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A telemetry system based on Bluetooth Low Energy (BLE) was constructed to simultaneously collect locomotor activity and physiological signals of small animal cohorts for circadian rhythm experiments; it consists of miniature transmitters and mobile phone with customized App. The continuous sampling signals obtained from the 3-axis acceleration and temperature sensors in the transmitters are sent to the mobile phone in real-time through Internet of Things (IoT) for temporary storage and then imported into the computer for summary and rhythm analysis by the general open-source mathematical software. Unlike expensive and complicated commercial telemetry systems with industrial wireless standards, no special data receivers and software are needed. In our validation experiment, six rats were divided into two groups under natural dark and light-dark cycles. For two consecutive weeks, the transmitter mounted on the head of the rat-recorded locomotor activity, skin temperature, and ambient temperature of each rat at a frequency of 6 Hz. After processing with Local Weighted Regression Scatter Smoothing (LOWESS) and Fast Fourier Transform (FFT) filtering, single cosinor and multi-components cosinor were then used to assess and characterize the circadian rhythm. The results showed that the rhythm values of the two groups of rats coincided with the corresponding light-dark cycle, and that the system was robust to data loss and error from BLE communication failures. Therefore, the proposed system provides a light-weight framework for long-term circadian rhythm monitoring in free-moving rodents to further simplify and promote experimental chronobiology animal studies.

Emerging evidence has demonstrated that long noncoding RNAs (lncRNAs) play critical roles in the epigenetic and transcriptional regulation of mammalian circadian systems. Circadian rhythmicity regulates many aspects of our immune system, and perturbation of the circadian clock can augment the inflammatory response. However, knowledge of the precise functions of lncRNAs in the regulation of immune functions within the circadian system is relatively limited. In this study, differentially expressed lncRNAs induced by Clock knockdown were screened via mRNA/lncRNA microarray and bioinformatic prediction analysis. We identified a Clock -regulated lncRNA, AK028245, which was correlated with the activation of the immune response. The expression levels of AK028245 were decreased in the spleen of immunosuppressed mice and elevated in immune-activated mice treated with lipopolysaccharide (LPS). Further, Clock knockdown decreased the expression of OTUD7B and A20, 2 early immune response factors acting on the NF-κB signaling pathway. Interestingly, inhibition of AK028245 increased their expression, mitigating the effects of Clock knockdown. In addition, inhibition of AK028245 downregulated the expression of tumor necrosis factor-α and interleukin-6 in the late stages of LPS stimulation and the expression of interferon-γ and Cxcl12 in the peak stages. We conclude that this newly identified lncRNA plays a role in the crosstalk between Clock and immune response regulators, likely resulting in a proinflammatory response targeting OTUD7B and A20. The lncRNA AK028245 has revealed a new mechanism of the immune response and provided new targets for the treatment of immune disorders.

Hand, foot, and mouth disease (HFMD) with central nerve system complications may rapidly progress to fulminated cardiorespiratory failure, with higher mortality and worse prognosis. It has been reported that circadian rhythms of heart rate (HR) and respiratory rate are useful in predicting prognosis of severe cardiovascular and neurological diseases. The present study aims to investigate the characteristics of the circadian rhythms of HR, respiratory rate, and temperature in HFMD patients with neurological complications. Hospitalized HFMD patients including 33 common cases (common group), 61 severe cases (severe group), and 9 critical cases (critical group) were contrasted retrospectively. Their HR, respiratory rate, and temperatures were measured every 4 hours during the first 48-hour in the hospital. Data were analyzed with the least-squares fit of a 24-hour cosine function by the single cosinor and population-mean cosinor method. Results of population-mean cosinor analysis demonstrated that the circadian rhythm of HR, respiratory rate, and temperature was present in the common and severe group, but absent in the critical group. The midline-estimating statistic of rhythm (MESOR) (P = 0.016) and acrophase (P < 0.01) of temperature and respiratory rate were significantly different among 3 groups. But no statistical difference of amplitude in temperature and respiratory rate was observed among the 3 groups (P = 0.14). MESOR value of HR (P < 0.001) was significantly different in 3 groups. However, amplitude and acrophase revealed no statistical difference in circadian characteristics of HR among 3 groups. Compared with the common group, the MESOR of temperature and respiratory rate was significantly higher, and acrophase of temperature and respiratory rate was 2 hours ahead in the severe group, critical HFMD patients lost their population-circadian rhythm of temperature, HR, and respiratory rate. The high values of temperature and respiratory rate for the common group were concentrated between 3 and 9 PM, whereas those for the severe group were more dispersive. And the high values for the critical group were equally distributed in 24 hours of the day. Circadian rhythm of patients' temperature in the common group was the same as the normal rhythm of human body temperature. Circadian rhythm of patients' temperature, HR and respiratory rate in 3 groups were significantly different.

Mice homozygous for a dominant-negative allele of the Clock gene (Clock (Δ19/Δ19)) have slightly but significantly decreased male fertility. The molecular mechanism for this reduction in fertility is unknown. In the present study, we used a small hairpin RNA (shRNA) strategy to specifically knock down the Clock gene expression in the testes of male mice and determined its effect on male fertility. Clock knockdown led to smaller litter size, a lower in vitro fertility rate, lower blastula formation rate, and lower acrosin activity of the knockdown sperm. Locomotor activity analysis of the Clock knockdown mice revealed that Clock knockdown in testes did not alter their circadian rhythm. Taken together, these results provide the first evidence that Clock gene expression in round spermatids is essential for maintaining male reproductivity and suggest that acrosin may be a novel regulatory target of the Clock gene that would regulate the fertilization and early embryonic development to blastula. These findings may provide new clues for development of novel male contraceptive strategies.

Objective: To study the influence of different surgery on heart rate's circadian rhythm in postoperation of intracerebral hemorrhage., Methods: One hundren cases of hypertensive intracerebral hemorrhage at basal ganglia had been collected. According to methods of operation, cases were divided into two groups. Forty-three cases and 57 cases were treated respectively by standard craniotomy (Craniotomy group) and microinvasion of puncturation (Microinvasion group) to remove hematoma. Value of heart rate (HR) was recorded by monitor every 1 hour for 9 days. Circadian rhythm of every day was analyzed by single cosinor and population mean-cosinor., Results: Circadian rhythm of postoperative HR appeared (P < 0.001) in microinvasion group after operation. Circadian rhythm of postoperative HR had not been appearing (P < 0.001) in craniotomy group until to the 8th day. All amplitudes in microinvasion group are higher than that in craniotomy group. Both of amplitudes in two groups begin to be low during 4-7 d after operation and recover the level till the 8th day., Conclusion: Influence of circadian rhythm in craniotomy was more severe than microinvasion puncturation. There would be a intimate relationship between encephaledema and recovery of circadian rhythm after neurosurgery.

The synchronization of the circadian signals to external or suprachiasmatic nucleus stimulation in the peripheral clocks is essential for maintaining the usual function of human body. However, aging will disrupt the synchronization of peripheral circadian rhythms, thus leading to some age-associated diseases. Up to now, little is known about the modification of the oscillatory rhythms in aged cells. A recent report showed that cell senescence in vascular human smooth muscle cells (HSMCs) altered circadian rhythms by a dysregulation of rhythmic gene expression. Furthermore, this alteration could be reversed by telomerase reconstitution. To test whether telomerase reconstitution can restore disrupted circadian rhythm in other types of senescent cells, we used fibroblasts as cell models to profoundly investigate the relationship between cell senescence and circadian rhythm modulation. We found that the response of rhythmic gene expression to serum stimulation was markedly attenuated in senescent fibroblasts, telomerase-reconstituted fibroblasts reset the circadian oscillation of rhythmic gene expression, and the activation of pERK-CREB and p38-CREB pathways might be involved in the circadian rhythm resetting. These findings suggested that telomerase reconstitution might be a good way to reset synchronization of peripheral circadian rhythms disrupted in senescent tissues.

The circadian clock is the central timing system that controls numerous physiologic processes. The current model of these oscillators is based on autoregulatory transcription and translation feedback loops of these circadian genes in which Period1 (Per1) gene occupies a central position. The laminin receptor 1 (Lamr1) and its precursor are expressed in most tissues and play important roles in several physiologic and pathologic processes, including cell differentiation, growth, migration and cancer invasion. The present study showed that Lamr1 was a novel protein that interacted with human circadian clock protein hPer1 by the yeast two-hybrid system and co-immunoprecipitation, which was expressed in many tissues and did not display circadian rhythm. The expression of hPer1 was knocked down to 84.9% by the hPer1 RNA interfering test, but the expression levels of Lamr1 was not depressed by the hPer1 RNA interfering test. The results suggest that Lamr1 is a novel protein that interacts with human circadian clock protein hPer1 and Lamr1 is not a direct efferent element of circadian clock.

Circadian rhythm has been involved in the regulation of many physiological activities. Autophagy is the metabolic process that transports substances in the cytoplasm to the lysosomes for degradation, and involved in the process of many diseases, including cancer. Studies have shown that autophagy activity has a circadian rhythm feature. As the core gene of the circadian rhythm system, clock has participated in the occurrence and development of cancer. The expression of clock whether regulates tumor development by autophagy has not been illustrated at present. In this study, we established a stable clock-knockdown strain of mouse breast cancer cell 4T1 to explore the changes in autophagy activity. The results showed that autophagy-related proteins ATG9A, ATG7, LC3B and Beclin1 (ATG6) were down-regulated with clock-knockdown, and the p62, the key factors of autophagy, was up-regulated. Moreover, clock-knockdown caused significant up-regulation of p65, and inhibition of IκBα and p-AKT，which were the core factors of several signaling pathways involved in autophagy. Serum rhythm-inducing experiments behaved that the expression of LC3B and Beclin1 had rhythmic characteristics，differed from LC3A. The regulative gene clock was initially established to regulate the autophagy-related genes, and the suggested direction of its regulatory pathways. [ABSTRACT FROM AUTHOR]

Social isolation usually affects the physical and mental health of the human and produces depression, whether it may relate with circadian rhythm still remains to be investigated. Two groups of mice were employed to this experiment to continuously monitoring their activities in a dark environment. Immunofluorescence was used to show the expression of the circadian clock gene in two groups of mice. The results demonstrate that the social isolation leads to decrease of mouse activities, phase advance of rhythm relatively and down-regulated of clock gene expression. We conclude that social isolation can affect the physiological function and the normal physiological rhythm of mice, which may be related to depression. [ABSTRACT FROM AUTHOR]

Transactivator (Tat), a regulatory protein of HIV-1, plays a very important role in HIV-1 infection by promoting the rapid replication of HIV. Research surrounding Tat protein function has mainly focused on inhibition of the immune system, promotion of growth of vascular endothelial cells, and nervous system damage. To date, very little research has addressed the role of Tat in circadian rhythms. Previous studies in our lab have found that the concentration of Tat protein in HIV patients’ blood was positively correlated with patients’ sleep quality and melatonin concentrations. In this study, we applied a Tat expression plasmid in mice. Result demonstrated that the locomotor activities of mice and the concentration of melatonin were significantly increased. Alternatively, the expression ofClockgene was markedly decreased. On the other hand, the expression of theCry1gene was significantly increased, while the expression ofBmal1andPer1genes exhibited no significant difference. [ABSTRACT FROM PUBLISHER]

The circadian rhythm is one of the basic systems in an organism. It helps the organism maintain harmony with the daily changes of the external environment to ensure proper physiological activities. Previous studies from our laboratory have indicated that the miR-29a/b/c can bind to the circadian clock gene hPer1 at the 3 ′UTR region and regulate its mRNA and protein expression, affecting various organismal physiological processes. Meanwhile, it has been reported that the circadian gene Per plays a role in the regulation of the early growth response gene Egr2, which plays an important role during midbrain development. Here, we confirmed that miR-29a/b/c regulates Egr2 function through mPer1 binding, which elucidates a novel connection between mPer1 and Egr2. [ABSTRACT FROM PUBLISHER]

Gravity heavily influences living organisms on earth including their circadian rhythm, which is fundamentally important for coordinately physiology in organisms as diverse as cyanobacteria, fungus and humans. Numerous researches have revealed that microgravity in outer space can affect circadian rhythm of astronauts and rodent animals, but the mechanism remains unknown. Using rotary cell culture system to simulate microgravity environment, we investigated the role of simulated microgravity in regulating the circadian rhythm of NIH3T3 cells. Our experiments found that simulated microgravity can not only influence the mRNA level of some core circadian genes, but also modify the circadian rhythm ofPer1andPer2synchronized after phorbol myristate acetate treatment. Remarkably, MEK/ERK pathway was transiently activated after a 2-h simulated microgravity treatment, with a significant upregulation ofKras,Raf1and p-ERK1/ERK2. Moreover, U0126, a selective inhibitor of MEK/ERK pathway, could disrupt the circadian rhythm ofPer1andPer2synchronized after simulated microgravity treatment. Together, our results unveil that simulated microgravity could act like a zeitgeber to influence the circadian rhythm of NIH3T3 by acting on MEK/ERK pathway, indicating that MEK/ERK pathway may act as a bridge which connects cells mechanotransduction pathway and circadian rhythm regulation. [ABSTRACT FROM PUBLISHER]

Traumatic brain injury (TBI) generally influences circadian rhythms and has been implicated changes in circadian rhythm. Whether TBI-induced changes in circadian rhythm may affect the prognosis or recovery from TBI remains to be investigated. Sixty-two patients with TBI were continuously monitored for intracranial pressure (ICP) during the first 24 hours after the implantation of ICP monitor. The data from each patient were analyzed using the least squares fit of a 24-h cosine function by single cosinor method. Parameters of circadian A (Amplitude)/M (MESOR) were used to evaluate the circadian rhythm of the patients. Student’st-test and Pearson’s chi-squared test were utilized to analyze the differences between good prognosis group and poor prognosis. A linear regression analysis was then applied to calculate the correlation between circadian A/M of ICP and Glasgow Coma Scale (GCS) before discharge, the Extended Glasgow Outcome Scale (GOS-E), the dosage of mannitol, and time spent in the intensive care unit (ICU), respectively. The results demonstrated that circadian A/M of patients’ ICP exhibited a positive correlation with GCS scores taken before discharge, GOS-E scores, and was negatively correlated with the amount of mannitol, and time spent in the ICU. We conclude that changes in the ICP circadian rhythm in TBI patients could reflect an internal signal of brain damage and, therefore, may be useful to predict a patient’s prognosis and recovery from TBI. [ABSTRACT FROM PUBLISHER]

Emerging evidences have shown that circadian rhythm can also be regulated by a nontranscriptional mechanism. In this study, we investigated thein vitrorhythmic characters of peroxiredoxin2 (PRX2) and nicotinamide adenine dinucleotide (NAD) in red blood cells (RBCs) and/or in heparin anticoagulated plasma under different light conditions. Our results showed that NAD had a free-running rhythm in accordance with that of PRX2 in RBCs, and both rhythms were liable to light entrainment. A decreased expression of PRX2 in RBCs and a notable phase delay of locomotor activity were found in mice intravenously injected with NAD, suggesting that NAD in nontranscriptional oscillating systems could receive the light zeitgeber signal and influence the expression of PRX2. NAD may also act as a nonphotic zeitgeber, which could make an advanced phase shift ofper2, entraining the transcriptional oscillating system to make these two oscillating systems synchronized in a very short time. This zeitgeber might be of great significance as a mediator between these two oscillating systems. [ABSTRACT FROM AUTHOR]

To study the effect of L-arginine on circadian system, ICR mice and AR4-2J cells were treated with L-arginine. The locomotor activity of the mice intraperitoneally injected with a high dose of L-arginine was observed. We also detected effects of arginine on Per1 and Per2 gene expressions in AR4-2J cells. Double-plotted actograms were applied to analyze the locomotor activity of the mice, and reverse transcription-polymerase chain reaction (RT-PCR) was used in analysis of the expression of the major circadian genes Per1 and Per2 in AR4-2J cells. The results presented here indicate that the intraperitoneal injection of L-arginine would strongly disrupt the circadian rhythm of locomotor activity in mice. While the co-culture of L-arginine would also disturb the circadian rhythm of Per1 and Per2 genes which were induced by PMA in AR4-2J cells, the expression levels of both genes greatly decreased. [ABSTRACT FROM AUTHOR]