Circadian gene period1b regulates proinflammatory cytokine expression through NF-κB signalling in zebrafish.

The circadian clock plays a critical role in regulating the immune system. Our previous publication revealed that a mutation in the circadian gene period1b (per1b) in zebrafish significantly decreased proinflammatory gene expression, particularly under constant darkness (DD) conditions; however, the underlying mechanisms remain unclear. In this study, using per1b-null mutant zebrafish and a larval tail fin injury model, we observed that the loss of per1b resulted in the downregulation expression of proinflammatory cytokines, such as IL-6 and TNF-α, at protein level. Furthermore, the loss of per1b downregulated ERK phosphorylation and inhibited p65 phosphorylation, leading to reduced NF-κB activation, which could downregulate the expression of proinflammatory cytokines, such as IL-6 and TNF-α, in zebrafish. These results provided insight into the communication between the circadian clock and immune functions.

Circadian genes period1b and period2 differentially regulate inflammatory responses in zebrafish.

The circadian clock has been shown to regulate various immune processes in different animals. Our previous report demonstrated that the innate immune responses in zebrafish show significant rhythmicity that could be regulated by melatonin. Here, we used diurnal zebrafish to determine the role of circadian genes in the inflammatory responses. Our results indicate that circadian genes exhibit rhythmic oscillations in zebrafish leukocytes, and mutations of the clock genes period1b (per1b) and period2 (per2) considerably affect these oscillations. Using a wounded zebrafish inflammation model, we found that under constant dark conditions (DD), the expression of pro-inflammatory cytokines is significantly downregulated in per1b gene mutant zebrafish and significantly upregulated in the per2 gene mutant zebrafish. Furthermore, using real-time imaging technology, we found that the per1b gene markedly disturbs the rhythmic recruitment of neutrophils toward the injury, whereas the per2 gene does not show a significant effect. Taken together, our results reveal differential functions of the circadian genes per1b and per2 in the inflammatory responses, serving as evidence that circadian rhythms play a vital role in immune processes.

Distribution of PERIOD-immunoreactive neurons and temporal change of the immunoreactivity under long-day and short-day conditions in the larval brain of the flesh fly Sarcophaga similis.

The flesh fly Sarcophaga similis show a clear photoperiodic response; they develop into adults under long days, whereas they arrest their development at the pupal stage under short days. Although the involvement of a circadian clock in photoperiodic time measurement is suggested in this species, the anatomical location of the clock neurons responsible for the time measurement has been unknown. We detected two PERIOD-immunoreactive cell clusters in the larval brain; one cluster was located at the dorsoanterior region and the other at the medial region. We further investigated their temporal changes in PERIOD-immunoreactivity and compared their patterns under different photoperiods.

Positive association between physical activity and PER3 expression in older adults.

The circadian clock regulates many physiological functions including physical activity and feeding patterns. In addition, scheduled exercise and feeding themselves can affect the circadian clock. The purpose of the present study was to investigate the relationship between physical/feeding activity and expression of clock genes in hair follicle cells in older adults. Twenty adult men (age, 68 ± 7 years, mean ± SE) were examined in this cross-sectional study. Prior to hair follicle cell collection, the participants were asked to wear a uniaxial accelerometer for one week. The timings of breakfast, lunch, and dinner were also recorded. Hair follicle cells were then collected over a 24 h period at 4 h intervals. The amplitude of PER3 expression was positively correlated with moderate and vigorous physical activity (r = 0.582, p = 0.007) and peak oxygen uptake (r = 0.481, p = 0.032), but these correlations were not observed for NR1D1 or NR1D2. No association was noted between meal times and the amplitude or the acrophase for any of these three clock genes. These findings suggest that rhythmic expression of the circadian clock gene PER3 is associated with the amount of daily physical activity and physical fitness in older adults.

Antimicrobial capacity of the freshwater planarians against S. aureus is under the control of Timeless.

Planarians, which are non-parasitic flatworms, are highly resistant to bacterial infections. To better understand the mechanisms underlying this resistance, we investigated the role of the circadian machinery in the anti-bacterial response of the freshwater planarian Schmidtea mediterranea. We identified Smed-Tim from S. mediterranea as a homolog of the mammalian clock gene Tim. We showed via RNA interference that Smed-Tim is required for the anti-microbial activities of Schmidtea mediterranea against Staphylococcus aureus infection during the light/dark cycle. Indeed, S. aureus infection leads to the expression of Smed-Tim, which in turn promotes Smed-Traf6 and Smed-morn2, but not Smed-p38 MAPK expression, 2 master regulators of planarian anti-microbial responses.

An epithelial circadian clock controls pulmonary inflammation and glucocorticoid action.

The circadian system is an important regulator of immune function. Human inflammatory lung diseases frequently show time-of-day variation in symptom severity and lung function, but the mechanisms and cell types underlying these effects remain unclear. We show that pulmonary antibacterial responses are modulated by a circadian clock within epithelial club (Clara) cells. These drive circadian neutrophil recruitment to the lung via the chemokine CXCL5. Genetic ablation of the clock gene Bmal1 (also called Arntl or MOP3) in bronchiolar cells disrupts rhythmic Cxcl5 expression, resulting in exaggerated inflammatory responses to lipopolysaccharide and an impaired host response to Streptococcus pneumoniae infection. Adrenalectomy blocks rhythmic inflammatory responses and the circadian regulation of CXCL5, suggesting a key role for the adrenal axis in driving CXCL5 expression and pulmonary neutrophil recruitment. Glucocorticoid receptor occupancy at the Cxcl5 locus shows circadian oscillations, but this is disrupted in mice with bronchiole-specific ablation of Bmal1, leading to enhanced CXCL5 expression despite normal corticosteroid secretion. The therapeutic effects of the synthetic glucocorticoid dexamethasone depend on intact clock function in the airway. We now define a regulatory mechanism that links the circadian clock and glucocorticoid hormones to control both time-of-day variation and the magnitude of pulmonary inflammation and responses to bacterial infection.

The circadian clock controls toll-like receptor 9-mediated innate and adaptive immunity.

Circadian rhythms refer to biologic processes that oscillate with a period of ~24 hr. These rhythms are sustained by a molecular clock and provide a temporal matrix that ensures the coordination of homeostatic processes with the periodicity of environmental challenges. We demonstrate the circadian molecular clock controls the expression and function of Toll-like receptor 9 (TLR9). In a vaccination model using TLR9 ligand as adjuvant, mice immunized at the time of enhanced TLR9 responsiveness presented weeks later with an improved adaptive immune response. In a TLR9-dependent mouse model of sepsis, we found that disease severity was dependent on the timing of sepsis induction, coinciding with the daily changes in TLR9 expression and function. These findings unveil a direct molecular link between the circadian and innate immune systems with important implications for immunoprophylaxis and immunotherapy.