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Trastuzumab emtansine (T-DM1) is a targeted therapy combining trastuzumab and emtansine for human epidermal growth factor receptor 2(HER2)-positive breast cancer, with common side effects including fatigue, nausea, pain, headache, low platelet count, and elevated liver enzymes. Hereditary Hemorrhagic Telangiectasia (HHT) is an autosomal dominant vascular dysplasia characterized by vascular malformations and telangiectasias in various organs. We present a case of a female patient with advanced breast cancer who developed HHT-like symptoms while on T-DM1 treatment. A 59-year-old woman treated with radiotherapy and T-DM1 every 21 days developed recurring nosebleeds and mucocutaneous and liver telangiectasias indistinguishable from HHT three months after receiving the first dose of T-DM1. Other organ vascular malformations were ruled out through screening protocols. The patient had no previous HHT symptoms or family history. Nasal care measures like lubrication and antifibrinolytics (tranexamic acid) were provided. In addition, propranolol was also prescribed due to its antiangiogenic and antitumoral properties, leading to significantly decreased epistaxis and telangiectasias. Microtubule disruptions caused by T-DM1, along with other angiogenic mechanisms may contribute to the development of telangiectasias resembling HHT. The use of propranolol, an initial approach for HHT, proved to be effective in this case. It is crucial for oncologists and HHT specialists to be aware of this rare adverse event associated with T-DM1 and to implement appropriate management strategies.
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OBJECTIVE: To study whether male factor infertility and insomnia share genetic risk variants and identify any molecular, cellular, and biologic interactions between these traits. DESIGN: The in silico study was performed. Two lists of genetic variants were manually curated through a literature review, one of those associated with male factor infertility and the other with insomnia. Genes were assigned to these variants to compose male factor infertility-associated (454 genes) and insomnia-associated (921 genes) gene lists. SETTING: Not applicable. PATIENT(S): Not applicable. INTERVENTION(S): Not applicable. MAIN OUTCOME MEASURE(S): Enrichment of biologic pathways and protein-protein interaction analysis. RESULT(S): Twenty-eight genes were common to both lists, representing a greater overlap than would be expected by chance. In the 28 genes contained in the intersection list, there was a significant enrichment of pathways related to kinesin binding. A protein-protein interaction analysis using the intersection list as input retrieved 25 nodes and indicated that two of them were kinesin-related proteins (PLEKHM2 and KCL1). CONCLUSION(S): The shared male factor infertility and insomnia genes, and the biologic pathways highlighted in this study, suggest that further functional investigations into the interplay between fertility and sleep are warranted.
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Infertilidad Masculina , Cinesinas , Trastornos del Inicio y del Mantenimiento del Sueño , Masculino , Humanos , Trastornos del Inicio y del Mantenimiento del Sueño/genética , Trastornos del Inicio y del Mantenimiento del Sueño/metabolismo , Infertilidad Masculina/genética , Cinesinas/genética , Unión Proteica , Mapas de Interacción de Proteínas , Predisposición Genética a la EnfermedadRESUMEN
Microtubules are components of the cytoskeleton that perform essential functions in eukaryotes, such as those related to shape change, motility and cell division. In this context some characteristics of these filaments are essential, such as polarity and dynamic instability. In trypanosomatids, microtubules are integral to ultrastructure organization, intracellular transport and mitotic processes. Some species of trypanosomatids co-evolve with a symbiotic bacterium in a mutualistic association that is marked by extensive metabolic exchanges and a coordinated division of the symbiont with other cellular structures, such as the nucleus and the kinetoplast. It is already established that the bacterium division is microtubule-dependent, so in this work, it was investigated whether the dynamism and remodeling of these filaments is capable of affecting the prokaryote division. To this purpose, Angomonas deanei was treated with Trichostatin A (TSA), a deacetylase inhibitor, and mutant cells for histone deacetylase 6 (HDAC6) were obtained by CRISPR-Cas9. A decrease in proliferation, an enhancement in tubulin acetylation, as well as morphological and ultrastructural changes, were observed in TSA-treated protozoa and mutant cells. In both cases, symbiont filamentation occurred, indicating that prokaryote cell division is dependent on microtubule dynamism.
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División Celular , Microtúbulos , Simbiosis , Microtúbulos/metabolismo , Microtúbulos/ultraestructura , Microtúbulos/efectos de los fármacos , Trypanosomatina/genética , Trypanosomatina/metabolismo , Trypanosomatina/ultraestructura , Trypanosomatina/fisiología , Ácidos Hidroxámicos/farmacología , Tubulina (Proteína)/metabolismo , Tubulina (Proteína)/genética , Bacterias/metabolismo , Bacterias/genética , Acetilación , Inhibidores de Histona Desacetilasas/farmacología , Histona Desacetilasa 6/metabolismo , Histona Desacetilasa 6/genética , Citoesqueleto/metabolismo , Citoesqueleto/ultraestructuraRESUMEN
Rapid hypocotyl elongation allows buried seedlings to emerge, where light triggers de-etiolation and inhibits hypocotyl growth mainly by photoreceptors. Phosphorylation/dephosphorylation events regulate many aspects of plant development. Only recently we have begun to uncover the earliest phospho-signaling responders to light. Here, we reported a large-scale phosphoproteomic analysis and identified 20 proteins that changed their phosphorylation pattern following a 20 min light pulse compared to darkness. Microtubule-associated proteins were highly overrepresented in this group. Among them, we studied CIP7 (COP1-INTERACTING-PROTEIN 7), which presented microtubule (MT) localization in contrast to the previous description. An isoform of CIP7 phosphorylated at Serine915 was detected in etiolated seedlings but was undetectable after a light pulse in the presence of photoreceptors, while CIP7 transcript expression decays with long light exposure. The short hypocotyl phenotype and rearrangement of MTs in etiolated cip7 mutants are complemented by CIP7-YFP and the phospho-mimetic CIP7S915D-YFP, but not the phospho-null CIP7S915A-YFP suggesting that the phosphorylated S915CIP7 isoform promotes hypocotyl elongation through MT reorganization in darkness. Our evidence on Serine915 of CIP7 unveils phospho-regulation of MT-based processes during skotomorphogenic hypocotyl growth.
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Proteínas de Arabidopsis , Arabidopsis , Oscuridad , Hipocótilo , Proteínas Asociadas a Microtúbulos , Hipocótilo/crecimiento & desarrollo , Hipocótilo/genética , Hipocótilo/metabolismo , Arabidopsis/crecimiento & desarrollo , Arabidopsis/metabolismo , Arabidopsis/genética , Arabidopsis/efectos de la radiación , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Fosforilación , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Asociadas a Microtúbulos/genética , Microtúbulos/metabolismo , Luz , Regulación de la Expresión Génica de las Plantas , Plantones/crecimiento & desarrollo , Plantones/genética , Plantones/metabolismo , Plantones/efectos de la radiaciónRESUMEN
Amyloid-ß (Aß) and hyperphosphorylated tau (P-tau) are Alzheimer's disease (AD) biomarkers that interact in a complex manner to induce most of the cognitive and brain alterations observed in this disease. Since the neuronal cytoskeleton is a common downstream pathological target of tau and Aß, which mostly lead to augmented microtubule instability, the administration of microtubule stabilizing agents (MSAs) can protect against their pathological actions. However, the effectiveness of MSAs is still uncertain due to their state-dependent negative effects; thus, evaluating their specific actions in different pathological or physiological conditions is required. We evaluated whether epothilone-D (Epo-D), a clinically used MSA, rescues from the functional and behavioral alterations produced by intracerebroventricular injection of Aß, the presence of P-tau, or their combination in rTg4510 mice. We also explored the side effects of Epo-D. To do so, we evaluated hippocampal-dependent spatial memory with the Hebb-Williams maze, hippocampal CA1 integrity and the intrinsic and synaptic properties of CA1 pyramidal neurons with the patch-clamp technique. Aß and P-tau mildly impaired memory retrieval, but produced contrasting effects on intrinsic excitability. When Aß and P-tau were combined, the alterations in excitability and spatial reversal learning (i.e., cognitive flexibility) were exacerbated. Interestingly, Epo-D prevented most of the impairments induced Aß and P-tau alone and combined. However, Epo-D also exhibited some side effects depending on the prevailing pathological or physiological condition, which should be considered in future preclinical and translational studies. Although we did not perform extensive histopathological evaluations or measured microtubule stability, our findings show that MSAs can rescue the consequences of AD-like conditions but otherwise be harmful if administered at a prodromal stage of the disease.
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Giardia lamblia is a globally distributed protozoan parasite that causes intestinal disease. Recently, there is an increase in refractory cases of giardiasis to chemotherapeutic agents, and drugs available cause side effects that may limit its use or cause therapeutic non-compliance. Therefore, search for alternative and less harmful drugs to treat giardiasis is an important task. In this sense, resveratrol (RSV) is a polyphenol with a wide range of pharmacological effects such as antimicrobial, anticarcinogenic and antioxidant. The aim of this study was to evaluate the effects of RSV on Giardia lamblia trophozoites in vitro and in silico, focusing on tubulin affectation, a major protein of the Giardia cytoskeleton which participates in relevant processes for cell survival. In vitro determinations showed that RSV inhibits parasite growth and adherence, causes morphological changes, and induces apoptosis-like cell death through tubulin alterations demonstrated by immunolocalization and Western blot assays. Bioinformatic analysis by molecular docking suggested that RSV binds to Giardia tubulin interface heterodimer, sharing binding residues to those reported with depolymerization inhibitors. These findings suggest that RSV affects microtubular dynamics and make it an interesting compound to study for its safety and antigiardiasic potential.
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Giardia lamblia , Giardiasis , Animales , Humanos , Giardiasis/tratamiento farmacológico , Giardiasis/parasitología , Tubulina (Proteína)/metabolismo , Tubulina (Proteína)/farmacología , Tubulina (Proteína)/uso terapéutico , Resveratrol/farmacología , Trofozoítos , Simulación del Acoplamiento MolecularRESUMEN
CONTEXT: Phytocompounds xanthatin and 8-epi-xanthatin, obtained from Xanthium chinese Mill, showed antitumoral activity in vitro related to the microtubules destabilizing properties of these phytocompounds. Five binding sites for microtubule destabilizing agents have been characterized on tubulin by high-resolution X-ray crystallography: vinca domain, colchicine, pironetin, maytansine site, and more recently, the seventh site. This work aims to develop a comprehensive computational strategy to understand and eventually predict the interaction between xanthatin and 8-epi-xanthatin with the destabilizing-antimitotic binding domain of the tubulin heterodimer. In addition, we propose a putative binding site for these phytocompounds into the microtubule destabilizing binding sites on the tubulin heterodimer. Xanthanolides showed higher stability in the colchicine and pironetin binding sites, whit a greater affinity for the former. In addition, we found that xanthanolides and non-classical colchicine binding site inhibitors share a high structural similarity. METHODS: The 3D structures for xanthatin and 8-epi-xanthatin were obtained using DFT with the hybrid functional B3LYP and the base 6-31G (d,p), implemented in Gaussian 09. The 3D coordinates for tubulin proteins were downloaded from PDB. The complexes tubulin-xanthanolides were predicted using a Monte-Carlo iterated search combined with the BFGS gradient-based optimizer implemented in the AutoDock Vina. The xanthanolides-tubulin complexes were energy minimized by molecular dynamics simulations at vacuum, and their stabilities were evaluated by solvated molecular dynamics simulations during 100 ns. All molecular dynamics simulations were performed using the conjugate gradient method implemented in NAMD2 and CHARMM36 forcefield.
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Antineoplásicos , Colchicina , Colchicina/farmacología , Colchicina/química , Tubulina (Proteína)/metabolismo , Furanos/farmacología , Sitios de Unión , Microtúbulos , Antineoplásicos/farmacologíaRESUMEN
Dengue virus (DENV) infection is the most arbovirosis in the world. However, medications have not been approved for its treatment. Drug discovery based on the host-targeted antiviral (HTA) constitutes a new promising strategy, considering their high genetic barrier to resistance and the low probability of selecting drug resistance strains. In this study, we have tested fifty-seven podophyllotoxin-related cyclolignans on DENV-2 infected cells and found the most promising compound was S.71. Using cellular and molecular biology experiments, we have discovered that the new lignan altered the distribution of microtubules, induced changes in cell morphology, and caused retraction of the rough endoplasmic reticulum. In addition, the compound alters the viral envelope protein and the double-stranded RNA, while there is a decrease in negative-strand RNA synthesis; especially when the compound was added between 6- and 12-hours post-infection. Altogether, S.71 decreases the viral yield through an HTA-related mechanism of action, possibly altering the DENV genome replication and/or polyprotein translation, through the alteration of microtubule distribution and endoplasmic reticulum deterioration. Finally, pharmacokinetic predictors show that S.71 falls within the standard ranges established for drugs.
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Virus del Dengue , Dengue , Virosis , Humanos , Virus del Dengue/genética , Antivirales/uso terapéutico , Replicación Viral , Técnicas de Cultivo de Célula , Virosis/tratamiento farmacológico , Dengue/tratamiento farmacológicoRESUMEN
Despite the advances in understanding the biology of hematologic neoplasms which has resulted in the approval of new drugs, the therapeutic options are still scarce for relapsed/refractory patients. Eribulin is a unique microtubule inhibitor that is currently being used in the therapy for metastatic breast cancer and soft tissue tumors. Here, we uncover eribulin's cellular and molecular effects in a molecularly heterogeneous panel of hematologic neoplasms. Eribulin reduced cell viability and clonogenicity and promoted apoptosis and cell cycle arrest. The minimal effects of eribulin observed in the normal leukocytes suggested selectivity for malignant blood cells. In the molecular scenario, eribulin induces DNA damage and apoptosis markers. The ABCB1, ABCC1, p-AKT, p-NFκB, and NFκB levels were associated with responsiveness to eribulin in blood cancer cells, and a resistance eribulin-related target score was constructed. Combining eribulin with elacridar (a P-glycoprotein inhibitor), but not with PDTC (an NFkB inhibitor), increases eribulin-induced apoptosis in leukemia cells. In conclusion, our data indicate that eribulin leads to mitotic catastrophe and cell death in blood cancer cells. The expression and activation of MDR1, PI3K/AKT, and the NFκB-related targets may be biomarkers of the eribulin response, and the combined treatment of eribulin and elacridar may overcome drug resistance in these diseases.
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Background: Diarrheal diseases caused by protozoa have a great impact on human health around the world. Giardia lamblia is one of the most common flagellates in the intestinal tract. Factors such as adverse effects to first-line drugs or the appearance of drug-resistant strains, make it necessary to identify new treatment alternatives. Agroindustry waste, like pomegranate peel, are a source of phenolic compounds, which possess antiparasitic activities. In vivo studies demonstrated antigiardiasic potential by reducing cyst shedding and protecting intestinal cells; however, they did not identify the compounds or elucidate any mechanism of action in the parasite. The objective of this study is to identify potential molecular targets and to test the in vitro effects of polyphenols from Punica granatum on Giardia lamblia. Methods: The in vitro antigiardial potential of polyphenolic extract from pomegranate peel (Punica granatum L.) obtained using microwave-ultrasound methodology was evaluated on Giardia lamblia trophozoites. Extract phytochemical identification was performed by HPLC/MS analysis. The effect of polyphenolic extract on growth and adhesion capacity was determined by parasite kinetics; morphological damage was evaluated by SEM, alteration on α-tubulin expression and distribution were analyzed by western blot and immunofluorescence, respectively. Results: The pomegranate peel extract showed the presence of ellagitannins (punicalin and punicalagin, galloyl-dihexahydroxydiphenoyl-hexoside), flavones (luteolin), and ellagic acid, that caused an inhibitory effect on growth and adhesion capacity, particularly on cells treated with 200 µg/mL, where growth inhibition of 74.36%, trophozoite adherence inhibition of 46.8% and IC50 of 179 µg/mL at 48 h were demonstrated. The most important findings were that the extract alters α-tubulin expression and distribution in Giardia trophozoites in a concentration-independent manner. Also, an increase in α-tubulin expression at 200 µg/mL was observed in western blot and diffuse or incomplete immunolabeling pattern, especially in ventral disk. In addition, the extract caused elongation, disturbance of normal shape, irregularities in the membrane, and flagella abnormalities. Discussion: The pomegranate peel extract affects Giardia trophozoites in vitro. The damage is related to the cytoskeleton, due to expression and distribution alterations in α-tubulin, particularly in the ventral disk, a primordial structure for adhesion and pathogenesis. Microtubule impairment could explain morphological changes, and inhibition of adhesion capacity and growth. Besides, this is the first report that suggests that ellagic acid, punicalin, punicalagin and luteolin could be interactioning with the rich-tubulin cytoskeleton of Giardia. Further investigations are needed in order to elucidate the mechanisms of action of the isolated compounds and propose a potential drug alternative for the giardiasis treatment.
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Giardia lamblia , Giardiasis , Granada (Fruta) , Animales , Humanos , Granada (Fruta)/metabolismo , Trofozoítos , Tubulina (Proteína)/metabolismo , Ácido Elágico/metabolismo , Luteolina/metabolismo , Microtúbulos/metabolismo , Citoesqueleto , Giardiasis/tratamiento farmacológico , Extractos Vegetales/farmacologíaRESUMEN
Prostate cancer is the second most common malignancy in men and the development of effective therapeutic strategies remains challenging when more advanced, androgen-independent or insensitive forms are involved. Accordingly, we have evaluated, using flow cytometry, confocal microscopy and image analysis, the anti-proliferative effects of (+)-2,3,9-trimethoxypterocarpan [(+)-PTC, 1] on relevant human prostate cancer cells as well as its capacity to control mitosis within them. In particular, the studies reported herein reveal that (+)-PTC exerts anti-proliferative activity against the PC-3â cell lines by regulating cell-cycle progression with mitosis being arrested in the prophase or prometaphase. Furthermore, it emerges that treatment of the target cells with this compound results in the formation of monopolar spindles, disorganized centrosomes and extensively disrupted γ-tubulin distributions while centriole replication remains unaffected. Such effects suggest (+)-PTC should be considered as a possible therapy for androgen-insensitive/independent prostate cancer.
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Microtúbulos , Neoplasias de la Próstata , Andrógenos , Línea Celular , Humanos , Masculino , Mitosis , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/metabolismo , Huso Acromático/metabolismo , Tubulina (Proteína)/metabolismoRESUMEN
The cytoskeleton is the main intracellular structure that determines the morphology of neurons and maintains their integrity. Therefore, disruption of its structure and function may underlie several neurodegenerative diseases. This review summarizes the current literature on the tau protein, microtubule-associated protein 2 (MAP2) and neurofilaments as common denominators in pathological conditions such as Alzheimer's disease (AD), cerebral ischemia, and multiple sclerosis (MS). Insights obtained from experimental models using biochemical and immunocytochemical techniques highlight that changes in these proteins may be potentially used as protein targets in clinical settings, which provides novel opportunities for the detection, monitoring and treatment of patients with these neurodegenerative diseases.
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The ubiquitin-proteasome system (UPS) degrades intracellular proteins through the 26S proteasome. We analysed how cold stress affects the UPS in glial cells. Together with a reduction in the 20S proteolytic activity and increased levels of polyubiquitinated proteins, exposure of glial cell cultures to cold induces a partial disassembly of the 26S proteasome. In particular, we found that Rpt5, a subunit of the 19S proteasome, relocates to cold-stable microtubules, although no apparent cytoskeletal redistribution was detected for other analysed subunits of the 19S or 20S complexes. Furthermore, we demonstrate that both the expression of the microtubule-associated protein MAP6 and the post-translational acetylation of α-tubulin modulate the association of Rpt5 with microtubules. This reversible association could be related to functional preservation of the proteolytic complex during cold stress.
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Complejo de la Endopetidasa Proteasomal , Ubiquitina , Microtúbulos/metabolismo , Neuroglía/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteínas , TemperaturaRESUMEN
The endoplasmic reticulum (ER)-to-Golgi intermediate compartment (ERGIC) is a membranous organelle that mediates protein transport between the ER and the Golgi apparatus. In neurons, clusters of these vesiculotubular structures are situated throughout the cell in proximity to the ER, passing cargo to the cis-Golgi cisternae, located mainly in the perinuclear region. Although ERGIC markers have been identified in neurons, the distribution and dynamics of neuronal ERGIC structures have not been characterized yet. Here, we show that long-distance ERGIC transport occurs via an intermittent mechanism in dendrites, with mobile elements moving between stationary structures. Slow and fast live-cell imaging have captured stable ERGIC structures remaining in place over long periods of time, as well as mobile ERGIC structures advancing very short distances along dendrites. These short distances have been consistent with the lengths between the stationary ERGIC structures. Kymography revealed ERGIC elements that moved intermittently, emerging from and fusing with stationary ERGIC structures. Interestingly, this movement apparently depends not only on the integrity of the microtubule cytoskeleton, as previously reported, but on the actin cytoskeleton as well. Our results indicate that the dendritic ERGIC has a dual nature, with both stationary and mobile structures. The neural ERGIC network transports proteins via a stop-and-go movement in which both the microtubule and the actin cytoskeletons participate.
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Retículo Endoplásmico , Aparato de Golgi , Citoesqueleto de Actina/metabolismo , Retículo Endoplásmico/metabolismo , Aparato de Golgi/metabolismo , Microtúbulos/metabolismo , Transporte de Proteínas/fisiologíaRESUMEN
Neocortex development comprises of a complex series of time- and space-specific processes to generate the typical interconnected six-layered architecture of adult mammals. Axon growth is required for the proper establishment of cortical circuits. Malformations in axonal growth and pathfinding might lead to severe neuropathologies, such as corpus callosum dysgenesis. Cenpj, a microcephaly gene, encodes a scaffold protein that regulates centrosome biogenesis and microtubule stabilization. During corticogenesis, Cenpj regulates progenitor division and neuronal migration. Since microtubule stabilization is crucial for axon extension, we investigated the role of Cenpj in axon growth during cortical development in a mouse model. Through loss- and gain-of-function assays ex vivo and in utero, we quantified callosal axonal length, branching, and growth cone size compared to controls. We observed that silencing Cenpj results in an increased axonal length. Ex vivo, we assessed the number of branches, the area of growth cones and the stability of microtubules. In silenced Cenpj axons, there were more branches, larger growth cone area, and more stable microtubules. Rescue experiments confirmed that neurons present axonal length comparable to controls. Here we propose that Cenpj regulates axon growth by destabilizing microtubules during cortical development. Finally, our findings suggest that Cenpj might be a novel target for axonal regeneration.
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Microcefalia , Proteínas Asociadas a Microtúbulos , Animales , Axones/metabolismo , Células Cultivadas , Conos de Crecimiento/metabolismo , Mamíferos/metabolismo , Ratones , Microcefalia/genética , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Microtúbulos/metabolismo , Neuronas/metabolismoRESUMEN
Microtubule organizing centers (MTOCs) perform critical cellular tasks by nucleating, stabilizing, and anchoring microtubule's minus ends. These capacities impact tremendously a wide array of cellular functions ranging from ascribing cell shape to orchestrating cell division and generating motile structures, among others. The phylum Apicomplexa comprises over 6000 single-celled obligate intracellular parasitic species. Many of the apicomplexan are well known pathogens such as Toxoplasma gondii and the Plasmodium species, causative agents of toxoplasmosis and malaria, respectively. Microtubule organization in these parasites is critical for organizing the cortical cytoskeleton, enabling host cell penetration and the positioning of large organelles, driving cell division and directing the formation of flagella in sexual life stages. Apicomplexans are a prime example of MTOC diversity displaying multiple functional and structural MTOCs combinations within a single species. This diversity can only be fully understood in light of each organism's specific MT nucleation requirements and their evolutionary history. Insight into apicomplexan MTOCs had traditionally been limited to classical ultrastructural work by transmission electron microscopy. However, in the past few years, a large body of molecular insight has emerged. In this work we describe the latest insights into nuclear MTOC biology in two major human and animal disease causing Apicomplexans: Toxoplasma gondii and Plasmodium spp.
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Microtubules (MTs) are important structures of the cytoskeleton in neurons. Mammalian brain MTs act as biomolecular transistors that generate highly synchronous electrical oscillations. However, their role in brain function is largely unknown. To gain insight into the MT electrical oscillatory activity of the brain, we turned to the honeybee (Apis mellifera) as a useful model to isolate brains and MTs. The patch clamp technique was applied to MT sheets of purified honeybee brain MTs. High resistance seal patches showed electrical oscillations that linearly depended on the holding potential between ± 200 mV and had an average conductance in the order of ~9 nS. To place these oscillations in the context of the brain, we also explored local field potential (LFP) recordings from the Triton X-permeabilized whole honeybee brain unmasking spontaneous oscillations after but not before tissue permeabilization. Frequency domain spectral analysis of time records indicated at least two major peaks at approximately ~38 Hz and ~93 Hz in both preparations. The present data provide evidence that MT electrical oscillations are a novel signaling mechanism implicated in brain wave activity observed in the insect brain.
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Ultraviolet (UV) exposure has been linked to skin damage and carcinogenesis, but recently UVB has been proposed as a therapeutic approach for cancer. Herein, we investigated the cellular and molecular effects of UVB in immortal and tumorigenic HPV positive and negative cells. Cells were irradiated with 220.5 to 1102.5 J/m2 of UVB and cell proliferation was evaluated by crystal violet, while cell cycle arrest and apoptosis analysis were performed through flow cytometry. UVB effect on cells was recorded at 661.5 J/m2 and it was exacerbated at 1102.5 J/m2. All cell lines were affected by proliferation inhibition, cell cycle ablation and apoptosis induction, with different degrees depending on tumorigenesis level or HPV type. Analysis of the well-known UV-responsive p53, E2F1 and microtubules system proteins was performed in SiHa cells in response to UVB through Western-blotting assays. E2F1 and the Microtubule-associated protein 2 (MAP2) expression decrease correlated with cellular processes alteration while p53 and Microtubule-associated Protein 1S (MAP1S) expression switch was observed since 882 J/m2, suggesting they were required under more severe cellular damage. However, expression transition of α-Tubulin3C and ß-Tubulin was abruptly noticed until 1102.5 J/m2 and particularly, γ-Tubulin protein expression remained without alteration. This study provides insights into the effect of UVB in cervical cancer cell lines.
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Factor de Transcripción E2F1/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de la radiación , Microtúbulos/efectos de la radiación , Proteína p53 Supresora de Tumor/metabolismo , Rayos Ultravioleta , Neoplasias del Cuello Uterino/patología , Apoptosis , Ciclo Celular , Proliferación Celular , Factor de Transcripción E2F1/genética , Femenino , Humanos , Células Tumorales Cultivadas , Proteína p53 Supresora de Tumor/genética , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/metabolismo , Neoplasias del Cuello Uterino/radioterapiaRESUMEN
Glyphosate, the most commonly used pesticide worldwide, blocks aromatic amino acid biosynthetic pathways and inhibits growth in plants. Although the specific mode of action of glyphosate in animals remains unclear, adverse effects during embryonic development have been reported, including epiboly delays, morphological alterations, and changes in central nervous system development and cardiogenesis. In this study, we suggest a possible toxicity mechanism for this herbicide related to changes in microtubule stability, which could alter the distribution and dynamics of cytoskeleton components. Using zebrafish embryos to evaluate in vivo effects of glyphosate exposure (5, 10, and 50 µg/ml), we found significant reductions in the levels of acetylated α-tubulin (50 µg/ml) and in the polymeric tubulin percentage in zebrafish embryos that had been exposed to 10 and 50 µg/ml glyphosate, without any changes in either the expression patterns of α-tubulin or the stability of actin filaments. These results indicate that high concentrations of glyphosate were associated with reduced levels of acetylated α-tubulin and altered microtubule stability, which may explain some of the neurotoxic and cardiotoxic effects that have been attributed to this herbicide.
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Acute promyelocytic leukemia (APL) is associated with PML-RARα oncogene, which is treated using all-trans retinoic acid (ATRA)-based chemotherapy. However, chemoresistance is observed in 20-30% of treated patients and represents a clinical challenge, raising the importance of the development of new therapeutic options. In the present study, the effects of three synthetic cyclopenta[b]indoles on the leukemia phenotype were investigated using NB4 (ATRA-sensitive) and NB4-R2 (ATRA-resistant) cells. Among the tested synthetic cyclopenta[b]indoles, compound 2, which contains a heterocyclic nucleus, was the most active, presenting time-dependent cytotoxic activity in the µM range in APL cells, without cytotoxicity for normal leukocytes, and was selected for further characterization. Compound 2 significantly decreased clonogenicity, increased apoptosis, and caused cell cycle arrest at S and G2/M phases in a drug concentration-dependent manner. Morphological analyses indicated aberrant mitosis and diffuse tubulin staining upon compound 2 exposure, which corroborates cell cycle findings. In the molecular scenario, compound 2 reduced STMN1 expression and activity, and induced PARP1 cleavage and H2AX and CHK2 phosphorylation, and modulated CDKN1A, PMAIP1, GADD45A, and XRCC3 expressions, indicating reduction of cell proliferation, apoptosis, and DNA damage. Moreover, in the in vivo tubulin polymerization assay, NB4 and NB4-R2 cells showed a reduction in the levels of polymerized tubulin upon compound 2 exposure, which indicates tubulin as a target of the drug. Molecular docking supports this hypothesis. Taken together, these data indicated that compound 2 exhibits antileukemic effects through disrupting the microtubule dynamics, identifying a possible novel potential antineoplastic agent for the treatment of ATRA-resistant APL.