RESUMO
Wolfram syndrome (WS) is a rare genetic disease characterized by diabetes, optic atrophy and deafness. Patients die at 35 years of age, mainly from respiratory failure or dysphagia. Unfortunately, there is no treatment to block the progression of symptoms and there is an urgent need for adequate research models. Here, we report on the phenotypical characterization of two loss-of-function zebrafish mutant lines: wfs1aC825X and wfs1bW493X. We observed that wfs1a deficiency altered the size of the ear and the retina of the fish. We also documented a decrease in the expression level of unfolded protein response (UPR) genes in basal condition and in stress condition, i.e. after tunicamycin treatment. Interestingly, both mutants lead to a decrease in their visual function measured behaviorally. These deficits were associated with a decrease in the expression level of UPR genes in basal and stress conditions. Interestingly, basal, ATP-linked and maximal mitochondrial respirations were transiently decreased in the wfs1b mutant. Taken together, these zebrafish lines highlight the critical role of wfs1a and wfs1b in UPR, mitochondrial function and visual physiology. These models will be useful tools to better understand the cellular function of Wfs1 and to develop novel therapeutic approaches for WS.
Assuntos
Atrofia Óptica , Síndrome de Wolfram , Animais , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mutação , Atrofia Óptica/genética , Fenótipo , Síndrome de Wolfram/genética , Síndrome de Wolfram/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismoRESUMO
Purpose: In recent years, the increase in the biopesticides synthesis for alternative agricultural uses has required their impacts study. Among these compounds, several of them are known to exert endocrinedisrupting (EDs) effects causing deregulation of physiological functions affecting cell signaling pathways involved in neural cell differentiation leading to developmental neurotoxicity. The objective of our study was to determine the impact of the biopesticide A6 structurally related to estrogenic EDs on zebrafish larvae, to define its toxicity, the mechanisms responsible, and to monitor the locomotors activity at nanomolar concentrations (0. 0.5, 5 and 50 nM).Materials and methods: Using imaging analysis tools, immunohistochemistry, quantitative PCR, and an automated behavior recording system (Zebrabox) we were able to assess these effects.Results: We have shown through its blue fluorescence properties that it accumulates in different parts of the body such as the intestine, adipose tissue, muscles, yolk sac and head. A6 also disrupted swimming behavior by affecting the expression of tyrosine hydroxylase (TH) in dopaminergic neurons.Conclusions: In conclusion, our study provided a mechanistic understanding of the A6 neurotoxic effect which could be the result of its binding to the estrogen receptor.
Assuntos
Neuroquímica , Praguicidas , Animais , Expressão Gênica , Larva , Tirosina 3-Mono-Oxigenase/genética , Tirosina 3-Mono-Oxigenase/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismoRESUMO
Tauopathies represent a vast family of neurodegenerative diseases, the most well-known of which is Alzheimer's disease. The symptoms observed in patients include cognitive deficits and locomotor problems and can lead ultimately to dementia. The common point found in all these pathologies is the accumulation in neural and/or glial cells of abnormal forms of Tau protein, leading to its aggregation and neurofibrillary tangles. Zebrafish transgenic models have been generated with different overexpression strategies of human Tau protein. These transgenic lines have made it possible to highlight Tau interacting factors or factors which may limit the neurotoxicity induced by mutations and hyperphosphorylation of the Tau protein in neurons. Several studies have tested neuroprotective pharmacological approaches. On few-days-old larvae, modulation of various signaling or degradation pathways reversed the deleterious effects of Tau mutations, mainly hTauP301L and hTauA152T. Live imaging and live tracking techniques as well as behavioral follow-up enable the analysis of the wide range of Tau-related phenotypes from synaptic loss to cognitive functional consequences.
Assuntos
Modelos Animais de Doenças , Tauopatias , Peixe-Zebra , Proteínas tau/metabolismo , Animais , Animais Geneticamente Modificados , Humanos , Terapia de Alvo Molecular , Proteínas tau/genéticaRESUMO
The sigma-1 receptor (S1R) is a highly conserved transmembrane protein highly enriched in mitochondria-associated endoplasmic reticulum (ER) membranes, where it interacts with several partners involved in ER-mitochondria Ca2+ transfer, activation of the ER stress pathways, and mitochondria function. We characterized a new S1R deficient zebrafish line and analyzed the impact of S1R deficiency on visual, auditory and locomotor functions. The s1r+25/+25 mutant line showed impairments in visual and locomotor functions compared to s1rWT. The locomotion of the s1r+25/+25 larvae, at 5 days post fertilization, was increased in the light and dark phases of the visual motor response. No deficit was observed in acoustic startle response. A critical role of S1R was shown in ER stress pathways and mitochondrial activity. Using qPCR to analyze the unfolded protein response genes, we observed that loss of S1R led to decreased levels of IRE1 and PERK-related effectors and increased over-expression of most of the effectors after a tunicamycin challenge. Finally, S1R deficiency led to alterations in mitochondria bioenergetics with decreased in basal, ATP-linked and non-mitochondrial respiration and following tunicamycin challenge. In conclusion, this new zebrafish model confirmed the importance of S1R activity on ER-mitochondria communication. It will be a useful tool to further analyze the physiopathological roles of S1R.
Assuntos
Mitocôndrias/metabolismo , Receptores sigma/metabolismo , Resposta a Proteínas não Dobradas , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/metabolismo , Animais , Animais Geneticamente Modificados/metabolismo , Sistemas CRISPR-Cas/genética , Edição de Genes , Larva/fisiologia , Locomoção , Proteínas de Membrana/metabolismo , Fenótipo , Receptores sigma/química , Receptores sigma/genética , Peixe-Zebra/crescimento & desenvolvimento , Proteínas de Peixe-Zebra/química , Proteínas de Peixe-Zebra/genética , Receptor Sigma-1RESUMO
Brain-derived neurotrophic factor (BDNF) dysregulations contribute to the neurotoxicity in neurodegenerative pathologies and could be efficiently targeted by therapies. In Alzheimer's disease (AD), although the relationship between BDNF and amyloid load has been extensively studied, how Tau pathology affects BDNF signaling remains unclear. Using the TAU-P301L transgenic zebrafish line, we investigated how early Tau-induced neurotoxicity modifies BDNF signaling. Alterations in BDNF expression levels were observed as early as 48 h post fertilization in TAU-P301L zebrafish embryos while TrkB receptor expression was not affected. Decreasing BDNF expression, using a knockdown strategy in wild-type embryos to mimic Tau-associated decrease, did not modify TrkB expression but promoted neurotoxicity as demonstrated by axonal outgrowth shortening and neuronal cell death. Moreover, the TrkB antagonist ANA-12 reduced the length of axonal projections. Rescue experiments with exogenous BDNF partially corrected neuronal alterations in TAU-P301L by counteracting primary axonal growth impairment but without effect on apoptosis. Importantly, the axonal rescue was proved functionally effective in a behavioral test, at a similar level as obtained with the GSK3ß inhibitor LiCl, known to decrease TAU phosphorylation. Finally, treatment with a TrkB agonist, 7,8-dihydroxyflavone, led to comparable results and allowed full rescue of locomotor response. We provided here strong evidence that Tau neurotoxicity provoked alterations in BDNF system and that BDNF pathway might represent an efficient therapeutic target.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/genética , Fármacos Neuroprotetores , Tauopatias/tratamento farmacológico , Peixe-Zebra , Animais , Axônios/efeitos dos fármacos , Axônios/ultraestrutura , Comportamento Animal/efeitos dos fármacos , Fator Neurotrófico Derivado do Encéfalo/agonistas , Fator Neurotrófico Derivado do Encéfalo/biossíntese , Fator Neurotrófico Derivado do Encéfalo/farmacologia , Morte Celular , Humanos , Larva , Cultura Primária de Células , Receptor trkB/biossíntese , Proteínas Recombinantes/farmacologia , Tauopatias/genéticaRESUMO
In previous work [Opt. Lett.44, 2827 (2019)OPLEDP0146-959210.1364/OL.44.002827], we presented a method based on digital holography and orthogonal matching pursuit, which is able to determine the 3D positions of small objects moving within a larger motionless object. Indeed, if the scattering density is sparse in direct 3D space, compressive sensing algorithms can be used. The method was validated by imaging red blood cell trajectories in the trunk vascular system of a zebrafish (Danio rerio) larva. We give here further details on the reconstruction technique and present a more robust version of the algorithm based on multiple illuminations.
RESUMO
REG-1α, a secreted protein containing a C-type lectin domain, is expressed in various organs and plays different roles in digestive system cells in physiological and pathological conditions. Like other members of the Reg family, REG-1α is expressed also in the brain where it has different functions. For instance, we previously reported that REG-1α regulates neurite outgrowth and is overexpressed during the very early stages of Alzheimer's disease (AD). However, REG-1α function in neural cells during neural degeneration remains unknown. First, REG-1α and phosphorylated tau expression were assessed in tissue sections from the hippocampus, representing neurofibrillary tangles (NFTs), from patients with AD, and from basal ganglia, representing subcortical NFTs, from patients with progressive supranuclear palsy (PSP). We found an association between REG-1α expression, tau hyperphosphorylation and NFTs in human brain samples from patients with these neurodegenerative diseases. Then, the effects of REG-1α overexpression on tau phosphorylation and axonal morphology were investigated i) in primary cultures of rat neurons that express human tau P301L and ii) in a transgenic zebrafish model of tauopathy that expresses human tau P301L. In the tau P301L cell model, REG-1α overexpression increased tau phosphorylation at the S202/T205 and S396 residues (early and late stages of abnormal phosphorylation, respectively) through the AKT/GSK3-ß pathway. This effect was associated with axonal defects both in tau P301L-expressing rat neurons and zebrafish embryos. Our findings suggest a functional role for REG-1α during tauopathy development and progression and, specifically, its involvement in the modification of tau phosphorylation temporal sequence.
Assuntos
Modelos Animais de Doenças , Litostatina/biossíntese , Tauopatias/metabolismo , Proteínas tau/biossíntese , Idoso , Idoso de 80 Anos ou mais , Animais , Animais Geneticamente Modificados , Encéfalo/metabolismo , Encéfalo/patologia , Células Cultivadas , Feminino , Humanos , Litostatina/genética , Masculino , Pessoa de Meia-Idade , Fosforilação/fisiologia , Ratos , Tauopatias/genética , Tauopatias/patologia , Peixe-Zebra , Proteínas tau/genéticaRESUMO
The primordium that generates the embryonic posterior lateral line of zebrafish migrates from the head to the tip of the tail along a trail of SDF1-producing cells. This migration critically depends on the presence of the SDF1 receptor CXCR4 in the leading region of the primordium and on the presence of a second SDF1 receptor, CXCR7, in the trailing region of the primordium. Here we show that inactivation of the estrogen receptor ESR1 results in ectopic expression of cxcr4b throughout the primordium, whereas ESR1 overexpression results in a reciprocal reduction in the domain of cxcr4b expression, suggesting that ESR1 acts as a repressor of cxcr4b. This finding could explain why estrogens significantly decrease the metastatic ability of ESR-positive breast cancer cells. ESR1 inactivation also leads to extinction of cxcr7b expression in the trailing cells of the migrating primordium; this effect is indirect, however, and due to the down-regulation of cxcr7b by ectopic SDF1/CXCR4 signaling in the trailing region. Both ESR1 inactivation and overexpression result in aborted migration, confirming the importance of this receptor in the control of SDF1-dependent migration.
Assuntos
Movimento Celular , Receptor alfa de Estrogênio/metabolismo , Sistema da Linha Lateral/embriologia , Sistema da Linha Lateral/metabolismo , Receptores CXCR4/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Peixe-Zebra/metabolismo , Animais , Sequência de Bases , Receptor alfa de Estrogênio/genética , Regulação da Expressão Gênica no Desenvolvimento , Dados de Sequência Molecular , Receptores CXCR/genética , Receptores CXCR/metabolismo , Receptores CXCR4/genética , Transdução de Sinais , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genéticaRESUMO
Photodynamic therapy (PDT) and photochemical internalization (PCI) are two methods that use light to provoke cell death or disturbance of cellular membranes, respectively, via excitation of a photosensitizer and the formation of reactive oxygen species (ROS). In this context, two-photon excitation (TPE) is of high interest for PCI and/or PDT due to spatiotemporal resolution of two-photon light and deeper penetration of near-infrared light in biological tissues. Here, we report that Periodic Mesoporous Ionosilica Nanoparticles (PMINPs) containing porphyrin groups allow the complexation of pro-apoptotic siRNA. These nano-objects were incubated with MDA-MB-231 breast cancer cells, and TPE-PDT led to significant cell death. Finally, MDA-MB-231 breast cancer cells were pre-incubated with the nanoparticles and then injected in the pericardial cavity of zebrafish embryos. After 24 h, the xenografts were irradiated with femtosecond pulsed laser and the size monitoring by imaging showed a decrease 24 h after irradiation. Pro-apoptotic siRNA was complexed with the nanoparticles and incubation with MDA-MB-231 cells did not lead to cancer cell death in dark conditions, but with two-photon irradiation, TPE-PCI was observed and a synergic effect between pro-apoptotic siRNA and TPE-PDT was noticed, leading to 90% of cancer cell death. Therefore, PMINPs represent an interesting system for nanomedicine applications.
Assuntos
Neoplasias da Mama , Nanopartículas , Fotoquimioterapia , Animais , Humanos , Feminino , Peixe-Zebra , RNA Interferente Pequeno/uso terapêutico , Fármacos Fotossensibilizantes/uso terapêutico , Fotoquimioterapia/métodos , Neoplasias da Mama/tratamento farmacológico , Inativação Gênica , Linhagem Celular TumoralRESUMO
DFNA68 is a rare subtype of autosomal dominant nonsyndromic hearing impairment caused by heterozygous alterations in the HOMER2 gene. To date, only 5 pathogenic or likely pathogenic coding variants, including two missense substitutions (c.188 C > T and c.587 G > C), a single base pair duplication (c.840dupC) and two short deletions (c.592_597delACCACA and c.832_836delCCTCA) have been described in 5 families. In this study, we report a novel HOMER2 variation, identified by massively parallel sequencing, in a Sicilian family suffering from progressive dominant hearing loss over 3 generations. This novel alteration is a nonstop substitution (c.1064 A > G) that converts the translational termination codon (TAG) of the gene into a tryptophan codon (TGG) and is predicted to extend the HOMER2 protein by 10 amino acids. RNA analyses from the proband suggested that HOMER2 transcripts carrying the nonstop variant escaped the non-stop decay pathway. Finally, in vivo studies using a zebrafish animal model and behavioral tests clearly established the deleterious impact of this novel HOMER2 alteration on hearing function. This study identifies the fourth causal variation responsible for DFNA68 and describes a simple in vivo approach to assess the pathogenicity of candidate HOMER2 variants.
Assuntos
Surdez , Perda Auditiva Neurossensorial , Perda Auditiva , Animais , Códon de Terminação , Surdez/genética , Perda Auditiva/genética , Perda Auditiva Neurossensorial/genética , Mutação , Linhagem , Peixe-Zebra/genéticaRESUMO
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease affecting upper and lower motor neurons. As a consequence, ALS patients display a locomotor disorder related to muscle weakness and progressive paralysis. Pathological mechanisms that participate in ALS involve deficient unfolded protein response, mitochondrial dysfunction and oxidative stress, among others. Finding a therapeutic target to break the vicious circle is particularly challenging. Sigma-1 receptor (S1R) is an endoplasmic reticulum (ER) chaperone that may be one of those targets. We here address and decipher the efficiency of S1R activation on a key ALS gene, TDP43, in zebrafish vertebrate model. While expression of mutant TDP43 (TDP43G348C) led to locomotor defects, treatment with the reference S1R agonist PRE-084 rescued motor performances in a zebrafish model. Treatment with the agonist ameliorated maximal mitochondrial respiration in the TDP43 context. We observed that TDP43G348C exacerbated ER stress induced by tunicamycin, resulting in increased levels of ER stress chaperone BiP and pro-apoptotic factor CHOP. Importantly, PRE-084 treatment in the same condition further heightened BiP levels but also EIF2α/ATF4 and NRF2 signalling cascades, both known to promote antioxidant protection during ER stress. Moreover, we showed that increasing NRF2 levels directly or by sulforaphane treatment rescued locomotor defects of TDP43G348C zebrafish. For the first time, we here provide the proof of concept that PRE-084 prevents mutant TDP43 toxicity by boosting ER stress response and antioxidant cascade through NRF2 signalling.
Assuntos
Esclerose Lateral Amiotrófica , Doenças Neurodegenerativas , Animais , Peixe-Zebra/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Antioxidantes/uso terapêutico , Esclerose Lateral Amiotrófica/genética , Proteínas de Ligação a DNA/genética , Estresse do Retículo Endoplasmático , Receptor Sigma-1RESUMO
Wolfram syndrome (WS) is a rare neurodegenerative disease resulting in deafness, optic atrophy, diabetes, and neurological disorders. Currently, no treatment is available for patients. The mutated gene, WFS1, encodes an endoplasmic reticulum (ER) protein, Wolframin. We previously reported that Wolframin regulated the ER-mitochondria Ca2+ transfer and mitochondrial activity by protecting NCS1 from degradation in patients' fibroblasts. We relied on a zebrafish model of WS, the wfs1ab KO line, to analyze the functional and behavioral impact of NCS1 overexpression as a novel therapeutic strategy. The wfs1ab KO line showed an increased locomotion in the visual motor and touch-escape responses. The absence of wfs1 did not impair the cellular unfolded protein response, in basal or tunicamycin-induced ER stress conditions. In contrast, metabolic analysis showed an increase in mitochondrial respiration in wfs1ab KO larvae. Interestingly, overexpression of NCS1 using mRNA injection restored the alteration of mitochondrial respiration and hyperlocomotion. Taken together, these data validated the wfs1ab KO zebrafish line as a pertinent experimental model of WS and confirmed the therapeutic potential of NCS1. The wfs1ab KO line therefore appeared as an efficient model to identify novel therapeutic strategies, such as gene or pharmacological therapies targeting NCS1 that will correct or block WS symptoms.
RESUMO
The Wolfram syndrome is a rare autosomal recessive disease affecting many organs with life-threatening consequences; currently, no treatment is available. The disease is caused by mutations in the WSF1 gene, coding for the protein wolframin, an endoplasmic reticulum (ER) transmembrane protein involved in contacts between ER and mitochondria termed as mitochondria-associated ER membranes (MAMs). Inherited mutations usually reduce the protein's stability, altering its homeostasis and ultimately reducing ER to mitochondria calcium ion transfer, leading to mitochondrial dysfunction and cell death. In this study, we found that activation of the sigma-1 receptor (S1R), an ER-resident protein involved in calcium ion transfer, could counteract the functional alterations of MAMs due to wolframin deficiency. The S1R agonist PRE-084 restored calcium ion transfer and mitochondrial respiration in vitro, corrected the associated increased autophagy and mitophagy, and was able to alleviate the behavioral symptoms observed in zebrafish and mouse models of the disease. Our findings provide a potential therapeutic strategy for treating Wolfram syndrome by efficiently boosting MAM function using the ligand-operated S1R chaperone. Moreover, such strategy might also be relevant for other degenerative and mitochondrial diseases involving MAM dysfunction.
Assuntos
Receptores sigma , Síndrome de Wolfram , Animais , Cálcio/metabolismo , Feminino , Humanos , Masculino , Camundongos , Receptores sigma/agonistas , Peixe-Zebra/metabolismo , Receptor Sigma-1RESUMO
The embryonic development of the posterior lateral line of zebrafish involves the migration from head to tail of a primordium comprising approximately 100 cells, and the deposition at regular intervals of presumptive mechanosensory organs (neuromasts). Migration depends on the presence of chemokine SDF1 along the pathway, and on the asymmetrical distribution of chemokine receptors CXCR4 and CXCR7 in the primordium. Primordium polarization depends on Wnt signaling in the leading region. Here, we examine the role of a major effector of Wnt signaling, lef1, in this system. We show that, although its inactivation has no overt effect on the expression of cxcr4b and cxcr7b, lef1 contributes to their control. We also show that cell proliferation, which ensures constant primordium size despite successive rounds of cell deposition, is reduced upon lef1 inactivation. Because of this defect, the primordium runs short of cells and vanishes before the line has been completed. We conclude that lef1-mediated Wnt signaling is involved in various aspects of primordium migration, although part of this implication is masked by a high level of developmental redundancy.
Assuntos
Padronização Corporal/fisiologia , Sistema da Linha Lateral/embriologia , Fatores de Transcrição/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Animais , Padronização Corporal/genética , Proliferação de Células , Hibridização In Situ , Reação em Cadeia da Polimerase , Regiões Promotoras Genéticas/genética , Receptores CXCR/genética , Receptores CXCR4/genética , Fatores de Transcrição/genética , Peixe-Zebra , Proteínas de Peixe-Zebra/genéticaRESUMO
Retinitis pigmentosa (RP) is one of the most common forms of inherited retinal degeneration with 1/4,000 people being affected. The vision alteration primarily begins with rod photoreceptor degeneration, then the degenerative process continues with cone photoreceptor death. Variants in 71 genes have been linked to RP. One of these genes, PDE6a is responsible for RP43. To date no treatment is available and patients suffer from pronounced visual impairment in early childhood. We used the novel zebrafish pde6aQ70X mutant, generated by N-ethyl-N-nitrosourea at the European Zebrafish Resource Centre, to better understand how PDE6a loss of function leads to photoreceptor alteration. Interestingly, zebrafish pde6aQ70X mutants exhibited impaired visual function at 5 dpf as evidenced by the decrease in their visual motor response (VMR) compared to pde6a WT larvae. This impaired visual function progressed with time and was more severe at 21 dpf. These modifications were associated with an alteration of rod outer segment length at 5 and 21 dpf. In summary, these findings suggest that rod outer segment shrinkage due to Pde6a deficiency begins very early in zebrafish, progresses with time. The zebrafish pde6aQ70X mutant represents an ideal model of RP to screen relevant active small molecules that will block the progression of the disease.
RESUMO
The ubiquitous use of ethinylestradiol (EE2), an active constituent of birth control preparations, results in continuous release of this synthetic estrogen to surface waters. Many studies document the untoward effects of EE2 on the endocrine system of aquatic organisms. Effects of environmental EE2 on the nervous system are still poorly documented. We studied effects of pico- to nanomolar concentrations of EE2 on early nervous system development of zebrafish larvae. EE2 disrupted axonal nerve regeneration and hair cell regeneration up to 50%. Gene expression in larval brain tissues showed significantly upregulated expression of target genes, such as estrogen and progesterone receptors, and aromatase B. In contrast, downregulation of the tyrosine hydroxylase, involved in the synthesis of neurotransmitters, occurred concomitant with diminution of proliferating cells. Overall, the size of exposed fish larvae decreased by 25% and their swimming behavior was modified compared to non-treated larvae. EE2 interferes with nervous system development, both centrally and peripherally, with negative effects on regeneration and swimming behavior. Survival of fish and other aquatic species may be at risk in chronically EE2-contaminated environments.
Assuntos
Etinilestradiol , Poluentes Químicos da Água , Animais , Anticoncepção , Etinilestradiol/toxicidade , Larva , Sistema Nervoso , Natação , Poluentes Químicos da Água/toxicidade , Peixe-ZebraRESUMO
We investigated a series of Mn2+-Prussian blue (PB) nanoparticles NazMnxFe1-x[Fe(CN)6]1-yâ¡y·nH2O of similar size, surface state and cubic morphology with various amounts of Mn2+ synthesized through a one step self-assembly reaction. We demonstrated by a combined experimental-theoretical approach that during the synthesis, Mn2+ substituted Fe3+ up to a Mn/Na-Mn-Fe ratio of 32 at% in the PB structure, while for higher amounts, the Mn2[Fe(CN)6] analogue is obtained. For comparison, the post-synthetic insertion of Mn2+ in PB nanoparticles was also investigated and completed with Monte-Carlo simulations to probe the plausible adsorption sites. The photothermal conversion efficiency (η) of selected samples was determined and showed a clear dependence on the Mn2+amount with a maximum efficiency for a Mn/Na-Mn-Fe ratio of 10 at% associated with a dependence on the nanoparticle concentration. Evaluation of the in vitro photothermal properties of these nanoparticles performed on triple negative human breast adenocarcinoma (MDA-MB-231) cells by using continuous and pulsed laser irradiation confirm their excellent PTT efficiency permitting low dose use.
Assuntos
Antineoplásicos/uso terapêutico , Ferrocianetos/uso terapêutico , Manganês/química , Nanopartículas/uso terapêutico , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Animais , Antineoplásicos/síntese química , Antineoplásicos/efeitos da radiação , Linhagem Celular Tumoral , Ferrocianetos/química , Ferrocianetos/efeitos da radiação , Humanos , Ferro/química , Ferro/efeitos da radiação , Manganês/efeitos da radiação , Nanopartículas/química , Nanopartículas/efeitos da radiação , Processos Fotoquímicos , Terapia Fototérmica , Ensaios Antitumorais Modelo de Xenoenxerto , Peixe-ZebraRESUMO
Migration of facial motoneurons in the zebrafish hindbrain depends on SDF1/CXCL12 signaling. Recent studies demonstrated that SDF1 can bind two chemokine receptors, CXCR4 and CXCR7. Here we explore the expression and function of the cxcr7b gene in zebrafish hindbrain development. By the time cxcr4b-expressing motoneurons migrate from rhombomere (r) r4 to r6, expression of cxcr7b is rapidly restricted to the ventral part of r5. Inactivation of either cxcr7b or cxcr4b impairs motoneuron migration, with however different phenotypes. Facial motoneurons preferentially accumulate in r5 in cxcr7b morphant embryos, while they are distributed between r4, r5 and r6 in cxcr4b morphants. Simultaneous inactivation of both receptors leads to yet a third phenotype, with motoneurons mostly distributed between r4 and r5. The latter phenotype resembles that of sdf1a morphant embryos. Double inactivation of sdf1a and cxcr7b indeed did not lead to a complete arrest of migration but rather to a partial rescue of r5 arrest of motoneuron migration. This result is in accordance with the functional hypothesis that SDF1 might interact with CXCR7 and that they have an antagonistic effect within r5. The ectopic expression of a truncated CXCR7 receptor leads to a motoneuron migration defect. Altogether, we show that CXCR7 is required, for proper tangential migration of facial motoneurons, by determining a permissive migration pathway through r5.
Assuntos
Movimento Celular/fisiologia , Nervo Facial/citologia , Neurônios Motores/fisiologia , Receptores CXCR4/metabolismo , Receptores CXCR/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Animais , Quimiocina CXCL12/genética , Quimiocina CXCL12/metabolismo , Humanos , Metaloproteinases da Matriz/metabolismo , Neurônios Motores/citologia , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo , Receptores CXCR/genética , Receptores CXCR4/genética , Rombencéfalo/citologia , Rombencéfalo/embriologia , Peixe-Zebra/anatomia & histologia , Proteínas de Peixe-Zebra/genéticaRESUMO
The posterior lateral line (PLL) of zebrafish comprises seven to eight sense organs at the end of embryogenesis, arranged in a single antero-posterior line that extends along the horizontal myoseptum from the ear to the tip of the tail. At the end of larval life, four antero-posterior lines extend on the trunk and tail, comprising together around 60 sense organs. The embryonic pattern is largely conserved among teleosts, although adult patterns are very diverse. Here we describe the transition from embryonic to juvenile pattern in the zebrafish, to provide a framework for understanding how the diversity of adult patterns comes about. We show that the four lines that extend over the adult body originate from latent precursors laid down by migrating primordia that arise during embryogenesis. We conclude that, in zebrafish, the entire development of the PLL system up to adulthood can be traced back to events that took place during the first 2 days of life. We also show that the transition from embryonic to adult pattern involves few distinct operations, suggesting that the diversity of patterns among adult teleosts may be due to differential control of these few operations acting upon common embryonic precursors.
Assuntos
Padronização Corporal , Sistema da Linha Lateral/crescimento & desenvolvimento , Peixe-Zebra/crescimento & desenvolvimento , Animais , Embrião não Mamífero/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Peixe-Zebra/anatomia & histologia , Peixe-Zebra/genética , Peixe-Zebra/fisiologiaRESUMO
BACKGROUND: The formation of the posterior lateral line of teleosts depends on the migration of a primordium that originates near the otic vesicle and moves to the tip of the tail. Groups of cells at the trailing edge of the primordium slow down at regular intervals and eventually settle to differentiate as sense organs. The migration of the primordium is driven by the chemokine SDF1 and by its receptor CXCR4, encoded respectively by the genes sdf1a and cxcr4b. cxcr4b is expressed in the migrating cells and is down-regulated in the trailing cells of the primordium. sdf1a is expressed along the path of migration. There is no evidence for a gradient of sdf1a expression, however, and the origin of the directionality of migration is not known. RESULTS: Here we document the expression of a second chemokine receptor gene, cxcr7, in the migrating primordium. We show that cxcr7 is highly expressed in the trailing cells of the primordium but not at all in the leading cells, a pattern that is complementary to that of cxcr4b. Even though cxcr7 is not expressed in the cells that lead primordium migration, its inactivation results in impaired migration. The phenotypes of cxcr4b, cxcr7 double morphant embryos suggest, however, that CXCR7 does not contribute to the migratory capabilities of primordium cells. We also show that, in the absence of cxcr4b, expression of cxcr7 becomes ubiquitous in the stalled primordium. CONCLUSION: Our observations suggest that CXCR7 is required to provide directionality to the migration. We propose that directionality is imposed on the primordium as soon as it comes in contact with the stripe of SDF1, and is maintained throughout migration by a negative interaction between the two receptors.