SummaryLymphocytes circulate through lymph nodes (LN) in search for antigen in what is believed to be a continuous process. Here, we show that lymphocyte migration through lymph nodes and lymph occurred in a non-continuous, circadian manner. Lymphocyte homing to lymph nodes peaked at night onset, with cells leaving the tissue during the day. This resulted in strong oscillations in lymphocyte cellularity in lymph nodes and efferent lymphatic fluid. Using lineage-specific genetic ablation of circadian clock function, we demonstrated this to be dependent on rhythmic expression of promigratory factors on lymphocytes. Dendritic cell numbers peaked in phase with lymphocytes, with diurnal oscillations being present in disease severity after immunization to induce experimental autoimmune encephalomyelitis (EAE). These rhythms were abolished by genetic disruption of T cell clocks, demonstrating a circadian regulation of lymphocyte migration through lymph nodes with time-of-day of immunization being critical for adaptive immune responses weeks later.

Though it has been shown that immunological functions of CD4+ T cells are time of day-dependent, the underlying molecular mechanisms remain largely obscure. To address the question whether T cells themselves harbor a functional clock driving circadian rhythms of immune function, we analyzed clock gene expression by qPCR in unstimulated CD4+ T cells and immune responses of PMA/ionomycin stimulated CD4+ T cells by FACS analysis purified from blood of healthy subjects at different time points throughout the day. Molecular clock as well as immune function was further analyzed in unstimulated T cells which were cultured in serum-free medium with circadian clock reporter systems. We found robust rhythms of clock gene expression as well as, after stimulation, IL-2, IL-4, IFN-γ production and CD40L expression in freshly isolated CD4+ T cells. Further analysis of IFN-γ and CD40L in cultivated T cells revealed that these parameters remain rhythmic in vitro. Moreover, circadian luciferase reporter activity in CD4+ T cells and in thymic sections from PER2::LUCIFERASE reporter mice suggest that endogenous T cell clock rhythms are self-sustained under constant culture conditions. Microarray analysis of stimulated CD4+ T cell cultures revealed regulation of the NF-κB pathway as a candidate mechanism mediating circadian immune responses. Collectively, these data demonstrate for the first time that CD4+ T cell responses are regulated by an intrinsic cellular circadian oscillator capable of driving rhythmic CD4+ T cell immune responses.

The mammalian circadian timing system consists of a master pacemaker in the suprachiasmatic nucleus (SCN), which is thought to synchronize peripheral clocks in various organs with each other and with external time. Our knowledge about the role of the SCN clock is based mainly on SCN lesion and transplantation studies. We have now directly deleted the SCN clock using the Cre/LoxP system and investigated how this affects synchronization of peripheral rhythms. Impaired locomotor activity and arrhythmic clock gene expression in the SCN confirm that the SCN clockwork was efficiently abolished in our mouse model. Nonetheless, under light-dark (LD) conditions, peripheral clocks remained rhythmic and synchronized to the LD cycle, and phase relationships between peripheral clocks were sustained. Adaptation to a shifted LD cycle was accelerated in SCN clock-deficient mice. Moreover, under zeitgeber-free conditions, rhythmicity of the peripheral clock gene expression was initially dampened, and after several days peripheral clocks were desynchronized. These findings suggest that the SCN clock is dispensable for the synchronization of peripheral clocks to the LD cycle. A model describing an SCN clock-independent pathway that synchronizes peripheral clocks with the LD cycle is discussed.-Husse, J., Leliavski, A., Tsang, A. H., Oster, H., Eichele, G. The light-dark cycle controls peripheral rhythmicity in mice with a genetically ablated suprachiasmatic nucleus clock. FASEB J. 28, 4950 -4960 (2014). www.fasebj.org Key Words: circadian ⅐ synaptotagmin10Circadian clocks coordinate a broad range of physiological processes such as the sleep-wake cycle, endocrine functions, locomotion, and various aspects of metabolism. The mammalian circadian timing system is hierarchically organized with a master pacemaker residing in the suprachiasmatic nucleus (SCN) and subordinate peripheral clocks present in most tissues (1). How different tissue clocks interact to orchestrate circadian rhythms and how this network of clocks is synchronized to external time remains one of the most intensely investigated areas of chronobiology.Circadian clockwork in the SCN and in the periphery is based on interlocked transcriptional-translational feedback loops driven by a common set of circadian clock genes (2). The circadian clockwork drives the rhythmic expression of tissue-specific clock output genes, some of which encode signaling molecules, which, in the case of the SCN, include neuropeptides (e.g., prokineticin 2 or arginine vasopressin; refs. 3, 4). SCN neurons are tightly coupled via gap junctions and neuropeptidergic signaling, resulting in coherent rhythms of electrical activity (5, 6). Both electrical activity and neuropeptide rhythms serve to transmit time information from the SCN to subordinate clocks in the brain and in the periphery (7,8).The function of the circadian timing system is to adapt an organism's physiology to changes in the environment caused by the rotation of the Earth. Thus, synchronization of the circadian system to the...

van der Horst GT, Oster H. High-fat diet-induced hyperinsulinemia and tissue-specific insulin resistance in Cry-deficient mice.