Tanja Lange

Sniffing neuropeptides: a transnasal approach to the human brain

Neuropeptides act as neuronal messengers in the brain, influencing many neurobehavioral functions. Their experimental and therapeutic use in humans has been hampered because, when administered systemically, these compounds do not readily pass the blood–brain barrier, and they evoke potent hormone-like side effects when circulating in the blood. We administered three peptides, melanocortin(4–10) (MSH/ACTH(4–10)), vasopressin and insulin, intranasally and found that they achieved direct access to the cerebrospinal fluid (CSF) within 30 minutes, bypassing the bloodstream.

Sleep and immune function

Sleep and the circadian system exert a strong regulatory influence on immune functions. Investigations of the normal sleep–wake cycle showed that immune parameters like numbers of undifferentiated naïve T cells and the production of pro-inflammatory cytokines exhibit peaks during early nocturnal sleep whereas circulating numbers of immune cells with immediate effector functions, like cytotoxic natural killer cells, as well as anti-inflammatory cytokine activity peak during daytime wakefulness. Although it is difficult to entirely dissect the influence of sleep from that of the circadian rhythm, comparisons of the effects of nocturnal sleep with those of 24-h periods of wakefulness suggest that sleep facilitates the extravasation of T cells and their possible redistribution to lymph nodes. Moreover, such studies revealed a selectively enhancing influence of sleep on cytokines promoting the interaction between antigen presenting cells and T helper cells, like interleukin-12. Sleep on the night after experimental vaccinations against hepatitis A produced a strong and persistent increase in the number of antigen-specific Th cells and antibody titres. Together these findings indicate a specific role of sleep in the formation of immunological memory. This role appears to be associated in particular with the stage of slow wave sleep and the accompanying pro-inflammatory endocrine milieu that is hallmarked by high growth hormone and prolactin levels and low cortisol and catecholamine concentrations.

Effects of sleep and circadian rhythm on the human immune system.

Many immune parameters show systematic fluctuations over the 24‐h day in human blood. Circulating naive T‐cells and production of proinflammatory cytokines, like interleukin‐12 (IL‐12), peak during nighttime, whereas cytotoxic effector leukocytes and production of the anti‐inflammatory cytokine IL‐10 peak during daytime. These temporal changes originate from a combined influence of the circadian system and sleep. Both brain functions act synergistically and share neuroendocrine effector mechanisms to convey control over immune functions. Sympathetic tone and cortisol levels show a circadian nadir during nighttime and are further suppressed by sleep, whereas growth hormone and prolactin show a circadian peak during nighttime and are further enhanced by sleep. Thus, the circadian system and sleep jointly evoke a unique endocrine constellation that is extremely effective in inducing changes in leukocyte traffic and a shift toward proinflammatory type 1‐cytokines during the nocturnal period of sleep, that is, an action with strong clinical implications.

Sleep Enhances the Human Antibody Response to Hepatitis A Vaccination

Objective The common belief that sleep supports immune defense has received surprisingly little direct experimental support. The antibody response to vaccination provides a valid tool to assess the influence of sleep on adaptive immune functioning in humans, which is also clinically relevant. Methods Two groups of healthy humans (N = 19) not previously infected with hepatitis A virus (HAV) were studied. On the night after primary vaccination with inactivated HAV, which took place at 0900 hours, one group had regular sleep. The other group stayed awake, and did not sleep before 2100 hours the following day. HAV antibody titers were measured repeatedly until 28 days after vaccination. Plasma hormone concentrations and white blood cell (WBC) subset counts were determined on the night and day after vaccination. Results Subjects who had regular sleep after vaccination, displayed a nearly two-fold higher HAV antibody titer after 4 weeks than subjects staying awake on this night (p =.018). Compared with wakefulness, sleep after vaccination distinctly increased release of several immune-stimulating hormones including growth hormone, prolactin, and dopamine (p < .01). Concentrations of thyrotropin, norepinephrine, and epinephrine were lowered by sleep (p < .02), whereas sleep only marginally influenced WBC subset counts. Conclusions Data suggest that sleep compared with sleep deprivation on the night after vaccination improves the formation of antigen-specific immune defense as reflected by antibody production in humans. Sleep presumably acts by inducing a hormonal environment in secondary lymphoid tissues, enhancing lymphocyte proliferation and differentiation and finally antibody synthesis. Results underscore the importance of sleep for immunocompetence.

Cortisol and epinephrine control opposing circadian rhythms in T cell subsets

Pronounced circadian rhythms in numbers of circulating T cells reflect a systemic control of adaptive immunity whose mechanisms are obscure. Here, we show that circadian variations in T cell subpopulations in human blood are differentially regulated via release of cortisol and catecholamines. Within the CD4(+) and CD8(+) T cell subsets, naive cells show pronounced circadian rhythms with a daytime nadir, whereas (terminally differentiated) effector CD8(+) T cell counts peak during daytime. Naive T cells were negatively correlated with cortisol rhythms, decreased after low-dose cortisol infusion, and showed highest expression of CXCR4, which was up-regulated by cortisol. Effector CD8(+) T cells were positively correlated with epinephrine rhythms, increased after low-dose epinephrine infusion, and showed highest expression of beta-adrenergic and fractalkine receptors (CX3CR1). Daytime increases in cortisol via CXCR4 probably act to redistribute naive T cells to bone marrow, whereas daytime increases in catecholamines via beta-adrenoceptors and, possibly, a suppression of fractalkine signaling promote mobilization of effector CD8(+) T cells from the marginal pool. Thus, activation of the major stress hormones during daytime favor immediate effector defense but diminish capabilities for initiating adaptive immune responses.

Acute sleep deprivation reduces energy expenditure in healthy men

BACKGROUND Epidemiologic evidence indicates that chronic sleep curtailment increases risk of developing obesity, but the mechanisms behind this relation are largely unknown. OBJECTIVE We examined the influence of a single night of total sleep deprivation on morning energy expenditures and food intakes in healthy humans. DESIGN According to a balanced crossover design, we examined 14 normal-weight male subjects on 2 occasions during a regular 24-h sleep-wake cycle (including 8 h of nocturnal sleep) and a 24-h period of continuous wakefulness. On the morning after regular sleep and total sleep deprivation, resting and postprandial energy expenditures were assessed by indirect calorimetry, and the free-choice food intake from an opulent buffet was tested in the late afternoon at the end of the experiment. Circulating concentrations of ghrelin, leptin, norepinephrine, cortisol, thyreotropin, glucose, and insulin were repeatedly measured over the entire 24-h session. RESULTS In comparison with normal sleep, resting and postprandial energy expenditures assessed on the subsequent morning were significantly reduced after sleep deprivation by ≈5% and 20%, respectively (P < 0.05 and P < 0.0001). Nocturnal wakefulness increased morning plasma ghrelin concentrations (P < 0.02) and nocturnal and daytime circulating concentrations of thyreotropin, cortisol, and norepinephrine (P < 0.05) as well as morning postprandial plasma glucose concentrations (P < 0.05). Changes in food intakes were variable, and no differences between wake and sleep conditions were detected. CONCLUSION Our findings show that one night of sleep deprivation acutely reduces energy expenditure in healthy men, which suggests that sleep contributes to the acute regulation of daytime energy expenditure in humans.

Sleep associated regulation of T helper 1/T helper 2 cytokine balance in humans.

Recent human studies suggested a supportive influence of regular nocturnal sleep on immune responses to experimental infection (vaccination). We hypothesized here that sleep could ease such responses by shifting the balance between T helper 1 (Th1) and T helper 2 (Th2) cytokine activity towards Th1 dominance thereby favoring cellular over humoral responses to infection. We compared the Th1/Th2 cytokine balance in 14 healthy men during regular nocturnal sleep (between 23:00 and 07:00 h) and while remaining awake during the same nocturnal interval, in a within-subject cross-over design. Blood was collected every 2 h. Production of T cell derived cytokines--interferon-gamma (IFN-gamma), interleukin-2 (IL-2), interleukin-4 (IL-4), and tumor necrosis factor-alpha (TNF-alpha)--was measured at the single cell level using multiparametric flow cytometry. Also, several immunoactive hormones--prolactin, growth hormone (GH), thyroid stimulating hormone (TSH), cortisol, and melatonin--were measured, the release of which is known to be regulated by sleep. Compared with wakefulness, early nocturnal sleep induced a shift in the Th1/Th2 cytokine balance towards increased Th1 activity, as indicated by an increased (p <.05) ratio of IFN-gamma/IL-4 producing T helper cells. However, the Th1 shift was only of moderate size and replaced by Th2 dominance during late sleep (p <.05). It could be mediated via release of prolactin and GH which both were distinctly increased during sleep (p <.001). Though unexpected, the most pronounced effect of sleep on T cell cytokine production was a robust decrease in TNF-alpha producing CD8+ cells probably reflecting increased extravasation of cytotoxic effector and memory T cells.

Selective Mobilization of Cytotoxic Leukocytes by Epinephrine

It is well-known that acute stress, presumably as a first defense against pathogens, enhances PBMC counts by mobilizing these β2-adrenoceptor positive cells from the marginal pool. Yet, only select leukocyte subsets participate in this phenomenon of adrenergic leukocytosis and underlying mechanisms are obscure. In this study, we analyzed in human blood adhesion molecule and chemokine receptor profiles in 14 leukocyte subsets, and responsiveness of subsets to epinephrine in vivo and in vitro. Five subsets, namely, CCR7−CD45RA+CD8+ effector T cells, CD4−CD8− γ/δ T cells, CD3+CD56+ NKT-like cells, CD16+CD56dim cytotoxic NK cells, and CD14dimCD16+ proinflammatory monocytes showed a rapid and transient increase after infusion of epinephrine at physiological concentrations. These cells were characterized by a CD62L−CD11abrightCX3CRbright phenotype, whereby expression of both CD11a and CX3CR1 was strongly correlated with adrenergic leukocytosis in vivo (r = 0.86 and 0.78, p < 0.005). The same subsets showed highest adherence to activated endothelium in vitro, which (except for proinflammatory monocytes) was reversed by epinephrine. We conclude that these five cytotoxic effector leukocyte subsets comprise the marginal pool by a CD11a/CX3CR1-mediated attachment to the endothelium. Epinephrine rapidly attenuates this attachment to allow demargination and release of the cells into the circulation that, because of their cytotoxic effector function, provide immediate protection from invading pathogens.

Crosstalk between the circadian clock circuitry and the immune system

Various features, components, and functions of the immune system present daily variations. Immunocompetent cell counts and cytokine levels present variations according to the time of day and the sleep-wake cycle. Moreover, different immune cell types, such as macrophages, natural killer cells, and lymphocytes, contain a circadian molecular clockwork. The biological clocks intrinsic to immune cells and lymphoid organs, together with inputs from the central pacemaker of the suprachiasmatic nuclei via humoral and neural pathways, regulate the function of cells of the immune system, including their response to signals and their effector functions. Consequences of this include, for example, the daily variation in the response to an immune challenge (e.g., bacterial endotoxin injection) and the circadian control of allergic reactions. The circadian-immune connection is bidirectional, because in addition to this circadian control of immune functions, immune challenges and immune mediators (e.g., cytokines) were shown to have strong effects on circadian rhythms at the molecular, cellular, and behavioral levels. This tight crosstalk between the circadian and immune systems has wide-ranging implications for disease, as shown by the higher incidence of cancer and the exacerbation of autoimmune symptoms upon circadian disruption. (Author correspondence: g.mazzoccoli@operapadrepio.it)

Sleep-dependent activity of T cells and regulatory T cells.

A number of immunological functions are dependent on circadian rhythms and regular sleep. This has impact on the type and magnitude of immune responses following antigenic challenge, for example in vaccination. Little is known about the underlying mechanisms. One possibility may be the circadian and sleep‐dependent modulation of CD4+CD25‐ T cell responses by CD4+CD25+ natural regulatory T cells (nTreg). In a variety of studies, nTreg have been shown to regulate T cell responses negatively. Thus, we investigated the influence of sleep and circadian rhythm on the number and function of nTreg as well as on the function of CD4+CD25‐ T cells. Seven healthy young men were examined under defined conditions on two occasions, i.e. during sleep and sleep deprivation. Venous blood was drawn periodically; numbers of nTreg, suppressive activity of nTreg, interleukin‐2 production and proliferation of CD4+CD25‐ T cells were explored in vitro. nTreg counts revealed a significant circadian rhythm with highest levels during the night (mean 95 nTreg/µl) and lowest levels during the day (mean 55 nTreg/µl). During normal sleep, the suppressive activity of nTreg was highest at 02.00 h and somewhat lower at 15.00 h. Surprisingly, almost no suppressive activity was present at 07.00 h. Deprivation of sleep abrogated this rhythm. CD4+CD25– T cell proliferation was dampened significantly by sleep deprivation. This is the first study in human cells to show that nTreg number and function follow a rhythm across the 24‐h period. Furthermore, sleep deprivation severely disturbs the functional rhythm of nTreg and CD4+CD25– T cells.