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1.
Adv Exp Med Biol ; 1185: 295-299, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31884627

RESUMO

The degeneration of photoreceptors is a common hallmark of ocular diseases like retinitis pigmentosa (RP) or age-related macular degeneration (AMD). To experimentally induce photoreceptor degeneration, the light damage paradigm is frequently used. In this study we show that the exposure to high amounts of cool white light (10,000 lux, 1 h) resulted in a more than 11-fold higher apoptotic rate in the retina compared to light exposure with 5000 lux for 30 min. Consequently, exposure to intense light resulted in a significant downregulation of retinal mRNA expression levels of the reference genes Gapdh, Gnb2l, Rpl32, Rps9, Actb, Ubc or Tbp compared to untreated controls. Investigators performing light-induced photoreceptor degeneration should be aware of the fact that higher light intensities will result in a dysregulation of reference genes.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Luz , Células Fotorreceptoras de Vertebrados/efeitos da radiação , Retina/efeitos da radiação , Apoptose , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Células Fotorreceptoras de Vertebrados/patologia , Retina/citologia , Degeneração Retiniana/patologia , Retinite Pigmentosa/patologia
2.
Life Sci ; 238: 116981, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31639394

RESUMO

AIM: Endothelial cell damage is critical to understand since its presence in the entire body makes the damage widespread instead of being localized. Being a major component of stem cell niche in bone marrow, deems it essential to gain knowledge of the damage to endothelium associated with bone marrow. Since radiation exposure has become common to numerous therapeutic modalities, its effects on bone marrow and its endothelial cells are crucial to understand. MATERIAL & METHODS: Microarray analysis was performed on irradiated human bone marrow endothelial cells (hBMECs) with and without prior treatment with radioprotectant amifostine to assess the effects of radiation on signalling pathways and the subsequent changes in pathways when treated with radioprotectant prior to radiation exposure. KEY FINDINGS: It was seen that adhesion pathways that were usually inactivated under normal circumstances were stimulated post radiation. However, where in the case of radiation exposure, these adhesion pathways included leukocyte adhesion and migration; in the case of radioprotected conditions the pathways revolve around cell-substrate adhesion and cell spreading. Genes like ROCK1, FLNA, RAC1, PRKCZ and MAP3K8 were seen to regulate the molecular switch between leukocyte-cell adhesion to cell-substrate adhesion. SIGNIFICANCE: Our study demonstrated that irradiated endothelium supports leukocyte adhesion and migration but shifts to substrate adhesion dependent cell spreading under radioprotected conditions in order to repair the monolayer damage from the radiation. The genes responsible for the shift were identified and can be employed to manipulate cell adhesion characteristics for the treatment of diseases caused by radiation or inflammation.


Assuntos
Amifostina/farmacologia , Biomarcadores/metabolismo , Medula Óssea/metabolismo , Adesão Celular , Endotélio Vascular/metabolismo , Raios gama , Leucócitos/metabolismo , Medula Óssea/efeitos dos fármacos , Medula Óssea/efeitos da radiação , Células Cultivadas , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/efeitos da radiação , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Redes Reguladoras de Genes/efeitos dos fármacos , Redes Reguladoras de Genes/efeitos da radiação , Humanos , Leucócitos/efeitos dos fármacos , Leucócitos/efeitos da radiação , Protetores contra Radiação/farmacologia
3.
Int J Radiat Biol ; 95(12): 1627-1639, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31509479

RESUMO

Purpose: Widespread medical use of radiation in diagnosis, imaging and treatment of different central nervous system malignancies lead to various consequences. Aim of this study was to further elucidate mechanism of cell response to radiation and possible consequence on neural differentiation.Materials and methods: NT2/D1 cells that resemble neural progenitors were used as a model system. Undifferentiated NT2/D1 cells and NT2/D1 cells in the early phase of neural differentiation were irradiated with low (0.2 Gy) and moderate (2 Gy) doses of γ radiation. The effect was analyzed on apoptosis, cell cycle, senescence, spheroid formation and the expression of genes and miRNAs involved in the regulation of pluripotency or neural differentiation.Results: Two grays of irradiation induced apoptosis, senescence and cell cycle arrest of NT2/D1 cells, accompanied with altered expression of several genes (SOX2, OCT4, SOX3, PAX6) and miRNAs (miR-219, miR-21, miR124-a). Presented results show that 2 Gy of radiation significantly affected early phase of neural differentiation in vitro.Conclusions: These results suggest that 2 Gy of radiation significantly affected early phase of neural differentiation and affect the population of neural progenitors. These findings might help in better understanding of side effects of radiotherapy in treatments of central nervous system malignancies.


Assuntos
Diferenciação Celular/efeitos da radiação , Células-Tronco Neurais/citologia , Células-Tronco Neurais/efeitos da radiação , Apoptose/efeitos da radiação , Biomarcadores/metabolismo , Contagem de Células , Senescência Celular/efeitos da radiação , Relação Dose-Resposta à Radiação , Regulação da Expressão Gênica/efeitos da radiação , Humanos , MicroRNAs/genética , Células-Tronco Neurais/metabolismo , Fatores de Tempo
4.
Int J Mol Sci ; 20(16)2019 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-31412584

RESUMO

Melatonin exerts oncostatic actions and sensitizes tumor cells to chemotherapeutics or radiation. In our study, we investigated the effects of docetaxel, vinorelbine, and radiation on human breast fibroblasts and its modulation by melatonin. Docetaxel or vinorelbine inhibits proliferation and stimulates the differentiation of breast preadipocytes, by increasing C/EBPα and PPARγ expression and by downregulating tumor necrosis factor α (TNFα), interleukin 6 (IL-6), and IL-11 expression. Radiation inhibits both proliferation and differentiation through the downregulation of C/EBPα and PPARγ and by stimulating TNFα expression. In addition, docetaxel and radiation decrease aromatase activity and expression by decreasing aromatase promoter II and cyclooxygenases 1 and 2 (COX-1 and COX-2) expression. Melatonin potentiates the stimulatory effect of docetaxel and vinorelbine on differentiation and their inhibitory effects on aromatase activity and expression, by increasing the stimulatory effect on C/EBPα and PPARγ expression and the downregulation of antiadipogenic cytokines and COX expression. Melatonin also counteracts the inhibitory effect of radiation on differentiation of preadipocytes, by increasing C/EBPα and PPARγ expression and by decreasing TNFα expression. Melatonin also potentiates the inhibitory effect exerted by radiation on aromatase activity and expression by increasing the downregulation of promoter II, and COX-1 and COX-2 expression. Our findings suggest that melatonin modulates regulatory effects induced by chemotherapeutic drugs or radiation on preadipocytes, which makes it a promising adjuvant for chemotherapy and radiotherapy sensibilization.


Assuntos
Antineoplásicos/farmacologia , Fibroblastos Associados a Câncer/efeitos dos fármacos , Fibroblastos Associados a Câncer/efeitos da radiação , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/efeitos da radiação , Melatonina/farmacologia , Radiação Ionizante , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Adipócitos/efeitos da radiação , Aromatase/metabolismo , Neoplasias da Mama , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Fibroblastos Associados a Câncer/metabolismo , Docetaxel/farmacologia , Ativação Enzimática/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Glândulas Mamárias Humanas/citologia , PPAR gama/genética , PPAR gama/metabolismo , Prostaglandina-Endoperóxido Sintases/genética , Prostaglandina-Endoperóxido Sintases/metabolismo , Vinorelbina/farmacologia
5.
Radiat Res ; 192(4): 410-421, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31390312

RESUMO

Radiation-induced acute skin injury and consequent fibrosis are common complications of cancer radiotherapy and radiation accidents. Stromal cell-derived factor-1α (SDF-1α) and its receptor, CXC chemokine receptor 4 (CXCR4) have been shown to be involved in multiple cellular events. However, the role of SDF-1α/CXCR4 axis in radiation-induced acute injury and fibrosis of skin has not been reported. In this study, we found that the expression of SDF-1α and CXCR4 was significantly increased in irradiated skin tissues of humans, monkeys and rats, compared to their nonirradiated counterparts. Mice with keratinocyte-specific ablation of CXCR4 showed less severe skin damage than wild-type mice after receiving a 35 Gy dose of radiation. Consistently, subcutaneous injection of AMD3100, an FDA approved SDF-1α/CXCR4 inhibitor, attenuated skin injury and fibrosis induced by exposure to radiation in a rat model. Mechanically, the SDF-1α/CXCR4 axis promotes pro-fibrotic TGF-b/Smad signaling through the PI3K-MAPK signaling cascade in human keratinocyte HaCaT cells and skin fibroblast WS1 cells. AMD3100 inhibited Smad2 nuclear translocation and transcriptional activity of Smad2/3 induced by radiation, which suppressed the pro-fibrotic TGF-b/Smad signaling pathway activated by exposure. Taken together, these findings demonstrate the involvement of SDF-1α/CXCR4 axis in radiation-induced acute injury and fibrosis of skin, and indicate that AMD3100 would be an effective countermeasure against these diseases.


Assuntos
Quimiocina CXCL12/metabolismo , Lesões por Radiação/metabolismo , Receptores CXCR4/metabolismo , Pele/patologia , Pele/efeitos da radiação , Animais , Fibrose , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Técnicas de Inativação de Genes , Compostos Heterocíclicos/farmacologia , Humanos , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Queratinócitos/efeitos da radiação , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos da radiação , Camundongos , Fosfatidilinositol 3-Quinases/metabolismo , Lesões por Radiação/patologia , Ratos , Receptores CXCR4/deficiência , Receptores CXCR4/genética , Pele/lesões , Pele/metabolismo , Proteína Smad2/metabolismo , Fator de Crescimento Transformador beta/metabolismo
6.
Artif Cells Nanomed Biotechnol ; 47(1): 3058-3066, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31339375

RESUMO

Repairing the lost or damaged mandible is very difficult and time-consuming, so there is a great hope for tissue engineering to accelerate it. At the present study, electrospinning was applied to fabricate polyvinylidene fluoride (PVDF) and PVDF-polyaniline (PANI) composite scaffolds. In addition, extremely low frequency pulsed electromagnetic field (PEMF) was applied for treating the stem cells derived from dental pulp (DPSCs) when cultured on the nanofibrous scaffolds. Osteoinductive property of the fabricated PVDF, PVDF-PANI scaffold at the presence and absence of the PEMF was investigated by evaluating the common osteogenic differentiation markers in seeded-DPSCs on the scaffold. Results demonstrated that cell attachment, protein adsorption and cells viability were increased when PEMF was applied. In addition, ALP activity, calcium content, osteogenic genes and protein evaluations confirmed that PEMF could significantly increase osteoinductivity of the PVDF while composite with PANI. According to the results, the use of polymers with piezoelectricity and conductivity features plus PEMF exposure has a promising potential to improve the current treatment methods in bone and mandibular defects.


Assuntos
Compostos de Anilina/farmacologia , Diferenciação Celular , Campos Eletromagnéticos , Células-Tronco Mesenquimais/citologia , Osteogênese , Polivinil/química , Tecidos Suporte/química , Fosfatase Alcalina/metabolismo , Cálcio/metabolismo , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/efeitos da radiação , Polpa Dentária/citologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/efeitos da radiação , Osteogênese/efeitos dos fármacos , Osteogênese/efeitos da radiação , Resistência à Tração , Engenharia Tecidual
7.
J Photochem Photobiol B ; 198: 111560, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31336216

RESUMO

Previous studies revealed significant impact on cancer cell by mid-infrared (MIR) radiation. However, the effects of narrow band MIR on immune reaction and infectious disease are still unknown. In this study, an enhanced innate immune response was observed through the interaction between Leptospiral outer membrane protein (LipL32) and toll-like receptor 2 (TLR2). Thereafter, human kidney proximal tubular cells (HK-2 cells) initiated a serial reaction of enhanced MCP-1 production. The 6 µm narrow bandwidth light source emitted by waveguide thermal emitter (WTE) was applied to induce carbonyl group (CO bond) stretching vibration during the stage of antigen-receptor complex formation. The amount of MCP-1 gene expression had 2.5 folds increase after narrow band MIR illumination comparing to non-MIR illumination at low dose LipL32 condition. Besides, both ELISA and confocal microscopy results also revealed that the chemokine concentration increased significantly after narrow band MIR illumination either at low or high concentration of LipL32. Furthermore, a specific phenomenon that narrow band MIR can amplify the signal of weak immune response by enhancing sensitivity of the interaction between antigen and receptor was observed. This study exhibits clear evidence that the narrow band MIR exposure can modulate the early immune response of infectious disease and play a potential role to develop host-directed therapy in the future.


Assuntos
Proteínas da Membrana Bacteriana Externa/farmacologia , Raios Infravermelhos , Lipoproteínas/farmacologia , Proteínas da Membrana Bacteriana Externa/imunologia , Linhagem Celular , Sobrevivência Celular/efeitos da radiação , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Túbulos Renais Proximais/citologia , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/metabolismo , Leptospira/metabolismo , Lipoproteínas/imunologia
8.
Nucleic Acids Res ; 47(14): 7703-7714, 2019 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-31269201

RESUMO

Gene autorepression is widely present in nature and is also employed in synthetic biology, partly to reduce gene expression noise in cells. Optogenetic systems have recently been developed for controlling gene expression levels in mammalian cells, but most have utilized activator-based proteins, neglecting negative feedback except for in silico control. Here, we engineer optogenetic gene circuits into mammalian cells to achieve noise-reduction for precise gene expression control by genetic, in vitro negative feedback. We build a toolset of these noise-reducing Light-Inducible Tuner (LITer) gene circuits using the TetR repressor fused with a Tet-inhibiting peptide (TIP) or a degradation tag through the light-sensitive LOV2 protein domain. These LITers provide a range of nearly 4-fold gene expression control and up to 5-fold noise reduction from existing optogenetic systems. Moreover, we use the LITer gene circuit architecture to control gene expression of the cancer oncogene KRAS(G12V) and study its downstream effects through phospho-ERK levels and cellular proliferation. Overall, these novel LITer optogenetic platforms should enable precise spatiotemporal perturbations for studying multicellular phenotypes in developmental biology, oncology and other biomedical fields of research.


Assuntos
Retroalimentação Fisiológica , Regulação da Expressão Gênica/genética , Redes Reguladoras de Genes/genética , Optogenética/métodos , Algoritmos , Animais , Células Cultivadas , Regulação da Expressão Gênica/efeitos da radiação , Redes Reguladoras de Genes/efeitos da radiação , Células HEK293 , Humanos , Luz , Modelos Genéticos , Proteínas Proto-Oncogênicas p21(ras)/genética , Reprodutibilidade dos Testes , Biologia Sintética/métodos
9.
Radiat Res ; 192(2): 121-134, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31161966

RESUMO

Radiation-induced fibrosis (RIF) is a common delayed effect of acute ionizing radiation exposure (DEARE) affecting diverse tissues including the heart, lungs, liver and skin, leading to reduced tissue function and increased morbidity. Monocytes, which may be classified into classical (CD14++, CD16-), intermediate (CD14++, CD16+) and non-classical (CD14+/low, CD16++) subtypes in humans and non-human primates (NHPs), and monocyte-derived macrophages may play an integral role in the pathogenesis of RIF. We tested the hypothesis that moderate to high levels of total-body exposure to radiation would alter monocyte polarization and produce phenotypes that could promote multi-organ fibrosis in a wellestablished NHP model of DEARE. Subjects were 16 young adult male rhesus macaques, ten of which were exposed to high-energy, 4 Gy X-ray total-body irradiation (TBI) and six that received sham irradiation (control). Total monocytes assessed by complete blood counts were 89% depleted in TBI animals by day 9 postirradiation (P < 0.05), but recovered by day 30 postirradiation and did not differ from control levels thereafter. Monocytes were isolated from peripheral blood mononuclear cells (PBMCs) and sorted into classical, intermediate and non-classical subsets using fluorescence-activated cell sorting (FACS) prior to and at 6 months post-TBI. At 6 months postirradiation, monocyte polarization shifted towards lower classical (92% → 86%) and higher intermediate (7% → 12%) and non-classical monocyte subsets (0.6% → 2%) (all P < 0.05) in TBI animals compared to baseline. No change in monocyte subsets was observed in control animals. Transcriptional profiles in classical and intermediate monocyte subsets were assessed using RNAseq. Classical monocyte gene expression did not change significantly over time or differ cross-sectionally between TBI and control groups. In contrast, significant numbers of differentially expressed genes (DEGs) were detected in intermediate monocyte comparisons between the TBI animals and all animals at baseline (304 DEGs), and in the TBI versus control animals at 6 months postirradiation (67 DEGs). Intermediate monocytes also differed between baseline and 6 months in control animals (147 DEGs). Pathway analysis was used to identify genes within significant canonical pathways, yielding 52 DEGs that were specific to irradiated intermediate monocytes. These DEGs and significant canonical pathways were associated with pro-fibrotic and anti-inflammatory signaling pathways that have been noted to induce M2 macrophage polarization. These findings support the hypothesis that TBI may alter monocyte programming and polarization towards a profibrotic phenotype, providing a novel target opportunity for therapies to inhibit or prevent RIF.


Assuntos
Monócitos/citologia , Monócitos/efeitos da radiação , Irradiação Corporal Total/efeitos adversos , Animais , Polaridade Celular/efeitos da radiação , Regulação da Expressão Gênica/efeitos da radiação , Macaca mulatta , Masculino , Monócitos/metabolismo , Fatores de Tempo , Transcrição Genética/efeitos da radiação
10.
Int J Mol Sci ; 20(12)2019 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-31212732

RESUMO

Light influences a wide range of physiological processes from prokaryotes to mammals. Neurospora crassa represents an important model system used for studying this signal pathway. At molecular levels, the WHITE COLLAR Complex (WCC), a heterodimer formed by WC-1 (the blue light photo-sensor) and WC-2 (the transcriptional activator), is the critical positive regulator of light-dependent gene expression. GATN (N indicates any other nucleotide) repeats are consensus sequences within the promoters of light-dependent genes recognized by the WCC. The distal GATN is also known as C-box since it is involved in the circadian clock. However, we know very little about the role of the proximal GATN, and the molecular mechanism that controls the transcription of light-induced genes during the dark/light transition it is still unclear. Here we showed a first indication that mutagenesis of the proximal GATA sequence within the target promoter of the albino-3 gene or deletion of the WC-1 zinc finger domain led to a rise in expression of light-dependent genes already in the dark, effectively decoupling light stimuli and transcriptional activation. This is the first observation of cis-/trans-acting repressive machinery, which is not consistent with the light-dependent regulatory mechanism observed in the eukaryotic world so far.


Assuntos
Sítios de Ligação , Escuridão , Fatores de Transcrição GATA/metabolismo , Regulação da Expressão Gênica/efeitos da radiação , Luz , Elementos de Resposta , Fatores de Transcrição/metabolismo , Sequência de Bases , Cromatina/genética , Cromatina/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Fatores de Transcrição GATA/química , Mutação , Neurospora/genética , Neurospora/metabolismo , Neurospora/efeitos da radiação , Motivos de Nucleotídeos , Fenótipo , Regiões Promotoras Genéticas , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Ativação Transcricional , Dedos de Zinco/genética
11.
Int J Mol Sci ; 20(12)2019 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-31216644

RESUMO

Tissue and cell damage caused by ionizing radiation is often highly genotoxic. The swift repair of DNA damage is crucial for the maintenance of genomic stability and normal cell fitness. Long noncoding RNAs (lncRNAs) have been reported to play an important role in many physiological and pathological processes in cells. However, the exact function of lncRNAs in radiation-induced DNA damage has yet to be elucidated. Therefore, this study aimed to analyze the potential role of lncRNAs in radiation-induced DNA damage. We examined the expression profiles of lncRNAs and mRNAs in 293T cells with or without 8 Gy irradiation using high-throughput RNA sequencing. We then performed comprehensive transcriptomic and bioinformatic analyses of these sequencing results. A total of 18,990 lncRNAs and 16,080 mRNAs were detected in all samples. At 24 h post irradiation, 49 lncRNAs and 323 mRNAs were differentially expressed between the irradiation group and the control group. qRT-PCR was used to verify the altered expression of six lncRNAs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses indicated that the predicted genes were mainly involved in the histone mRNA metabolic process and Wnt signaling pathways. This study may provide novel insights for the study of lncRNAs in radiation-induced DNA damage.


Assuntos
Regulação da Expressão Gênica/efeitos da radiação , RNA Longo não Codificante/genética , RNA Mensageiro/genética , Radiação Ionizante , Biologia Computacional/métodos , Dano ao DNA , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Células HEK293 , Humanos , MicroRNAs/genética , Interferência de RNA , Reprodutibilidade dos Testes , Transcriptoma
12.
Reprod Biol Endocrinol ; 17(1): 48, 2019 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-31226998

RESUMO

BACKGROUND: Puberty in male Atlantic salmon in aquaculture can start as early as after the first winter in seawater, stunts growth and entails welfare problems due to the maturation-associated loss of osmoregulation capacity in seawater. A better understanding of the regulation of puberty is the basis for developing improved cultivation approaches that avoid these problems. Our aim here was to identify morphological and molecular markers signaling the initiation of, and potential involvement in, testis maturation. METHODS: In the first experiment, we monitored for the first time in large Atlantic salmon males several reproductive parameters during 17 months including the first reproductive cycle. Since testicular growth accelerated after the Winter solstice, we focused in the second experiment on the 5 months following the winter solstice, exposing fish from February 1 onwards to the natural photoperiod (NL) or to continuous additional light (LL). RESULTS: In the first experiment, testis weight, plasma androgens and pituitary gonadotropin transcript levels increased with the appearance of type B spermatogonia in the testis, but testicular transcript levels for gonadotropin or androgen receptors did not change while being clearly detectable. In the second experiment, all males kept under NL had been recruited into puberty until June. However, recruitment into puberty was blocked in ~ 40% of the males exposed to LL. The first morphological sign of recruitment was an increased proliferation activity of single spermatogonia and Sertoli cells. Irrespective of the photoperiod, this early sign of testis maturation was accompanied by elevated pituitary gnrhr4 and fshb and testicular igf3 transcript levels as well as increased plasma androgen levels. The transition into puberty occurred again with stable testicular gonadotropin and androgen receptor transcript levels. CONCLUSIONS: The sensitivity to reproductive hormones is already established before puberty starts and up-regulation of testicular hormone receptor expression is not required to facilitate entry into puberty. The increased availability of receptor ligands, on the other hand, may result from an up-regulation of pituitary Gnrh receptor expression, eventually activating testicular growth factor and sex steroid release and driving germ and Sertoli cell proliferation and differentiation.


Assuntos
Hormônios Esteroides Gonadais/metabolismo , Receptores de Esteroides/metabolismo , Salmo salar/metabolismo , Maturidade Sexual , Testículo/metabolismo , Animais , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Regulação da Expressão Gênica/efeitos da radiação , Masculino , Fotoperíodo , Hipófise/metabolismo , Receptores do FSH/genética , Receptores do FSH/metabolismo , Receptores de Esteroides/genética , Reprodução/genética , Reprodução/fisiologia , Salmo salar/genética , Estações do Ano , Água do Mar
13.
Yale J Biol Med ; 92(2): 241-250, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31249485

RESUMO

Circadian clocks drive biological rhythms in physiology and behavior, providing a selective advantage by enabling organisms to synchronize to the 24 h environmental day. This process depends on light-dark transitions as the main signal that shifts the phase of the clock. In mammals, the light input reaches the master circadian clock in the hypothalamic suprachiasmatic nucleus through glutamatergic afferents from the retina, resulting in phase-shifts of the overt rhythms which depend on the time of the day at which light is applied, leading to changes in the activity of circadian core clock genes (i.e., Per1). This circadian gating of the synchronizing effect of light is dependent on the specific activation of signal transduction pathways involving several kinases acting on protein effectors. Protein phosphorylation is also an important regulatory mechanism essential for the generation and maintenance of circadian rhythms and plays a crucial role in the degradation and the appropriate turnover of PER proteins. In this work, we review the role of the main kinases implicated in the function of the master clock, with emphasis in those involved in circadian photic entrainment.


Assuntos
Relógios Circadianos/fisiologia , Transdução de Sinal Luminoso/fisiologia , Mamíferos/fisiologia , Proteínas Quinases/metabolismo , Animais , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Luz , Mamíferos/genética , Mamíferos/metabolismo , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Fosforilação/efeitos da radiação
14.
Yale J Biol Med ; 92(2): 259-270, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31249487

RESUMO

Circadian disruption has been linked to markers for poor health outcomes in humans and animal models. What is it about circadian disruption that is problematic? One hypothesis is that phase resetting of the circadian system, which occurs in response to changes in environmental timing cues, leads to internal desynchrony within the organism. Internal desynchrony is understood as acute changes in phase relationships between biological rhythms from different cell groups, tissues, or organs within the body. Do we have strong evidence for internal desynchrony associated with or caused by circadian clock resetting? Here we review the literature, highlighting several key studies from measures of gene expression in laboratory rodents. We conclude that current evidence offers strong support for the premise that some protocols for light-induced resetting are associated with internal desynchrony. It is important to continue research to test whether internal desynchrony is necessary and/or sufficient for negative health impact of circadian disruption.


Assuntos
Relógios Circadianos/fisiologia , Ritmo Circadiano/fisiologia , Periodicidade , Fotoperíodo , Animais , Relógios Circadianos/genética , Relógios Circadianos/efeitos da radiação , Ritmo Circadiano/genética , Ritmo Circadiano/efeitos da radiação , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Luz , Núcleo Supraquiasmático/metabolismo , Núcleo Supraquiasmático/fisiopatologia , Núcleo Supraquiasmático/efeitos da radiação
15.
J Photochem Photobiol B ; 197: 111537, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31247384

RESUMO

Light is the most prominent zeitgeber of the circadian system, which contains central and peripheral oscillators. Our previous studies found that light wavelength could influence the rhythms of melatonin synthesis and clock gene expression in the central oscillator of chicks. However, the effect of monochromatic light on the peripheral oscillator and the role of melatonin have yet to be clarified. In this study, 216 newly hatched chicks were divided into three groups (intact, sham operation and pinealectomy) and were raised under white (WL), red (RL), green (GL) or blue (BL) light for 14 days. Their plasma and livers were sampled at 6 time points with 4-h intervals. Plasma melatonin concentration and liver clock gene expression (cClock, cBmal1, cBmal2, cCry1, cCry2, cPer2, cPer3) were measured for circadian rhythm analysis. In intact and sham operation chicks under WL, all liver clock genes showed circadian expression along with oscillations in plasma melatonin. However, positive clock genes peaked at subjective night along with melatonin, while negative clock genes peaked at subjective day or the shifting time of day-night. Chick exposure to monochromatic light led to an unaltered circadian rhythmicity in plasma melatonin and liver clock genes; however, their rhythmic parameters were notably influenced. Compared to WL, GL enhanced the mesor and amplitude of melatonin and all kinds of clock genes, whereas RL had the opposite effect. Pinealectomy significantly decreased expression of liver clock genes, which was consistent with the reduction in plasma melatonin concentration, especially for the GL group, and resulted in the expression of liver clock genes showing low-mesor and low-amplitude oscillations as well as no statistically significant differences among the monochromatic light groups. Thus, we speculated that melatonin plays a key role in the effects of light wavelength on clock gene rhythm in the chick liver.


Assuntos
Proteínas CLOCK/metabolismo , Ritmo Circadiano/genética , Regulação da Expressão Gênica/efeitos da radiação , Luz , Fígado/metabolismo , Melatonina/sangue , Animais , Proteínas CLOCK/genética , Galinhas , Ritmo Circadiano/efeitos da radiação
16.
J Photochem Photobiol B ; 196: 111511, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31129510

RESUMO

Prolonged exposure of the skin to ultraviolet radiation (UV) leads to its damage and loss of protective properties. This condition called photoaging of the skin is caused by a number of destructive factors, such as reactive oxygen species (ROS) and proteolytic enzymes that cause damage to the extracellular matrix, e.g. collagen fibers. Many cells of the immune system, including neutrophils, are involved in the photoaging process. The presence of neutrophils in the skin exposed to UV irradiation is known; however, the mechanism of neutrophil activity at these conditions remains unclear. In our study, we focused on the ability of neutrophils to release neutrophil extracellular traps (NETs) and the role of these structures in the photoaging process. NET release occurs in response to various stimuli; however, we hereby showed that the UVA and UVB radiation that reaches the Earth's surface could activate the mechanism of netosis. UV-induced netosis was much faster than that activated by chemical or biological factors; however, it also occurred due to the production of ROS, known signal mediators in netosis. In this work, we also identified the probable netosis signaling pathway involved in the neutrophil response to UV. The participation of NET components may explain the ongoing process of skin photoaging, but it is also important to indicate netosis as a potential target for skin protection therapy. Antioxidants tested in this work, such as N-acetylcysteine, ethamsylate, as well as vitamin B1 (thiamine), can successfully inhibit UV-induced netosis, and thus be used as protective components against the negative effects of solar radiation.


Assuntos
Armadilhas Extracelulares/metabolismo , Neutrófilos/efeitos da radiação , Raios Ultravioleta , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Células Cultivadas , Armadilhas Extracelulares/química , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Interleucinas/genética , Interleucinas/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Neutrófilos/citologia , Neutrófilos/metabolismo , Fosforilação/efeitos da radiação , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Envelhecimento da Pele/efeitos da radiação , Quinase Syk/metabolismo
17.
Int J Oncol ; 54(6): 1907-1920, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31081062

RESUMO

The p53 protein is a tumour suppressor and transcription factor that regulates the expression of target genes involved in numerous stress responses systems. In this study, we designed a screening strategy using DNA damage­induced mouse and human transcriptome data to identify novel downstream targets of p53. Our method selected genes with an induced expression in multiple organs of X­ray­irradiated p53 wild­type mice. The expression of inka box actin regulator 2 gene, known as Inka2, was upregulated in 12 organs when p53 expression was induced. Similarly, INKA2 was induced in a p53­dependent manner at both the mRNA and protein level in human cells treated with adriamycin. Reporter assays confirmed that p53 directly regulated INKA2 through an intronic binding site. The overexpression of INKA2 produced a slight decrease in cancer cell growth in the colony formation assay. Moreover, the analysis of The Cancer Genome Atlas (TCGA) data revealed a decreased INKA2 expression in tumour samples carrying p53 mutations compared with p53 wild­type samples. In addition, significantly higher levels of DNA methylation were observed in the INKA2 promoter in tumour samples, concordant with the reduced INKA2 expression in tumour tissues. These results demonstrate the potential of INKA2 as a cancer cell growth inhibitor. Furthermore, INKA2 protein interacts with the serine/threonine­protein kinase, p21 (RAC1) activated kinase (PAK)4, which phosphorylates ß­catenin to prevent ubiquitin­proteasomal degradation. As ß­catenin was downregulated in a stable INKA2­expressing cell line, the findings of this study suggest that INKA2 is a novel, direct downstream target of p53 that potentially decreases cell growth by inhibiting the PAK4­ß­catenin pathway.


Assuntos
Perfilação da Expressão Gênica/métodos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias/genética , Proteína Supressora de Tumor p53/metabolismo , Quinases Ativadas por p21/metabolismo , Animais , Linhagem Celular Tumoral , Metilação de DNA , Doxorrubicina/farmacologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Células HCT116 , Humanos , Camundongos , Mutação , Neoplasias/metabolismo , Fosforilação , Regiões Promotoras Genéticas , Análise de Sequência de RNA/métodos , Transdução de Sinais , Proteína Supressora de Tumor p53/genética , beta Catenina/metabolismo
18.
Genes (Basel) ; 10(4)2019 04 03.
Artigo em Inglês | MEDLINE | ID: mdl-30987199

RESUMO

Fluorescent light (FL) has been utilized for ≈60 years and has become a common artificial light source under which animals, including humans, spend increasing amounts of time. Although the solar spectrum is quite dissimilar in both wavelengths and intensities, the genetic consequences of FL exposure have not been investigated. Herein, we present comparative RNA-Seq results that establish expression patterns within skin, brain, and liver for Danio rerio, Oryzias latipes, and the hairless mouse (Mus musculus) after exposure to FL. These animals represent diurnal and nocturnal lifestyles, and ≈450 million years of evolutionary divergence. In all three organisms, FL induced transcriptional changes of the acute phase response signaling pathway and modulated inflammation and innate immune responses. Our pathway and gene clustering analyses suggest cellular perception of oxidative stress is promoting induction of primary up-stream regulators IL1B and TNF. The skin and brain of the three animals as well as the liver of both fish models all exhibit increased inflammation and immune responses; however, the mouse liver suppressed the same pathways. Overall, the conserved nature of the genetic responses observed after FL exposure, among fishes and a mammal, suggest the presence of light responsive genetic circuitry deeply embedded in the vertebrate genome.


Assuntos
Encéfalo/metabolismo , Perfilação da Expressão Gênica/veterinária , Redes Reguladoras de Genes/efeitos da radiação , Fígado/metabolismo , Pele/metabolismo , Animais , Encéfalo/efeitos da radiação , Fluorescência , Regulação da Expressão Gênica/efeitos da radiação , Imunidade Inata/efeitos da radiação , Fígado/efeitos da radiação , Masculino , Camundongos , Especificidade de Órgãos , Oryzias , Análise de Sequência de RNA/veterinária , Pele/efeitos da radiação , Peixe-Zebra
19.
Biomed Environ Sci ; 32(3): 177-188, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30987692

RESUMO

OBJECTIVE: Pyroptosis is an inflammatory form of programmed cell death. This phenomenon has been recently reported to play an important role in radiation-induced normal tissue injury. Connexin43 (Cx43) is a gap junction protein that regulates cell growth and apoptosis. In this study, we investigated the effect of Cx43 on X-ray-induced pyroptosis in the human umbilical vein endothelial cells (HUVECs). METHODS: HUVECs, Cx43 overexpression, and Cx43 knockdown strains were irradiated with 10 Gy. Proteins were detected using western blot analysis. Cell pyroptosis was evaluated using the fluorescence-labeled inhibitor of caspase assay (FLICA) and propidium iodide staining through flow cytometry and confocal microscopy. Cell morphology and cytotoxicity were detected by scanning electron microscopy and lactate dehydrogenase release assay, respectively. RESULTS: Irradiation with 10 Gy X-ray induced pyroptosis in the HUVECs and reduced Cx43 expression. The pyroptosis in the HUVECs was significantly attenuated by overexpression of Cx43 as it decreased the level of active caspase-1. However, interference of Cx43 expression with siRNA significantly promoted pyroptosis by increasing the active caspase-1 level. Pannexin1 (Panx1), a gap junction protein regulates pyroptosis, and its cleaved form is used to evaluate channel opening and active state. The level of cleaved Panx1 in the HUVECs and Cx43 knockdown strains increased in the presence of X-ray, but decreased in the Cx43 overexpression strains. Furthermore, interference of Panx1 with siRNA alleviated the upregulation of pyroptosis caused by Cx43 knockdown. CONCLUSION: Results suggest that single high-dose X-ray irradiation induces pyroptosis in the HUVECs. In addition, Cx43 regulates pyroptosis directly by activating caspase-1 or indirectly by cleaving Panx1.


Assuntos
Caspase 1/genética , Conexina 43/genética , Regulação da Expressão Gênica/efeitos da radiação , Células Endoteliais da Veia Umbilical Humana/efeitos da radiação , Piroptose , Raios X/efeitos adversos , Caspase 1/metabolismo , Conexina 43/metabolismo , Conexinas/genética , Conexinas/metabolismo , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo
20.
PLoS One ; 14(4): e0215250, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30998706

RESUMO

Radiotherapy is an important therapeutic approach against cervical cancer but associated with adverse effects including vaginal fibrosis and dyspareunia. We here assessed the immunological and oxidative responses to cervical irradiation in an animal model for radiation-induced cervicitis. Rats were sedated and either exposed to 20 Gy of ionising radiation given by a linear accelerator or only sedated (controls) and euthanized 1-14 days later. The expressions of toll-like receptors (TLRs) and coupled intracellular pathways in the cervix were assessed with immunohistofluorescence and western blot. Expression of cytokines were analysed with the Bio-Plex Suspension Array System (Bio-Rad). We showed that TLRs 2-9 were expressed in the rat cervix and cervical irradiation induced up-regulation of TLR5, TRIF and NF-κB. In the irradiated cervical epithelium, TLR5 and TRIF were increased in concert with an up-regulation of oxidative stress (8-OHdG) and antioxidant enzymes (SOD-1 and catalase). G-CSF, M-CSF, IL-10, IL- 17A, IL-18 and RANTES expressions in the cervix decreased two weeks after cervical irradiation. In conclusion, the rat uterine cervix expresses the TLRs 2-9. Cervical irradiation induces immunological changes and oxidative stress, which could have importance in the development of adverse effects to radiotherapy.


Assuntos
Colo do Útero/imunologia , Raios gama/efeitos adversos , Regulação da Expressão Gênica/efeitos da radiação , Lesões Experimentais por Radiação/imunologia , Receptores Toll-Like/imunologia , Cervicite Uterina/imunologia , Proteínas Adaptadoras de Transporte Vesicular/imunologia , Animais , Colo do Útero/patologia , Citocinas/imunologia , Feminino , Regulação da Expressão Gênica/imunologia , NF-kappa B/imunologia , Estresse Oxidativo/imunologia , Estresse Oxidativo/efeitos da radiação , Lesões Experimentais por Radiação/patologia , Ratos , Ratos Sprague-Dawley , Cervicite Uterina/patologia
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