Effects of long-term noise exposure during sleep on cognitive function and biological clock-related mechanisms in mice / 环境与职业医学

Background Environmental noise pollution is serious, and there are few studies on the effects of long-term noise exposure during sleep on cognitive function and possible biological clock mechanism. Objective To explore the cognitive impairment induced by noise exposure during sleep in mice and possible biological clock mechanism, and to provide a theoretical basis for the protection against noise exposure. Methods Twenty male C57BL/6J mice were randomly divided into a control group and a noise-exposed group, 10 mice in each group. The noise-exposed group was exposed to sleep-period noise using a noise generator for 12 h (08:00–20:00) per day for a total of 30 d. The calibrated noise intensity was set at 90 dB. No intervention was imposed on the control group. At the end of the noise exposure, cognitive function of mice was examined using the new object recognition experiment and the open field test, and the hippocampal tissue damage of mice were evaluated by Nissl staining, ionized calcium binding adaptor molecule 1 (Iba1) immunofluorescence staining, and real-time fluorescence quantitative PCR for inflammatory factors and biological clock genes. Oxidative stress indicators in the hippocampus of mice were also detected by assay kit. Results After noise exposure during sleep period, the results of new object recognition experiment showed that the discrimination index of mice in the noise-exposed group was 0.06±0.04, which was significantly lower than that of the control group (0.65±0.13) (P<0.05). The results of open field test showed that the central activity distance of the noise-exposed group was (242.20±176.10) mm, which was significantly lower than that of the control group, (1548.00±790.30) mm (P < 0.05), and the central activity time of the noise-exposed group was (0.87±0.64) s, which was significantly lower than that of the control group, (6.00±2.86) s (P < 0.05). The Nissl staining results showed that compared with the control group, neurons in the hippocampus of the noise-exposed mice were shrunken, deeply stained, disorganized, and loosely connected. The immunofluorescence results showed that microglia in the hippocampus of the noise-exposed mice were activated and the expression of Iba1 was significantly increased compared with those of the control group (P<0.05). The real-time PCR results of showed that the mRNA levels of the biological clock genes Clock, Per2, and Rev-erbα were significantly increased compared with those of the control group (P<0.05), and the mRNA level of Per1 was significantly decreased compared with that of the control group (P<0.05); and the mRNA levels of IL-18, IL-6, iNOS, and NLRP3 in the hippocampal tissues of mice were significantly increased compared with those of the control group (P<0.05). The results of oxidative stress evaluation showed that compared with the control group, reduced glutathione content was significantly reduced in the noise-exposed group (P<0.001). Conclusion Noise exposure during sleep period can lead to the destabilization of biological clock genes in hippocampal tissues and trigger hippocampal neuroinflammation, which can lead to the activation of microglia and cause cognitive impairment in mice.

Effects of long-term noise exposure during sleep on liver circadian clock and lipid metabolism / 环境与职业医学

Background Long-term exposure to noise during sleep may has adverse effects on metabolic system, and liver lipid metabolism is closely related to circadian clock genes. Objective To investigate the effects of long-term noise exposure during sleep on liver circadian clock and lipid metabolism in mice and its related mechanism. Methods Twenty C57BL/6J male mice were randomly divided into two groups: a noise exposure group and a control group with 10 mice in each group. The mice in the noise exposure group were exposed to white noise at 90 dB sound pressure level (SPL) for 30 consecutive days, 8 h a day, from 9:00 to 17:00. The mice in the control group were exposed to background noise ≤40 dB SPL. After noise exposure, the animals were neutralized at 14:00 (ZT6) and 2:00 (ZT18), 5 animals at each time spot, and the liver tissues were collected. Total cholesterol and triglyceride in liver were determined by cholesterol oxidase method and glycerol phosphate oxidase method respectively. The expressions of circadian clock genes (Clock, Bmal1, Rev-erbα, and Rev-erbβ) and lipid metabolism genes (Srebp1c, Hmgcr, Fasn, Lxrα, Acc1, and Chrebp) in liver were detected by quantitative real-time PCR. Results Compared with the control group, the content of total cholesterol in liver in the noise exposure group increased by 48% (P<0.05) and the content of liver triglyceride increased by 61% (P<0.05) at ZT18. The mRNA expression levels of circadian clock genes Clock and Bmal1 in the noise exposure group was significantly increased at ZT18 and decreased at ZT6 (P<0.05). The mRNA expression level of Rev-erbα decreased at both ZT6 and ZT18 (P<0.05). The mRNA expression level of Rev-erbβ had no significant change at ZT6 and ZT18. The mRNA expression levels of liver lipid metabolism related genes Srebp1c, Hmgcr, Chrebp, and Lxrα in the noise exposure group were higher than those in the control group at ZT18 (P<0.05). The mRNA expression levels of Acc1 and Fasn showed no significant change at ZT6, then an upward trend at ZT18, but no significant difference between the two time spots (P>0.05). Conclusion Long-term noise exposure during sleep can cause circadian clock and lipid metabolism disorders in mice. Among them, suppression of key circadian clock genes may be associated with Rev-erbα-mediated upregulation of the nuclear receptors Srebp1c and Chrebp for lipid synthesis and deposition in the liver, resulting in lipid metabolism disorder.

Difference in ketone bodies levels between diurnal and nocturnal fasting in children: observational study

Introduction Children are susceptible to developing preoperative ketonemia, which can be affected by changes in the circadian rhythm and counter-regulatory hormones. It is unclear whether ketonemia depends on the timing of fasting. Objective To assess the effect of preoperative fasting time (diurnal vs. night) on the preoperative concentration of ketone bodies in children. Methods We conducted a prospective-observational clinical study between September 2020 and March 2021, including children under 48 months of age scheduled for elective surgery. Two groups were identified based on fasting time, as follows: diurnal fasting (group A, n = 40) and nocturnal fasting (group B, n = 52). Demographic data, duration of fasting, time of excess fasting, type of food intake, the concentration of ketone bodies and capillary blood glucose, level of anxiety, and dehydration were analyzed in both groups. Results Diurnal fasting was associated with higher incidence of ketonemia compared with nocturnal fasting (Group A: 62.5% (95% CI 48.1-82.0); group B: 38,5% (95% CI 26.5-52.5), P=0.02). Most of the patients exceeded the duration of fasting recommended by preoperative fasting guidelines (95.6%). The type of food eaten before surgery was significantly associated with the presence of ketonemia (P=0.01). Conclusions Preoperative ketonemia is relatively common in patients under 48 months of age, especially among those who undergo diurnal fasting compared to nocturnal fasting.

Introducción Los niños son susceptibles a desarrollar cetonemia preoperatoria que puede verse afectada por cambios en el ritmo circadiano y las hormonas contrarreguladoras. No está claro si la cetonemia depende de la hora del ayuno. Objetivo Evaluar el efecto del momento del ayuno preoperatorio (diurno vs. nocturno) sobre la concentración preoperatoria de los cuerpos cetónicos en niños. Métodos Llevamos a cabo un estudio clínico observacional entre septiembre de 2020 y marzo de 2021, en niños menores de 48 meses, programados para cirugía electiva. Se identificaron dos grupos basados en la hora del ayuno, como sigue: ayuno diurno (grupo A, n = 40) y ayuno nocturno (grupo B, n = 52). En ambos grupos se analizaron los datos demográficos, la duración del ayuno, el tiempo excesivo de ayuno, el tipo de ingesta de alimentos, la concentración de cuerpos cetónicos, la glicemia capilar, el nivel de ansiedad y la deshidratación. Resultados El ayuno diurno se asocio con una mayor incidencia de cenotemia en comparación con el ayuno nocturno (Grupo A: 62,5% (IC 95% 48,1-82,0); grupo B: 38,5% (95% CI 26.5-52.5), P=0.02). La mayoría de los pacientes excedieron el tiempo de ayuno recomendado según las guías de ayuno preoperatorio (95,6%). El tipo de alimentos ingeridos antes de la cirugía se asoció de manera importante con la presencia de cetonemia (P=0,01). Conclusiones La cetonemia preoperatoria es relativamente común en pacientes menores de 48 meses de edad, especialmente entre quienes se someten a ayuno diurno en comparación con ayuno nocturno.

The Influence of Biological Clock System on the Circadian Rhythm of Sleep Based on Essence-Qi-Spirit Theory / 中医杂志

The essence-qi-spirit theory is an important part of traditional Chinese medicine， whose steady state is the material and functional basis for the balance of yin and yang in the body， making the essence， qi and spirit integrated， and body and spirit harmonized. Based on this theory， it is proposed that essence and qi depletion， spirit dissipation and qi dispersion， disharmony between yin and yang is the main pathogenesis of sleep disorders. Therefore， the method of regulating and harmonzing yin and yang by essence gathering， qi nourishing and spirit storing can be used to treat sleep disorder. The biological clock system of the circadian rhythm of sleep is regulated by the molecular oscillation that is generated by the transcription of the biological clock gene， and is a clock gradually formed by orga-nisms constantly adapting to the laws of nature. As the material basis， power， and embodiment of sound and peaceful sleep， essence， qi and spirit can perceive and transmit natural signals， whose functions are similar to what is recognized by modern science that oscillation amplifies the rhythm signal， and synchronously regulates the expression signal of the biological clock gene， thereby forming a biological clock system with “input-oscillation-output” as the feedback cycle. It is believed that the regulation method of yin and yang by essence gathering， qi nourishing and spirit storing may comprehensively regulate the physiological activities through brain/ muscle aryl hydrocarbon receptor nuclear translocator-like protein 1 （BMAL1）/circadian locomotor output cycles kaput （CLOCK）-period protein （PER）/ cryptochrome （CRY） transcriptional feedback loop， thereby adapting to the natural environment changes， playing an active role in the treatment of sleep disorders， and provideing a new idea for traditional Chinese medicine to reshape the molecular regulation system of the endogenous biological clock to prevent and treat sleep disorders.

Research progress on the biological clock genes and diabetic retinopathy / 中华眼底病杂志

Diabetic retinopathy (DR) is one of the most common and serious complication of diabetes mellitus, which is the main cause of vision loss in adults. Biological clock genes produce circadian rhythms and control its operation, while the disorder of the expression causes the occurrence and development of a series of diseases. It has been demonstrated that biological clock genes might take effects in the development and progression of DR. On the one hand, circadian rhythm disorder-related behavior disrupts the circadian oscillation of clock genes, and the change in its expression level is prone to unbalanced regulation of glucose metabolism, ultimately increasing the risk of type 2 diabetes mellitus and DR pathogenesis. On the other hand, DR patients exhibit symptoms of circadian rhythm disorders, and it has been suggested that the clock genes may control the development and progression of DR by affecting a variety of retinal pathophysiological processes. Therefore, maintaining normal circadian rhythm can be used as a disease prevention strategy, and studying the molecular mechanism of clock genes in DR can provide new ideas for more comprehensive elaboration of the pathogenesis of DR and search for new therapeutic targets.

Biological clock rhythm disorder and hypertension / 国际脑血管病杂志

Biological clock proteins are involved in the regulation of many important physiological processes, including blood pressure. The deletion or mutation of core circadian clock genes may cause elevated blood pressure levels and disrupted blood pressure rhythms, exacerbating vascular function damage, and ultimately leading to the occurrence, development and poor outcome of ischemic stroke. This article reviews the molecular mechanism of biological clock rhythm, the relationship between biological clock gene and blood pressure regulation mechanism, the mechanism of circadian rhythm disorder in the occurrence and development of hypertension, and the relationship between blood pressure rhythm disorder and stroke.

Research progress of circadian rhythm and circadian clock gene regulation in the development of myopia / 国际眼科杂志(Guoji Yanke Zazhi)

With the development of society, the incidence of myopia and the population of myopia has increased year by year, which has become a major public health problem. Therefore, the research on the pathogenesis and prevention and control measures of myopia is imminent. In recent years, the role of the biological clock in the development of myopia has gradually attracted scholars interest. Now the author starts from the impact of the biological clock on the axial length, retina and choroid in the development of myopia. In order to provide new ideas for the study of prevention and control measures and the pathogenesis of myopia, a brief review is made from the perspective of contemporary society and disrupted body clock.

Role of epigenetic regulation of biological clock genes in tumors / 中国药理学通报

The circadian clock is regulated at the molecular level by transcriptional-translational feedback loop of clock genes, which ensures that a variety of physiological processes have a-round 24 h circadian rhythms, including cell metabolism, cell proliferation, cell apoptosis and tumorigenesis, to maintain the homeostasis. Thus, the disturbance of circadian clock will disrupt homeostasis, causing various diseases, including neoplasm, metabolic syndrome, Parkinson's disease, COPD and cardiovascular diseases. Disturbance of circadian clock is closely related with tumorigenesis, and acts on various molecules and pathways leading to tumorigenesis, including oncogene and tumor suppressor gene, cell cycle, metabolic reprogramming, immune escape, endocrine disruption, alteration of gastrointestinal microbiome. This review focuses on changes in clock genes expression which disrupt cell cycle and may play a role in tumorigenesis, and epi-geneties, an important way to regulate gene expression, which can alter clock gene expression, thus playing an important role in the process of " the alternation of clock gene expression-disruption of cell cycle-tumorigenesis".

The Circadian System Is Essential for the Crosstalk of VEGF-Notch-mediated Endothelial Angiogenesis in Ischemic Stroke / 神经科学通报·英文版

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.

Cognitive profile in mild cognitive impairment with Lewy bodies

INTRODUCTION@#This study aimed to elucidate the cognitive profile of patients with mild cognitive impairment with Lewy bodies (MCI-LB) and to compare it to that of patients with mild cognitive impairment due to Alzheimer's disease (MCI-AD).@*METHODS@#Subjects older than 60 years with probable MCI-LB (n = 60) or MCI-AD (n = 60) were recruited. All patients were tested with Mini-Mental State Examination (MMSE) and Montreal Cognitive Assessment (MoCA) to assess their global cognitive profile.@*RESULTS@#The MCI-AD and MCI-LB patients did not differ in total MMSE and MoCA scores. However, some sub-items in MMSE and MoCA were shown to be screening markers for differentiating MCI-LB from MCI-AD. In the visuoconstructive test, the total score and hands subitem score in the clock-drawing test were significantly lower in MCI-LB than in MCI-AD. As for the executive function, the 'animal fluency test', 'repeat digits backward test' and 'take paper by your right hand' in MMSE all showed lower scores in MCI-LB compared with MCI-AD. As for memory, 'velvet' and 'church' in MoCA and 'ball' and 'national flag' in MMSE had lower scores in MCI-AD than in MCI-LB.@*CONCLUSION@#This study presents the cognitive profile of patients with MCI-LB. In line with the literature on Dementia with Lewy bodies, our results showed lower performance on tests for visuoconstructive and executive function, whereas memory remained relatively spared in the early period.

Circadian rhythm and health: dialogue between traditional Chinese medicine and modern medicine / 中国中药杂志

Circadian rhythm refers to the daily rhythmic variations in an organism. The irregular lifestyles of modern humans have led to a high incidence of chronic diseases, highlighting an inseparable relationship between disrupted circadian rhythm and disease development. TCM has long discussed rhythmic variations, with records dating back to the Yellow Emperor's Inner Canon(Huang Di Nei Jing), which laid a rich theoretical foundation for the research on circadian rhythm. Modern medical research has provided a more comprehensive explanation of its molecular mechanisms. This article integrated the current understanding of circadian rhythm in both Chinese and western medicine, emphasizing the crucial relationship between rhythm regulation and disease treatment. By highlighting the interdisciplinary nature of the two fields, it offers new directions for exploring the field of chronomedicine.

Research progress on the correlation between circadian rhythm and clock genes and the pathogenesis of diabetic retinopathy / 国际眼科杂志(Guoji Yanke Zazhi)

Diabetic retinopathy(DR)is the most common microvascular complication of patients with diabetes mellitus, and it has become one of the leading causes of visual impairment among working-age people worldwide. The pathogenesis of DR is complicated with multiple mechanisms. Plenty of studies have indicated that circadian rhythm and clock genes are closely related to the pathogenesis of DR. Circadian rhythm is a physiological process regulated by clock genes, which takes 24h as a cycle and is consistent with the changes of light and dark outside. Circadian rhythm regulates various physiological activities of the body. The disturbance of circadian rhythm induces DR by affecting the blood glucose level and the physiological homeostasis of the eye in patients with diabetes mellitus, and clock genes may be involved in the pathogenesis of DR by regulating oxidative stress response, inflammatory response, retinal autophagy rhythm, mitochondrial dysfunction and endothelial progenitor cell function. This paper will introduce the generation and regulation mechanism of circadian rhythm, as well as the internal circadian rhythm of retina, and further discuss the influence of circadian rhythm and clock genes on the occurrence and development of DR, aiming to provide a reference for the prevention and treatment of DR.

Circadian rhythm in prostate cancer: time to take notice of the clock / 亚洲男科学杂志(英文版)

The circadian clock is an evolutionary molecular product that is associated with better adaptation to changes in the external environment. Disruption of the circadian rhythm plays a critical role in tumorigenesis of many kinds of cancers, including prostate cancer (PCa). Integrating circadian rhythm into PCa research not only brings a closer understanding of the mechanisms of PCa but also provides new and effective options for the precise treatment of patients with PCa. This review begins with patterns of the circadian clock, highlights the role of the disruption of circadian rhythms in PCa at the epidemiological and molecular levels, and discusses possible new approaches to PCa therapy that target the circadian clock.

Teste do Desenho do Relógio: dados normativos para idosos / Clock Drawing Test: standard data for older adults / Prueba de Diseño de Reloj: datos normativos para ancianos

Trata-se de um estudo quantitativo, retrospectivo, correlacional e de corte transversal, com objetivo de fornecer dados normativos do TDR para idosos, levando em consideração diferentes faixas etárias e níveis de escolaridade. Duzentos e trinta e cinco foram entrevistados individualmente, distribuídos em cinco grupos etários e quatro níveis de escolaridade. Os instrumentos foram Ficha de Dados Sociodemográficos, Miniexame do Estado Mental (MEEM), Escala de Depressão Geriátrica, versão reduzida (GDS-15), Tarefa de Fluência Verbal Semântica (TFVS) e o TDR. Utilizou-se estatísticas descritivas, correlação de Pearson e análise univariada (one-way ANOVA) com post hoc Scheffe. Os escores do TDR apresentaram associações significativas com os anos de idade, anos de escolaridade, MEEM, TFVS e GDS-15. Houve diferença de desempenho no TDR ao considerarem os grupos por idade. O estudo fornece valores normativos para o TDR em uma amostra de idosos do sul do Brasil que foram influenciados pela idade, escolaridade, sintomatologia depressiva e fluência verbal. (AU)

This was a quantitative, retrospective, correlational, cross-sectional study that aimed to provide normative CDT (Clock-Drawing Test) data for older adults, taking into account different age groups and educational levels. The sample included 235 older adults distributed among five age groups and four levels of education. The instruments were Sociodemographic Data Sheet, the Mini-Mental State Examination (MMSE), the Geriatric Depression Scale reduced version (GDS-15), the Semantic Verbal Fluency Task (TFVS), and the CDT. Descriptive statistics, Pearson's correlation, and univariate analysis (one-way ANOVA) with Scheffe post hoc were used. The CDT scores showed significant associations with age, years of schooling, MMSE, TFVS, and GDS-15. There was a difference in performance in CDT when considering age groups. The present study was able to provide normative values ​​for CDT in a sample of older adults in southern Brazil that ​​were influenced by age, education, depressive symptoms, and verbal fluency. (AU)

Se trata de un estudio cuantitativo, retrospectivo, correlacional y transversal, con el objetivo de aportar datos normativos sobre el TDR para ancianos, teniendo en cuenta diferentes grupos de edad y niveles educativos. La muestra incluyó a 235 ancianos distribuidos en cinco grupos de edad y cuatro niveles de educación. Los instrumentos utilizados fueron Ficha de Datos Sociodemográficos, Mini Examen del Estado Mental (MMSE), Escala de Depresión Geriátrica, versión reducida (GDS-15), Tarea de Fluidez Verbal Semántica (TFVS) y TDR. Se emplearon estadísticas descriptivas, correlación de Pearson y análisis univariante (one-way ANOVA) con post hoc Scheffe. Los puntajes de TDR mostraron asociaciones significativas con la edad, años de escolaridad, MMSE, TFVS y GDS-15. Hubo diferencia en el desempeño en el TDR al considerar los grupos por edad. El presente estudio fue capaz de proporcionar valores normativos para TDR en una muestra de ancianos en el sur de Brasil influenciados por la edad, la escolaridad, los síntomas depresivos y la fluidez verbal. (AU)

Anmeidan Modulates Circadian Rhythm of Spontaneous Activity and Influences Clock Proteins in Sleep-deprived Rats / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Anmeidan （AMD） on biological rhythm and related protein expression in sleep-deprived rats. MethodA total of 80 SD rats were randomized into control group （Ctrl， equivalent volume of saline）， model group （SD， equivalent volume of saline）， AMD group （9.09 g·kg-1·d-1）， and melatonin group （MT， 0.27 g·kg-1·d-1）. Insomnia was induced in rats by self-made sleep deprivation box （4 weeks）. Circadian rhythm of spontaneous activity was evaluated by spontaneous activity video analysis system. Morphology of hypothalamus was observed based on hematoxylin-eosin （HE） staining， and the histomorphology of hypothalamus neurons and the Nissl's bodies based on Nissl staining. Western blotting was employed to detect the expression of hypothalamic proteins in cAMP-response element binding protein （CREB）/clock gene period （Per） pathway， and immunohistochemistry the expression of brain and muscle ARNT-like protein 1 （Bmal1）， Clock， Per1， and cryptochrome circadian regulator 1 （Cry1）. ResultThe model group demonstrated circadian rhythm disorder， as manifested by the significant increase in activity time in 6 designated time periods compared with the control group， and the rise in the activity speed and frequency （P<0.01）. Moreover， model group showed decrease in number of neurons which were sparsely arranged with shrunken or fragmented nuclei， reduction in number and loss of Nissl's bodies with light color， and drop in the relative expression of p-CREB and Per1， and the positive rate of Bmal1， Clock， Per1， and Cry1 （P<0.01）. Compared with model group， AMD group demonstrated reduction in time， speed， and frequency of activity （P<0.01）. Moreover， the AMD group also showed alleviation of neuronal damage （P<0.01）， and increase in the number of neurons with clear nuclei and cytoplasm in some， and the number of Nissl's bodies. AMD raised the expression of p-CREB and Per1 proteins， and the positive rate of Bmal1， Clock， Per1， and Cry1 （P<0.01）. ConclusionAMD ameliorated spontaneous circadian rhythm of sleep-deprived rats by regulating CREB/Per signaling pathway and further increasing the expression of Bmal1， Clock， Per1， and Cry1.

Effect of Jiangtang Xiaozhi Tablet on Liver Circadian Clock-related Genes CLOCK， BMAL1， REV-ERBα， and REV-ERBβ in Mice with Metabolic Dysfunction-associated Fatty Liver Disease / 中国实验方剂学杂志

ObjectiveTo explore the effect of Jiangtang Xiaozhi tablet （JTXZT） on metabolic dysfunction-associated fatty liver disease and to study the mechanism from the perspective of circadian clock-related genes such as circadian locomotor output cycles kaput （CLOCK）， brain and muscle aryl hydrocarbon receptor nuclear translocator-like protein 1 （BMAL1）， reverse-eritroblastosis receptor （REV-ERB）α and β. MethodA total of 50 male SPF C57BL/6J mice were randomized into normal group （n=10） and modeling group （n=40）. The normal group was fed with normal diet， and the modeling group with high-fat diet for 4 weeks. Then the model mice were randomly classified into model group， high-dose （12.5 g·kg-1） and low-dose （6.25 g·kg-1） Jiangtang Xiaozhi tablet groups， and orlistat group （70 mg·kg-1）， with 10 mice in each group. The normal group and model group received equivalent volume of distilled water （8 weeks）. Then， the body weight of mice was measured， and the content of serum triglyceride （TG）， total cholesterol （TC）， low-density lipoprotein cholesterol （LDL-C）， high-density lipoprotein cholesterol （HDL-C）， aspartate aminotransferase （AST）， and alanine aminotransferase （ALT） was determined with biochemical method. Serum content of free fatty acid （FFA） and leptin was detected by enzyme-linked immunosorbent assay （ELISA）. Pathological changes of liver tissue and epididymal adipose tissue were observed based on hematoxylin-eosin （HE） staining. Liver fibrosis was examined based on Masson's trichrome staining， and changes of lipids based on oil red O staining. The expression of CLOCK， BMAL1， REV-ERBα， and REV-ERBβ was detected by Western blot and immunohistochemistry assay. ResultCompared with the normal group， the model group had high content of TG， TC， LDL-C， HDL-C， AST， ALT， FFA， and leptin （P<0.05， P<0.01）， showed ballooning degeneration and focal microvesicular steatosis of liver cells， enlarged adipocytes， and inflammatory cell clusters and fibrous tissue hyperplasia， and displayed increased protein expression of sterol regulatory element binding protein （SREBP） 1 and peroxisome proliferators-activated receptor （PPAR）γ （P<0.01） and decreased protein expression of PPARα （P<0.05）， CLOCK， BMAL1， REV-ERBα and β （P<0.05， P<0.01）. Compared with the model group， JTXZT-H group down-regulated the content of TG， TC， LDL-C， HDL-C， AST， ALT， FFA， and leptin in mice （P<0.05， P<0.01）， and the JTXZT groups demonstrated reduction in the degree and range of ballooning degeneration of liver tissue， alleviation of the compression of hepatic sinusoidal tissue， unobvious inflammatory cell infiltration and fibrous tissue proliferation， reduction in the expression of SREBP1 and PPARγ （P<0.05， P<0.01）， and rise of the protein expression of PPARα （P<0.01）， CLOCK， BMAL1， REV-ERBα， and REV-ERBβ （P<0.05， P<0.01）. ConclusionJTXZT can significantly alleviate the metabolic dysfunction-associated fatty liver disease in mice caused by high-fat diet. The mechanism is the likelihood that it regulates downstream related lipid metabolism proteins （such as SREBP1， PPARγ， and PPARα）.

Effect of Jiangtang Xiaozhi Tablet on Liver Circadian Clock-related Genes CLOCK， BMAL1， REV-ERBα， and REV-ERBβ in Mice with Metabolic Dysfunction-associated Fatty Liver Disease / 中国实验方剂学杂志

ObjectiveTo explore the effect of Jiangtang Xiaozhi tablet （JTXZT） on metabolic dysfunction-associated fatty liver disease and to study the mechanism from the perspective of circadian clock-related genes such as circadian locomotor output cycles kaput （CLOCK）， brain and muscle aryl hydrocarbon receptor nuclear translocator-like protein 1 （BMAL1）， reverse-eritroblastosis receptor （REV-ERB）α and β. MethodA total of 50 male SPF C57BL/6J mice were randomized into normal group （n=10） and modeling group （n=40）. The normal group was fed with normal diet， and the modeling group with high-fat diet for 4 weeks. Then the model mice were randomly classified into model group， high-dose （12.5 g·kg-1） and low-dose （6.25 g·kg-1） Jiangtang Xiaozhi tablet groups， and orlistat group （70 mg·kg-1）， with 10 mice in each group. The normal group and model group received equivalent volume of distilled water （8 weeks）. Then， the body weight of mice was measured， and the content of serum triglyceride （TG）， total cholesterol （TC）， low-density lipoprotein cholesterol （LDL-C）， high-density lipoprotein cholesterol （HDL-C）， aspartate aminotransferase （AST）， and alanine aminotransferase （ALT） was determined with biochemical method. Serum content of free fatty acid （FFA） and leptin was detected by enzyme-linked immunosorbent assay （ELISA）. Pathological changes of liver tissue and epididymal adipose tissue were observed based on hematoxylin-eosin （HE） staining. Liver fibrosis was examined based on Masson's trichrome staining， and changes of lipids based on oil red O staining. The expression of CLOCK， BMAL1， REV-ERBα， and REV-ERBβ was detected by Western blot and immunohistochemistry assay. ResultCompared with the normal group， the model group had high content of TG， TC， LDL-C， HDL-C， AST， ALT， FFA， and leptin （P<0.05， P<0.01）， showed ballooning degeneration and focal microvesicular steatosis of liver cells， enlarged adipocytes， and inflammatory cell clusters and fibrous tissue hyperplasia， and displayed increased protein expression of sterol regulatory element binding protein （SREBP） 1 and peroxisome proliferators-activated receptor （PPAR）γ （P<0.01） and decreased protein expression of PPARα （P<0.05）， CLOCK， BMAL1， REV-ERBα and β （P<0.05， P<0.01）. Compared with the model group， JTXZT-H group down-regulated the content of TG， TC， LDL-C， HDL-C， AST， ALT， FFA， and leptin in mice （P<0.05， P<0.01）， and the JTXZT groups demonstrated reduction in the degree and range of ballooning degeneration of liver tissue， alleviation of the compression of hepatic sinusoidal tissue， unobvious inflammatory cell infiltration and fibrous tissue proliferation， reduction in the expression of SREBP1 and PPARγ （P<0.05， P<0.01）， and rise of the protein expression of PPARα （P<0.01）， CLOCK， BMAL1， REV-ERBα， and REV-ERBβ （P<0.05， P<0.01）. ConclusionJTXZT can significantly alleviate the metabolic dysfunction-associated fatty liver disease in mice caused by high-fat diet. The mechanism is the likelihood that it regulates downstream related lipid metabolism proteins （such as SREBP1， PPARγ， and PPARα）.

Effects of circadian misalignment induced by abnormal photoperiod on expression rhythm of clock genes and glucose uptake related genes in gastrocnemius in rats / 中国康复理论与实践

ObjectiveTo investigate the impacts of circadian misalignment on glucose uptake of skeletal muscle and metabolism in rats. MethodsA total of 36 male Wistar rats were divided into circadian alignment (CA, normal light-dark cycles, n = 18) and circadian misalignment (CM, shifted light-dark cycles, n = 18) groups. ClockLab behavior analysis was performed for 18 days (61 to 78 days after modeling). Intraperitoneal injection glucose tolerance test and physiologic measures were performed 85 days after modeling. They were euthanized 91 to 92 days after modeling, at 8:00, 12:00, 16:00, 20:00, 24:00 and 4:00 next day. Gastrocnemius tissue was collected and measured for Bmal1, Clock, Per2, Tbc1d1, Glut4 and Pgc1α by reverse transcription real-time quantitative polymerase chain reaction (RT-qPCR). ResultsThe circadian cycle increased (t = -6.557, P < 0.001), the amplitude decreased (t = 2.326, P = 0.030) and the area under curve (AUC) of the blood glucose of intraperitoneal glucose tolerance test decreased (t = -2.622, P = 0.016) in CM group. In gastrocnemius, there was difference in the expression of the Bmal1 (F = 6.691, P < 0.001), Clock (F = 4.188, P = 0.007), Per2 (F = 10.893, P < 0.001), Tbc1d1 (F = 3.411, P = 0.018), Glut4 (F = 5.439, P = 0.002) and Pgc1α (F = 15.376, P < 0.001) across different time; meanwhile, there was difference in the expression of Bmal1 (F = 5.020, P = 0.035), Per2 (F = 8.996, P = 0.006), Tbc1d1 (F = 51.111, P < 0.001) and Pgc1α (F = 10.177, P = 0.004) between groups, and the total Tbc1d1 expression decreased in CM group (t = 4.349, P < 0.001). ConclusionCM induced by shifted light-dark cycles may lead to glucose tolerance impairment in rats, which may be related to the decreased expression of Tbc1d1 and the changes of transcription rhythm of Bmal1, Per2 and Pgc1α in gastrocnemius.

Co-regulation of circadian clock genes and microRNAs in bone metabolism / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

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.

Effect of circadian rhythm changes on the expression of RORs and the RORs agonist SR1078 on corneal epithelial wound repair / 中华实验眼科杂志

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.