RESUMO
Low level light therapy uses light of specific wavelengths in red and near-infrared spectral range to treat various pathological conditions. This light is able to modulate biochemical cascade reactions in cells that can have important health implications. In this study, the effect of low intensity light at 650, 808 and 1064 nm on neurons and two types of cancer cells (neuroblastoma and HeLa) is reported, with focus on the photoinduced change of intracellular level of Ca2+ ions and corresponding signaling pathways. The obtained results show that 650 and 808 nm light promotes intracellular Ca2+ elevation regardless of cell type, but with different dynamics due to the specificities of Ca2+ regulation in neurons and cancer cells. Two origins responsible for Ca2+ elevation are determined to be: influx of exogenous Ca2+ ions into cells and Ca2+ release from endoplasmic reticulum. Our investigation of the related cellular processes shows that light-induced membrane depolarization is distinctly involved in the mechanism of Ca2+ influx. Ca2+ release from endoplasmic reticulum activated by reactive oxygen species generation is considered as a possible light-dependent signaling pathway. In contrast to the irradiation with 650 and 808 nm light, no effects are observed under 1064 nm irradiation. We believe that the obtained insights are of high significance and can be useful for the development of drug-free phototherapy.
Assuntos
Sinalização do Cálcio/efeitos da radiação , Cálcio/efeitos da radiação , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/efeitos da radiação , Cálcio/fisiologia , Membrana Celular/metabolismo , Eletrofisiologia , Corantes Fluorescentes/química , Células HeLa , Humanos , Raios Infravermelhos , Terapia com Luz de Baixa Intensidade , Neurônios/efeitos da radiação , Imagem Óptica , Espécies Reativas de Oxigênio/efeitos da radiaçãoRESUMO
After 50 years of studies on photobiomodulation (PBM), there is still so much to investigate to understand the laser light-nonplant cells interactions. The current scientific knowledge allows to say that the phenomena induced by PBM are based on cellular pathways that are the key points of cell life. The mitochondria chromophores, also present on the bacterial membrane, the calcium channels, ion that regulates the life-and-death cellular processes, as well as the TRP family, whose genes have been found in protozoa and suggest that its basic mechanism evolved long before the appearance of animals, seem to be elective targets in photobiomodulatory events by wavelengths from 600 up to 980 nm. The ambiguous resulting cellular communication way, mediated by ATP, ROS and/or calcium, leads to cell manipulation, which modifies its metabolism and whose response connects all life-forms from bacteria to vertebrates. Because of the Giano-Bifronte features of ROS and calcium, as well as the fine balance of energetic mitochondrial processes, whose alteration is responsible for several diseases, the PBM can show unpredictable results and it requires scrupulous approach to avoid cellular damages. However, when carefully applied, PBM is able to improve nonhealthy cell's responses and represents a reliable support in human and veterinary medicine.
Assuntos
Sinalização do Cálcio/efeitos da radiação , Cálcio/metabolismo , Terapia com Luz de Baixa Intensidade , Redes e Vias Metabólicas/efeitos da radiação , Trifosfato de Adenosina/metabolismo , Animais , Canais de Cálcio/metabolismo , Humanos , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Espécies Reativas de Oxigênio/metabolismoRESUMO
Changes in illumination can rapidly influence behavior that is normally controlled by the circadian clock. This effect is termed masking. In mice, masking requires melanopsin-expressing retinal ganglion cells that detect blue light and project to the thalamus. It is not known whether masking is wavelength-dependent in other vertebrates, nor is it known whether the thalamus is also involved or how it influences masking. Here, we address these questions in zebrafish. We find that diel vertical migration, a circadian behavior in larval zebrafish, is effectively triggered by blue, but not by red light. Two-photon calcium imaging reveals that a thalamic nucleus and a downstream structure, the habenula, have a sustained response to blue but not to red light. Lesioning the habenula reduces light-evoked climbing. These data suggest that the thalamo-habenula pathway is involved in the ability of blue light to influence a circadian behavior.
Assuntos
Ritmo Circadiano , Habenula/metabolismo , Transdução de Sinais , Tálamo/metabolismo , Peixe-Zebra/fisiologia , Animais , Cálcio/metabolismo , Sinalização do Cálcio/efeitos da radiação , Ritmo Circadiano/efeitos da radiação , Larva , Luz , Transdução de Sinais/efeitos da radiaçãoRESUMO
This research investigated the influence of extremely low frequency magnetic fields (ELF-MF; 50 Hz, 8 mT, 4 h per day, for 28 days) on calcium ion signaling and the double messenger system in the hippocampus of mice. Messengers that were studied included: G-protein, Ins(1,4,5)P3 (IP3 ), diacylglycerol (DAG), cAMP-dependent protein kinase (PKA), and Ca2+ -dependent protein kinase C (PKC). The results showed that ELF-MF caused an increase in the levels of Gi protein, IP3, DAG, PKA and PKC beta, calcium and calmodulin-dependent protein phosphatase calcineuring (PP2B), and intracellular Ca2+ content, and a decrease in calcium/calmodulin-dependent protein kinase II (CaMK II) and PKC alpha. In addition, ELF-MF exposure decreased the level of brain-derived neurotrophic factor (BDNF), which played a key role in hippocampal neuronal cell death. However, oral administration of procyanidins from lotus seedpod (LSPCs) (especially 90 mg kg-1 ) significantly recovered these changes, and nearly reached normal levels. All these showed that LSPCs may mediate calcium signal and double messenger system through Ca2+ /CaMK II/CREB/BDNF and DG/PKC/MAPK signaling pathways to reverse the alteration caused by ELF-MF. Bioelectromagnetics. 38:436-446, 2017. © 2017 Wiley Periodicals, Inc.
Assuntos
Biflavonoides/farmacologia , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/efeitos da radiação , Catequina/farmacologia , Hipocampo/citologia , Campos Magnéticos/efeitos adversos , Magnoliopsida/química , Proantocianidinas/farmacologia , Sementes/química , Animais , Biflavonoides/isolamento & purificação , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Catequina/isolamento & purificação , Diglicerídeos/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Hipocampo/efeitos dos fármacos , Hipocampo/efeitos da radiação , Inositol 1,4,5-Trifosfato/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos ICR , Proantocianidinas/isolamento & purificação , Proteínas Quinases/metabolismoRESUMO
The application of current channelrhodopsin-based optogenetic tools is limited by the lack of strict ion selectivity and the inability to extend the spectra sensitivity into the near-infrared (NIR) tissue transmissible range. Here we present an NIR-stimulable optogenetic platform (termed 'Opto-CRAC') that selectively and remotely controls Ca(2+) oscillations and Ca(2+)-responsive gene expression to regulate the function of non-excitable cells, including T lymphocytes, macrophages and dendritic cells. When coupled to upconversion nanoparticles, the optogenetic operation window is shifted from the visible range to NIR wavelengths to enable wireless photoactivation of Ca(2+)-dependent signaling and optogenetic modulation of immunoinflammatory responses. In a mouse model of melanoma by using ovalbumin as surrogate tumor antigen, Opto-CRAC has been shown to act as a genetically-encoded 'photoactivatable adjuvant' to improve antigen-specific immune responses to specifically destruct tumor cells. Our study represents a solid step forward towards the goal of achieving remote and wireless control of Ca(2+)-modulated activities with tailored function.
Assuntos
Sinalização do Cálcio/efeitos da radiação , Imunomodulação , Raios Infravermelhos , Optogenética/métodos , Animais , Células Dendríticas/fisiologia , Células Dendríticas/efeitos da radiação , Modelos Animais de Doenças , Macrófagos/fisiologia , Macrófagos/efeitos da radiação , Melanoma/imunologia , Melanoma/terapia , Camundongos , Linfócitos T/fisiologia , Linfócitos T/efeitos da radiaçãoRESUMO
One challenge in biology is signal transduction monitoring in a physiological context. Intravital imaging techniques are revolutionizing our understanding of tumor and host cell behaviors in the tumor environment. However, these deep tissue imaging techniques have not yet been adopted to investigate the second messenger calcium (Ca²âº). In the present study, we established conditions that allow the in vivo detection of Ca²âº signaling in three-dimensional tumor masses in mouse models. By combining intravital imaging and a skinfold chamber technique, we determined the ability of photodynamic cancer therapy to induce an increase in intracellular Ca²âº concentrations and, consequently, an increase in cell death in a p53-dependent pathway.
Assuntos
Sinalização do Cálcio/fisiologia , Microscopia Intravital/métodos , Neoplasias Experimentais/terapia , Fototerapia/métodos , Proteína Supressora de Tumor p53/metabolismo , Animais , Apoptose/fisiologia , Apoptose/efeitos da radiação , Sinalização do Cálcio/efeitos da radiação , Morte Celular/fisiologia , Morte Celular/efeitos da radiação , Feminino , Células HeLa , Humanos , Camundongos , Camundongos Nus , Camundongos Transgênicos , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologiaRESUMO
High mobility group box 1 (HMGB1) protein is a unique non histone nuclear protein that acts extracellularly as a mediator of delayed inflammation. Sub lethal dose of UVB triggers the release of cytokines from macrophages (MΦs). Adding to the panoply of UVB induced cytokines; it is reported that UVB induces HMGB1 release from mouse peritoneal MΦs in time and partially dose dependent manner, independent of TNF-α. UVB also enhanced the transcription of HMGB1 gene and expression of cellular protein, which influences its subsequent release. HMGB1 is secreted by an unconventional secretion pathway of unknown mechanism. Caspase-1 has been shown to function as a general regulator of stress induced unconventional secretion for a number of cytokines. In the present study, we have observed that pharmacological inhibitors specific for caspase-1 (ZVAD and YVAD) abrogated UVB induced HMGB1 release from MΦs. This effect was most likely mediated via physical interaction between HMGB1 and active caspase-1 (p10 and p20) as demonstrated by immunoprecipitation. In addition, it was found that HMGB1 and active caspase-1 p20 release depends on UVB mediated enhancement of intracellular Ca(2+). Thus our data suggests that optimal dose of UVB (50 mJ/cm(2)) induces HMGB1 upregulation and active release from mouse peritoneal MΦs which is mediated by caspase-1 in a Ca(2+) dependent manner.
Assuntos
Caspase 1/metabolismo , Proteína HMGA1a/metabolismo , Macrófagos Peritoneais/efeitos da radiação , Clorometilcetonas de Aminoácidos/farmacologia , Animais , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/efeitos da radiação , Células Cultivadas , Inibidores de Cisteína Proteinase/farmacologia , Proteína HMGA1a/genética , Macrófagos Peritoneais/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Oligopeptídeos/farmacologia , Via Secretória/efeitos dos fármacos , Via Secretória/efeitos da radiação , Ativação Transcricional/efeitos da radiação , Raios Ultravioleta/efeitos adversos , Regulação para Cima/efeitos da radiaçãoRESUMO
There is increasing emphasis on the use of systems biology approaches to define radiation-induced responses in cells and tissues. Such approaches frequently rely on global screening using various high throughput 'omics' platforms. Although these methods are ideal for obtaining an unbiased overview of cellular responses, they often cannot reflect the inherent heterogeneity of the system or provide detailed spatial information. Additionally, performing such studies with multiple sampling time points can be prohibitively expensive. Imaging provides a complementary method with high spatial and temporal resolution capable of following the dynamics of signaling processes. In this review, we utilize specific examples to illustrate how imaging approaches have furthered our understanding of radiation-induced cellular signaling. Particular emphasis is placed on protein colocalization, and oscillatory and transient signaling dynamics.
Assuntos
Regulação da Expressão Gênica/efeitos da radiação , Imagem Molecular/métodos , Transdução de Sinais/efeitos da radiação , Animais , Sinalização do Cálcio/efeitos da radiação , Dano ao DNA , Reparo do DNA , Ativação Enzimática/efeitos da radiação , Previsões , Humanos , Peroxidação de Lipídeos , Sistema de Sinalização das MAP Quinases/efeitos da radiação , Mapeamento de Interação de Proteínas , Proteínas Quinases/metabolismo , Espécies Reativas de Oxigênio , Análise de Célula ÚnicaRESUMO
Lepidopteran insects/insect cells display 50-100 times higher radioresistance than humans, and are evolutionarily closest to mammals amongst all radioresistant organisms known. Compared to mammalian cells, Lepidopteran cells (TN-368, Sf9) display more efficient antioxidant system and DNA repair and suffer considerably less radiation-induced DNA/cytogenetic damage and apoptosis. Recent studies indicate that a considerably lower radiation-induced oxidative stress may significantly reduce macromolecular damage in Lepidopteran cells. Since nitrosative stress contributes in radiation-induced cellular damage, we investigated its nature in the γ-irradiated Sf9 cells (derived from Spodoptera frugiperda; order Lepidoptera; family Noctuidae) and compared with BMG-1 human cell line having significant NOS expression. Radiation induced considerably less ROS/RNS in Sf9 cells, which remained unchanged on treatment with NOS inhibitor l-NMMA. Surprisingly, growth of Sf9 cultures or irradiation could not induce NO or its metabolites, indicating negligible basal/radiation-induced NOS activity that remained unchanged even after supplementation with arginine. Cytosolic calcium release following high-dose (1000-2000Gy at 61.1cGys(-1)) γ-irradiation or H(2)O(2) (250µM) treatment also failed to generate NO in Sf9 cells having high constitutive levels of calmodulin, whereas BMG-1 cells displayed considerable calcium-dependent NO generation even following 10Gy dose. These results strongly imply the lack of calcium-mediated NOS activity in Sf9 cells. Addition of exogenous NO from GSH-NO caused considerable increase in radiation-induced apoptosis, indicating significant contribution of constitutively attenuated nitrosative stress response into the radioresistance of Lepidopteran cells. Our study demonstrates for the first time that a calcium-insensitive, attenuated nitrosative stress response may contribute significantly in the unusual radioresistance displayed by Lepidopteran insect cells.
Assuntos
Cálcio/metabolismo , Lepidópteros/citologia , Tolerância a Radiação , Espécies Reativas de Nitrogênio/metabolismo , Animais , Arginina/metabolismo , Sinalização do Cálcio/efeitos da radiação , Calmodulina/metabolismo , Linhagem Celular , Hemina/farmacologia , Humanos , Proteínas de Insetos/metabolismo , Lepidópteros/enzimologia , Lepidópteros/efeitos da radiação , Potencial da Membrana Mitocondrial , Nitratos/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico/farmacologia , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/metabolismo , Nitritos/metabolismo , Estresse Oxidativo/efeitos da radiação , Espécies Reativas de Oxigênio/metabolismo , ômega-N-Metilarginina/farmacologiaRESUMO
Er,Cr:YSGG lasers are used clinically in dentistry. The advantages of laser therapy include minimal thermal damage and the alleviation of pain. This study examined whether the Er,Cr:YSGG laser has in vivo and in vitro antinociceptive effects in itself. In capsaicin-evoked acute licking/shaking tests and Hargreaves tests, laser irradiation with an aerated water spray suppressed nociceptive behavior in mice. Laser irradiation attenuated TRPV1 activation by capsaicin in Ca(2+) imaging experiments with TRPV1-overexpressing cells and cultured trigeminal neurons. Therefore, the laser-induced behavioral changes are probably due to the loss of TRPV1 activity. TRPV4 activity was also attenuated, but limited mechanical antinociception by the laser was observed. The laser failed to alter the other receptor functions, which indicates that the antinociceptive effect of the laser is dependent on TRPV1. These results suggest that the Er,Cr:YSGG laser has analgesic effects via TRPV1 inhibition. Such mechanistic approaches may help define the laser-sensitive pain modality and increase its beneficial uses.
Assuntos
Lasers de Estado Sólido/uso terapêutico , Nociceptores/efeitos da radiação , Dor/prevenção & controle , Canais de Cátion TRPV/efeitos da radiação , Animais , Comportamento Animal/efeitos dos fármacos , Comportamento Animal/efeitos da radiação , Bloqueadores dos Canais de Cálcio/farmacologia , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/efeitos da radiação , Capsaicina/farmacologia , Dinoprostona/farmacologia , Células HEK293 , Células HeLa , Temperatura Alta , Humanos , Masculino , Camundongos , Camundongos Endogâmicos ICR , Camundongos Knockout , Neurônios/efeitos dos fármacos , Neurônios/efeitos da radiação , Nociceptores/efeitos dos fármacos , Limiar da Dor/efeitos da radiação , Tempo de Reação/efeitos da radiação , Rutênio Vermelho/farmacologia , Fármacos do Sistema Sensorial/farmacologia , Limiar Sensorial/efeitos da radiação , Canais de Cátion TRPV/efeitos dos fármacos , Sensação Térmica/efeitos da radiação , Tato/efeitos da radiação , Nervo Trigêmeo/efeitos dos fármacos , Nervo Trigêmeo/efeitos da radiaçãoRESUMO
Low-level laser therapy (LLLT) has been found to produce anti-inflammatory effects in a variety of disorders. Bronchial smooth muscle (BSM) hyperreactivity is associated with increased Ca+2 sensitivity and increased RhoA mRNA expression. In the current study, we investigated if LLLT could reduce BSM contraction force and RhoA mRNA expression in tumor necrosis factor-alpha (TNF-alpha)-induced BSM hyperreactivity. In the study, 112 male Wistar rats were divided randomly into 16 groups, and BSM was harvested and suspended in TNF-alpha baths for 6 and 24 h, respectively. Irradiation with LLLT was performed with a wavelength of 660 nm for 42 s with a dose of 1.3 J/cm2. This LLLT dose was administered once in the 6-h group and twice in the 24-h group. LLLT significantly decreased contraction force in BSM at 6 h (TNF-alpha + LLLT: 11.65+/-1.10 g/100 mg of tissue) (F=3115) and at 24 h (TNF-alpha+ LLLT: 14.15+/-1.1 g/100 mg of tissue) (F=3245, p<0.05) after TNF-alpha, respectively, when compared to vehicle-bathed groups (control). LLLT also significantly decreased the expression of RhoA mRNA in BSM segments at 6 h (1.22+/-0.20) (F=2820, p<0.05) and 24 h (2.13+/-0.20) (F=3324, p<0.05) when compared to BSM segments incubated with TNF-alpha without LLLT irradiation. We conclude that LLLT administered with this protocol, reduces RhoA mRNA expression and BSM contraction force in TNF-alpha-induced BSM hyperreactivity.
Assuntos
Brônquios/efeitos dos fármacos , Brônquios/efeitos da radiação , Terapia com Luz de Baixa Intensidade , Músculo Liso/efeitos dos fármacos , Músculo Liso/efeitos da radiação , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Proteína rhoA de Ligação ao GTP/genética , Amidas/farmacologia , Animais , Brônquios/metabolismo , Hiper-Reatividade Brônquica/induzido quimicamente , Hiper-Reatividade Brônquica/genética , Hiper-Reatividade Brônquica/radioterapia , Sinalização do Cálcio/efeitos da radiação , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/efeitos da radiação , Técnicas In Vitro , Masculino , Músculo Liso/metabolismo , Piridinas/farmacologia , Ratos , Ratos WistarRESUMO
Transient receptor potential vanilloid type 1 (TRPV1) is a molecular sensor for detecting adverse stimuli, such as capsaicin, heat, and acid. TRPV1 has been localized in keratinocytes and is suggested to be a mediator of heat-induced matrix metalloproteinase-1 (MMP-1). With regard to the multimodal activation of TRPV1, we hypothesize that TRPV1 might also mediate UV-induced MMP-1 in keratinocytes. In HaCaT, a human keratinocyte cell line, we initially confirmed capsaicin-induced membrane current and Ca(2+) influx. UV irradiation induced slow and persistent calcium influx and increased membrane current, which was inhibited by TRPV1 inhibitors (capsazepine and ruthenium red). The UV-induced MMP-1 expression in HaCaT was also decreased by TRPV1 inhibitors and was facilitated by capsaicin. Knock-down of TRPV1 using siRNA transfection also decreased MMP-1 expression, as well as UV-induced Ca(2+) influx in HaCaT. UV failed to induce MMP-1 expression in HaCaT cells cultured in Ca(2+)-free media. Both the UV-induced increase in [Ca(2+)](i) and MMP-1 were suppressed by Gö6976 (a calcium-dependent PKC inhibitor), but not by rottlerin (a calcium-independent PKC inhibitor). In addition to a plausible role of TRPV1 in UV-induced MMP-1 expression, we showed that UV increased TRPV1 expression in both HaCaT cells and human skin in vivo. From these results, we suggest that UV-induced MMP-1 expression might be mediated in part by PKC-dependent activation of TRPV1 and subsequent Ca(2+)-influx in human keratinocytes. J. Cell. Physiol. 219: 766-775, 2009. (c) 2009 Wiley-Liss, Inc.