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1.
Anal Chim Acta ; 1178: 338803, 2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34482878

RESUMO

The use of biologics in the therapeutic landscape has increased exponentially since the last 3 decades. Nevertheless, patients with central nervous system (CNS) related disorders could not yet benefit from this revolution because the blood-brain barrier (BBB) severely hampers biologics from entering the brain. Considerable effort has been put into generating methods to modulate or circumvent the BBB for delivery of therapeutics to the CNS. A promising strategy is receptor-mediated transcytosis (RMT). Recently, Wouters et al. (2020) discovered a mouse anti-transferrin receptor nanobody that is able to deliver a biologically active peptide to the brain via RMT. The present study aims to sample a derivative of this brain-penetrating nanobody (Nb105) in the CNS. Therefore, we compared the applicability of cerebral open flow microperfusion (cOFM) and microdialysis as sampling techniques to directly obtain high molecular weight substances from the cerebral interstitial fluid. A custom AlphaScreen™ assay was validated to quantify nanobody concentrations in the samples. In vitro microdialysis probe (AtmosLM™, 1 MDa cut-off) recovery by gain and by loss for Nb105 was 18.3 ± 3.2% and 27.0 ± 2.5% respectively, whereas for cOFM it was 87.2 ± 4.0% and 97.3 ± 1.6%. Although a large difference in in vitro recovery is observed between cOFM and microdialysis, in vivo similar results were obtained. Immunohistochemical stainings showed an astrocytic and microglial reaction in the immediate vicinity along the implantation track for both probe types. Coronal sections showed higher fluorescein isothiocyanate-dextran and immunoglobulin G extravasation around the microdialysis probe track than after cOFM sampling experiments, however this leakage was clearly limited compared to a positive control where the BBB was disrupted. This is the first study that samples a bispecific nanobody in the brain's interstitial fluid in function of time, providing a pharmacokinetic profile of nanobodies in the CNS. Furthermore, this is the first time a cOFM study is performed in awake freely moving mice, providing data on inflammation and blood-brain barrier integrity in the mouse brain. Overall, this work demonstrates that, while taking into account the (bio)analytical considerations, both microdialysis and cOFM are suitable in vivo sampling techniques for quantification of nanobodies in the CNS.


Assuntos
Barreira Hematoencefálica , Encéfalo , Animais , Transporte Biológico , Líquido Extracelular , Humanos , Camundongos , Microdiálise
2.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 38(9): 907-911, 2021 Sep 10.
Artigo em Chinês | MEDLINE | ID: mdl-34487542

RESUMO

ABCC1 gene is expressed in various tissues and organs of the human body, and can transport substrates including drugs, heavy metals, toxic substances and organic anions. Previous research on ABCC1 gene has mostly focused on tumor multidrug resistance. Recently, ABCC1 has been proposed as a candidate gene for hereditary hearing impairment, which has attracted much attention. ABCC1-associated deafness may be related to its role in biological barriers. This article has summarized recent progress in the study of the role of ABCC1 in the blood-testis barrier, placental barrier, blood-brain barrier, blood-labyrinth barrier, which may provide insight into its biological functions.


Assuntos
Surdez , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Placenta , Transporte Biológico , Surdez/genética , Feminino , Humanos , Masculino , Gravidez
3.
Nat Commun ; 12(1): 4662, 2021 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-34341345

RESUMO

Impaired cellular cholesterol efflux is a key factor in the progression of renal, cardiovascular, and autoimmune diseases. Here we describe a class of 5-arylnicotinamide compounds, identified through phenotypic drug discovery, that upregulate ABCA1-dependent cholesterol efflux by targeting Oxysterol Binding Protein Like 7 (OSBPL7). OSBPL7 was identified as the molecular target of these compounds through a chemical biology approach, employing a photoactivatable 5-arylnicotinamide derivative in a cellular cross-linking/immunoprecipitation assay. Further evaluation of two compounds (Cpd A and Cpd G) showed that they induced ABCA1 and cholesterol efflux from podocytes in vitro and normalized proteinuria and prevented renal function decline in mouse models of proteinuric kidney disease: Adriamycin-induced nephropathy and Alport Syndrome. In conclusion, we show that small molecule drugs targeting OSBPL7 reveal an alternative mechanism to upregulate ABCA1, and may represent a promising new therapeutic strategy for the treatment of renal diseases and other disorders of cellular cholesterol homeostasis.


Assuntos
Transportador 1 de Cassete de Ligação de ATP/metabolismo , Colesterol/metabolismo , Nefropatias Diabéticas/metabolismo , Compostos Orgânicos/farmacologia , Podócitos/metabolismo , Proteinúria/metabolismo , Receptores de Esteroides/antagonistas & inibidores , Transportador 1 de Cassete de Ligação de ATP/genética , Animais , Transporte Biológico/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Células HEK293 , Humanos , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Camundongos da Linhagem 129 , Camundongos Knockout , Estrutura Molecular , Niacinamida/química , Niacinamida/farmacologia , Compostos Orgânicos/síntese química , Compostos Orgânicos/química , Podócitos/citologia , Interferência de RNA , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Células THP-1
4.
Int J Mol Sci ; 22(15)2021 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-34360593

RESUMO

Water transport in epithelia occurs transcellularly (aquaporins) and paracellularly (claudin-2, claudin-15). Recently, we showed that downregulated tricellulin, a protein of the tricellular tight junction (tTJ, the site where three epithelial cells meet), increased transepithelial water flux. We now check the hypothesis that another tTJ-associated protein, angulin-1 (alias lipolysis-stimulated lipoprotein receptor, LSR) is a direct negative actuator of tTJ water permeability depending on the tightness of the epithelium. For this, a tight and an intermediate-tight epithelial cell line, MDCK C7 and HT-29/B6, were stably transfected with CRISPR/Cas9 and single-guide RNA targeting angulin-1 and morphologically and functionally characterized. Water flux induced by an osmotic gradient using 4-kDa dextran caused water flux to increase in angulin-1 KO clones in MDCK C7 cells, but not in HT-29/B6 cells. In addition, we found that water permeability in HT-29/B6 cells was not modified after either angulin-1 knockout or tricellulin knockdown, which may be related to the presence of other pathways, which reduce the impact of the tTJ pathway. In conclusion, modulation of the tTJ by knockout or knockdown of tTJ proteins affects ion and macromolecule permeability in tight and intermediate-tight epithelial cell lines, while the transepithelial water permeability was affected only in tight cell lines.


Assuntos
Células Epiteliais/metabolismo , Receptores de Lipoproteínas/metabolismo , Junções Íntimas/metabolismo , Fatores de Transcrição/metabolismo , Água/metabolismo , Animais , Transporte Biológico , Cães , Células Epiteliais/citologia , Células HT29 , Humanos , Células Madin Darby de Rim Canino , Receptores de Lipoproteínas/genética , Fatores de Transcrição/genética
5.
Sheng Li Xue Bao ; 73(4): 681-689, 2021 Aug 25.
Artigo em Chinês | MEDLINE | ID: mdl-34405223

RESUMO

Prostaglandin E2 (PGE2), a bioactive lipid mediator, is one of the most important locally acting factors involved in a variety of physiological and pathophysiological processes. PGE2 binds with four EP receptors (EP1-4) to activate G protein-coupled receptor signaling responses. Recent functional and molecular studies have revealed that PGE2 plays an essential role in regulation of renal fluid transport via a variety of mechanisms. The water balance mainly depends on the regulation of aquaporin-2 (AQP2) by arginine vasopressin (AVP) in renal collecting duct principal cells. In recent years, increasing evidence suggests that PGE2 plays an important role in renal water reabsorption in the collecting ducts. In this paper, we reviewed the role of PGE2 and its receptors in the regulation of water reabsorption in the kidney, which may provide a new therapeutic strategy for many diseases especially nephrogenic diabetes insipidus.


Assuntos
Aquaporina 2 , Diabetes Insípido Nefrogênico , Aquaporina 2/genética , Aquaporina 2/metabolismo , Transporte Biológico , Dinoprostona , Humanos , Água/metabolismo
6.
FASEB J ; 35(9): e21810, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34390520

RESUMO

Copper (Cu) is an essential micronutrient required for the activity of redox-active enzymes involved in critical metabolic reactions, signaling pathways, and biological functions. Transporters and chaperones control Cu ion levels and bioavailability to ensure proper subcellular and systemic Cu distribution. Intensive research has focused on understanding how mammalian cells maintain Cu homeostasis, and how molecular signals coordinate Cu acquisition and storage within organs. In humans, mutations of genes that regulate Cu homeostasis or facilitate interactions with Cu ions lead to numerous pathologic conditions. Malfunctions of the Cu+ -transporting ATPases ATP7A and ATP7B cause Menkes disease and Wilson disease, respectively. Additionally, defects in the mitochondrial and cellular distributions and homeostasis of Cu lead to severe neurodegenerative conditions, mitochondrial myopathies, and metabolic diseases. Cu has a dual nature in carcinogenesis as a promotor of tumor growth and an inducer of redox stress in cancer cells. Cu also plays role in cancer treatment as a component of drugs and a regulator of drug sensitivity and uptake. In this review, we provide an overview of the current knowledge of Cu metabolism and transport and its relation to various human pathologies.


Assuntos
Cobre/metabolismo , Homeostase/fisiologia , Animais , Transporte Biológico/fisiologia , ATPases Transportadoras de Cobre/metabolismo , Humanos , Doenças Metabólicas/metabolismo , Doenças Mitocondriais/metabolismo , Doenças Neurodegenerativas/metabolismo
7.
Nat Commun ; 12(1): 4988, 2021 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-34404781

RESUMO

Glycans are fundamental cellular building blocks, involved in many organismal functions. Advances in glycomics are elucidating the essential roles of glycans. Still, it remains challenging to properly analyze large glycomics datasets, since the abundance of each glycan is dependent on many other glycans that share many intermediate biosynthetic steps. Furthermore, the overlap of measured glycans can be low across samples. We address these challenges with GlyCompare, a glycomic data analysis approach that accounts for shared biosynthetic steps for all measured glycans to correct for sparsity and non-independence in glycomics, which enables direct comparison of different glycoprofiles and increases statistical power. Using GlyCompare, we study diverse N-glycan profiles from glycoengineered erythropoietin. We obtain biologically meaningful clustering of mutant cell glycoprofiles and identify knockout-specific effects of fucosyltransferase mutants on tetra-antennary structures. We further analyze human milk oligosaccharide profiles and find mother's fucosyltransferase-dependent secretor-status indirectly impact the sialylation. Finally, we apply our method on mucin-type O-glycans, gangliosides, and site-specific compositional glycosylation data to reveal tissues and disease-specific glycan presentations. Our substructure-oriented approach will enable researchers to take full advantage of the growing power and size of glycomics data.


Assuntos
Vias Biossintéticas , Glicômica , Polissacarídeos/biossíntese , Transporte Biológico , Vias Biossintéticas/genética , Análise por Conglomerados , Análise de Dados , Eritropoetina/metabolismo , Fucosiltransferases/genética , Gangliosídeos , Técnicas de Inativação de Genes , Glicosilação , Humanos , Mucinas
8.
Molecules ; 26(16)2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34443333

RESUMO

The latest data link the chronic consumption of large amounts of fructose present in food with the generation of hypertension and disturbances in carbohydrate and lipid metabolism, which promote the development of obesity, non-alcoholic fatty liver disease, insulin resistance, and type 2 diabetes. This effect is possible after fructose is absorbed by the small intestine cells and, to a lesser extent, by hepatocytes. Fructose transport is dependent on proteins from the family of glucose transporters (GLUTs), among which GLUT5 selectively absorbs fructose from the intestine. In this study, we examined the effect of four phenolic-rich extracts obtained from A. graveolens, B. juncea, and M. chamomilla on fructose uptake by Caco-2 cells. Extracts from B. juncea and M. chamomilla most effectively reduced fluorescent fructose analogue (NBDF) accumulation in Caco-2, as well as downregulated GLUT5 protein levels. These preparations were able to decrease the mRNA level of genes encoding transcription factors regulating GLUT5 expression-thioredoxin-interacting protein (TXNIP) and carbohydrate-responsive element-binding protein (ChREBP). Active extracts contained large amounts of apigenin and flavonols. The molecular docking simulation suggested that some of identified phenolic constituents can play an important role in the inhibition of GLUT5-mediated fructose transport.


Assuntos
Dieta , Frutose/metabolismo , Transportador de Glucose Tipo 5/metabolismo , Fenóis/análise , Extratos Vegetais/farmacologia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Transporte Biológico/efeitos dos fármacos , Células CACO-2 , Proteínas de Transporte/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , RNA Mensageiro/genética
9.
ACS Appl Mater Interfaces ; 13(33): 39112-39125, 2021 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-34384220

RESUMO

Autophagy inhibition could hinder the underlying protective mechanisms in the course of tumor treatment. The advances in autophagy inhibition have driven focus on the functionalized nanoplatforms by combining the current treatment paradigms with complementary autophagy inhibition for enhanced efficacy. Furthermore, Ca2+ overload is also a promising adjuvant target for the tumor treatment by augmenting mitochondrial damage. In this view, complementary mitochondrial Ca2+ overload and autophagy inhibition were first demonstrated as a novel strategy suitable for homing in on the shortage of photodynamic therapy (PDT). We constructed biodegradable tumor-targeted inorganic/organic hybrid nanocomposites (DPGC/OI) synchronously encapsulating IR780 and Obatoclax by biomineralization of the nanofilm method, which consists of pH-triggered calcium phosphate (CP), long circulation phospholipid block copolymers 1,2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE)-poly(ethylene glycol) (PEG)2000-glucose (DPG). In the presence of the hydrophilic PEG chain and glucose transporter 1 (Glut-1) ligands, DPGC would become an effectively tumor-oriented nanoplatform. Subsequently, IR780 as an outstanding photosensitizer could produce increased amounts of toxic reactive oxygen species (ROS) after laser irradiation. Calcium phosphate (CP) as the Ca2+ nanogenerator could generate Ca2+ at low pH to induce mitochondrial Ca2+ overload. The dysfunction of mitochondria could enhance increased amounts of ROS. Based on the premise that autophagy would degrade dysfunctional organelles to sustain metabolism and homeostasis, which might participate in resistance to PDT, Obatoclax as an autophagy inhibitor would hinder the protective mechanism from cancer cells with negligible toxicity. Such an enhanced PDT via mitochondrial Ca2+ overload and autophagy inhibition could be realized by DPGC/OI.


Assuntos
Autofagia/efeitos dos fármacos , Fosfatos de Cálcio/química , Glucose/química , Indóis/química , Nanocompostos/química , Fosfatidiletanolaminas/química , Fármacos Fotossensibilizantes/química , Polietilenoglicóis/química , Animais , Transporte Biológico , Melhoramento Biomédico , Feminino , Humanos , Indóis/metabolismo , Indóis/farmacologia , Camundongos Endogâmicos BALB C , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Neoplasias/diagnóstico por imagem , Neoplasias/terapia , Fosfolipídeos/química , Fotoquimioterapia , Fármacos Fotossensibilizantes/farmacologia , Pirróis/química , Pirróis/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Propriedades de Superfície , Distribuição Tecidual
10.
Nat Commun ; 12(1): 5042, 2021 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-34413297

RESUMO

Food production must increase significantly to sustain a growing global population. Reducing plant water loss may help achieve this goal and is especially relevant in a time of climate change. The plant cuticle defends leaves against drought, and so understanding water movement through the cuticle could help future proof our crops and better understand native ecology. Here, via mathematical modelling, we identify mechanistic properties of water movement in cuticles. We model water sorption in astomatous isolated cuticles, utilising three separate pathways of cellulose, aqueous pores and lipophilic. The model compares well to data both over time and humidity gradients. Sensitivity analysis shows that the grouping of parameters influencing plant species variations has the largest effect on sorption, those influencing cellulose are very influential, and aqueous pores less so but still relevant. Cellulose plays a significant role in diffusion and adsorption in the cuticle and the cuticle surfaces.


Assuntos
Celulose/metabolismo , Plantas/metabolismo , Água/metabolismo , Adsorção , Transporte Biológico , Difusão , Secas , Umidade , Modelos Biológicos , Permeabilidade , Epiderme Vegetal/metabolismo , Folhas de Planta/metabolismo
11.
Int J Mol Sci ; 22(16)2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34445115

RESUMO

The SWEET (Sugars Will Eventually be Exported Transporter) proteins are a novel family of sugar transporters that play key roles in sugar efflux, signal transduction, plant growth and development, plant-pathogen interactions, and stress tolerance. In this study, 22 ClaSWEET genes were identified in Citrullus lanatus (Thunb.) through homology searches and classified into four groups by phylogenetic analysis. The genes with similar structures, conserved domains, and motifs were clustered into the same groups. Further analysis of the gene promoter regions uncovered various growth, development, and biotic and abiotic stress responsive cis-regulatory elements. Tissue-specific analysis showed most of the genes were highly expressed in male flowers and the roots of cultivated varieties and wild cultivars. In addition, qRT-PCR results further imply that ClaSWEET proteins might be involved in resistance to Fusarium oxysporum infection. Moreover, a significantly higher expression level of these genes under various abiotic stresses suggests its multifaceted role in mediating plant responses to drought, salt, and low-temperature stress. The genome-wide characterization and phylogenetic analysis of ClaSWEET genes, together with the expression patterns in different tissues and stimuli, lays a solid foundation for future research into their molecular function in watermelon developmental processes and responses to biotic and abiotic stresses.


Assuntos
Transporte Biológico/genética , Citrullus/genética , Genoma de Planta/genética , Família Multigênica/genética , Proteínas de Plantas/genética , Estresse Fisiológico/genética , Açúcares/metabolismo , Fusarium/genética , Regulação da Expressão Gênica de Plantas/genética , Estudo de Associação Genômica Ampla/métodos , Filogenia , Raízes de Plantas/genética , Regiões Promotoras Genéticas/genética
12.
BMC Plant Biol ; 21(1): 372, 2021 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-34388971

RESUMO

BACKGROUND: Oilseed rape (B. napus L.) has great potential for phytoremediation of cadmium (Cd)-polluted soils due to its large plant biomass production and strong metal accumulation. Soil properties and the presence of other soluble compounds or ions, cause a heterogeneous distribution of Cd. RESULTS: The aim of our study was to reveal the differential responses of B. napus to different Cd abundances. Herein, we found that high Cd (50 µM) severely inhibited the growth of B. napus, which was not repressed by low Cd (0.50 µM) under hydroponic culture system. ICP-MS assays showed that the Cd2+ concentrations in both shoots and roots under 50 µM Cd were over 10 times higher than those under 0.50 µM Cd. Under low Cd, the concentrations of only shoot Ca2+/Mn2+ and root Mn2+ were obviously changed (both reduced); under high Cd, the concentrations of most cations assayed were significantly altered in both shoots and roots except root Ca2+ and Mg2+. High-throughput transcriptomic profiling revealed a total of 18,021 and 1408 differentially expressed genes under high Cd and low Cd conditions, respectively. The biological categories related to the biosynthesis of plant cell wall components and response to external stimulus were over-accumulated under low Cd, whereas the terms involving photosynthesis, nitrogen transport and response, and cellular metal ion homeostasis were highly enriched under high Cd. Differential expression of the transporters responsible for Cd uptake (NRAMPs), transport (IRTs and ZIPs), sequestration (HMAs, ABCs, and CAXs), and detoxification (MTPs, PCR, MTs, and PCSs), and some other essential nutrient transporters were investigated, and gene co-expression network analysis revealed the core members of these Cd transporters. Some Cd transporter genes, especially NRAMPs and IRTs, showed opposite responsive patterns between high Cd and low Cd conditions. CONCLUSIONS: Our findings would enrich our understanding of the interaction between essential nutrients and Cd, and might also provide suitable gene resources and important implications for the genetic improvement of plant Cd accumulation and resistance through molecular engineering of these core genes under varying Cd abundances in soils.


Assuntos
Brassica napus/genética , Brassica napus/metabolismo , Cádmio/metabolismo , Transporte Biológico , Brassica napus/crescimento & desenvolvimento , Quelantes/metabolismo , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Íons/metabolismo , Solo/química , Tetraploidia , Transcriptoma
13.
Int J Mol Sci ; 22(15)2021 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-34360673

RESUMO

Auxin is a key regulator of plant development affecting the formation and maturation of reproductive structures. The apoplastic route of auxin transport engages influx and efflux facilitators from the PIN, AUX and ABCB families. The polar localization of these proteins and constant recycling from the plasma membrane to endosomes is dependent on Rab-mediated vesicular traffic. Rab proteins are anchored to membranes via posttranslational addition of two geranylgeranyl moieties by the Rab Geranylgeranyl Transferase enzyme (RGT), which consists of RGTA, RGTB and REP subunits. Here, we present data showing that seed development in the rgtb1 mutant, with decreased vesicular transport capacity, is disturbed. Both pre- and post-fertilization events are affected, leading to a decrease in seed yield. Pollen tube recognition at the stigma and its guidance to the micropyle is compromised and the seed coat forms incorrectly. Excess auxin in the sporophytic tissues of the ovule in the rgtb1 plants leads to an increased tendency of autonomous endosperm formation in unfertilized ovules and influences embryo development in a maternal sporophytic manner. The results show the importance of vesicular traffic for sexual reproduction in flowering plants, and highlight RGTB1 as a key component of sporophytic-filial signaling.


Assuntos
Arabidopsis/enzimologia , Sementes/enzimologia , Transdução de Sinais , Alquil e Aril Transferases/metabolismo , Alquil e Aril Transferases/fisiologia , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Transporte Biológico , Regulação da Expressão Gênica de Plantas , Ácidos Indolacéticos/metabolismo , Mutação , Tubo Polínico/fisiologia , Sementes/crescimento & desenvolvimento , Sementes/metabolismo
14.
Int J Mol Sci ; 22(16)2021 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-34445480

RESUMO

Multivesicular body (MVB)-mediated endosomal sorting and macroautophagy are the main pathways mediating the transport of cellular components to the vacuole and are essential for maintaining cellular homeostasis. The interplay of these two pathways remains poorly understood in plants. In this study, we show that FYVE DOMAIN PROTEIN REQUIRED FOR ENDOSOMAL SORTING 1 (FREE1), which was previously identified as a plant-specific component of the endosomal sorting complex required for transport (ESCRT), essential for MVB biogenesis and plant growth, can be transported to the vacuole for degradation in response to iron deficiency. The vacuolar transport of ubiquitinated FREE1 protein is mediated by the autophagy pathway. As a consequence, the autophagy deficient mutants, atg5-1 and atg7-2, accumulate more endogenous FREE1 protein and display hypersensitivity to iron deficiency. Furthermore, under iron-deficient growth condition autophagy related genes are upregulated to promote the autophagic degradation of FREE1, thereby possibly relieving the repressive effect of FREE1 on iron absorption. Collectively, our findings demonstrate a unique regulatory mode of protein turnover of the ESCRT machinery through the autophagy pathway to respond to iron deficiency in plants.


Assuntos
Proteínas de Arabidopsis/química , Arabidopsis/metabolismo , Proteínas Relacionadas à Autofagia/genética , Ferro/metabolismo , Proteínas de Transporte Vesicular/química , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Autofagia , Transporte Biológico , Complexos Endossomais de Distribuição Requeridos para Transporte , Endossomos/metabolismo , Mutação , Proteólise , Ubiquitinação
15.
Int J Mol Sci ; 22(15)2021 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-34360935

RESUMO

Proximal tubules (PTs) take up most of the glucose in the glomerular filtrate and return it to peritubular capillary blood. Sodium-glucose cotransporter 2 (SGLT2) at the apical membrane takes up glucose into the cell. Glucose then flows across the cells and is transported to the interstitium via glucose transporter 2 (GLUT2) at the basolateral membrane. However, glucose transport under SGLT2 inhibition remains poorly understood. In this study, we evaluated the dynamics of a fluorescent glucose analog, 2-NBDG, in the PTs of live mice treated with or without the SGLT2 inhibitor, luseogliflozin. We employed real-time multiphoton microscopy, in which insulin enhanced 2-NBDG uptake in skeletal muscle. Influx and efflux of 2-NBDG in PT cells were compared under hypo-, normo-, and hyperglycemic conditions. Luseogliflozin did not exert significant effects on glucose influx parameters under any level of blood glucose. Our results suggest that blood glucose level per se does not alter glucose influx or efflux kinetics in PTs. In conclusion, neither SGLT2 inhibition nor blood glucose level affect glucose uptake kinetics in PTs. The former was because of glucose influx through basolateral GLUT2, which is an established bidirectional transporter.


Assuntos
Transporte Biológico/efeitos dos fármacos , Glicemia/efeitos dos fármacos , Glucose/metabolismo , Túbulos Renais Proximais , Sorbitol/análogos & derivados , Animais , Linhagem Celular , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Sorbitol/farmacologia
16.
Postepy Biochem ; 67(1): 16-27, 2021 03 31.
Artigo em Polonês | MEDLINE | ID: mdl-34378902

RESUMO

Rab proteins are necessary for membrane fusion and fission and as such are key regulators of intracellular transport in eukaryotic cells. They also control other aspects of cell functioning, including the cytoskeleton rearrangements, determination of cell polarity or signal transduction. Rab proteins exert their control both indirectly, because they decide whether all necessary proteins and other cargo reach their correct destinations in the cell, and directly, through interactions of their active forms with effector proteins. Therefore, the results of Rab dysfunctions manifest themselves on all levels of biological organization â from cells, through tissues and organs, to whole organisms. In plants, Rab-dependent processes are important for cell architecture, differentiation, reactions to biotic and abiotic stress, as well as for the efficiency of agricultural production.


Assuntos
Plantas , Proteínas rab de Ligação ao GTP , Transporte Biológico , Células Eucarióticas/metabolismo , Plantas/metabolismo , Transdução de Sinais , Proteínas rab de Ligação ao GTP/metabolismo
17.
Nat Commun ; 12(1): 4847, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34381031

RESUMO

Circulating phosphate levels are tightly controlled within a narrow range in mammals. By using a novel small-molecule inhibitor, we show that the enzymatic activity of inositol hexakisphosphate kinases (IP6K) is essential for phosphate regulation in vivo. IP6K inhibition suppressed XPR1, a phosphate exporter, thereby decreasing cellular phosphate export, which resulted in increased intracellular ATP levels. The in vivo inhibition of IP6K decreased plasma phosphate levels without inhibiting gut intake or kidney reuptake of phosphate, demonstrating a pivotal role of IP6K-regulated cellular phosphate export on circulating phosphate levels. IP6K inhibition-induced decrease in intracellular inositol pyrophosphate, an enzymatic product of IP6K, was correlated with phosphate changes. Chronic IP6K inhibition alleviated hyperphosphataemia, increased kidney ATP, and improved kidney functions in chronic kidney disease rats. Our results demonstrate that the enzymatic activity of IP6K regulates circulating phosphate and intracellular ATP and suggest that IP6K inhibition is a potential novel treatment strategy against hyperphosphataemia.


Assuntos
Fosfatos/sangue , Fosfotransferases (Aceptor do Grupo Fosfato)/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Homeostase/efeitos dos fármacos , Humanos , Hiperfosfatemia/tratamento farmacológico , Fosfatos de Inositol/metabolismo , Mamíferos , Fosfatos/metabolismo , Fosfotransferases (Aceptor do Grupo Fosfato)/antagonistas & inibidores , Ratos , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores Virais/genética , Receptores Virais/metabolismo , Insuficiência Renal Crônica/tratamento farmacológico
18.
Nat Commun ; 12(1): 4682, 2021 08 03.
Artigo em Inglês | MEDLINE | ID: mdl-34344886

RESUMO

A key impediment to studying water-related mechanisms in plants is the inability to non-invasively image water fluxes in cells at high temporal and spatial resolution. Here, we report that Raman microspectroscopy, complemented by hydrodynamic modelling, can achieve this goal - monitoring hydrodynamics within living root tissues at cell- and sub-second-scale resolutions. Raman imaging of water-transporting xylem vessels in Arabidopsis thaliana mutant roots reveals faster xylem water transport in endodermal diffusion barrier mutants. Furthermore, transverse line scans across the root suggest water transported via the root xylem does not re-enter outer root tissues nor the surrounding soil when en-route to shoot tissues if endodermal diffusion barriers are intact, thereby separating 'two water worlds'.


Assuntos
Raízes de Plantas/metabolismo , Água/metabolismo , Arabidopsis/anatomia & histologia , Arabidopsis/citologia , Arabidopsis/genética , Arabidopsis/metabolismo , Transporte Biológico , Hidrodinâmica , Modelos Biológicos , Mutação , Raízes de Plantas/anatomia & histologia , Raízes de Plantas/citologia , Raízes de Plantas/genética , Brotos de Planta/metabolismo , Estômatos de Plantas/metabolismo , Análise Espectral Raman , Xilema/metabolismo
19.
Viruses ; 13(7)2021 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-34202098

RESUMO

Inhibition of the binding of enveloped viruses surface glycoproteins to host cell receptor(s) is a major target of vaccines and constitutes an efficient strategy to block viral entry and infection of various host cells and tissues. Cellular entry usually requires the fusion of the viral envelope with host plasma membranes. Such entry mechanism is often preceded by "priming" and/or "activation" steps requiring limited proteolysis of the viral surface glycoprotein to expose a fusogenic domain for efficient membrane juxtapositions. The 9-membered family of Proprotein Convertases related to Subtilisin/Kexin (PCSK) serine proteases (PC1, PC2, Furin, PC4, PC5, PACE4, PC7, SKI-1/S1P, and PCSK9) participate in post-translational cleavages and/or regulation of multiple secretory proteins. The type-I membrane-bound Furin and SKI-1/S1P are the major convertases responsible for the processing of surface glycoproteins of enveloped viruses. Stefan Kunz has considerably contributed to define the role of SKI-1/S1P in the activation of arenaviruses causing hemorrhagic fever. Furin was recently implicated in the activation of the spike S-protein of SARS-CoV-2 and Furin-inhibitors are being tested as antivirals in COVID-19. Other members of the PCSK-family are also implicated in some viral infections, such as PCSK9 in Dengue. Herein, we summarize the various functions of the PCSKs and present arguments whereby their inhibition could represent a powerful arsenal to limit viral infections causing the present and future pandemics.


Assuntos
Regulação Viral da Expressão Gênica , Pró-Proteína Convertases/metabolismo , Viroses/virologia , Internalização do Vírus , Vírus/genética , Transporte Biológico , Furina/metabolismo , Humanos , Pró-Proteína Convertase 9/metabolismo , Pró-Proteína Convertases/genética , Proteólise , SARS-CoV-2/enzimologia , SARS-CoV-2/metabolismo , Serina Endopeptidases/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Envelope Viral/metabolismo , Vírus/metabolismo
20.
Molecules ; 26(12)2021 Jun 09.
Artigo em Inglês | MEDLINE | ID: mdl-34207619

RESUMO

Trypanosoma cruzi is the etiologic agent for Chagas disease, which affects 6-7 million people worldwide. The biological diversity of the parasite reflects on inefficiency of benznidazole, which is a first choice chemotherapy, on chronic patients. ABC transporters that extrude xenobiotics, metabolites, and mediators are overexpressed in resistant cells and contribute to chemotherapy failure. An ABCC-like transport was identified in the Y strain and extrudes thiol-conjugated compounds. As thiols represent a line of defense towards reactive species, we aimed to verify whether ABCC-like transport could participate in the regulation of responses to stressor stimuli. In order to achieve this, ABCC-like activity was measured by flow cytometry using fluorescent substrates. The present study reveals the participation of glutathione and ceramides on ABCC-like transport, which are both implicated in stress. Hemin modulated the ABCC-like efflux which suggests that this protein might be involved in cellular detoxification. Additionally, all strains evaluated exhibited ABCC-like activity, while no ABCB1-like activity was detected. Results suggest that ABCC-like efflux is not associated with natural resistance to benznidazole, since sensitive strains showed higher activity than the resistant ones. Although benznidazole is not a direct substrate, ABCC-like efflux increased after prolonged drug exposure and this indicates that the ABCC-like efflux mediated protection against cell stress depends on the glutathione biosynthesis pathway.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Doença de Chagas/tratamento farmacológico , Glutationa/metabolismo , Nitroimidazóis/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Animais , Transporte Biológico , Doença de Chagas/parasitologia , Resistência a Medicamentos , Estresse Oxidativo/fisiologia , Tripanossomicidas/farmacologia , Trypanosoma cruzi/metabolismo
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