2.Feeding rhythm entrains circadian metabolism genes, but not the circadian clock, in brown adipose tissue.
Jiang-Hui CHEN ; Mei-Yu ZHOU ; Rong-Feng HUANG ; Hao-Ran XIN ; Shu-Ting CHENG ; Min-Dian LI ; Shi-Fei TONG
Acta Physiologica Sinica 2022;74(5):726-736
The central circadian clock and feeding rhythm coordinately reset peripheral circadian clocks. Emerging evidence suggests that feeding rhythm resets peripheral circadian clocks in a tissue-specific manner. This study aimed to determine whether and how feeding rhythm regulates circadian rhythms of the circadian clock and metabolic genes in brown adipose tissue (BAT). We applied different regimens of time-restricted feeding (TRF) in wildtype and Per1/2 deficient C57BL/6 mice, and quantified the effects of sex, treatment duration, constant light, and circadian clock on circadian rhythms of the BAT circadian clock and metabolic genes by RT-qPCR; Representative circadian clock genes are Bmal1, Nr1d1, Dbp, and Per2, and representative metabolic genes are uncoupling protein 1 (Ucp1), 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (Pfkfb3) that controls the flux through glycolysis, pyruvate dehydrogenase kinase isozyme 4 (Pdk4) gating the tricarboxylic acid cycle, and carnitine palmitoyltransferase 1A (Cpt1a) that controls mitochondrial fatty acid oxidation. The results showed that, daytime-restricted feeding (DRF) moderately shifted the phase of the BAT circadian clock in female mice within 7 or 36 d, and resulted in the loss of circadian rhythm in Dbp and Per2 transcripts in males. DRF induced de novo oscillation of the Ucp1 transcript, and shifted the phase of representative metabolic genes, such as Pfkfb3, Pdk4, and Cpt1a, more than 7 h. Constant light is known to disrupt the synchrony of the central circadian clock. The results showed that constant light promoted phase entrainment of the circadian clock by DRF in BAT, but abolished the oscillation of the metabolic genes (except for Pdk4). Despite combined treatment with Per1/2 deficiency and constant darkness, DRF was sufficient to drive circadian rhythms of Bmal1 and Dbp, but not those of Nr1d1, Ucp1, Pfkfb3, and Cpt1a. Overall, the circadian clock of BAT has weak adaptation to altered feeding rhythms and sex differences. The central circadian clock antagonizes DRF in the entrainment of the BAT circadian clock, whereas DRF resets circadian rhythms of metabolic genes, such as Ucp1, Pfkfb3, and Cpt1a, in a circadian clock-dependent manner.
Female
;
Male
;
Animals
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Mice
;
Mice, Inbred C57BL
;
Circadian Clocks
;
Adipose Tissue, Brown
;
ARNTL Transcription Factors
;
Circadian Rhythm
3.BMAL1 functions as a cAMP-responsive coactivator of HDAC5 to regulate hepatic gluconeogenesis.
Jian LI ; Sihan LV ; Xinchen QIU ; Jiamin YU ; Junkun JIANG ; Yalan JIN ; Wenxuan GUO ; Ruowei ZHAO ; Zhen-Ning ZHANG ; Chao ZHANG ; Bing LUAN
Protein & Cell 2018;9(11):976-980
ARNTL Transcription Factors
;
deficiency
;
metabolism
;
Animals
;
Cyclic AMP
;
metabolism
;
Gluconeogenesis
;
Glucose
;
biosynthesis
;
HEK293 Cells
;
Histone Deacetylases
;
metabolism
;
Humans
;
Liver
;
metabolism
;
Mice
;
Mice, Knockout
4.The role of clock gene BMAL1 in exercise-induced skeletal muscle injury recovery.
Ze-Ting FU ; Yu XIA ; Hai-Li DING
Chinese Journal of Applied Physiology 2022;38(3):220-226
Objective: To investigate the role of clock gene BMAL1 in exercise-induced skeletal muscle injury recovery. Methods: Two hundred and eight 8-week-old SD rats were randomly divided into the control group (Group C, n=104) and the exercise group (Group E, n=104). Group E performed a 90-minute downhill run on the treadmill. After exercise, the gastrocnemius muscle of 8 rats in Group C and Group E were collected at 0 h, 6 h, 12 h, 18 h, 24 h, 30 h, 36 h, 42 h, 48 h, 54 h, 60 h, 66 h and 72 h. The expression of skeletal muscle core clock gene, BMAL1 was measured by real-time fluorescence quantitative PCR. The parameters of fitting cosine curve were obtained by cosine analysis software circacompare (R package), and the change trend of rhythmic oscillation was analyzed. The ultrastructure of skeletal muscle fibers was observed by transmission electron microscope. The expressions of skeletal muscle BMAL1 and DESMIN were detected by Western blot; Immunofluorescence was used to observe the localization and contents of BMAL1 and DESMIN. Results: In Group C, three complete circadian rhythm cycles of mRNA BMAL1 were observed within 72 hours; in Group E, the circadian rhythm of BMAL1 mRNA disappeared at 0 h~24 h. Compared with Group C, the expression level of BMAL1 mRNA was significantly increased at 0 h, 6 h, 12 h, and 18 h after exercise in Group E (P<0.05), and the expression of BMAL1 protein was significantly increased at 0 h and 12 h after exercise(P<0.05), and recovered to the level of that in Group C from 24 h to 72 h(P>0.05). The expression of DESMIN protein was decreased at 0 h and 12 h after exercise(P<0.05), gradually increased at 24 h, increased significantly at 48 h(P<0.01), and recovered to the control level at 72 h (P>0.05). In Group E, BMAL1 and DESMIN were co-localized at 0 h, 12 h, and 24 h after exercise; the colocalization at 0 h~24 h showed a trend of first decreasing and then increasing, and the fluorescence intensity at 24 h reached the highest value. Conclusion: The post-exercise clock gene BMAL1 may be involved in the enhanced synergy of regulating the cytoskeletal protein DESMIN, it is thus related to the promotion of muscle fiber structure recovery.
ARNTL Transcription Factors/metabolism*
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Animals
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Desmin/metabolism*
;
Muscle, Skeletal/physiology*
;
Physical Conditioning, Animal/adverse effects*
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RNA, Messenger/metabolism*
;
Rats
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Rats, Sprague-Dawley
5.Correlation analysis of clock genes and MEN2 medullary thyroid carcinoma.
Ya Kui MOU ; Chao REN ; Yu Mei LI ; Guo Hua YU ; Gui Bin ZHENG ; Hong SONG ; Cong Xian LU ; Ru Xian TIAN ; Xin Cheng SONG
Chinese Journal of Otorhinolaryngology Head and Neck Surgery 2022;57(9):1079-1086
Objective: To investigate the correlation between CLOCK and BMAL1 genes and MEN2 medullary thyroid carcinoma (MTC). Methods: Thirteen cases with MEN2 MTC and thirteen cases with non-MEN2 MTC were selected who were treated in the Yantai Yuhuangding Hospital between January 2013 and September 2021. Clinical indicators such as blood calcitonin level, tumor diameter and metastatic lymph node of patients were collected. The expression differences of CLOCK and BMAL1 between MEN2 MTC and para-carcinoma tissue as well as between MEN2 MTC and non-MEN2 MTC were detected by immunohistochemistry and qPCR. The correlation between lymph node metastasis and CLOCK or BMAL1 expression was analyzed. Protein-protein interaction (PPI) network analysis combined with qPCR and correlation analysis was used to explore the expression regulation relationship between RET and circadian clock genes. The rhythm disorder of MEN2 cells was verified by lipopolysaccharide cell stimulation experiment after dexamethasone rhythm synchronization. Results: MEN2 MTC exhibited typical RET gene mutation. The mean blood calcitonin level, the tumor diameter and the number of metastatic lymph nodes of patients with MEN2 MTC were higher than those of patients with non-MEN2 MTC (t value was 2.76, 2.53, 2.26, all P<0.05). Immunohistochemical results showed that the expression levels of CLOCK and BMAL1 in MEN2 MTC were higher than those in non-MEN2 MTC, while negatively expressed in para-cancerous thyroid follicle. qPCR displayed that the expression of CLOCK gene in cancer tissues was higher than that in non-MEN2 MTC and para-cancerous tissues (t value was 2.68 and 2.86, all P<0.05); the expression of BMAL1 gene in MEN2 MTC was higher than that in non-MEN2 MTC and para-cancerous tissues (t value was 2.21 and 2.35, all P<0.05). Correlation analysis showed that the expression levels of CLOCK and BMAL1 genes were positively correlated with the number of lymph node metastases in patients with MEN2 MTC (r=0.65, P<0.001; r=0.52, P=0.005). PPI network analysis indicated that the expression of CLOCK gene was positively correlated with the abnormal expression of RET gene (r=0.96, P<0.001). With lipopolysaccharide to stimulate cultured cells in vitro after dexamethasone rhythm synchronization, the expressions of CLOCK and BMAL1 in MEN2 MTC cells (0.47±0.22 and 2.60±1.48) at 12 hours of synchronization were significantly lower than those in para-cancerous tissues (1.70±1.62 and 8.23±2.52), the difference was statistically significant(t=5.04, P=0.007; t=3.34, P=0.029). Conclusion: CLOCK and BMAL1 are correlated with the occurrence and development of MEN2 MTC, and may be potential targets for the development of new therapeutic strategies for MEN2 MTC.
ARNTL Transcription Factors/genetics*
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CLOCK Proteins/genetics*
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Calcitonin
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Carcinoma, Neuroendocrine/genetics*
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Dexamethasone
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Humans
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Lipopolysaccharides
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Lymphatic Metastasis
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Multiple Endocrine Neoplasia Type 2a/genetics*
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Thyroid Neoplasms/surgery*
6.The Circadian System Is Essential for the Crosstalk of VEGF-Notch-mediated Endothelial Angiogenesis in Ischemic Stroke.
Yuxing ZHANG ; Xin ZHAO ; Chun GUO ; Ying ZHANG ; Fukang ZENG ; Qian YIN ; Zhong LI ; Le SHAO ; Desheng ZHOU ; Lijuan LIU
Neuroscience Bulletin 2023;39(9):1375-1395
Ischemic stroke is a major public health problem worldwide. Although the circadian clock is involved in the process of ischemic stroke, the exact mechanism of the circadian clock in regulating angiogenesis after cerebral infarction remains unclear. In the present study, we determined that environmental circadian disruption (ECD) increased the stroke severity and impaired angiogenesis in the rat middle cerebral artery occlusion model, by measuring the infarct volume, neurological tests, and angiogenesis-related protein. We further report that Bmal1 plays an irreplaceable role in angiogenesis. Overexpression of Bmal1 promoted tube-forming, migration, and wound healing, and upregulated the vascular endothelial growth factor (VEGF) and Notch pathway protein levels. This promoting effect was reversed by the Notch pathway inhibitor DAPT, according to the results of angiogenesis capacity and VEGF pathway protein level. In conclusion, our study reveals the intervention of ECD in angiogenesis in ischemic stroke and further identifies the exact mechanism by which Bmal1 regulates angiogenesis through the VEGF-Notch1 pathway.
Rats
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Animals
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Vascular Endothelial Growth Factor A/pharmacology*
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Brain Ischemia/metabolism*
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Ischemic Stroke
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Signal Transduction
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ARNTL Transcription Factors/pharmacology*
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Neovascularization, Physiologic/physiology*
7.Basic science review on circadian rhythm biology and circadian sleep disorders.
Annals of the Academy of Medicine, Singapore 2008;37(8):662-668
The sleep-wake cycle displays a characteristic 24-hour periodicity, providing an opportunity to dissect the endogenous circadian clock through the study of aberrant behaviour. This article surveys the properties of circadian clocks, with emphasis on mammals. Information was obtained from searches of peer-reviewed literature in the PUBMED database. Features that are highlighted include the known molecular components of clocks, their entrainment by external time cues and the output pathways used by clocks to regulate metabolism and behaviour. A review of human circadian rhythm sleep disorders follows, including recent discoveries of their genetic basis. The article concludes with a discussion of future approaches to the study of human circadian biology and sleep-wake behaviour.
ARNTL Transcription Factors
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Animals
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Basic Helix-Loop-Helix Transcription Factors
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physiology
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CLOCK Proteins
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Circadian Rhythm
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genetics
;
physiology
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Humans
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Neurons, Afferent
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physiology
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Neurons, Efferent
;
physiology
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Polymorphism, Single Nucleotide
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Sleep Disorders, Circadian Rhythm
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genetics
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physiopathology
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Suprachiasmatic Nucleus
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cytology
;
physiology
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Trans-Activators
;
physiology
8.Changes of biological clock protein in neonatal rats with hypoxic-ischemic brain damage.
Yong-Fu LI ; Mei-Fang JIN ; Bin SUN ; Xing FENG
Chinese Journal of Contemporary Pediatrics 2013;15(1):62-66
OBJECTIVETo study the effects of biological clock protein on circadian disorders in hypoxic-ischemic brain damage (HIBD) by examining levels of CLOCK and BMAL1 proteins in the pineal gland of neonatal rats.
METHODSSeventy-two 7-day-old Sprague-Dawley (SD) rats were randomly divided into sham-operated and HIBD groups. HIBD model was prepared according to the modified Levine method. Western blot analysis was used to measure the levels of CLOCK and BMAL1 in the pineal gland at 0, 2, 12, 24, 36 and 48 hours after operation.
RESULTSBoth CLOCK and BMAL levels in the pineal gland increased significantly 48 hours after HIBD compared with the sham-operated group (P<0.05). There were no significant differences in levels of CLOCK and BMAL proteins between the two groups at 0, 2, 12, 24 and 36 hours after operation (P>0.05).
CONCLUSIONSLevels of CLOCK and BMAL1 proteins in the pineal gland of rats increase significantly 48 hours after HIBD, suggesting that both CLOCK and BMAL1 may be involved the regulatory mechanism of circadian disorders in rats with HIBD.
ARNTL Transcription Factors ; analysis ; physiology ; Animals ; Animals, Newborn ; CLOCK Proteins ; analysis ; physiology ; Chronobiology Disorders ; etiology ; Female ; Hypoxia-Ischemia, Brain ; metabolism ; Male ; Pineal Gland ; chemistry ; Rats ; Rats, Sprague-Dawley ; Time Factors
9.BMAL1 gene regulates the osteogenic differentiation of bone marrow mesenchymal stem cells.
Xiaoguang LI ; Xiao-long GUO ; Bin GUO
West China Journal of Stomatology 2016;34(3):312-316
Periodontitis is a chronic infective disease characterized as the destruction of the supporting tissues of the teeth. Bone marrow mesenchymal stem cells, which are ideal adult stem cells for the regeneration of supporting tissues, may play important roles in restoring the structure and function of the periodontium and in promoting the treatment of periodontal disease. As a consequence, the characteristics, especially osteogenic differentiation mechanism, of these stem cells have been extensively investigated. The regulation of the physiological behavior of these stem cells is associated with BMAL1 gene. This gene is a potential treatment target for periodontal disease, although the specific mechanism remains inconclusive. This study aimed to describe the characteristics of BMAL1 gene and its ability to regulate the osteogenic differentiation of stem cells.
ARNTL Transcription Factors
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genetics
;
Adult
;
Adult Stem Cells
;
Bone Marrow Cells
;
physiology
;
Cell Differentiation
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Hematopoietic Stem Cells
;
Humans
;
Mesenchymal Stromal Cells
;
physiology
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Osteogenesis
;
physiology
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Periodontal Ligament
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Periodontitis
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Periodontium
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Regeneration
;
Tooth
10.Circadian regulation of low density lipoprotein receptor promoter activity by CLOCK/BMAL1, Hes1 and Hes6.
Yeon Ju LEE ; Dong Hee HAN ; Youngmi Kim PAK ; Sehyung CHO
Experimental & Molecular Medicine 2012;44(11):642-652
Low density lipoprotein receptor (LDLR) plays an important role in the cholesterol homeostasis. We examined the possible circadian regulation of LDLR and mechanism(s) underlying it. In mice, blood glucose and plasma triglyceride, total and high density lipoprotein cholesterol varied distinctively throughout a day. In addition, LDLR mRNA oscillated in the liver in a functional clock-dependent manner. Accordingly, analysis of human LDLR promoter sequence revealed three putative E-boxes, raising the possible regulation of LDLR expression by E-box-binding transcription factors. To test this possibility, human LDLR promoter reporter constructs were transfected into HepG2 cells and the effects of CLOCK/BMAL1, Hes1, and Hes6 expression were analyzed. It was found that positive circadian transcription factor complex CLOCK/BMAL1 upregulated human LDLR promoter activity in a serum-independent manner, while Hes family members Hes1 and Hes6 downregulated it only under serum-depleted conditions. Both effects were mapped to proximal promoter region of human LDLR, where mutation or deletion of well-known sterol regulatory element (SRE) abolished only the repressive effect of Hes1. Interestingly, hes6 and hes1 mRNA oscillated in an anti-phasic manner in the wild-type but not in the per1-/-per2-/- mouse. Comparative analysis of mouse, rat and human hes6 genes revealed that three E-boxes are conserved among three species. Transfection and site-directed mutagenesis studies with hes6 reporter constructs confirmed that the third E-box in the exon IV is functionally induced by CLOCK/BMAL1. Taken together, these results suggest that LDLR expression is under circadian control involving CLOCK/BMAL1 and Hes family members Hes1 and Hes6.
ARNTL Transcription Factors/physiology
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Animals
;
Base Sequence
;
Basic Helix-Loop-Helix Transcription Factors/*genetics/metabolism/physiology
;
CLOCK Proteins/physiology
;
Cholesterol/blood
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*Circadian Rhythm
;
E-Box Elements
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Exons
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*Gene Expression Regulation
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Hep G2 Cells
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Homeodomain Proteins/*genetics/metabolism/physiology
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Homeostasis
;
Humans
;
Liver/metabolism
;
Male
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Mice
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Mice, Inbred C57BL
;
*Promoter Regions, Genetic
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Receptors, LDL/*genetics/metabolism
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Repressor Proteins/*genetics/metabolism/physiology
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Transcription, Genetic