene expression analysis revealed that NE induced an elevated expression of inflammatory cytokines and chemokines in resting and activated memory CD8 T cells in addition to a reduced expression of growth-related cytokines. The effects of NE on memory CD8 T cells were primarily mediated by ADRB2 as confirmed by the adrenergic receptor agonist and antagonist assays. Finally, individuals with high serum levels of NE had similar elevated gene expressions observed in vitro compared to the low NE group. Our results demonstrate that NE preferentially modulates the functions of memory CD8 T cells by inducing inflammatory cytokine production and reducing activation-induced memory CD8 T cell expansion. Chemokines play a pivotal role in regulating the immune response through a tightly controlled expression. Elevated levels of inflammatory chemokines commonly occur with aging but the mechanism underlying this age-associated change is not fully understood. Here we report the role of microRNA-125b (miR-125b) in regulating inflammatory CC chemokine 4 (CCL4) expression in human immune cells and its altered expression with aging. We first analyzed the mRNA level of CCL4 in 8 different types of immune cells including CD4 and CD8 T cell subsets (nave, central and effector memory), B cells and monocytes in blood from both young (≤ 42 yr) and old (≥ 70 yr) adults. We observed that monocytes and nave CD8 T cells expressed higher levels of CCL4 and exhibited an age-related increase in CCL4. We then found the level of miR-125b was inversely correlated with the level of CCL4 in these cells, and the level of miR-125b was reduced in monocytes and nave CD8 T cells of the old compared to the young adults. Knock-down of miR-125b by shRNA in monocytes and nave CD8 T cells led to an increase of CCL4 protein whereas enhanced miR-125b expression by transfection in nave CD8 T cells resulted in a reduction of the CCL4 protein in response to stimulation. Finally, we demonstrated that miR-125b action requires the seed sequence in 3UTR of CCL4. Together these findings demonstrated that miR-125b is a negative regulator of CCL4 and its reduction is partially responsible for the age-related increase of CCL4. The TCR repertoire serves as a reservoir of TCRs for recognizing all potential pathogens. Two major types of T cells, CD4+ and CD8+, that use the same genetic elements and process to generate a functional TCR, differ in their recognition of peptide bound to MHC class II and I, respectively. However, it is currently unclear to what extent the TCR repertoire of CD4+ and CD8+ T cells is different. Here, we report a comparative analysis of the TCRβ repertoires of CD4+ and CD8+ T cells using a 5-RACE/PCR/sequencing method. We found that TCRβ diversity of CD4+ T cells ranges from 1.8-8.2 x105 and is 3-4 times greater on average than that of CD8+ T cells in each study subject. Furthermore, there was little overlap in TCRβ sequences between CD4+ (0.3%) and CD8+ (1.3%) T cells. Further analysis showed that CD4+ and CD8+ T cells exhibited distinct preferences for certain amino acids in the CDR3, and this was further confirmed by a support vector machine classifier, suggesting there are distinct and discernible differences between TCRβ CDR3 in CD4+ and CD8+ T cells. Finally, we identified 6-12% of the unique TCRβs that share an identical CDR3 with different V genes. Together, our findings reveal the distinct features of the TCRβ repertoire between CD4+ and CD8+ T cells and could potentially be used to evaluate the competency of T cell immunity.
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