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1.
Neurosci Bull ; 2021 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-33528793

RESUMO

Glioma-associated microglial cells, a key component of the tumor microenvironment, play an important role in glioma progression. In this study, the mouse glioma cell line GL261 and the mouse microglia cell line BV2 were chosen. First, circadian gene expression in glioma cells co-cultured with either M1 or M2 microglia was assessed and the exosomes of M2-polarized and unpolarized BV-2 microglia were extracted. Subsequently, we labeled the exosomes with PKH67 and treated GL261 cells with them to investigate the exosome distribution. GL261 cell phenotypes and related protein expression were used to explore the role of M2 microglial exosomes in gliomas. Then a specific miR-7239-3p inhibitor was added to verify miR-7239-3p functions. Finally, the mouse subcutaneous tumorigenic model was used to verify the tumorigenic effect of M2 microglial exosomes in vivo. Our results showed that in gliomas co-cultured with M2 microglia, the expression of the BMAL1 protein was decreased (P < 0.01), while the expression of the CLOCK protein was increased (P < 0.05); opposite results were obtained in gliomas co-cultured with M1 microglia. After treatment with M2 microglial exosomes, the apoptosis of GL261 cells decreased (P < 0.001), while the viability, proliferation, and migration of GL261 cells increased. Increased expression of N-cadherin and Vimentin, and decreased E-cadherin expression occurred upon treatment with M2 microglial exosomes. Addition of an miR-7239-3p inhibitor to M2 microglial exosomes reversed these results. In summary, we found that miR-7239-3p in the glioma microenvironment is recruited to glioma cells by exosomes and inhibits Bmal1 expression. M2 microglial exosomes promote the proliferation and migration of gliomas by regulating tumor-related protein expression and reducing apoptosis.

2.
Chronobiol Int ; 38(3): 451-465, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33435737

RESUMO

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

3.
J Hazard Mater ; 401: 123794, 2021 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-33113737

RESUMO

In this work, an ultrasensitive and selective electrochemiluminescence (ECL) aptasensor with Au-tetrahedral aptamer nanostructure (Au-TAN) for acetamiprid detection was developed, which employed luminescence property of luminol and hydrogen peroxide (H2O2) as a co-reactant to apply the prepared Au-TAN to the luminescence systems. Au-TAN was prepared to modify an electrode surface via an Au-S bond to form a stable tetrahedral nanostructure. Fixed on the surface of the working electrode, Au-TAN could not only enhance the function of the aptamer but also boost the sensing performance. At the same time, Au nanoparticles (AuNPs) of the Au-TAN could also catalyze H2O2, thereby enhancing the luminescence performance of this aptasensor. The pH of the buffer solution, the concentration of H2O2 and the concentration of Au-TAN were optimized. Under the optimal conditions, the aptasensor had a detection limit of 0.0576 pM (S/N = 3), which was lower than those of other aptasensors for acetamiprid detection. Moreover, the weak alkaline environment explored in the experiment could expand its application range. Above all, the proposed method presented a high accuracy and sensitivity.

4.
J Biol Rhythms ; 35(6): 542-554, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32748687

RESUMO

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

5.
Cell Death Discov ; 6: 58, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32695477

RESUMO

This study was conducted to identify genes that are differentially expressed in paracancerous tissue and to determine the potential predictive value of selected gene panel. Gene transcriptome data of bladder tissue was downloaded from UCSC Xena browser and NCBI GEO repository, including GTEx (the Genotype-Tissue Expression project) data, TCGA (The Cancer Genome Atlas) data, and GEO (Gene Expression Omnibus) data. Differentially Expressed Genes (DEGs) analysis was performed to identify tumor-DEGs candidate genes, using the intersection of tumor-paracancerous DEGs genes and paracancerous-normal DEGs genes. The survival-related genes were screened by Kaplan-Meier (KM) survival analysis and univariable Cox regression with the cutoff criteria of KM < 0.05 and cox p-value < 0.05. The risk model was developed using Lasso regression. The clinical data were analyzed by univariate and multivariate Cox regression analysis. Gene Ontology (GO) and KEGG enrichment analysis were performed in the DEGs genes between the high-risk and low-risk subgroups. We identified six survival-related genes, EMP1, TPM1, NRP2, FGFR1, CAVIN1, and LATS2, found in the DEG analyses of both, tumor-paracancerous and paracancerous-normal differentially expressed data sets. Then, the patients were classified into two clusters, which can be distinguished by specific clinical characteristics. A three-gene risk prediction model (EMP1, FGFR1, and CAVIN1) was constructed in patients within cluster 1. The model was applied to categorize cluster 1 patients into high-risk and low-risk subgroups. The prognostic risk score was considered as an independent prognostic factor. The six identified survival-related genes can be used in molecular characterization of a specific subtype of bladder cancer. This subtype had distinct clinical features of T (topography), N (lymph node), stage, grade, and survival status, compared to the other subtype of bladder cancer. Among the six identified survival-related genes, three-genes, EMP1, FGFR1, and CAVIN1, were identified as potential independent prognostic markers for the specific bladder cancer subtype with clinical features described.

6.
Sci Total Environ ; 737: 139785, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32516665

RESUMO

In order to effectively monitor the residue of kanamycin (KAN), a dual-signal-amplified electrochemiluminescence (ECL) aptasensor based on multi-walled carbon nanotubes@titanium dioxide/thionine (MWCNTs@TiO2/Thi) was proposed. MWCNTs@TiO2 with large specific surface area and favorable biocompatibility could accelerate charge transfer and enable high loading of luminol to enhance ECL response. As a perfect electronic mediator, Thi could also accelerate electron conductivity to further enhance ECL intensity. The ECL intensity of MWCNTs@TiO2/Thi was enhanced for 3.6-fold compared with that of individual Thi because Thi could strongly interact with MWCNTs through π-π stacking force to enhance the electronic transmission. With the outstanding electron transfer property of MWCNTs@TiO2 and Thi, ECL intensity of the proposed aptasensor was obviously increased. Upon addition of KAN, the aptamer bound to its target, which caused that the ECL intensity decrease significantly. Therefore, KAN concentration could be monitored on the basis of signal intensity. Under optimal conditions, the constructed aptasensor exhibited a sensitive response towards KAN and a low detection limit of 0.049 ng mL-1 was obtained. It also possessed the excellent specificity, favorable stability and good reproducibility. Importantly, the application of proposed ECL aptasensor provides an efficient approach for highly sensitive detection of various small molecular contaminants.


Assuntos
Aptâmeros de Nucleotídeos , Técnicas Biossensoriais , Nanopartículas Metálicas , Nanotubos de Carbono , Técnicas Eletroquímicas , Canamicina , Medições Luminescentes , Reprodutibilidade dos Testes
7.
Mol Cell Biochem ; 467(1-2): 65-75, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32067140

RESUMO

Circadian rhythms help organisms adapt to changes of external environment by regulating energy metabolism and remaining the balance of homeostasis. Numerous researches have proved that the physiological function of liver was precisely controlled by circadian rhythms. Clock, one of core circadian genes, has been demonstrated to regulate the oxidative phosphorylation process of mitochondrial, which provides energy for living cells and acts as one of the hub for apoptosis. However, whether Clock gene regulates mitochondrial apoptosis pathways in liver cells remains less explored. In the present study, we used lentiviral vector to establish a stable AML12 cell lines which were capable of expressing specific shRNA to interfere the expression of Clock gene and investigated the effect of Clock on mitochondrial apoptosis pathways. Herein, we found that the interference of Clock gene could significantly suppress mitochondrial apoptosis pathways by stabilizing mitochondrial membrane potential and inhibiting mitochondria out membrane permeablization, which might be a result of lower expression of BAD and BIM proteins. Moreover, the interference of Clock gene could downregulate the expression of mitochondrial apoptosis factors, i.e. AIF, CYCS, APAF-1 and SMAC, which will suppress the formation of apoptosome and the process of DNA degradation to further inhibit apoptosis process. This work provides an insight on the important role of Clock gene participating in mitochondrial apoptosis pathways of hepatocytes and unveils a probable pathogenesis of how circadian rhythm regulates liver diseases.


Assuntos
Proteínas CLOCK/genética , Proteínas CLOCK/metabolismo , Hepatócitos/citologia , Mitocôndrias/metabolismo , Animais , Apoptose , Linhagem Celular , Permeabilidade da Membrana Celular , Técnicas de Silenciamento de Genes , Redes Reguladoras de Genes , Hepatócitos/metabolismo , Potencial da Membrana Mitocondrial , Camundongos , Membranas Mitocondriais/metabolismo
8.
Sci Total Environ ; 712: 136410, 2020 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-32050375

RESUMO

In order to solve the problem that the sensor cannot be reused due to the passivation of the electrode surface, a refreshable electrochemical aptasensor based on a hydrophobic electrode and a magnetic nanocomposite had been developed. Therein, the hydrophobic electrode was formed by modifying a screen-printed carbon electrode (SPCE) with polydimethylsiloxane (PDMS), which could avoid adsorption of molecules on the electrode surface due to its hydrophobicity. Combined with aptamer (Apt), the synthesized graphene oxide-ferroferric oxide (GO-Fe3O4) was used as a magnetic catcher to capture specific organophosphorus pesticides (OPs), which could be removed to the working area of SPCE with a magnet for electrochemical detection. The performance analysis of hydrophobic electrode showed that the SPCE could be used twice. When the electrochemical signals of Apt/GO-Fe3O4 and OPs/Apt/GO-Fe3O4 were recorded using the same SPCE, the current differences between them were directly related to the concentrations of OPs. Through the contrast test between the spiked vegetable samples and the OPs standard solutions, it was found that the OPs concentrations could be qualitatively evaluated by comparing the current differences. At the same time, the characteristic of collecting target with magnetic catcher was helpful for detecting OPs with a low concentration. Therefore, the refreshable aptasensor provided a huge potential to small molecule target evaluation.

9.
J Comput Biol ; 26(12): 1418-1426, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31305134

RESUMO

Approximately half of the unexplained recurrent spontaneous abortions remain unexplained (URSAs). We aimed to provide novel insights into the biological characteristics and related pathways of differentially expressed genes (DE-genes), DE-methylated genes, and DE-miRNAs in URSA, and construct a molecular miRNAs-mRNAs network. Four data sets (GSE22490, GSE121950, GSE73025, and GSE43256) were gained from GEO data sets. We identified the DE-genes, DE-methylated genes, and DE-miRNAs using the LIMMA package in R software. Function and enrichment analyses were conducted using DAVID. A protein-protein network was performed by STRING. We predicted the target genes of DE-miRNA using DIANA-microT-CDS. Then, we constructed miRNAs-mRNAs network. There were 137 genes that overlapped in two expression profile data sets (GSE121950 and GSE22490). We found 10 overlapping DE-methylated genes and DE-genes with opposite expression alteration trends. All those 10 genes were hypermethylated lowly expressed genes. Pathway analysis illustrated that DE-genes were enriched in osteoclast differentiation, leishmaniasis, NF-kappa B signaling pathway, Toll-like receptor signaling pathway, and tuberculosis. Based on protein-protein interaction analysis, TLR8, TLR2, CD86, TLR4, IL10, CD163, FCGR1A, CXCL8, FCGR3A, HCK, PLEK, and MNDA were identified as hub genes for DE-genes. We screened out 47 DE-miRNAs and 42 overlapping DE-genes between predicted target genes of DE-miRNAs and the 137 DE-genes. We then constructed miRNAs-mRNAs network. This study identified several genes and miRNAs involved in the development and progression of URSA, including FCGR1A, FCGR3A, CXCL8, HCK, PLEK, IL10, hsa-miR-498, and hsa-miR-4530. Although further in vivo and in vitro validations are required, our results may provide a theoretical basis for future studies.


Assuntos
Aborto Espontâneo/genética , Biologia Computacional/métodos , Metilação de DNA/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , MicroRNAs/genética , Feminino , Redes Reguladoras de Genes , Humanos , Gravidez , Mapas de Interação de Proteínas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
10.
Biochem Biophys Res Commun ; 517(1): 111-117, 2019 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-31303273

RESUMO

Doxorubicin (DOX) is a potent anti-neoplastic agent with cumulative cardiotoxicity. DOX-induced cardiotoxicity has been shown to depend on the different dosing times. However, the basis for determining the dosing time to minimize DOX-induced cardiotoxicity and the underlying mechanisms remain incompletely understood. Here we first showed that SIRT3, the major mitochondrial deacetylase, is negatively correlated to DOX-induced cardiotoxicity through the regulation of ATP production, mitochondrial membrane potential (MMP) level and ROS level in human pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Then, we used in vivo experiments to demonstrate that DOX significantly reduced the SIRT3 expression and the SIRT3 activity as reflected by the increased AcK68MnSOD/MnSOD ratio in rats after six weeks of treatment. Notably, the activity of SIRT3 had an obvious diurnal rhythm pattern in the myocardium of healthy rats. More importantly, an obvious lower AcK68MnSOD/MnSOD ratio was observed in rat hearts with DOX administrated at Zeitgeber time (ZT) 9 (ZT 0 was the time lights were turned on) than ZT1, which represent the peak and trough of SIRT3 activity. Moreover, DOX ZT9 reduced the body weight loss, extended the survival period, improved the heart function and alleviated the myocardial lesions compared to DOX ZT1. Mechanistic investigations demonstrated that DOX ZT1 significantly reduced ATP production, oxygen consumption rate (OCR) at various respiration states, MMP level and MnSOD activity and enhanced the H2O2 level compared with CON ZT1, whereas there was no significant effect for DOX ZT9 compared with CON ZT9. Taken together, dosing at the peak time of SIRT3 activity reduced DOX-induced cardiotoxicity, which may be related to the increased endogenous tolerance against the mitochondrial dysfunction and oxidative stress caused by DOX.


Assuntos
Antibióticos Antineoplásicos/efeitos adversos , Cardiotoxicidade/prevenção & controle , Doxorrubicina/efeitos adversos , Estresse Oxidativo/efeitos dos fármacos , Sirtuína 3/metabolismo , Animais , Antibióticos Antineoplásicos/administração & dosagem , Cardiotoxicidade/etiologia , Cardiotoxicidade/metabolismo , Cardiotoxicidade/patologia , Linhagem Celular , Relação Dose-Resposta a Droga , Doxorrubicina/administração & dosagem , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias Cardíacas/efeitos dos fármacos , Mitocôndrias Cardíacas/metabolismo , Mitocôndrias Cardíacas/patologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Ratos Sprague-Dawley
11.
Oncol Lett ; 15(5): 7097-7103, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29731875

RESUMO

The circadian rhythm regulates numerous physiological activities, including sleep and wakefulness, behavior, immunity and metabolism. Previous studies have demonstrated that circadian rhythm disorder is associated with the occurrence of tumors. Responsible for regulating a number of functions, the Circadian locomotor output cycles kaput (Clock) gene is one of the core regulatory genes of circadian rhythm. The Clock gene has also been implicated in the occurrence and development of tumors in previously studies. The present study evaluated the role of the Clock gene in the proliferation and migration of mouse breast cancer 4T1 cells, and investigated its possible regulatory pathways and mechanisms. It was reported that downregulation of Clock facilitated the proliferation and migration of breast cancer cells. Further investigation revealed the involvement of IQ motif containing GTPase activating protein 1 (IQGAP1) protein expression in the Clock regulatory pathway, further influencing the expression of E-cadherin, a known proprietor of tumor cell migration and invasion. To the best of our knowledge, the present study is the first to report that Clock, acting through the regulation of the scaffolding protein IQGAP1, regulates the downstream expression of E-cadherin, thereby affecting tumor cell structure and motility. These results confirmed the role of Clock in breast cancer tumor etiology and provide insight regarding the molecular avenues of its regulatory nature, which may translate beyond breast cancer into other known functions of the gene.

12.
Exp Biol Med (Maywood) ; 241(2): 205-15, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26264441

RESUMO

Our previous study found that CLOCK knockdown in the testes of male mice led to a reduced fertility, which might be associated with the lower acrosin activity. In this present study, we examined the differential expression in proteins of CLOCK knockdown sperm. Clock gene expression was knocked down in cells to confirm those differentially expressions and serine protease inhibitor SERPINA3K was identified as a potential target. The up-regulated SERPINA3K revealed an inverse relationship with Clock knockdown. Direct treatment of normal sperm with recombinant SERPINA3K protein inhibited the acrosin activity and reduced in vitro fertilization rate. The luciferase reporter gene assay showed that the down-regulated of Clock gene could activate the Serpina3k promoter, but this activation was not affected by the mutation of E-box core sequence. Co-IP demonstrated a natural interaction between SERPIAN3K and RORs (α and ß). Taken together, these results demonstrated that SERPINA3K is involved in the Clock gene-mediated male fertility by regulating acrosin activity and provide the first evidence that SERPINA3K could be regulated by Clock gene via retinoic acid-related orphan receptor response elements.


Assuntos
Acrosina/biossíntese , Proteínas CLOCK/metabolismo , Relógios Circadianos , Regulação da Expressão Gênica , Serpinas/metabolismo , Espermatozoides/fisiologia , Animais , Proteínas CLOCK/deficiência , Feminino , Fertilidade , Técnicas de Inativação de Genes , Masculino , Camundongos Endogâmicos ICR
13.
Cell Physiol Biochem ; 37(3): 911-20, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26390085

RESUMO

BACKGROUND/AIMS: Circadian locomotor output cycles protein kaput (CLOCK) plays a key role in maintaining circadian rhythms and activation of downstream elements. However, its function on human female reproductive system remains unknown. METHODS: To investigate the potential role of CLOCK, CLOCK-shRNAs were transfected into mouse 129 ES cells or injected into the ovaries of adult female mice. Western blotting was utilized to analyze the protein interactions and flow cytometry was used to assess apoptosis. RESULTS: The expression of CLOCK peaked at the 6th week in the healthy fetuses. However, an abnormal expression of CLOCK was detected in fetuses from spontaneous miscarriage. To determine the effect of CLOCK on female fertility, a small hairpin RNA (shRNA) strategy was used to specifically knockdown the CLOCK gene expression in vitro and in vivo. Knockdown of CLOCK induced apoptosis in mouse embryonic stem (mES) cells and inhibited the proliferation in mES cells in vitro. CLOCK knockdown also led to decreased release of oocytes and smaller litter size compared with control in vivo. CONCLUSIONS: Collectively, theses findings indicate that CLOCK plays an important role in fertility and that the CLOCK knockdown leads to reduction in reproduction and increased miscarriage risk.


Assuntos
Aborto Espontâneo/metabolismo , Proteínas CLOCK/genética , Proteínas CLOCK/metabolismo , Ciclo Menstrual , Aborto Espontâneo/etiologia , Aborto Espontâneo/genética , Aborto Animal/metabolismo , Animais , Apoptose , Proliferação de Células , Relógios Circadianos , Feminino , Fertilidade , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos da Linhagem 129 , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Ovário/metabolismo , RNA Interferente Pequeno/metabolismo
14.
Acta Biochim Biophys Sin (Shanghai) ; 46(4): 313-7, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24578160

RESUMO

Several essential biological progresses in mammals are regulated by circadian rhythms. Though the molecular mechanisms of oscillating these circadian rhythms have been uncovered, the specific functions of the circadian genes are not very clear. It has been reported that knocking down circadian genes by microRNA is a useful strategy to explore the function of the circadian rhythms. In this study, through a forward bioinformatics screening approach, we identified miR-29a/b/c as potent inhibitors for the human circadian gene hPER1. We further found that miR-29a/b/c could directly target hPER1 3'untranslated region (UTR) and down-regulate hPER1 at both mRNA and protein expression levels in human A549 cells. Thus, our findings suggested that the expression of hPER1 is regulated by miR-29a/b/c, which may also provide a new clue for the function of hPER1.


Assuntos
Regiões 3' não Traduzidas , MicroRNAs/fisiologia , Proteínas Circadianas Period/genética , Algoritmos , Sequência de Bases , Western Blotting , Linhagem Celular Tumoral , Primers do DNA , Regulação para Baixo , Humanos , Reação em Cadeia da Polimerase Via Transcriptase Reversa
15.
J Biol Rhythms ; 28(3): 208-17, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23735500

RESUMO

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


Assuntos
Proteínas CLOCK/genética , Ritmo Circadiano/genética , Fertilidade/genética , Fertilidade/fisiologia , Animais , Western Blotting , DNA/genética , Fertilização In Vitro , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos ICR , Atividade Motora/fisiologia , Tamanho do Órgão/fisiologia , Plasmídeos/genética , Antígeno Prostático Específico/metabolismo , RNA Interferente Pequeno/biossíntese , RNA Interferente Pequeno/genética , Contagem de Espermatozoides , Motilidade Espermática/fisiologia , Espermatozoides/enzimologia , Testículo/anatomia & histologia , Testículo/fisiologia , Testosterona/sangue , Transfecção
16.
Brain Res ; 1491: 147-55, 2013 Jan 23.
Artigo em Inglês | MEDLINE | ID: mdl-23159716

RESUMO

Alterations in metabolism could be due to cell-autonomous effects associated with altered expression of Clock in central nervous system feeding centers and/or peripheral tissues involved in metabolism. Clock mutant mice are hyperphagic and obese, which indicates that Clock is related to obesity. In the present study, we used intracerebroventricular injection of recombinant adenoviral vector harboring Clock genes to explore the role of Clock on diet induced obesity and the mechanisms involved in leptin resistance and leptin signaling in mice. The results demonstrated that expression of Clock in the arcuate nucleus of diet induced obesity mice was down-regulated. The recombinant adenoviral vector harboring Clock genes could reduce obesity indexes of diet induced obesity mice including body weight, BMI and total fat mass, attenuate hyperleptinemia, increase leptin sensitivity and decrease accumulated suppressor of cytokine signaling-3 in the arcuate nucleus. These results indicate that Clock plays an important role on obesity, which may be involved in leptin resistance and regulation of suppressor of cytokine signaling-3 in arcuate nucleus.


Assuntos
Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/genética , Leptina/genética , Obesidade/genética , Animais , Western Blotting , Composição Corporal/fisiologia , Índice de Massa Corporal , Peso Corporal/genética , Peso Corporal/fisiologia , Ventrículos Cerebrais/fisiologia , Dependovirus/genética , Vetores Genéticos , Injeções Intraventriculares , Metabolismo dos Lipídeos/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Reação em Cadeia da Polimerase em Tempo Real , Receptores para Leptina , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina/genética
17.
Exp Biol Med (Maywood) ; 236(9): 1078-84, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21750017

RESUMO

As a main component of circadian genes, clock plays not only an important role in circadian rhythm but also in the regulation of many physiological systems. The dysfunction of clock genes is associated with the development of various disorders. Many studies have investigated the association between clock genes and blood coagulation and the fibrinolytic system. The present study was designed to investigate the effect of downregulation of circulatory Clock on blood coagulation and fibrinolysis at the initial stage of active phase in male mice. Downregulation of the expression of the Clock gene by siRNA and, subsequently, its effect on the thrombotic potential and the expression of relative coagulative and/or fibrinolytic factors were investigated. It was found that the Clock interfered mice were less liable to thrombosis and showed prolonged prothrombin time (PT) and activated partial thromboplastin time (APTT) at Zeitgeber time (ZT) 15. Meanwhile, these mice also showed an increase in factor VII (FVII) and a decrease in thrombomodulin (TM) and plasminogen activator inhibitor 1 (PAI-1) at ZT 15 at both transcriptional and translational levels. PT, APTT and mRNA expressions of fvii, tm and pai-1 were analyzed with the least-squares fit of a 24-h cosine function by single cosinor method; no circadian rhythm was determined in PT and APTT, and a higher amplitude of fvii in the Clock RNAi group was found with a circadian phase shift, while lower amplitudes of tm and pai-1 were found in the Clock RNAi group with nearly no phase shift. All these results suggest that downregulation of the Clock gene in circulatory system has an effect on factors involved in both blood coagulation and fibrinolysis resulting in an enhancement in mice. This may be considered as an indication that Clock regulates thrombotic homeostasis through the fibrinolytic system.


Assuntos
Proteínas CLOCK/fisiologia , Fibrinólise/genética , Animais , Aorta Abdominal/metabolismo , Coagulação Sanguínea/genética , Coagulação Sanguínea/fisiologia , Proteínas CLOCK/análise , Proteínas CLOCK/genética , Linhagem Celular , Regulação para Baixo/genética , Fator VII/análise , Fibrinólise/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Tempo de Tromboplastina Parcial , Inibidor 1 de Ativador de Plasminogênio/análise , Tempo de Protrombina , RNA Interferente Pequeno/genética
18.
Bioelectromagnetics ; 30(8): 613-20, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19572333

RESUMO

It has been suggested that some animals are much more capable of perceiving certain kinds of geophysical stimuli which may precede earthquakes than humans, but the anecdotal phenomena or stories about unusual animal behaviors prior to an earthquake should be interpreted with objective data. During the Wenchuan magnitude 8.0 earthquake that happened in Wenchuan county (31.0 degrees north latitude, 103.4 degrees east longitude) of Sichuan province, China, on May 12, 2008, eight mice were monitored for locomotor activity and circadian rhythm in constant darkness with temperature 22-24 degrees C and humidity 55-65% for 38 days. The ongoing monitoring of locomotor activity of mice in our laboratory made it possible to design a posteriori study investigating whether the earthquake was associated with any change in animal behavior. Based on analyzing the recorded data with single cosinor, we found that the locomotor activity dramatically decreased in six of these eight mice on day 3 before the earthquake, and the circadian rhythm of their locomotor activity was no longer detected. The behavioral change lasted for 6 days before the locomotor activity returned to its original state. Analyses of concurrent geomagnetic data showed a higher total intensity during the span when the circadian rhythm in locomotor activity weakened. These results indicated that the behaviors, including circadian rhythm and activity, in these mice indeed changed prior to the earthquake, and the behavioral change might be associated with a change of geomagnetic intensity.


Assuntos
Comportamento Animal , Terremotos , Animais , China , Ritmo Circadiano , Geologia , Locomoção , Magnetismo , Masculino , Camundongos
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