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1.
Carbohydr Polym ; 229: 115516, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31826493

RESUMO

Acute myocardial infarction (MI) is a common cardiovascular disease with high mortality. In this study, an injectable thermosensitive hydrogel of chitosan (CS)/dextran (DEX)/ß-glycerophosphate (ß-GP) loaded with umbilical cord mesenchymal stem cells (UCMSCs) was prepared for MI treatment. The good biocompatibility of hydrogels was confirmed by 3T3 cells and HUVECs culture study in vitro. HUVECs encapsulated in the hydrogels showed a cell delivery ability. Furthermore, the results indicated that hydrogel could encapsulate most of UCMSCs and the cells exhibited good viability in CS/ 1.0DEX/ß-GP hydrogels. The expression of cardiac markers of cTnI and Cx43 and signaling pathways of p-Akt and p-ERK1/2 were studied, and it showed that UCMSCs differentiate towards myocardium and has a great potential for therapeutic use of cardiac repair. In conclusion, the thermosensitive hydrogel of CS/1.0 DEX/ß-GP loaded UCMSCs was a promising candidate as cell delivery vehicle for cardiac repair to reconstitute damaged myocardium.


Assuntos
Quitosana/química , Dextranos/química , Glicerofosfatos/química , Hidrogéis/química , Células 3T3 , Animais , Materiais Biocompatíveis/química , Materiais Biocompatíveis/farmacologia , Diferenciação Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Conexina 43/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Camundongos , Infarto do Miocárdio/patologia , Infarto do Miocárdio/terapia , Reologia , Transdução de Sinais/efeitos dos fármacos , Tecidos Suporte/química , Troponina I/metabolismo
2.
Carbohydr Polym ; 229: 115529, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31826519

RESUMO

To improve the application of chitosan (CS)-based scaffold for tissue regeneration, in this study, silk sericin (SS) and silk fibroin (SF) were employed to strengthen the cross-linking of the CS/glycerophosphate (GP) scaffolds. The CS/GP, SS/CS/GP and SF/SS/CS/GP scaffolds were fabricated by collosol-gelling and freeze-drying. The SS and SF improved the physical cross-linking of CS/GP scaffolds, formed uniform layered structures, increased mechanical properties, and decreased degradation velocity. Silk proteins significantly promoted cell growth into the scaffold and increased the blood compatibility. When the SF/SS/CS/GP scaffold was implanted into SD rats, appeared an initial acute inflammatory response, host cells invaded, new muscle fibers grow and formed. And no systemic acute toxicity was observed. The findings demonstrated that silk proteins could enhance the physical and biological properties of chitosan scaffold.


Assuntos
Quitosana/química , Fibroínas/química , Sericinas/química , Animais , Materiais Biocompatíveis/química , Materiais Biocompatíveis/farmacologia , Proliferação de Células/efeitos dos fármacos , Módulo de Elasticidade , Fibroblastos/citologia , Fibroblastos/metabolismo , Glicerofosfatos/química , Hemólise/efeitos dos fármacos , Camundongos , Porosidade , Próteses e Implantes , Ratos , Ratos Sprague-Dawley
3.
Artigo em Inglês | MEDLINE | ID: mdl-30959116

RESUMO

The successive acylation of glycerol-3-phosphate (G3P) by glycerol-3-phosphate acyltransferases and acylglycerol-3-phosphate acyltransferases produces phosphatidic acid (PA), a precursor for CDP-diacylglycerol-dependent phospholipid synthesis. PA is further dephosphorylated by LIPINs to produce diacylglycerol (DG), a substrate for the synthesis of triglyceride (TG) by DG acyltransferases and a precursor for phospholipid synthesis via the CDP-choline and CDP-ethanolamine (Kennedy) pathways. The channeling of fatty acids into TG for storage in lipid droplets and secretion in lipoproteins or phospholipids for membrane biogenesis is dependent on isoform expression, activity and localization of G3P pathway enzymes, as well as dietary and hormonal and tissue-specific factors. Here, we review the mechanisms that control partitioning of substrates into lipid products of the G3P pathway.


Assuntos
Glicerol-3-Fosfato O-Aciltransferase/metabolismo , Glicerofosfatos/metabolismo , Glicerofosfolipídeos/metabolismo , Lipogênese , Transdução de Sinais , Acilação , Animais , Ácidos Graxos/metabolismo , Humanos , Gotículas Lipídicas/metabolismo , Triglicerídeos/metabolismo
4.
Mater Sci Eng C Mater Biol Appl ; 107: 110343, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31761212

RESUMO

The use of injectable hydrogels is currently restricted by the challenge of achieving fast gelation, good mechanical strength, and cytocompatibility. Polymeric self-assembly is a potent tool for generating functional materials that combine multiple characteristics and can react to external factors. In this study, we have developed fiber-reinforced composite hydrogels that exhibits significantly enhanced mechanical strength, reduced gelling time, and excellent cytocompatibility. The practicability of developing a chitosan-based thermogelling solution using hydroxyapatite and polyelectrolyte complex (PEC) self-assembled fibers were evaluated. The effect of ßGP concentration on gelation time was studied by varying the concentration of ßGP added to the chitosan solution. Various combinations were tested to create a suitable hydrogel environment for cell encapsulation, growth, and proliferation at physiological pH and temperature. Determination of Young modulus revealed that PEC fibers reinforced hydrogel was three times stiffer than chitosan-ßGP gels. The gelation time was reduced to 3 min, and the hydrogels had porous structures and gels at physiological pH, temperature, and showed >80% viability for MTT assay to MG63 cells. Moreover, confocal imaging of PEC fiber reinforced hydrogels showed noticeable viability and proliferation. The molecular interactions between gelling agents, polyelectrolytes, and hydroxyapatite were studied using FTIR. We investigated interfacial bonding between PEC fibers with ßGP, NaHCO3, and HAp. The combination of hydroxyapatite and polymer self-assembly technique improved the efficiency of injectable hydrogels that are helpful in minimally invasive applications.


Assuntos
Materiais Biocompatíveis/química , Hidrogéis/administração & dosagem , Hidrogéis/química , Engenharia Tecidual/métodos , Materiais Biocompatíveis/farmacologia , Osso e Ossos/fisiologia , Linhagem Celular Tumoral , Quitosana/química , Durapatita/química , Módulo de Elasticidade , Eletrólitos/química , Glicerofosfatos/química , Humanos , Concentração de Íons de Hidrogênio , Injeções , Teste de Materiais , Nanoestruturas/química , Espectroscopia de Infravermelho com Transformada de Fourier , Temperatura , Tecidos Suporte
5.
Hum Genomics ; 13(1): 67, 2019 12 11.
Artigo em Inglês | MEDLINE | ID: mdl-31829291

RESUMO

BACKGROUND: Aging is believed to have a close association with cardiovascular diseases, resulting in various pathological alterations in blood vessels, including vascular cell phenotypic shifts. In aging vessels, the microRNA(miRNA)-mediated mechanism regulating the vascular smooth muscle cell (VSMC) phenotype remains unclarified. MiRNA microarray was used to compare the expressions of miRNAs in VSMCs from old rats (oVSMCs) and young rats (yVSMCs). Quantitative reverse transcription real-time PCR (qRT-PCR) and small RNA transfection were used to explore the miR-542-3p expression in oVSMCs and yVSMCs in vitro. Calcification induction of yVSMCs was conducted by the treatment of ß-glycerophosphate (ß-GP). Alizarin red staining was used to detect calcium deposition. Western blot and qRT-PCR were used to investigate the expression of the smooth muscle markers, smooth muscle 22α (SM22α) and calponin, and the osteogenic markers, osteopontin (OPN), and runt-related transcription factor 2 (Runx2). Lentivirus was used to overexpress miR-542-3p and bone morphogenetic protein 7 (BMP7) in yVMSCs. Luciferase reporter assay was conducted to identify the target of miR-542-3p. RESULTS: Compared with yVSMCs, 28 downregulated and 34 upregulated miRNAs were identified in oVSMCs. It was confirmed by qRT-PCR that oVSMC expressed four times lower miR-542-3p than yVSMCs. Overexpressing miR-542-3p in yVSMCs suppressed the osteogenic differentiation induced by ß-GP. Moreover, miR-542-3p targets BMP7 and overexpressing BMP7 in miR-542-3p-expressing yVSMCs reverses miR-542-3p's inhibition of osteogenic differentiation. CONCLUSIONS: miR-542-3p regulates osteogenic differentiation of VSMCs through targeting BMP7, suggesting that the downregulation of miR-542-3p in oVSMCs plays a crucial role in osteogenic transition in the aging rat.


Assuntos
Envelhecimento/genética , Proteína Morfogenética Óssea 7/metabolismo , Regulação para Baixo/genética , MicroRNAs/genética , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/metabolismo , Osteogênese/genética , Animais , Sequência de Bases , Regulação para Baixo/efeitos dos fármacos , Glicerofosfatos/farmacologia , Modelos Biológicos , Miócitos de Músculo Liso/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Ratos
6.
Biomed Res Int ; 2019: 5768285, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31886229

RESUMO

We investigated the effects of a human umbilical cord mesenchymal stem cell-conditioned medium (MSC-CM)/chitosan/collagen/ß-glycerophosphate (ß-GP) thermosensitive hydrogel (MSC-CM/hydrogel) on mice with third-degree burns. MSC-CM was collected and mixed with chitosan, collagen, and ß-GP to generate the thermosensitive MSC-CM/hydrogel, which was stored in the liquid phase at 4°C. The wounds of established third-degree burned mice were then externally covered with the MSC-CM/hydrogel, which formed a gel when placed on the wounds at physiological temperature. Injured mice in three additional groups were treated with unconditioned MSC medium (UM), MSC-CM, or UM/chitosan/collagen/ß-GP thermosensitive hydrogels. Skin wound samples were obtained 4, 14, and 28 days after burning for further analysis by hematoxylin and eosin and Ki-67 staining. Wound healing rates and times, in addition to immunohistochemical results, were then compared and analyzed among the four groups. Application of the MSC-CM/hydrogel shortened healing time, limited the area of inflammation, enhanced reepithelialization, promoted the formation of high-quality, well-vascularized granulation tissue, and attenuated the formation of fibrotic and hypertrophic scar tissue. In summary, MSC-CM/hydrogel effectively promotes wound healing in third-degree burned mice.


Assuntos
Queimaduras/patologia , Meios de Cultivo Condicionados/farmacologia , Hidrogéis/farmacologia , Células-Tronco Mesenquimais , Cordão Umbilical , Cicatrização/efeitos dos fármacos , Animais , Células Cultivadas , Quitosana/química , Colágeno/química , Meios de Cultivo Condicionados/química , Glicerofosfatos/química , Humanos , Hidrogéis/química , Masculino , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Temperatura , Cordão Umbilical/citologia , Cordão Umbilical/metabolismo
7.
Biomed Res Int ; 2019: 3139496, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31886199

RESUMO

Background: Melatonin has been demonstrated to protect against calcification in cyclosporine nephrotoxicity. The wingless-type MMTV integration site family member 1 (Wnt1)/ß-catenin pathway is associated with cardiovascular calcification. This study aimed to explore whether melatonin could attenuate VSMC calcification through regulating the Wnt1/ß-catenin signaling pathway. Methods: The effects of melatonin on vascular calcification were investigated in vascular smooth muscle cells (VSMCs). Calcium deposits were visualized by Alizarin Red Staining. Calcium content and alkaline phosphatase (ALP) activity were used to evaluate osteogenic differentiation. Western blots were used to measure the expression of runt-related transcription factor 2 (Runx2), α-smooth muscle actin (α-SMA), and cleaved caspase-3. Results: Melatonin markedly ameliorated calcium deposition and ALP activity. Runx2 and cleaved caspase-3 were found to be reduced and α-SMA was found to be increased by melatonin, together with a decrease in apoptosis. Immunofluorescence assay revealed a lower Runx2 protein level in the melatonin group. Melatonin treatment significantly decreased the expression of Wnt1 and ß-catenin. Treatment with lithium chloride or transglutaminase 2 abrogated the protective effects of melatonin. Conclusion: Melatonin can attenuate ß-GP-induced VSMC calcification through the suppression of Wnt1/ß-catenin system.


Assuntos
Glicerofosfatos/efeitos adversos , Melatonina/farmacologia , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Calcificação Vascular , Via de Sinalização Wnt/efeitos dos fármacos , Proteína Wnt1/metabolismo , Animais , Glicerofosfatos/farmacologia , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/patologia , Ratos , Ratos Sprague-Dawley , Calcificação Vascular/induzido quimicamente , Calcificação Vascular/tratamento farmacológico , Calcificação Vascular/metabolismo , Calcificação Vascular/patologia , beta Catenina/metabolismo
8.
Nat Commun ; 10(1): 5303, 2019 11 22.
Artigo em Inglês | MEDLINE | ID: mdl-31757957

RESUMO

Glycerol-3-phosphate (G3P) is a well-known mobile regulator of systemic acquired resistance (SAR), which provides broad spectrum systemic immunity in response to localized foliar pathogenic infections. We show that G3P-derived foliar immunity is also activated in response to genetically-regulated incompatible interactions with nitrogen-fixing bacteria. Using gene knock-down we show that G3P is essential for strain-specific exclusion of non-desirable root-nodulating bacteria and the associated foliar pathogen immunity in soybean. Grafting studies show that while recognition of rhizobium incompatibility is root driven, bacterial exclusion requires G3P biosynthesis in the shoot. Biochemical analyses support shoot-to-root transport of G3P during incompatible rhizobia interaction. We describe a root-shoot-root signaling mechanism which simultaneously enables the plant to exclude non-desirable nitrogen-fixing rhizobia in the root and pathogenic microbes in the shoot.


Assuntos
Glicerofosfatos/imunologia , Imunidade Vegetal/imunologia , Proteínas de Plantas/genética , Raízes de Plantas/imunologia , Brotos de Planta/imunologia , Rhizobium/imunologia , Soja/imunologia , Simbiose/imunologia , Técnicas de Silenciamento de Genes , Glicerofosfatos/metabolismo , Proteínas de Plantas/metabolismo , Raízes de Plantas/metabolismo , Brotos de Planta/metabolismo , Rhizobium/metabolismo , Transdução de Sinais , Soja/metabolismo
9.
Int J Mol Sci ; 20(22)2019 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-31752206

RESUMO

The ability of bone-marrow-derived mesenchymal stem/stromal cells (BM-MSCs) to differentiate into osteoblasts makes them the ideal candidate for cell-based therapies targeting bone-diseases. Polyphosphate (polyP) is increasingly being studied as a potential inorganic source of phosphate for extracellular matrix mineralisation. The aim of this study is to investigate whether polyP can effectively be used as a phosphate source during the in vitro osteogenic differentiation of human BM-MSCs. Human BM-MSCs are cultivated under osteogenic conditions for 28 days with phosphate provided in the form of organic ß-glycerolphosphate (BGP) or calcium-polyP nanoparticles (polyP-NP). Mineralisation is demonstrated using Alizarin red staining, cellular ATP content, and free phosphate levels are measured in both the cells and the medium. The effects of BGP or polyP-NP on alkaline phosphatase (ALP) activity and gene expression of a range of osteogenic-related markers are also assessed. PolyP-NP supplementation displays comparable effects to the classical BGP-containing osteogenic media in terms of mineralisation, ALP activity and expression of osteogenesis-associated genes. This study shows that polyP-NP act as an effective source of phosphate during mineralisation of BM-MSC. These results open new possibilities with BM-MSC-based approaches for bone repair to be achieved through doping of conventional biomaterials with polyP-NP.


Assuntos
Cálcio/química , Células-Tronco Mesenquimais/citologia , Osteogênese/efeitos dos fármacos , Polifosfatos/farmacologia , Fosfatase Alcalina/metabolismo , Fosfatos de Cálcio , Técnicas de Cultura de Células , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Regulação da Expressão Gênica/efeitos dos fármacos , Glicerofosfatos/farmacologia , Humanos , Células-Tronco Mesenquimais/efeitos dos fármacos , Nanopartículas , Polifosfatos/química
10.
Int J Biochem Cell Biol ; 116: 105614, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31550547

RESUMO

Osteopontin (OPN) is an osteoblast-derived secretory protein that plays a role in bone remodeling, osteoblast responsiveness, and inflammation. We recently found that osteoblast differentiation is type-specific, with conditions of JNK inactivation inducing osteoblasts that preferentially express OPN (OPN-type). Since OPN-type osteoblasts highly express osteogenesis-inhibiting proteins and Rankl, an important inducer of osteoclastogenesis, an increased appearance of OPN-type osteoblasts may be associated with inefficient and poor-quality bone regeneration. However, whether specific osteogenic inducers can modulate OPN-type osteoblast differentiation is completely unknown. Here, we demonstrate that bone morphogenic protein 9 (BMP9) prevents induction of OPN-type osteoblast differentiation under conditions of JNK inhibition. Although JNK inactivation suppressed both BMP2- and BMP9-induced matrix mineralization and osteocalcin expression, the expression of Rankl and specific cytokines such as Gpha2, Esm1, and Sfrp1 under similar conditions was increased in all cells except those treated with BMP9. Increased expression of Id4, a critical transcriptional regulator of OPN-type osteoblast differentiation, was similarly prevented only in BMP9-treated cells. We also found that BMP9 specifically induces the expression of Hey1, a bHLH transcriptional repressor, and that Id4 inhibits the suppressive effects of Hey1 on Opn promoter activity by forming Id4-Hey1 complexes in osteoblasts. Using site-direct mutagenesis, ChIP, and immunoprecipitation, we elucidated that BMP9-induced overexpression of Hey1 can overcome the effects of Id4 and suppress OPN expression. We further found that p38 activation and JNK inactivation are involved in BMP9-induced Hey1 expression. Collectively, these data suggest that BMP9 is a unique osteogenic inducer that regulates OPN-type osteoblast differentiation.


Assuntos
Proteínas de Ciclo Celular/genética , Fator 2 de Diferenciação de Crescimento/farmacologia , Proteínas Inibidoras de Diferenciação/genética , Osteoblastos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Osteopontina/genética , Animais , Animais Recém-Nascidos , Proteína Morfogenética Óssea 2/farmacologia , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular/efeitos dos fármacos , Regulação da Expressão Gênica , Glicerofosfatos/farmacologia , Glicoproteínas/genética , Glicoproteínas/metabolismo , Proteínas Inibidoras de Diferenciação/metabolismo , MAP Quinase Quinase 4/antagonistas & inibidores , MAP Quinase Quinase 4/genética , MAP Quinase Quinase 4/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mutagênese Sítio-Dirigida , Osteoblastos/citologia , Osteoblastos/metabolismo , Osteocalcina/genética , Osteocalcina/metabolismo , Osteogênese/genética , Osteopontina/metabolismo , Cultura Primária de Células , Proteoglicanas/genética , Proteoglicanas/metabolismo , Ligante RANK/genética , Ligante RANK/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
11.
J Biosci ; 44(4)2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31502564

RESUMO

Vascular calcification is a common problem in the elderly with diabetes, heart failure and end-stage renal disease. The differentiation of vascular smooth muscle cells (VSMCs) into osteoblasts is the main feature, but the exact mechanism remains unclear. It is not clear whether adiponectin (APN) affects osteogenic differentiation of VSMCs. This study aims to explore the effect of APN on vascular calcification by using a cell model induced by beta-glycerophosphate (beta-GP). VSMCs were isolated and treated with beta-GP and APN in this study. The alkaline phosphatase (ALP) activity and expression levels of Runx2, BMP-2, collagen type I and osteocalcin were determined. The expression levels of STAT3 and p-STAT3 in nucleus and cytoplasm of VSMCs were analyzed. The results showed that APN significantly inhibited the expression of ALP, Runx2, BMP-2, collagen I, osteocalcin and the formation of the mineralized matrix in VSMCs induced by beta-GP. APN reduces the osteogenic differentiation of VSMCs induced by beta-GP and down-regulates the expression of the osteogenic transcription factor osterix by inhibiting STATS3 phosphorylation and nuclear transport. APN may be one of the potential candidates for clinical treatment of vascular calcification.


Assuntos
Adiponectina/genética , Osteogênese/genética , Fator de Transcrição STAT3/genética , Calcificação Vascular/genética , Animais , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Glicerofosfatos/farmacologia , Humanos , Janus Quinase 2/antagonistas & inibidores , Janus Quinase 2/genética , Camundongos , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Osteogênese/efeitos dos fármacos , RNA/genética , Fator de Transcrição STAT3/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição Sp7/genética , Calcificação Vascular/induzido quimicamente , Calcificação Vascular/patologia
12.
Nat Commun ; 10(1): 3813, 2019 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-31444353

RESUMO

Salicylic acid (SA)-mediated innate immune responses are activated in plants perceiving volatile monoterpenes. Here, we show that monoterpene-associated responses are propagated in feed-forward loops involving the systemic acquired resistance (SAR) signaling components pipecolic acid, glycerol-3-phosphate, and LEGUME LECTIN-LIKE PROTEIN1 (LLP1). In this cascade, LLP1 forms a key regulatory unit in both within-plant and between-plant propagation of immunity. The data integrate molecular components of SAR into systemic signaling networks that are separate from conventional, SA-associated innate immune mechanisms. These networks are central to plant-to-plant propagation of immunity, potentially raising SAR to the population level. In this process, monoterpenes act as microbe-inducible plant volatiles, which as part of plant-derived volatile blends have the potential to promote the generation of a wave of innate immune signaling within canopies or plant stands. Hence, plant-to-plant propagation of SAR holds significant potential to fortify future durable crop protection strategies following a single volatile trigger.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/imunologia , Resistência à Doença/imunologia , Doenças das Plantas/imunologia , Lectinas de Plantas/metabolismo , Compostos Orgânicos Voláteis/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Retroalimentação Fisiológica , Glicerofosfatos/imunologia , Glicerofosfatos/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Imunidade Inata , Monoterpenos/imunologia , Monoterpenos/metabolismo , Ácidos Pipecólicos/imunologia , Ácidos Pipecólicos/metabolismo , Doenças das Plantas/microbiologia , Lectinas de Plantas/genética , Plantas Geneticamente Modificadas , Pseudomonas syringae/imunologia , Ácido Salicílico/imunologia , Ácido Salicílico/metabolismo , Transdução de Sinais/imunologia , Compostos Orgânicos Voláteis/imunologia
13.
Insect Biochem Mol Biol ; 114: 103226, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31446033

RESUMO

The huge energy demand posed by insect flight activity is met by an efficient oxidative phosphorylation process that takes place within flight muscle mitochondria. In the major arbovirus vector Aedes aegypti, mitochondrial oxidation of pyruvate, proline and glycerol 3-phosphate (G3P) represent the major energy sources of ATP to sustain flight muscle energy demand. Although adenylates exert critical regulatory effects on several mitochondrial enzyme activities, the potential consequences of altered adenylate levels to G3P oxidation remains to be determined. Here, we report that mitochondrial G3P oxidation is controlled by adenylates through allosteric regulation of cytochrome c oxidase (COX) activity in A. aegypti flight muscle. We observed that ADP significantly activated respiratory rates linked to G3P oxidation, in a protonmotive force-independent manner. Kinetic analyses revealed that ADP activates respiration through a slightly cooperative mechanism. Despite adenylates caused no effects on G3P-cytochrome c oxidoreductase activity, COX activity was allosterically activated by ADP. Conversely, ATP exerted powerful inhibitory effects on respiratory rates linked to G3P oxidation and on COX activity. We also observed that high energy phosphate recycling mechanisms did not contribute to the regulatory effects of adenylates on COX activity or G3P oxidation. We conclude that mitochondrial G3P oxidation in A. aegypti flight muscle is regulated by adenylates through the allosteric modulation of COX activity, underscoring the bioenergetic relevance of this novel mechanism and the potential consequences for mosquito dispersal.


Assuntos
Aedes/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Glicerofosfatos/metabolismo , Mitocôndrias/metabolismo , Miofibrilas/metabolismo , Regulação Alostérica , Animais , Respiração Celular , Feminino , Oxirredução
14.
Cell Physiol Biochem ; 53(2): 323-336, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31359737

RESUMO

BACKGROUND/AIMS: Vascular calcification represents a huge clinical problem contributing to adverse cardiovascular events, with no effective treatment currently available. Upregulation of hepatocyte growth factor has been linked with vascular calcification, and thus, represent a potential target in the development of a novel therapeutic strategy. Glycomimetics have been shown to interrupt HGF-receptor signalling, therefore this study investigated the effect of novel glycomimetics on osteogenic signalling and vascular calcification in vitro. METHODS: Primary human vascular smooth muscle cells (HVSMCs) were induced by ß-glycerophosphate (ß-GP) and treated with 4 glycomimetic compounds (C1-C4). The effect of ß-GP and C1-C4 on alkaline phosphatase (ALP), osteogenic markers and c-Met/Notch3/HES1 signalling was determined using colorimetric assays, qRT-PCR and western blotting respectively. RESULTS: C1-C4 significantly attenuated ß-GP-induced calcification, as shown by Alizarin Red S staining and calcium content by day 14. In addition, C1-C4 reduced ALP activity and prevented upregulation of the osteogenic markers, BMP-2, Runx2, Msx2 and OPN. Furthermore, ß-GP increased c-Met phosphorylation at day 21, an effect ameliorated by C2 and C4 and the c-Met inhibitor, crizotinib. We next interrogated the effects of the Notch inhibitor DAPT and confirmed an inhibition of ß-GP up-regulated Notch3 protein by C2, DAPT and crizotinib compared to controls. Hes-1 protein upregulation by ß-GP, was also significantly downregulated by C2 and DAPT. GOLD docking analysis identified a potential binding interaction of C1-C4 to HGF which will be investigated further. CONCLUSION: These findings demonstrate that glycomimetics have potent anti-calcification properties acting via HGF/c-Met and Notch signalling.


Assuntos
Músculo Liso Vascular/citologia , Proteínas Proto-Oncogênicas c-met/metabolismo , Receptor Notch3/metabolismo , Fatores de Transcrição HES-1/metabolismo , Calcificação Vascular/metabolismo , Materiais Biomiméticos/farmacologia , Proteína Morfogenética Óssea 2/metabolismo , Linhagem Celular , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Glicerofosfatos/farmacologia , Proteínas de Homeodomínio/metabolismo , Humanos , Miócitos de Músculo Liso/metabolismo , Proteínas Recombinantes/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta/metabolismo
15.
Sci Total Environ ; 692: 219-232, 2019 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-31349163

RESUMO

The multi-barrier deep geological repository system is currently considered as one of the safest option for the disposal of high-level radioactive wastes. Indigenous microorganisms of bentonites may affect the structure and stability of these clays through Fe-containing minerals biotransformation and radionuclides mobilization. The present work aimed to investigate the behavior of bentonite and its bacterial community in the case of a uranium leakage from the waste containers. Hence, bentonite microcosms were amended with uranyl nitrate (U) and glycerol-2-phosphate (G2P) and incubated aerobically for 6 months. Next generation 16S rRNA gene sequencing revealed that the bacterial populations of all treated microcosms were dominated by Actinobacteria and Proteobacteria, accounting for >50% of the community. Additionally, G2P and nitrate had a remarkable effect on the bacterial diversity of bentonites by the enrichment of bacteria involved in the nitrogen and carbon biogeochemical cycles (e.g. Azotobacter). A significant presence of sulfate-reducing bacteria such as Desulfonauticus and Desulfomicrobium were detected in the U-treated microcosms. The actinobacteria Amycolatopsis was enriched in G2P­uranium amended bentonites. High-Angle Annular Dark-Field Scanning Transmission Electron Microscopy analyses showed the capacity of Amycolatopsis and a bentonite consortium formed by Bradyrhizobium-Rhizobium and Pseudomonas to precipitate U as U phosphate mineral phases, probably due to the phosphatase activity. The different amendments did not affect the mineralogy of the bentonite pointing to a high structural stability. These results would help to predict the impact of microbial processes on the biogeochemical cycles of elements (N and U) within the bentonite barrier under repository relevant conditions and to determine the changes in the microbial community induced by a uranium release.


Assuntos
Bactérias/metabolismo , Bentonita/análise , Glicerofosfatos/metabolismo , Microbiota/efeitos dos fármacos , Resíduos Radioativos/análise , Urânio/metabolismo , Bactérias/classificação
16.
Acta Crystallogr F Struct Biol Commun ; 75(Pt 7): 470-479, 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-31282866

RESUMO

(S)-3-O-Geranylgeranylglyceryl phosphate synthase (GGGPS) catalyzes the initial ether-bond formation between sn-glycerol 1-phosphate (G1P) and geranylgeranyl pyrophosphate to synthesize (S)-3-O-geranylgeranylglyceryl phosphate in the production of an archaeal cell-membrane lipid molecule. Archaeal GGGPS proteins are divided into two groups (group I and group II). In this study, the crystal structure of the archaeal group II GGGPS from Thermoplasma acidophilum (TaGGGPS) was determined at 2.35 Šresolution. The structure of TaGGGPS showed that it has a TIM-barrel fold, the third helix of which is disordered (α3*), and that it forms a homodimer, although a pre-existing structure of an archaeal group II GGGPS (from Methanothermobacter thermautotrophicus) showed a hexameric form. The structure of TaGGGPS showed the precise G1P-recognition mechanism of an archaeal group II GGGPS. The structure of TaGGGPS and molecular-dynamics simulation analysis showed fluctuation of the ß2-α2, α3* and α5a regions, which is predicted to be important for substrate uptake and/or product release by TaGGGPS.


Assuntos
Alquil e Aril Transferases/química , Proteínas Arqueais/química , Glicerofosfatos/química , Thermoplasma/enzimologia , Sítios de Ligação , Cristalização , Cristalografia por Raios X , Estrutura Secundária de Proteína , Homologia de Sequência de Aminoácidos , Especificidade por Substrato
17.
Mater Sci Eng C Mater Biol Appl ; 103: 109870, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31349408

RESUMO

Endoscopic submucosal dissection (ESD) is a useful tool for the treatment of early gastric neoplasms, however post-ESD bleeding and perforation restrict its widespread application. In this study, we developed the chitosan/ß-glycerophosphate/collagen (CS/GP/Col) thermo-sensitive systems to satisfy the requirements for the endoscopic treatment for ESD-induced ulcer. The results indicated that the addition of collagen to CS/GP system did not lead to remarkable changes on the physicochemical properties of the systems, which can transform from solution to hydrogels under physiological temperature within 90s, technically makes it suitable to be applied through catheter to gastric ulcer during ESD operation. Besides, hydrogels with high collagen concentration showed better biocompatibility, effectively protected L929, GES-1, HVSMC and CCD-18Co cells from acidic condition, induced more growth factors such as EGF, VEGF and FGF in those cells, and promoted coagulation. These results indicated that the CS/GP/Col thermo-sensitive hydrogel might be a promising biomaterial for the endoscopic treatment of ESD-induced ulcer, and further research can be carried out.


Assuntos
Quitosana , Colágeno , Mucosa Gástrica/cirurgia , Gastroscopia/efeitos adversos , Glicerofosfatos , Hidrogéis , Complicações Pós-Operatórias/tratamento farmacológico , Neoplasias Gástricas/cirurgia , Úlcera , Animais , Linhagem Celular , Quitosana/química , Quitosana/farmacologia , Colágeno/química , Colágeno/farmacologia , Mucosa Gástrica/patologia , Glicerofosfatos/química , Glicerofosfatos/farmacologia , Humanos , Hidrogéis/química , Hidrogéis/farmacologia , Camundongos , Ratos , Ratos Sprague-Dawley , Neoplasias Gástricas/patologia , Úlcera/tratamento farmacológico , Úlcera/etiologia
18.
Rheumatology (Oxford) ; 58(12): 2153-2161, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31143951

RESUMO

OBJECTIVES: Biologic treatment has recently revolutionized the management of RA. Despite this success, ∼30-40% of the patients undergoing biologic treatment respond insufficiently. The aim of this study was to identify several specific reliable metabolites for predicting the response of RA patients to TNF-α inhibitors (TNFi) and abatacept (ABT), using capillary electrophoresis-time-of-flight mass spectrometry (CE-TOFMS). METHODS: We collected serum from RA patients with moderate or high disease activity prior to biologic treatment, and obtained the serum metabolomic profiles of these samples using CE-TOFMS. The patients' response was determined 12 weeks after starting biologic treatment, according to the EULAR response criteria. We compared the metabolites between the response and non-response patient groups and analysed their discriminative ability. RESULTS: Among 43 total patients, 14 of 26 patients in the TNFi group and 6 of 17 patients in the ABT group responded to the biologic treatment. Of the metabolites separated by CE-TOFMS, 196 were identified as known substances. Using an orthogonal partial least-squares discriminant analysis, we identified five metabolites as potential predictors of TNFi responders and three as predictors of ABT responders. Receiver operating characteristic analyses for multiple biomarkers revealed an area under the curve (AUC) of 0.941, with a sensitivity of 85.7% and specificity of 100% for TNFi, and an AUC of 0.985, with a sensitivity of 100% and specificity of 90.9% for ABT. CONCLUSION: By metabolomic analysis, we identified serum biomarkers that have a high ability to predict the response of RA patients to TNFi or ABT treatment.


Assuntos
Abatacepte/uso terapêutico , Antirreumáticos/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Metabolômica , Inibidores do Fator de Necrose Tumoral/uso terapêutico , Alanina/análogos & derivados , Alanina/metabolismo , Aminobutiratos/metabolismo , Área Sob a Curva , Artrite Reumatoide/metabolismo , Biomarcadores/metabolismo , Caproatos/metabolismo , Ácido Cítrico/metabolismo , Eletroforese Capilar , Feminino , Glicerofosfatos/metabolismo , Humanos , Masculino , Espectrometria de Massas , Pessoa de Meia-Idade , Prognóstico , Ácido Quínico/metabolismo , Taurina/metabolismo
19.
Mar Biotechnol (NY) ; 21(4): 488-502, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31076921

RESUMO

Ammonia is toxic to aquatic animal. Currently, only limited works were reported on the responses of aquatic animals after ammonia exposure using "omics" technologies. Tilapia suffers from the stress of ammonia-nitrogen during intensive recirculating aquaculture. Optimizing ammonia stress tolerance has become an important issue in tilapia breeding. The molecular and biochemical mechanisms of ammonia-nitrogen toxicity have not been understood comprehensively in tilapia yet. In this study, using RNA-seq and gas chromatograph system coupled with a Pegasus HT time-of-flight mass spectrometer (GC-TOF-MS) techniques, we investigated differential expressed genes (DEGs) and metabolomes in the liver at 6 h post-challenges (6 hpc) and 24 h post-challenges (24 hpc) under high concentration of ammonia-nitrogen treatment. We detected 2258 DEGs at 6 hpc and 315 DEGs at 24 hpc. Functional enrichment analysis indicated that DEGs were significantly associated with cholesterol biosynthesis, steroid and lipid metabolism, energy conservation, and mitochondrial tissue organization. Metabolomic analysis detected 31 and 36 metabolites showing significant responses to ammonia-nitrogen stress at 6 and 24 hpc, respectively. D-(Glycerol 1-phosphate), fumaric acid, and L-malic acid were found significantly down-regulated at both 6 and 24 hpc. The integrative analysis of transcriptomics and metabolomics suggested considerable alterations and precise control of gene expression at both physiological and molecular levels in response to the stress of ammonia-nitrogen in tilapia.


Assuntos
Amônia/toxicidade , Proteínas de Peixes/genética , Fígado/efeitos dos fármacos , Metaboloma/genética , Tilápia/genética , Poluentes Químicos da Água/toxicidade , Animais , Colesterol/metabolismo , Exposição Ambiental , Proteínas de Peixes/classificação , Proteínas de Peixes/metabolismo , Fumaratos/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Ontologia Genética , Glicerofosfatos/metabolismo , Metabolismo dos Lipídeos , Fígado/metabolismo , Malatos/metabolismo , Anotação de Sequência Molecular , Estresse Fisiológico/genética , Tilápia/metabolismo , Transcriptoma
20.
Ren Fail ; 41(1): 220-228, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30973285

RESUMO

Vascular calcification (VC) is a pathological process characterized by abnormal deposition of calcium phosphate, hydroxyapatite and other mineral substances in the vascular wall. Hyperphosphorus is an important risk factor associated with VC in the general population and patients with chronic kidney disease (CKD). However, there is still a lack of early biomarkers for hyperphosphorus induced VC. We established a calcific rat aorta vascular smooth muscle cells (RASMCs) model by stimulating with ß-glycerophosphate. Then we performed label-free quantitative proteomics combined with liquid chromatograph-mass spectrometer/mass spectrometer (LC-2D-MS/MS)analysis and bioinformatics analysis to find the potential biomarkers for VC. In the current study, we identified 113 significantly proteins. Fifty six of these proteins were significantly up-regulated and the other 57 proteins were significantly decreased in calcific RASMCs, compared to that of normal control cells (fold-change (fc)>1.2, p < .05). Bioinformatics analysis indicated that these significant proteins mainly involved in the placenta blood vessel development and liver regeneration. Their molecule function was cell adhesion molecule binding. Among them, Smarca4 is significantly up-regulated in calcific RASMCs with fc = 2.72 and p = .01. In addition, we also established VC rat model. Real-time quantitative PCR analysis confirmed that the expression of Smarca4 was significantly increased in the aorta of calcified rat. Consistent with the up-regulation of Smarca4, the expression of VC associated microRNA such as miR-133b and miR-155 was also increased. Consequently, our study demonstrates that Smarca4 is involved in hyperphosphorus-induced VC. This finding may contribute to the early diagnosis and prevention of VC.


Assuntos
DNA Helicases/metabolismo , Hiperfosfatemia/metabolismo , Falência Renal Crônica/complicações , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Calcificação Vascular/metabolismo , Animais , Aorta/patologia , Biomarcadores/metabolismo , Linhagem Celular , Modelos Animais de Doenças , Glicerofosfatos/toxicidade , Humanos , Falência Renal Crônica/induzido quimicamente , Falência Renal Crônica/metabolismo , Masculino , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/patologia , Fosfatos/sangue , Fosfatos/metabolismo , Proteômica/instrumentação , Proteômica/métodos , Ratos , Ratos Wistar , Espectrometria de Massas em Tandem/instrumentação , Espectrometria de Massas em Tandem/métodos , Regulação para Cima , Calcificação Vascular/patologia
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