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1.
J Vis Exp ; (167)2021 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-33554972

RESUMO

Despite several advances in cardiac tissue engineering, one of the major challenges to overcome remains the generation of a fully functional vascular network comprising several levels of complexity to provide oxygen and nutrients within bioengineered heart tissues. Our laboratory has developed a three-dimensional in vitro model of the human heart, known as the "cardiac spheroid" or "CS". This presents biochemical, physiological, and pharmacological features typical of the human heart and is generated by co-culturing its three major cell types, such as cardiac myocytes, endothelial cells, and fibroblasts. Human induced pluripotent stem cells-derived cardiomyocytes (hiPSC-CMs or iCMs) are co-cultured at ratios approximating the ones found in vivo with human cardiac fibroblasts (HCFs) and human coronary artery endothelial cells (HCAECs) in hanging drop culture plates for three to four days. The confocal analysis of CSs stained with antibodies against cardiac Troponin T, CD31 and vimentin (markers for cardiac myocytes, endothelial cells and fibroblasts, respectively) shows that CSs present a complex endothelial cell network, resembling the native one found in the human heart. This is confirmed by the 3D rendering analysis of these confocal images. CSs also present extracellular matrix (ECM) proteins typical of the human heart, such as collagen type IV, laminin and fibronectin. Finally, CSs present a contractile activity measured as syncytial contractility closer to the one typical of the human heart compared to CSs that contain iCMs only. When treated with a cardiotoxic anti-cancer agent, such as doxorubicin (DOX, used to treat leukemia, lymphoma and breast cancer), the viability of DOX-treated CSs is significantly reduced at 10 µM genetic and chemical inhibition of endothelial nitric oxide synthase, a downstream target of DOX in HCFs and HCAECs, reduced its toxicity in CSs. Given these unique features, CSs are currently used as in vitro models to study heart biochemistry, pathophysiology, and pharmacology.


Assuntos
Bioengenharia/métodos , Coração/fisiopatologia , Esferoides Celulares/citologia , Animais , Cardiotoxinas/farmacologia , Contagem de Células , Separação Celular , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Cocultura , Colágeno/farmacologia , Doxorrubicina/farmacologia , Doxorrubicina/toxicidade , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Fibroblastos/citologia , Géis , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Miócitos Cardíacos/citologia , Miócitos Cardíacos/efeitos dos fármacos , Ratos , Fixação de Tecidos
2.
Nat Commun ; 12(1): 388, 2021 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-33452260

RESUMO

The practices of synthetic biology are being integrated into 'multiscale' designs enabling two-way communication across organic and inorganic information substrates in biological, digital and cyber-physical system integrations. Novel applications of 'bio-informational' engineering will arise in environmental monitoring, precision agriculture, precision medicine and next-generation biomanufacturing. Potential developments include sentinel plants for environmental monitoring and autonomous bioreactors that respond to biosensor signaling. As bio-informational understanding progresses, both natural and engineered biological systems will need to be reimagined as cyber-physical architectures. We propose that a multiple length scale taxonomy will assist in rationalizing and enabling this transformative development in engineering biology.


Assuntos
Bioengenharia/tendências , Previsões , Biologia Sintética/tendências , Bioengenharia/métodos , Biologia Sintética/métodos
4.
Nat Commun ; 11(1): 4440, 2020 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-32895374

RESUMO

Traditionally engineered genetic circuits have almost exclusively used naturally occurring transcriptional repressors. Recently, non-natural transcription factors (repressors) have been engineered and employed in synthetic biology with great success. However, transcriptional anti-repressors have largely been absent with regard to the regulation of genes in engineered genetic circuits. Here, we present a workflow for engineering systems of non-natural anti-repressors. In this study, we create 41 inducible anti-repressors. This collection of transcription factors respond to two distinct ligands, fructose (anti-FruR) or D-ribose (anti-RbsR); and were complemented by 14 additional engineered anti-repressors that respond to the ligand isopropyl ß-d-1-thiogalactopyranoside (anti-LacI). In turn, we use this collection of anti-repressors and complementary genetic architectures to confer logical control over gene expression. Here, we achieved all NOT oriented logical controls (i.e., NOT, NOR, NAND, and XNOR). The engineered transcription factors and corresponding series, parallel, and series-parallel genetic architectures represent a nascent anti-repressor based transcriptional programming structure.


Assuntos
Bioengenharia/métodos , Repressores Lac/antagonistas & inibidores , Proteínas de Escherichia coli/metabolismo , Expressão Gênica/fisiologia , Redes Reguladoras de Genes , Repressores Lac/síntese química , Ligantes , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/síntese química , Biologia Sintética/métodos , Fatores de Transcrição/síntese química , Fatores de Transcrição/metabolismo
5.
Nat Commun ; 11(1): 3658, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32694613

RESUMO

Biological systems organize multiple hierarchical structures in parallel, and create dynamic assemblies and functions by energy dissipation. In contrast, emerging artificial non-equilibrium self-assembling systems have remained relatively simplistic concerning hierarchical design, and non-equilibrium multi-component systems are uncharted territory. Here we report a modular DNA toolbox allowing to program transient non-equilibrium multicomponent systems across hierarchical length scales by introducing chemically fueled molecular recognition orchestrated by reaction networks of concurrent ATP-powered ligation and cleavage of freely programmable DNA building blocks. Going across hierarchical levels, we demonstrate transient side-chain functionalized nucleic acid polymers, and further introduce the concept of transient cooperative multivalency as a key to bridge length scales to pioneer fuel-driven encapsulation, self-assembly of colloids, and non-equilibrium transient narcissistic colloidal self-sorting on a systems level. The fully programmable and functionalizable DNA components pave the way to design chemically fueled 4D (3 space, 1 time) molecular multicomponent systems and autonomous materials.


Assuntos
Trifosfato de Adenosina/química , Bioengenharia/métodos , DNA/química , Nanotecnologia/métodos , Coloides , DNA Ligases/química , Desoxirribonucleases de Sítio Específico do Tipo II/química , Conformação de Ácido Nucleico , Polimerização , Polímeros/química
6.
Theranostics ; 10(16): 7034-7052, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32641977

RESUMO

This review provides an update for the international research community on the cell modeling tools that could accelerate the understanding of SARS-CoV-2 infection mechanisms and could thus speed up the development of vaccines and therapeutic agents against COVID-19. Many bioengineering groups are actively developing frontier tools that are capable of providing realistic three-dimensional (3D) models for biological research, including cell culture scaffolds, microfluidic chambers for the culture of tissue equivalents and organoids, and implantable windows for intravital imaging. Here, we review the most innovative study models based on these bioengineering tools in the context of virology and vaccinology. To make it easier for scientists working on SARS-CoV-2 to identify and apply specific tools, we discuss how they could accelerate the discovery and preclinical development of antiviral drugs and vaccines, compared to conventional models.


Assuntos
Antivirais/isolamento & purificação , Antivirais/farmacologia , Betacoronavirus , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/prevenção & controle , Pandemias/prevenção & controle , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/prevenção & controle , Vacinas Virais/isolamento & purificação , Vacinas Virais/farmacologia , Betacoronavirus/química , Betacoronavirus/genética , Betacoronavirus/imunologia , Bioengenharia/métodos , Bioengenharia/tendências , Reatores Biológicos , Técnicas de Cultura de Células , Simulação por Computador , Infecções por Coronavirus/imunologia , Descoberta de Drogas/métodos , Descoberta de Drogas/tendências , Avaliação de Medicamentos/métodos , Avaliação de Medicamentos/tendências , Farmacorresistência Viral , Interações entre Hospedeiro e Microrganismos/genética , Interações entre Hospedeiro e Microrganismos/imunologia , Humanos , Modelos Biológicos , Organoides/citologia , Organoides/virologia , Pneumonia Viral/imunologia , Nanomedicina Teranóstica
8.
Nat Commun ; 11(1): 2105, 2020 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-32355158

RESUMO

3D-printing networks of droplets connected by interface bilayers are a powerful platform to build synthetic tissues in which functionality relies on precisely ordered structures. However, the structural precision and consistency in assembling these structures is currently limited, which restricts intricate designs and the complexity of functions performed by synthetic tissues. Here, we report that the equilibrium contact angle (θDIB) between a pair of droplets is a key parameter that dictates the tessellation and precise positioning of hundreds of picolitre-sized droplets within 3D-printed, multi-layer networks. When θDIB approximates the geometrically-derived critical angle (θc) of 35.3°, the resulting networks of droplets arrange in regular hexagonal close-packed (hcp) lattices with the least fraction of defects. With this improved control over droplet packing, we can 3D-print functional synthetic tissues with single-droplet-wide conductive pathways. Our new insights into 3D droplet packing permit the fabrication of complex synthetic tissues, where precisely positioned compartments perform coordinated tasks.


Assuntos
Bioengenharia/instrumentação , Bicamadas Lipídicas/química , Impressão Tridimensional , Bioengenharia/métodos , Materiais Biomiméticos/química , Cinética , Lipídeos/química , Microscopia Confocal , Temperatura , Água/química
9.
Nat Commun ; 11(1): 2183, 2020 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-32366821

RESUMO

Coupling myoelectric and mechanical signals during voluntary muscle contraction is paramount in human-machine interactions. Spatiotemporal differences in the two signals intrinsically arise from the muscular excitation-contraction process; however, current methods fail to deliver local electromechanical coupling of the process. Here we present the locally coupled electromechanical interface based on a quadra-layered ionotronic hybrid (named as CoupOn) that mimics the transmembrane cytoadhesion architecture. CoupOn simultaneously monitors mechanical strains with a gauge factor of ~34 and surface electromyogram with a signal-to-noise ratio of 32.2 dB. The resolved excitation-contraction signatures of forearm flexor muscles can recognize flexions of different fingers, hand grips of varying strength, and nervous and metabolic muscle fatigue. The orthogonal correlation of hand grip strength with speed is further exploited to manipulate robotic hands for recapitulating corresponding gesture dynamics. It can be envisioned that such locally coupled electromechanical interfaces would endow cyber-human interactions with unprecedented robustness and dexterity.


Assuntos
Eletromiografia/métodos , Força da Mão/fisiologia , Contração Muscular/fisiologia , Músculo Esquelético/fisiologia , Amplitude de Movimento Articular/fisiologia , Membros Artificiais , Bioengenharia/instrumentação , Bioengenharia/métodos , Fenômenos Biomecânicos , Eletrônica Médica/instrumentação , Eletrônica Médica/métodos , Dedos/fisiologia , Antebraço/fisiologia , Mãos/fisiologia , Humanos , Desenho de Prótese/instrumentação , Desenho de Prótese/métodos
11.
Med Sci (Paris) ; 36(4): 382-388, 2020 Apr.
Artigo em Francês | MEDLINE | ID: mdl-32356715

RESUMO

As burden of chronic respiratory diseases is constantly increasing, improving in vitro lung models is essential in order to reproduce as closely as possible the complex pulmonary architecture, responsible for oxygen uptake and carbon dioxide clearance. The study of diseases that affect the respiratory system has benefited from in vitro reconstructions of the respiratory epithelium with inserts in air/liquid interface (2D) or in organoids able to mimic up to the arborescence of the respiratory tree (3D). Recent development in the fields of pluripotent stem cells-derived organoids and genome editing technologies has provided new insights to better understand pulmonary diseases and to find new therapeutic perspectives.


Assuntos
Técnicas de Cultura de Células , Pulmão/citologia , Organoides/citologia , Células Epiteliais Alveolares/citologia , Células Epiteliais Alveolares/fisiologia , Animais , Bioengenharia/métodos , Bioengenharia/tendências , Dióxido de Carbono/farmacologia , Dióxido de Carbono/fisiologia , Técnicas de Cultura de Células/métodos , Técnicas de Cultura de Células/tendências , Células Cultivadas , Edição de Genes/métodos , Edição de Genes/tendências , Humanos , Pulmão/patologia , Pulmão/fisiologia , Modelos Biológicos , Organoides/patologia , Organoides/fisiologia , Oxigênio/farmacologia , Oxigênio/fisiologia , Troca Gasosa Pulmonar/fisiologia , Mucosa Respiratória/citologia , Mucosa Respiratória/efeitos dos fármacos , Tecidos Suporte/química
12.
Med Sci (Paris) ; 36(3): 261-263, 2020 Mar.
Artigo em Francês | MEDLINE | ID: mdl-32228845

RESUMO

Organoids offer an elegant approach to model human diseases and test new drugs. Nonalcoholic fatty liver disease (NAFLD) whose incidence has dramatically increased in recent years with the rise of obesity, is defined by triglyceride accumulation in hepatocytes, inflammation, liver injury, and progression to fibrosis. There is currently no approved therapy but many pathways are being explored. Two American teams have created mini-steatotic livers using different approaches, both using induced pluripotent stem cells (iPS), thus offering new tools to test developing drugs.


Assuntos
Bioengenharia/tendências , Ciência de Laboratório Médico/tendências , Hepatopatia Gordurosa não Alcoólica/patologia , Técnicas de Cultura de Tecidos/tendências , Animais , Bioengenharia/métodos , Células Cultivadas , Progressão da Doença , Hepatócitos/citologia , Hepatócitos/patologia , Hepatócitos/fisiologia , Humanos , Fígado/patologia , Fígado/fisiologia , Cirrose Hepática/patologia , Ciência de Laboratório Médico/métodos , Modelos Biológicos , Técnicas de Cultura de Tecidos/métodos , Tecidos Suporte
13.
Plant Sci ; 294: 110445, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32234228

RESUMO

Phosphate acquisition by plants is an essential process that is directly implicated in the optimization of crop yields. Purple acid phosphatases (PAPs) are ubiquitous metalloenzymes, which catalyze the hydrolysis of a wide range of phosphate esters and anhydrides. While some plant PAPs display a preference for ATP as the substrate, others are efficient in hydrolyzing phytate or 2-phosphoenolpyruvate (PEP). PAP from red kidney bean (rkbPAP) is an efficient ATP- and ADPase, but has no activity towards phytate. Crystal structures of this enzyme in complex with ATP analogues (to 2.20 and 2.60 Å resolution, respectively) complement the recent structure of rkbPAP with a bound ADP analogue (ChemBioChem 20 (2019) 1536). Together these complexes provide the first structural insight of a PAP in complex with molecules that mimic biologically relevant substrates. Homology modeling was used to generate three-dimensional structures for the active sites of PAPs from tobacco (NtPAP) and thale cress (AtPAP26) that are efficient in hydrolyzing phytate and PEP as preferred substrates, respectively. The combining of crystallographic data, substrate docking simulations and a phylogenetic analysis of 49 plant PAP sequences (including the first PAP sequences reported from Eucalyptus) resulted in the identification of several active site residues that are important in defining the substrate specificities of plant PAPs; of particular relevance is the identification of a motif ("REKA") that is characteristic for plant PAPs that possess phytase activity. These results may inform bioengineering studies aimed at identifying and incorporating suitable plant PAP genes into crops to improve phosphorus acquisition and use efficiency. Organic phosphorus sources increasingly supplement or replace inorganic fertilizer, and efficient phosphorus use of crops will lower the environmental footprint of agriculture while enhancing food production.


Assuntos
Fosfatase Ácida/metabolismo , Fosfatase Ácida/genética , Bioengenharia/métodos , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Phaseolus/genética , Phaseolus/metabolismo , Especificidade por Substrato
15.
Sci Adv ; 6(12): eaaz0330, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32206719

RESUMO

Nanobiohybrids, synthesized by integrating functional nanomaterials with living systems, have emerged as an exciting branch of research at the interface of materials engineering and biological science. Nanobiohybrids use synthetic nanomaterials to impart organisms with emergent properties outside their scope of evolution. Consequently, they endow new or augmented properties that are either innate or exogenous, such as enhanced tolerance against stress, programmed metabolism and proliferation, artificial photosynthesis, or conductivity. Advances in new materials design and processing technologies made it possible to tailor the physicochemical properties of the nanomaterials coupled with the biological systems. To date, many different types of nanomaterials have been integrated with various biological systems from simple biomolecules to complex multicellular organisms. Here, we provide a critical overview of recent developments of nanobiohybrids that enable new or augmented biological functions that show promise in high-tech applications across many disciplines, including energy harvesting, biocatalysis, biosensing, medicine, and robotics.


Assuntos
Materiais Biocompatíveis/química , Bioengenharia , Nanoestruturas/química , Biocatálise , Materiais Biocompatíveis/síntese química , Bioengenharia/métodos , Sobrevivência Celular , Técnicas de Química Sintética , Nanotecnologia , Tecidos Suporte
16.
Int J Mol Sci ; 21(6)2020 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-32178385

RESUMO

Heart disease is the most common cause of death in developed countries, but the medical treatments for heart failure remain limited. In this context, the development of cardiac regeneration therapy for severe heart failure is important. Owing to their unique characteristics, including multiple differentiation and infinitive self-renewal, pluripotent stem cells can be considered as a novel source for regenerative medicine. Janus kinase/signal transducer and activator of transcription 3 (JAK/STAT3) signaling plays critical roles in the induction, maintenance, and differentiation of pluripotent stem cells. In the heart, JAK/STAT3 signaling has diverse cellular functions, including myocardial differentiation, cell cycle re-entry of matured myocyte after injury, and anti-apoptosis in pathological conditions. Therefore, regulating STAT3 activity has great potential as a strategy of cardiac regeneration therapy. In this review, we summarize the current understanding of STAT3, focusing on stem cell biology and pathophysiology, as they contribute to cardiac regeneration therapy. We also introduce a recently reported therapeutic strategy for myocardial regeneration that uses engineered artificial receptors that trigger endogenous STAT3 signal activation.


Assuntos
Coração/fisiopatologia , Células-Tronco Pluripotentes/metabolismo , Células-Tronco Pluripotentes/fisiologia , Regeneração/fisiologia , Medicina Regenerativa/métodos , Fator de Transcrição STAT3/metabolismo , Animais , Bioengenharia/métodos , Diferenciação Celular/fisiologia , Humanos , Miocárdio/metabolismo
17.
NPJ Syst Biol Appl ; 6(1): 6, 2020 03 13.
Artigo em Inglês | MEDLINE | ID: mdl-32170148

RESUMO

In biotechnology, the emergence of high-throughput technologies challenges the interpretation of large datasets. One way to identify meaningful outcomes impacting process and product attributes from large datasets is using systems biology tools such as metabolic models. However, these tools are still not fully exploited for this purpose in industrial context due to gaps in our knowledge and technical limitations. In this paper, key aspects restraining the routine implementation of these tools are highlighted in three research fields: monitoring, network science and hybrid modeling. Advances in these fields could expand the current state of systems biology applications in biopharmaceutical industry to address existing challenges in bioprocess development and improvement.


Assuntos
Bioengenharia/métodos , Produtos Biológicos/metabolismo , Biologia de Sistemas/métodos , Produtos Biológicos/farmacologia , Biotecnologia/métodos , Biotecnologia/tendências , Indústrias/tendências , Modelos Biológicos
18.
Occup Med (Lond) ; 70(4): 282-285, 2020 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-32009172

RESUMO

Protein contact dermatitis (PCD) often presents as chronic hand eczema (CHE) with an immediate hypersensitivity to protein proved by a positive skin prick test or by the presence of specific immunoglobulin E. This is frequently induced by occupational exposure to proteins in food workers, farmers, animal breeders, veterinarians and healthcare professionals. While skin barrier impairment is crucial in the pathogenesis of PCD, methods to assess skin barrier function such as trans-epidermal water loss and stratum corneum hydration are not widely used in clinical settings. We describe the diagnostic workup of occupational PCD due to Argentinean shrimps and discuss how the use of skin bioengineering methods including assessment of corneocytes morphology by Scanning Electron Microscopy provides with insightful information on skin barrier function. Diagnosis of PCD is time-consuming and a multidisciplinary team contributes to early diagnosis and proper occupational rehabilitation.


Assuntos
Bioengenharia/métodos , Dermatite Alérgica de Contato/diagnóstico , Dermatite Ocupacional/diagnóstico , Exposição Ocupacional/análise , Testes Cutâneos/métodos , Adulto , Humanos , Masculino , Exposição Ocupacional/efeitos adversos , Pele/imunologia
19.
Sci Rep ; 10(1): 1581, 2020 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-32005953

RESUMO

Predicting the effects of invasive ecosystem engineering species in new bioregions has proved elusive. In part this is because separating biological effects from purely physical mechanisms has been little studied and yet could help predict potentially damaging bioinvasions. Here we tested the effects of a large bio-engineering fanworm Sabella spallanzanii (Sabella) versus worm-like structures (mimics) on gas and nutrient fluxes in a marine soft bottom sediment. Experimental plots of sediment in Hauraki Gulf (New Zealand) were used to test the hypothesis that ecosystem engineers negatively influence benthic ecosystem function through autogenic mechanisms, facilitating activity by biofouling organisms and competitive exclusion of native infauna. Enhanced physical structure associated with Sabella and mimics increased nitrogen fluxes, community metabolism and reduced denitrification from 23 µmol m-2 h-1 to zero at densities greater than 25 m2. Sabella plots on average had greater respiration (29%), NH4 release (33%), and greater NO3 release (52%) compared to mimics, suggesting allogenic (biological) mechanisms occur, but play a secondary role to autogenic (physical) mechanisms. The dominance of autogenic mechanisms indicates that bio-engineers are likely to cause significant impacts when established, regardless of fundamental differences in recipient regions or identity of the introduced bio-engineer. In the case of Sabella spallanzanii, compromised denitrification has the potential to tip the balance of net solute and gas exchanges and cause further ecological degradation in an already eutrophic system.


Assuntos
Biodegradação Ambiental , Bioengenharia/métodos , Incrustação Biológica , Ecossistema , Espécies Introduzidas , Nitrogênio/metabolismo , Animais , Organismos Aquáticos/metabolismo , Incrustação Biológica/prevenção & controle , Sedimentos Geológicos , Oceanos e Mares , Poliquetos/metabolismo
20.
Int J Mol Sci ; 21(3)2020 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-32024292

RESUMO

A large proportion of the recombinant proteins manufactured today rely on microbe-based expression systems owing to their relatively simple and cost-effective production schemes. However, several issues in microbial protein expression, including formation of insoluble aggregates, low protein yield, and cell death are still highly recursive and tricky to optimize. These obstacles are usually rooted in the metabolic capacity of the expression host, limitation of cellular translational machineries, or genetic instability. To this end, several microbial strains having precisely designed genomes have been suggested as a way around the recurrent problems in recombinant protein expression. Already, a growing number of prokaryotic chassis strains have been genome-streamlined to attain superior cellular fitness, recombinant protein yield, and stability of the exogenous expression pathways. In this review, we outline challenges associated with heterologous protein expression, some examples of microbial chassis engineered for the production of recombinant proteins, and emerging tools to optimize the expression of heterologous proteins. In particular, we discuss the synthetic biology approaches to design and build and test genome-reduced microbial chassis that carry desirable characteristics for heterologous protein expression.


Assuntos
Bactérias/metabolismo , Bioengenharia/métodos , Biotecnologia/métodos , Regulação da Expressão Gênica , Engenharia Metabólica/métodos , Proteínas Recombinantes/biossíntese , Animais , Bactérias/genética , Bactérias/crescimento & desenvolvimento , Humanos , Proteínas Recombinantes/genética
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