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Objective:To investigate the effect of circadian rhythm changes on the expression of retinoic acid-related orphan receptors (RORs) and the RORs agonist SR1078 on corneal epithelial wound repair.Methods:A total of 228 SPF C57BL/6 female mice aged 6-8 weeks old were selected, and 180 mice were divided into the normal circadian rhythm group, full-day group, full-night group, 12-hour reversed circadian rhythm group and 3-week reversed circadian rhythm group, with 36 mice in each group.The remaining 48 mice were randomly divided into phosphate buffered saline (PBS) control group and SR1078 group by random number table method, with 24 mice in each group.According to grouping, the mice were placed in a light box where the light (light intensity of 300 lx) and dark time could be controlled.The light time of the normal circadian rhythm group, the PBS control group and the SR1078 group in the light box was from 7: 00 to 19: 00, and the dark time was from 19: 00 to 7: 00 the next day.According to the Zeitgeber Time method, the starting time of light at 7: 00 was recorded as ZT0, and the time of closing light at 19: 00 was recorded as ZT12.Real-time fluorescence quantitative polymerase chain reaction was used to detect the relative expression levels of RORα and RORγ mRNA at ZT1, ZT5, ZT9, ZT13, ZT17, ZT21 in the five groups.In the PBS control group and SR1078 group, a golf-like knife was used to establish the mouse corneal epithelial injury model, and the model eyes were administered with drugs once every 6 hours according to the grouping.The corneal epithelial defect area was measured with Adobe Photoshop CC2019 software, and the corneal epithelial defect rate was calculated and compared between the two groups.The correlation between the relative expression levels of RORα and RORγ mRNA in mice corneal epithelium of the five groups and corneal epithelial defect rate in the PBS control group and SR1078 group was analyzed.The corneal epithelium repair was observed by whole cornea spreading and immunofluorescence staining, and the number of corneal epithelial dividing cells in the PBS control group and the SR1078 group was calculated and compared.The use and care of animals complied with the ARVO statement.This study protocol was approved by the Laboratory Animal Ethics Committee of Jinan University (No.JN-A-2002-01).Results:Compared with the normal circadian rhythm group, the relative expression levels of RORα/RORγ mRNA in the full-day group, full-night group, 12-hour reversed cirdian rhythym group and 3-week reversed cirdian rhythym group showed an overall decreasing trend.There was a statistically significant difference in the corneal epithelial defect rate between the PBS control group and the SR1078 group at different time points after modeling ( Fgroup=74.01, P<0.001; Ftime=5 171.48, P<0.001). Twelve hours after modeling, the corneal epithelial defect rate in the SR1078 group was significantly lower than that in the PBS control group, and the difference was statistically significant ( P<0.05). The relative expression levels of RORα and RORγ mRNA in corneal tissue was moderately positively correlated with the corneal epithelial defect rate in mice ( r=0.614, 0.537; both at P<0.01); The regression equation of the straight line between the relative expression level of RORα mRNA and the change in corneal epithelial defect rate was Y=33.153X-43.052 ( F=20.58, P<0.001), and the linear regression equation between the relative expression level of RORγ mRNA and the change of corneal epithelial defect rate was Y=2.764X-1.364 ( F=13.11, P<0.001). There was a significant overall difference in the number of corneal epithelial dividing cells at various time points following modeling between the PBS control group and the SR1078 group ( Fgroup=160.55, P<0.001; Ftime=83.57, P<0.001). The number of dividing cells in the SR1078 group was significantly less than that in the PBS control group at 24, 30, and 36 hours following modeling, and the differences were statistically significant (all at P<0.05). Conclusions:Circadian rhythm changes reduce the expression of RORα and RORγ mRNA in the mouse cornea.SR1078 can promote the expression of RORα and RORγ mRNA in corneal epithelium to decrease the number of mouse corneal epithelial dividing cells, and inhibit the repair after corneal trauma.
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Objective:To investigate the effect of BMAL1 gene on the proliferation, migration and invasion ability of radiation-resistant nasopharyngeal carcinoma cell line (5-8FR) and the molecular mechanism. Methods:A multi-target click model was constructed for radiation-resistant nasopharyngeal carcinoma cell line 5-8FR by low-dose fractionated irradiation, and the results of clone formation assay were used to fit the multi-target click model and calculate the sensitization ratio of radiotherapy. The expression levels of PI3K/Akt/MMP-2/9 signaling pathway-related proteins in 5-8FR and control 5-8F cell lines were detected by Western blot. The overexpression and knockdown vectors of BMAL1 gene were constructed and transfected with 5-8F and 5-8F cell lines, respectively. The BMAL1 gene overexpression (pcDNA-BMAL1) and its control (pcDNA) and interference (BMAL1-shRNA) and control (con-shRNA) cell lines were stably transfected with nasopharyngeal carcinoma cell line 5-8F and radiation-resistant cell line 5-8FR, respectively. Western blot was performed to verify the infection efficiency and detect the changes of PI3K/Akt/MMP-2/9 signaling pathway-related proteins after overexpression or interference of BMAL1 gene in both groups of cells. CCK-8 assay, cell scratch test and Transwell chamber test were conducted to investigate the proliferation, migration and invasion capabilities of 5-8FR cell line after overexpression or interference of BMAL1 gene. Results:BMAL1 gene expression was down-regulated, and those of PI3K/Akt pathway proteins and downstream related molecules of MMP-2 and MMP-9 were up-regulated, and TIMP-2 and TIMP-1 expression was down-regulated in nasopharyngeal carcinoma radiation-resistant cell lines. Overexpression of BMAL1 gene inhibited the expression of PI3K/Akt pathway proteins and downstream related molecules of MMP-2 and MMP-9, promoted the expression of TIMP-2 and TIMP-1, and inhibited the proliferation, migration and invasion capabilities of radiation-resistant nasopharyngeal carcinoma cells, while interference with BMAL1 gene yielded the opposite results. Conclusions:BMAL1 gene can reverse the expression of PI3K/Akt/MMP-2/9 signaling pathway-related proteins in radiation-resistant nasopharyngeal carcinoma cell lines and inhibit the proliferation, migration and invasion capabilities of radiation-resistant nasopharyngeal carcinoma cell lines.
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Mammalian bone is constantly metabolized from the embryonic stage, and the maintenance of bone health depends on the dynamic balance between bone resorption and bone formation, mediated by osteoclasts and osteoblasts. It is widely recognized that circadian clock genes can regulate bone metabolism. In recent years, the regulation of bone metabolism by non-coding RNAs has become a hotspot of research. MicroRNAs can participate in bone catabolism and anabolism by targeting key factors related to bone metabolism, including circadian clock genes. However, research in this field has been conducted only in recent years and the mechanisms involved are not yet well established. Recent studies have focused on how to target circadian clock genes to treat some diseases, such as autoimmune diseases, but few have focused on the co-regulation of circadian clock genes and microRNAs in bone metabolic diseases. Therefore, in this paper we review the progress of research on the co-regulation of bone metabolism by circadian clock genes and microRNAs, aiming to provide new ideas for the prevention and treatment of bone metabolic diseases such as osteoporosis.
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Animals , Circadian Clocks/genetics , Circadian Rhythm/genetics , Mammals/genetics , MicroRNAs/genetics , Osteogenesis/genetics , Osteoporosis/geneticsABSTRACT
RESUMEN Objetivo: Comparar el estado nutricional, cronotipo y conducta alimentaria que presenta un sujeto que posee simultáneamente los polimorfismos rs3749474T y rs4864548A con respecto a grupos de sujetos que poseen solo uno de dichos polimorfismos. Métodos: La presencia de los polimorfismos se determinó mediante PCR. Se determinó IMC, riesgo cardiovascular y porcentaje de grasa según lo descrito por Durnin y Womersley y la ecuación de Siri. Se aplicó el cuestionario TFEQ-P19 adaptado a población chilena y el cuestionario Horne-Ostberg. Resultados: El sujeto con ambos polimorfismos presentó obesidad, riesgo cardiovascular y cronotipo trasnochador. Sus puntajes en las dimensiones de alimentación sin control y alimentación emocional fueron bajos. Su puntaje en cuanto a la restricción cognitiva fue el más alto. Conclusiones: La presencia del haplotipo TA (rs3749474T; rs4864548A) aumentaría la posibilidad de tener un cronotipo de tipo de tipo trasnochador, riesgo de obesidad y riesgo cardiovascular asociado a los centímetros de cintura.
ABSTRACT Objective: To compare the nutritional status, chronotype and eating behavior of a subject who simultaneously has the rs3749474T and rs4864548A polymorphisms to groups of subjects who have only one of these polymorphisms. Methods: The presence of the polymorphisms was established by PCR. BMI, cardiovascular risk, and fat percentage were determined as described by Durnin and Womersley and the Siri equation. The TFEQ-P19 questionnaire (adapted to the Chilean population) and the Hor-ne-Ostberg questionnaire were applied. Results: The subject with both polymorphisms presented obesity, cardiovascular risk, and late-night chronotype. Its scores on the dimensions of uncontrolled feeding and emotional feeding were low, and its score for cognitive restriction was the highest. Conclusions: The presence of the TA haplotype (rs3749474; rs4864548A) would increase the possibility of having a late-night type chronotype, risk of obesity, and cardiovascular risk associated with waist centimeters.
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Circadian rhythms have been shown to regulate several physiological functions including body temperature, sleep/wake cycle, physical activity, and cognition. These functions are controlled by circadian clock genes, and the circadian clock system in the body is classified into two clocks which are the central clock in the suprachiasmatic nucleus (SCN) of the hypothalamus and peripheral clocks in peripheral tissues such as the liver and skeletal muscle. Therefore, many researchers are conducting basic and applied research on the relationship between circadian rhythm in peripheral tissues and physiological functions including sports performance and effects of acute exercise and exercise training. On the other hand, it has been shown that abnormal circadian rhythms and disturbance of circadian rhythms can lead to the development of several diseases such as diabetes, cancer, sarcopenia, depression, and dementia. Thus, it is also important to regulate individual circadian rhythm by considering for timing of exercise and daily physical activity. Exercise and physical activity are found to have an influence on circadian rhythms regulation (Chrono-exercise) and accumulate evidences between timing of exercise and health outcomes. This review aims to introduce evidence for chrono-exercise and suggests the importance for considering the timing of exercise and physical activity.
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In mammals, the circadian rhythms have been shown to regulate several physiological functions, including body temperature, sleep-wake behavior, physical activity, hormonal secretions, and metabolism. These processes are controlled by circadian clock genes, and abnormal circadian rhythms are associated with the development of obesity, diabetes, and lifestyle-related diseases. In addition, the timing of behaviors such as food intake, exercise, and stress influence circadian rhythms, including clock gene expression in peripheral tissues. Therefore, the interaction between nutrition and the circadian clock is so-called “chrono-nutrition” is poised to become an important research field of chronobiology. In this review, we review the effects of a timed-nutrition on circadian clocks and their timing-dependent effects on physiological functions.
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OBJECTIVE：To study the improvement effects of isopimpinelline on p-chlorophenylalanine（PCPA）-induced pineal injury model rats and its effect on expression of biological clock gene. METHODS ：Totally 60 rats were divided into blank control group（2％ polysorbate solution），model control group （2％ polysorbate solution），positive control group （melatonin，10 mg/kg） and isopimpinelline high-dose ，medium-dose and low-dose groups （3，1.5，0.75 mg/kg）. Except for blank control group ，rats in other groups were given PCPA intraperitoneally （450 mg/kg）to establish pineal injury model. After modeling finished ，they were given relevant medicine intragastrically ，once a day ，for consecutive 7 d. On the 6th day of administration ，the sleep latency and sleep duration of rats in each group were investigated by pentobarbital sodium coordination sleep test ；after last administration ， ELISA assay was used to determine the serum level of melatonin in rats. Fluorescence microscope and electron microscope were used to observe the pathological tissue and cell ultrastructure changes of the pineal gland. RT-qPCR was used to detect the mRNA expressions of biological clock gene Clock，Bmal1，Per1，Per2，Per3，Cry1，Cry2 in pineal gland of rats. RESULTS ：Compared with blank control group ，model control group had significantly longer sleep latency （P＜0.05）；serum melatonin ，mRNA expressions of Bmal1 and Per1 in pineal gland were significantly decreased （P＜0.05 or P＜0.01）while mRNA expression of Per3 was increased significantly （P＜0.05）. The pineal gland cell arrangement disorder ，nuclear pyknosis ，vacuolar degeneration increased and cell number decreased significantly ；mitochondria swollen ，cristae broken and pyknosis were observed. Compared with model control group ，the sleep latency of isopimpinelline high-dose group was shortened significantly （P＜0.05），sleep duration time was prolonged significantly （P＜0.05）；the levels of melatonin in serum ，mRNA expressions of Clock，Bmal1， Per1，Cry1 and Cry2 in pineal gland of rats were increased significantly （P＜0.05 or P＜0.01）. In isopimpinelline medium-dose group，the sleep latency was shortened significantly （P＜0.05）；the levels of melatonin in serum and mRNA expressions of Clock， Bmal1，Per1，Cry1，Cry2 in pineal gland were increased significantly （P＜0.05 or P＜0.01），while mRNA expression of Per3 was decreased significantly （P＜0.05）. In isopimpinelline low-dose group ，the levels of mRNA expressions of Clock，Bmal1，Per2 and Cry2 were increased significantly （P＜0.05），while mRNA expression of Per3 was decreased significantly （P＜0.05）. Cell arrangement disorder was improved and nuclear pyknosis vacuole degeneration was decreased to some extent in isopimpinelline groups；mitochondria swelled ，cristae fractured ，and pyknosis decreased to some extent. CONCLUSIONS ：Isopimpinelline can improve PCPA-induced pineal gland injury in rats ；it can up-regulate the expressions of positive regulators Clock，Bmal1 and negative regulators Per1，Per2，Cry1，Cry2，while down-regulate the expression of negative regulator Per3.
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Objective To investigate the function of sorafenib on the growth of hepatocellular carcinoma by establishing subcutaneous transplantation tumor model with nude mice.To explore the effect of sorafenib on circadian clock gene expression in hepatoma cells.Methods Mouse tumor model was established by implanting hepatocarcinoma cell (HepG2) subcutaneously in Balb/C nude mice.Sixteen experimental mice were randomly divided into two groups:sorafenib treatment group (n =8) and solvent control group (n =8).The nude mice were treated with sorafenib (100 mg/kg) and DMSO daily by intragastric administration,respectively.The volume of tumors was recorded every 3 days.The expressions level of circadian clock genes (Per1,Per2,Per3,CLOCK,Cry1,Cry2,BMAL1 and CKIε) were detected by real-time polymerase chain reaction (Real-time PCR).The correlations between the size of the xenografts and the expressions of the circadian clock genes were further analyzed.Results Compared with the control group,the tumor size in the sorafenib treatment group were significantly smaller comparing with the control group.Results of Real-time PCR showed that the expression level of Per1,Cry1 and BMAL1 mRNA was remarkably decreased in the treatment group (Per1,P =0.02;Cry1,P =0.002;BMAL1,P =0.035),the differences were statistically significant.Correlation analysis showed that the size of subcutaneous transplantation tumorsin nude mice was negatively correlated with the expressions of Per1,Per2,Cry1 and Cry2 mRNA in control group.While,the size of subcutaneous transplantation tumors was negatively correlated with the expressions of Per2,Per3 and BMAL1 levels in the sorafenib treatment group.Conclusions There is a negative correlation between the expression levels of some biological clock genes and the size of transplantation tumor in nude mice.Sorafenib treatment significantly inhibited the growth of hepatocellular carcinoma in nude mice and down-regulation the expressions of Per1,Cry1 and BMAL1 mRNA in hepatoma cells.
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Objective@#To investigate the role of clock gene Bmal1, Per2 and Egr1 expression in learning and memory undergoing sevoflurane anesthesia after acute sleep deprivation.@*Methods@#72 male SD rats were equally divided into four groups using a random number table (n=18) : normal control group (group Control), do not do any processing; sleep deprivation group (group SD), acute sleep deprivation for 96 h; sevoflurane group (group Sev), suffering 2.5% sevoflurane for 3 h; sleep deprivation+sevoflurane group (group SD+Sev), 96 h sleep deprivation followed by 3 h 2.5% sevoflurane inhalation. The Morris water maze, for spatial memory acquisition test, was used to measure the time percent of target quadrant and numbers of platform-site crossovers before sleep deprivation (T0) and at 1 d (T1), 3 d (T2), 7 d (T3) after inhalation anesthesia. Rats were sacrificed after spatial memory acquisition test. Brain hippocampus samples were obtained for determination of Bmal1, Per2 and Egr1 expression by Western blot, and neuron morphology was observed by the Nissl staining.@*Results@#Compared with Control group, the percentage of time in target quadrant and the numbers of platform-site crossovers were significantly decreased at T1 and T2 in Sev group (P<0.05); and compared with Control group, the percentage of time in target quadrant and the numbers of platform-site crossovers were also significantly decreased at T1, T2 and T3 in SD group rats (P<0.05). Compared with Sev group rats (the percentages of time in target quadrant: T1: (32.37±1.36)%; T2: (30.91±1.26)%; T3: (33.78±2.20)%; the numbers of platform-site crossovers: T1: (4.55±0.39); T2: (3.11±0.37); T3: (3.95±0.34)), the percentages of time in target quadrant (T1: (27.20±1.42)%; T2: (28.19±1.04)%; T3: (30.06±1.22)%) and the numbers of platform-site crossovers (T1: (3.11±0.46); T2: (3.30±0.38); T3: ( 3.20±0.39)) in SD+Sev group rats were significantly decreased at T1, T2 and T3 (all P<0.05). Compared with control group, the levels of hippocampal proteins Bmal1, Per2 and Egr1 were significantly reduced at T1 in Sev group (P<0.05). Compared with control group, the level of hippocampal protein Per2 was significantly increased, but the levels of hippocampal proteins Bmal1 and Egr1 were significantly decreased at T1 and T2 in SD group (P<0.01). Compared with Sev group, the levels of hippocampal proteins Bmal1 and Egr1 were significantly reduced at T1, T2 and T3 in SD+Sev group (P<0.01), and the protein level of hippocampal Per2 was significantly decreased at T1, but then increased at T2 and T3 in SD+Sev group (P<0.01). The hippocampal Nissl staining in CA1 at T2 revealed that there were irregular distribution of pyramidal neurons existed in Sev group, and vacuolar degeneration with vague outlines of pyramidal neurons in SD group, while pyramidal neuron atrophy and few number of Nissl bodies, compared with control group, were observed in SD+Sev group.@*Conclusion@#Acute sleep deprivation following with sevoflurane anesthesia resulted in hippocampal memory impairment, which was associated with abnormal expression of hippocampal Bmal1, Per2 and Egr1.
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Objective To find the effect of alter-expressed PER2 on proliferation,apoptosis and other clock genes expression in human oral squamous cell carcinoma SCC15 cells.Methods Short hairpin RNA interference was used to knockdown PER2 in SCC15 human oral squamous cell carcinoma cells.Flow cytometry analysis was used to testify the cell proliferation and apoptosis.Quantitative real-time PCR was used to testify the mRNA expressions of PER3,BMAL1,DEC1,DEC2,CRY2,TIM,RORα,NPAS2,PER1 and REV-ERBα.Results The proliferation was enhanced and apoptosis was decreased after PER2 knockdown in SCC15 cells (P<0.05).The mRNA expression of PER3,BMAL1,DEC1,DEC2,CRY2,TIM,RORα and NPAS2 was significantly down-regulated,and the mRNA expression of PER1 and REV-ERBα was significantly up-regulated (P<0.05).Conclusions Clock gene PER2 plays an important role in regulating other clock genes of the clock gene network in cancer cells,PER2 knockdown can enhance proliferation and recede apoptosis of cancer cell.
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Objective To explore the effect of 1a,25(OH)2 D3 on circadian clock gene expressions in cardiac myocytes.Methods Cultured cardiac myocytes isolated from 7 -day -old Sprague -Dawley(SD)rats were identified by immunofluorescence.The medium including 1a,25 (OH)2 D3 (final concentrations were 0 nmol/L,1 nmol/L, 10 nmol/L,50 nmol/L and 100 nmol/L)were added to primary myocardial cells to culture for 2 h and then total RNA was extracted.Real -time polymerase chain reaction (RT -PCR)was applied to analyze myocardial cells circadian clock gene (Bmal1,Per2,Rev -erba)transcript levels to determine optimum concentration of 1a,25(OH)2 D3 .Then, the primary myocardial cells cultured for 72 h were divided into 3 groups:the control group was of serum -free culture medium;serum shock group was of DMEMcontaining 50%volume fraction of horse serum cultured 2 h;1a,25(OH)2 D3 treatment group receiving 1a,25 (OH)2 D3 at optimal concentration cultured 2 h.The cells were collected at 7 time points (0 h,4 h,8 h,12 h,16 h,20 h,24 h)and then total RNA was extracted.RT -PCR was applied to analyze circa-dian clock gene (Bmal1,Per2,Rev -erba)transcript levels in the myocardial cells.Results In the presence of 50 nmol/L 1 a,25(OH)2 D3 ,the Bmal1 mRNA expression showed the highest level,but the Per2 and Rev -erba mRNA expression levels were minimum.Compared with the control group,both 1a,25 (OH)2 D3 treatment group and serum shock group caused day -cycle rhythmic oscillation in circadian clock genes(Bmal1,Per2,Rev -erba)in the cardiac myocytes.And the expressions pattern of Bmal1 and Per2 genes were in the opposite phase.While Bmal1 gene expres-sion appeared at peak at 12 h,Per2 gene expression appeared in a trough.Expression of Rev -erba gene trend began to rise at 8 h,and the highest expression level appeared at 12 -16 h.Conclusions 1a,25(OH)2 D3 can affect Bmal1, Per2 and Rev -erba mRNA expressions of circadian clock genes in the cardiac myocytes.
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PURPOSE: Recent studies demonstrated disruption of the circadian clock gene is associated with the development of obesity and metabolic syndrome. Obesity is often caused by the high calorie intake, In addition, the chronic stress tends to contribute to the increased risk for obesity. To evaluate the molecular mechanisms, we examined the expression of circadian clock genes in high fat diet-induced mice models with the chronic stress. METHODS: C57BL/6J mice were fed with a 45% or 60% high fat diet for 8 weeks. Daily immobilization stress was applied to mice fed with a 45% high fat for 16 weeks. We compared body weight, food consumption, hormone levels and metabolic variables in blood. mRNA expression levels of metabolic and circadian clock genes in both fat and liver were determined by quantitative RT-PCR. RESULTS: The higher fat content induced more severe hyperglycemia, hyperlipidemia and hyperinsulinemia, and these results correlated with their relevant gene expressions in fat and liver tissues. Chronic stress had only minimal effects on metabolic variables, but it altered the expression patterns of metabolic and circadian clock genes. These results suggest that the fat metabolism regulates the function of the circadian clock genes in peripheral tissues, and stress hormones may contribute to its regulation.
Subject(s)
Animals , Mice , Body Weight , Circadian Clocks , Diet, High-Fat , Gene Expression , Hyperglycemia , Hyperinsulinism , Hyperlipidemias , Immobilization , Liver , Metabolism , Obesity , RNA, MessengerABSTRACT
Objective To explore the expression of circadian clock gene Per2 in the tissue of esophageal cancer and the relevance between VEGF,cyclin D1.Methods 80 cases with esophageal cancer who received surgical treatment and pathological specimens preserved were selected,and the normal tissues adjacent to carcinoma were used as control.The expression of Per2,VEGF and cyclin D1 were detected by immunohistochemistry,and the data were analyzed.Results Compared the tissue adjacent to carcinoma with carcinoma tissue,Per2,VEGF and cyclin D1 expression had statistically significant differences(x2 =26.86,24.34,23.72,all P ＜0.01).Per2 expression in different differentiation degree and the TMN stages had statistically significant differences (x2 =6.45,7.77,all P ＜ 0.05) ;cyclin D1 expression in lymph node metastasis and TMN stages had statistically significant differences(x2 =12.99,8.99,all P ＜ 0.01) ;VEGF expression in lymph node metastasis and TMN stages had statistically significant differences(x2 =6.68,6.14,all P ＜ 0.01).Per2 expression was negatively correlated with VEGF,cyclin D1 expression (r =-0.644,-0.523,all P ＜ 0.01).Conclusion Clock gene Per2 and VEGF,cyclin D1 expression in esophageal cancer has certain correlation,it can be a prognostic indicator of esophageal cancer.
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@#Per2 gene plays one of the most critical roles of clock gene which modulates circadian rhythm both in the physiological, biochemical and behavioral processes of organisms. The distributions of per2 gene include suprachiasmatic nucleus of the hypothalamus, central nucleus of amygdala, bed nucleus of the stria terminalis, hippocampus and other components of limbic system; it affects the emotional and visceral activities through participating in the system of circadian rhythm. The central per2 gene regulates the hypothalamus-pituitaryadrenal axis through integration of light input, steroid hormones and other neurotransmitters integration, acting on the target organs, and presentes a circadian rhythm of movement. This article reviewed the morphology and biology of per2 gene, and its participation in limbic system regulating the circadian rhythm of motional and visceral activities.
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Chronological nutrition is based on biological clocks that include clock genes and telomeres. Clock genes predict the day/night cycle to regulate both physical and mental activity in best condition, and prevent lifestyle-related diseases. Telomeres, the repeated series of DNA sequences that cap the ends of chromosomes, become shorter during cell division, thus determine lifespan of the individuals and organs. Even when dietary intake and exercise are adequate, disturbance of diurnal rhythm results in hypertension and hyperglycemia. Human activity is driven by NADH and ATP produced from nutrients, and the resulting NAD and AMP prevent telomere shortening by activating enzymes called SIRT1 and AMPK, respectively. Both enzymes collaborate in activating the master regulator PGC-1α that prevents oxidative stress and obesity. Physical activity increases PGC-1α and releases a hormone irisin from muscle that also prevents obesity. The dietary habit conforming good chronological nutrition are as follows: take nutritionally balanced breakfast every morning, distribute energy intake in the ratio breakfast: lunch: dinner = 3:3:4, and avoid dinner later than 21 o’clock or take earlier light dinner. Slow feeding and the intake of vegetables before carbohydrate are recommended to prevent rapid blood sugar increase. Regular 7 hour sleep is essential for the removal of metabolic wastes by “brain glymph system” to prevent dementia. The homeostatic and hedonic feeding and daily activity are controlled by human brain. Thus, lifestyle-related diseases will be prevented by moderation following the principles of chronological nutrition, irrespective of risk gene polymorphism.
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The Period 3 and Clock genes are important components of the mammalian molecular circadian system. Studies have shown association between polymorphisms in these clock genes and circadian phenotypes in different populations. Nevertheless, differences in the pattern of allele frequency and genotyping distribution are systematically observed in studies with different ethnic groups. To investigate and compare the pattern of distribution in a sample of Asian and Caucasian populations living in Brazil, we evaluated two well-studied polymorphisms in the clock genes: a variable number of tandem repeats (VNTR) in PER3 and a single nucleotide polymorphism (SNP) in CLOCK. The aim of this investigation was to search for clues about human evolutionary processes related to circadian rhythms. We selected 109 Asian and 135 Caucasian descendants. The frequencies of the shorter allele (4 repeats) in the PER3 gene and the T allele in the CLOCK gene among Asians (0.86 and 0.84, respectively) were significantly higher than among Caucasians (0.69 and 0.71, respectively). Our results directly confirmed the different distribution of these polymorphisms between the Asian and Caucasian ethnic groups. Given the genetic differences found between groups, two points became evident: first, ethnic variations may have implications for the interpretation of results in circadian rhythm association studies, and second, the question may be raised about which evolutionary conditions shaped these genetic clock variations.
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Adult , Female , Humans , Male , Asian People/genetics , CLOCK Proteins/genetics , Circadian Rhythm/genetics , White People/genetics , Genetic Variation/genetics , Period Circadian Proteins/genetics , Asian People/ethnology , Brazil , White People/ethnology , Gene Frequency , Genotype , Minisatellite Repeats/genetics , Phenotype , Polymorphism, Single Nucleotide/geneticsABSTRACT
Objective To explore the effects of light on the expression patterns of clock and clock-related genes in peripheral lymphocytes.To develop basic knowledge needed for clinical application of the circadian clock.Methods One hundred Sprague-Dawley rats were housed under constant dark(DD)or normal light-dark(LD 12:12)conditions for six weeks.Peripheral iymphoeytes were collected at different time points.The expression level of the clock gene and melatonin receptor genes mt1 and mt2 were detected using semi-quantitative RT-PCR.The data were analyzed with cosine software.Results Circadian expression of the genes was observed in both groups,but the peak phase,amplitude and strength of expression of each gene differed with the light conditions.Conclusion Light influences the expression of clock and clock-related genes in rats' peripheral lymphocytes.The clock gene might play an important role in regulating the expression of mt1 and mt2.
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AIM: To establish a PCR method for investigating the expression of clock genes in cultured rattus cardiac myocytes. METHODS: PCR was carried out using 3 primer pairs based on the published sequences of dbp, bmal1 and per2 genes of rattus. The conditions of PCR were optimized and the specificity of amplication was tested. RESULTS: In a volume of 20 ?l, the optimal PCR mixture of bmal1 gene contains 0.5 U Taq polymerase, 0.006 ?mol dNTP and 0.035 ?mol Mg 2+; the annealing temperature being 57 ℃; and circle times being 30. In a same volume, the optimal PCR mixture of dbp gene contains 0.5 U Taq polymerase, 0.006 ?mol dNTP and 0.03 ?mol Mg 2+; the annealing temperature being 58 ℃; and circle times being 32. The optimal PCR mixture of per2 gene contains 0.5 U Taq polymerase, 0.006 ?mol dNTP and 0.05 ?mol Mg 2+; the annealing temperature being 57 ℃; circle times being 30. The specificity of amplication was very high. CONCLUSION: The PCR method can successfully detect mRNA expression of clock genes in cultured rattus cardiac myocytes.
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Objective To investigate the effects of circadian clock gene Period2(Per2)on the proliferation,differentiation and apoptosis of K562 cells and its probable molecular mechanism.Methods The Per2 expression plasmid pcDNA3.1-Per2 and empty control plasmid were respectively transfected into K562 cells with cationic liposome,and the resistant cells stably expressing Per2 gene were obtained by G418 selection.Their morphological changes were observed under light microscope following Wright-Giemsa staining.Trypan blue excluding staining and MTT assay were employed to evaluate cell proliferation.Flow cytometry was performed to analyze cell cycle distribution and cell apoptosis,and electron microscopy was used to detect cell apoptosis.Meanwhile,the expressions of proliferation and apoptosis associated proteins,such as P53,Cyclin B1 and C-Myc,were respectively detected by RT-PCR and Western blot analysis at mRNA and protein level.Results The K562/Per2 cell line stably expressing Per2 gene was screened out.As compared with either the empty plasmid transfected group(K562/empty)or the untreated group(K562/untreated),K562/Per2 cells was smaller in volume and showed no obvious cellular differentiation.Circadian clock gene Per2 could significantly inhibit both growth and proliferation of K562 cells.The percentage of K562 cells in G2/M phase increased [K562/Per2 group(36.1?5.5)%,K562/empty group(12.5?2.9)%,untreated group(9.7?2.3)%,P