RESUMEN
Many established bioinks fulfill important requirements regarding fabrication standards and cytocompatibility. Current research focuses on development of functionalized bioinks with an improved support of tissue-specific cell differentiation. Many approaches primarily depend on decellularized extracellular matrices or blood components. In this study, we investigated the combination of a highly viscous alginate-methylcellulose (algMC) bioink with collagen-based artificial extracellular matrix (aECM) as a finely controllable and tailorable system composed of collagen type I (col) with and without chondroitin sulfate (CS) or sulfated hyaluronan (sHA). As an additional stabilizer, the polyphenol tannic acid (TA) was integrated into the inks. The assessment of rheological properties and printability as well as hydrogel microstructure revealed no adverse effect of the integrated components on the inks. Viability, adhesion, and proliferation of bioprinted immortalized human mesenchymal stem cells (hTERT-MSC) was improved indicating enhanced interaction with the designed microenvironment. Furthermore, chondrogenic matrix production (collagen type II and sulfated glycosaminoglycans) by primary human chondrocytes (hChon) was enhanced by aECM. Supplementing the inks with TA was required for these positive effects but caused cytotoxicity as soon as TA concentrations exceeded a certain amount. Thus, combining tailorable aECM with algMC and balanced TA addition proved to be a promising approach for promoting adhesion of immortalized stem cells and differentiation of chondrocytes in bioprinted scaffolds.
Asunto(s)
Alginatos , Células Madre Mesenquimatosas , Humanos , Células Madre Mesenquimatosas/metabolismo , Glicosaminoglicanos/química , Glicosaminoglicanos/metabolismo , Glicosaminoglicanos/farmacología , Colágeno Tipo I/metabolismo , Colágeno Tipo I/farmacología , Diferenciación Celular , Metilcelulosa/metabolismo , Metilcelulosa/farmacología , Taninos/metabolismo , Taninos/farmacologíaRESUMEN
Glycosaminoglycans (GAGs) with unique structures from marine animals show intriguing pharmacological activities and negligible biological risks, providing more options for us to explore safer agents. The swim bladder is a tonic food and folk medicine, and its GAGs show good anticoagulant activity. In this study, two GAGs, CMG-1.0 and GMG-1.0, were extracted and isolated from the swim bladder of Cynoscion microlepidotus and Gadus morhua. The physicochemical properties, precise structural characteristics, and anticoagulant activities of these GAGs were determined for the first time. The analysis results of the CMG-1.0 and GMG-1.0 showed that they were chondroitin sulfate (CS)/dermatan sulfate (DS) hybrid chains with molecular weights of 109.3 kDa and 123.1 kDa, respectively. They were mainly composed of the repeating disaccharide unit of -{IdoA-α1,3-GalNAc4S-ß1,4-}- (DS-A). The DS-B disaccharide unit of -{IdoA2S-α1,3-GalNAc4S-ß1,4-}- also existed in both CMG-1.0 and GMG-1.0. CMG-1.0 had a higher proportion of CS-O disaccharide unit -{-GlcA-ß1,3-GalNAc-ß1,4-}- but a lower proportion of CS-E disaccharide unit -{-GlcA-ß1,3-GalNAc4S6S-ß1,4-}- than GMG-1.0. The disaccharide compositions of the GAGs varied in a species-specific manner. Anticoagulant activity assay revealed that both CMG-1.0 and GMG-1.0 had potent anticoagulant activity, which can significantly prolong activated partial thromboplastin time. GMG-1.0 also can prolong the thrombin time. CMG-1.0 showed no intrinsic tenase inhibition activity, while GMG-1.0 can obviously inhibit intrinsic tenase with EC50 of 58 nM. Their significantly different anticoagulant activities may be due to their different disaccharide structural units and proportions. These findings suggested that swim bladder by-products of fish processing of these two marine organisms may be used as a source of anticoagulants.
Asunto(s)
Sulfatos de Condroitina , Dermatán Sulfato , Animales , Sulfatos de Condroitina/farmacología , Sulfatos de Condroitina/química , Dermatán Sulfato/farmacología , Dermatán Sulfato/análisis , Dermatán Sulfato/química , Vejiga Urinaria/química , Glicosaminoglicanos/química , Anticoagulantes/farmacología , DisacáridosRESUMEN
Stichopus monotuberculatus is a tropical sea cucumber species and used as a folk medicine and tonic food. In this study, a fucosylated glycosaminoglycan (SmFG), the depolymerized SmFG (dSmFG) and its oligosaccharide fractions were prepared. The SmFG and its depolymerized products were comprised of a chondroitin-sulfate-E backbone, and various sulfated fucose side chains, including an unusual disaccharide side chain connected to the C-3 position of D-glucuronic acid (GlcA) or GlcA-ol. A peeling reaction occurred during the deaminative depolymerization process. The dSmFG and its fractions showed strong anticoagulant activity by selectively inhibiting intrinsic tenase complex, and had no anti-factor IIa, Xa and VIIa activity. The anticoagulant activity reduced with the decrease of molecular weight, and the unusual branch and novel reducing end may enhance the anticoagulant activity. These findings can provide significant information for development and utilization of depolymerized products from SmFG in food and pharmaceutical industries.
Asunto(s)
Glicosaminoglicanos , Pepinos de Mar , Animales , Anticoagulantes/química , Anticoagulantes/farmacología , Sulfatos de Condroitina/química , Disacáridos , Fucosa/química , Ácido Glucurónico , Glicosaminoglicanos/química , Glicosaminoglicanos/farmacología , Oligosacáridos/química , Pepinos de Mar/química , SulfatosRESUMEN
Phosphate and sulfate groups are integral to energy metabolism and introduce negative charges into biological macromolecules. One purpose of such modifications is to elicit precise binding/activation of protein partners. The physico-chemical properties of the two groups, while superficially similar, differ in one important respect-the valency of the central (phosphorus or sulfur) atom. This dictates the distinct properties of their respective esters, di-esters and hence their charges, interactions with metal ions and their solubility. These, in turn, determine the contrasting roles for which each group has evolved in biological systems. Biosynthetic links exist between the two modifications; the sulfate donor 3'-phosphoadenosine-5'-phosphosulfate being formed from adenosine triphosphate (ATP) and adenosine phosphosulfate, while the latter is generated from sulfate anions and ATP. Furthermore, phosphorylation, by a xylosyl kinase (Fam20B, glycosaminoglycan xylosylkinase) of the xylose residue of the tetrasaccharide linker region that connects nascent glycosaminoglycan (GAG) chains to their parent proteoglycans, substantially accelerates their biosynthesis. Following observations that GAG chains can enter the cell nucleus, it is hypothesized that sulfated GAGs could influence events in the nucleus, which would complete a feedback loop uniting the complementary anionic modifications of phosphorylation and sulfation through complex, inter-connected signalling networks and warrants further exploration.
Asunto(s)
Vías Biosintéticas , Glicosaminoglicanos , Adenosina Trifosfato/metabolismo , Ésteres , Glicosaminoglicanos/química , Fosforilación , Sulfatos/metabolismoRESUMEN
Oviductus ranae (O.ran.) has been widely used as a tonic and a traditional animal-based Chinese medicine. O.ran. extracts have been reported to have numerous biological activities, including activities that are often associated with mammalian glycosaminoglycans such as anti-inflammatory, antiosteoperotic, and anti-asthmatic. Glycosaminoglycans are complex linear polysaccharides ubiquitous in mammals that possess a wide range of biological activities. However, their presence and possible structural characteristics within O.ran. were previously unknown. In this study, glycosaminoglycans were isolated from O.ran. and their disaccharide compositions were analyzed by liquid chromatography-ion trap/time-of-flight mass spectrometry (LC-MS-ITTOF). Heparan sulfate (HS)/heparin (HP), chondroitin sulfate (CS)/dermatan sulfate (DS) and hyaluronic acid (HA) were detected in O.ran. with varied disaccharide compositions. HS species contain highly acetylated disaccharides, and have various structures in their constituent chains. CS/DS chains also possess a heterogeneous structure with different sulfation patterns and densities. This novel structural information could help clarify the possible involvement of these polysaccharides in the biological activities of O.ran..
Asunto(s)
Glicosaminoglicanos/análisis , Glicosaminoglicanos/química , Materia Medica/química , Sulfatos de Condroitina/análisis , Cromatografía Liquida , Dermatán Sulfato/análogos & derivados , Dermatán Sulfato/análisis , Disacáridos/análisis , Disacáridos/aislamiento & purificación , Glicosaminoglicanos/aislamiento & purificación , Heparina/análisis , Heparitina Sulfato/análisis , Espectrometría de Masas/métodos , Sensibilidad y EspecificidadRESUMEN
Improved ocular delivery of a poorly soluble anti-glaucoma drug, acetazolamide (ACZ), in a stable nanosuspension (NS) was the main target of the study. The anionic polypeptide, poly-γ-glutamic acid (PG) and the glycosaminoglycan, hyaluronic acid, were used to stabilize ACZ-NS prepared using the antisolvent precipitation (AS-PT) coupled with sonication technique. To endue in site biocompatibility with high tolerability, soya lecithin (SL) phospholipid has been also combined with polyvinyl alcohol (PVA). NS with uniform PS in the range 100-300 nm, high ζ > ±20 mV, and enhanced saturation solubility were produced. Targeting solvent removal with control on future particle growth, post-production processing of NS was done using spray drying. The carriers' composition and amount relative to ACZ-NS were optimized to allow for the production of a redispersible dry crystalline powder. Particles crystallinity was confirmed using X-ray powder diffraction (XRPD) and differential scanning calorimetry (DSC) in liquid and spray dried NS. The modified Draize test proved the safety and tolerability following application to rabbit eyes accompanying an efficient ocular hypotensive activity using a steroid glaucoma model.
Asunto(s)
Acetazolamida , Materiales Biocompatibles/uso terapéutico , Portadores de Fármacos/uso terapéutico , Ojo/efectos de los fármacos , Glaucoma/tratamiento farmacológico , Nanopartículas/uso terapéutico , Acetazolamida/administración & dosificación , Acetazolamida/farmacocinética , Animales , Disponibilidad Biológica , Ojo/patología , Glicosaminoglicanos/química , Lecitinas/química , Péptidos/química , Alcohol Polivinílico/química , Conejos , Glycine max/químicaRESUMEN
Magnetic fields (MFs) have been used as an external stimulus to increase cell proliferation in chondrocytes and extracellular matrix (ECM) synthesis of articular cartilage. However, previously published studies have not shown that MFs are homogeneous through cell culture systems. In addition, variables such as stimulation times and MF intensities have not been standardized to obtain the best cellular proliferative rate or an increase in molecular synthesis of ECM. In this work, a stimulation device, which produces homogeneous MFs to stimulate cell culture surfaces was designed and manufactured using a computational model. Furthermore, an in vitro culture of primary rat chondrocytes was established and stimulated with two MF schemes to measure both proliferation and ECM synthesis. The best proliferation rate was obtained with an MF of 2 mT applied for 3 h, every 6 h for 8 days. In addition, the increase in the synthesis of glycosaminoglycans was statistically significant when cells were stimulated with an MF of 2 mT applied for 5 h, every 6 h for 8 days. These findings suggest that a stimulation with MFs is a promising tool that could be used to improve in vitro treatments such as autologous chondrocyte implantation, either to increase cell proliferation or stimulate molecular synthesis. Bioelectromagnetics. 2020;41:41-51 © 2019 Bioelectromagnetics Society.
Asunto(s)
Cartílago Articular/metabolismo , Condrocitos/citología , Condrocitos/metabolismo , Matriz Extracelular/metabolismo , Campos Magnéticos/efectos adversos , Animales , Proliferación Celular , Supervivencia Celular , Células Cultivadas , Células Inmovilizadas , Simulación por Computador , Glicosaminoglicanos/química , Ratas , Ratas Wistar , Propiedades de Superficie , Temperatura , Factores de TiempoRESUMEN
A nutraceutical is defined as a standardized pharmaceutical-grade nutrient. Among hundreds of nutraceuticals, polysaccharide or glycan-based products such as those containing chondroitin sulfate glycosaminoglycan isolated from animal cartilage have been on the top nutraceutical selling list for many years. It is expected that the nutraceutical market will reach $250 billion dollars worldwide by 2018. Glycans are most abundant biopolymers on earth those are synthesized by bacteria, fungi, plants, insects, and animals. Glycans that are synthesized by animals or from all marine sources can be modified with covalently linked sulfates or containing acidic monosaccharides whereas glycans that are synthesized by terrestrial plants or fungi usually do not contain sulfates. Glycans such as starch are common sources of energy for animals, therefore they are on the nutrient-side of nutraceuticals. Undigestible polysaccharides from plants could serve as dietary fiber for humans that change the contents of the gastrointestinal tract and affect how other nutrients and chemicals are absorbed, thus dietary fibers could be called nutraceuticals. Other intra- and extracellular glycans from different sources serve as biological active components that regulate a myriad of physiological functions. The reported biological functions for such glycans are not limited to immune system regulatory, anti-coagulating, anti-tumor, anti-viral, anti-oxidant, anti-aging, and lipid-lowing activities, which make them pharmaceutical-side nutraceuticals. This review will present the full spectrum of glycan-based nutraceuticals and summarize current knowledge (published data from 1960s to current) of the structure, biological function, and mechanisms of glycans from both terrestrial and marine sources.
Asunto(s)
Suplementos Dietéticos , Glicosaminoglicanos/farmacología , Polisacáridos/farmacología , Animales , Organismos Acuáticos/química , Glicosaminoglicanos/química , Humanos , Polisacáridos/químicaRESUMEN
Glutaraldehyde (GLUT) crosslinked bioprosthetic heart valves (BHVs) might fail due to progressive degradation and calcification. GLUT cannot stabilize glycosaminoglycans (GAGs), which are important for BHVs' life time. In this current study we developed a new BHVs preparation strategy using exogenous hyaluronic acid (HA)/chondroitin sulfate (CS) supplement and sodium trimetaphosphate (STP) crosslinking method. Exogenous HA and CS provide additional GAGs for pericardiums. STP could link two GAGs by reacting with hydroxyl groups in GAGs' repeating polysaccharides units. The feeding ratios of HA/CS were optimized. The GAGs content and long-term stability in vitro, biocompatibility, the in vivo GAGs stability and anti-calcification potential of GLUT/HA/CS and STP treated pericardiums were characterized. We demonstrated that GLUT/HA/CS and STP treated pericardiums had sufficiently increased GAGs' amount and stability and decreased calcification. This new exogenous hyaluronic acid/chondroitin sulfate supplement and sodium trimetaphosphate crosslinking strategy would be a promising method to make BHVs with better structural stability and anti-calcification properties.
Asunto(s)
Bioprótesis , Sulfatos de Condroitina/química , Reactivos de Enlaces Cruzados/química , Glicosaminoglicanos/química , Prótesis Valvulares Cardíacas , Ácido Hialurónico/química , Animales , Válvula Aórtica/cirugía , Materiales Biocompatibles , Coagulación Sanguínea , Glutaral/química , Masculino , Pericardio/patología , Adhesividad Plaquetaria , Polifosfatos/química , Ratas , Ratas Sprague-Dawley , Espectrofotometría Infrarroja , Sulfatos , PorcinosRESUMEN
Heparan sulfate glycosaminoglycans (HS GAGs) attached to proteoglycans harbor high affinity binding sites for various growth factors (GFs) and direct their organization and activity across the cell-matrix interface. Here, we describe a mild and efficient method for generating HS-protein conjugates. The two-step process utilizes a "copper-free click" coupling between differentially sulfated heparinoids primed at their reducing end with an azide handle and a bovine serum albumin protein modified with complementary cyclooctyne functionality. When adsorbed on tissue culture substrates, the glycoconjugates served as extracellular matrix proteoglycan models with the ability to sequester FGF2 and influence mesenchymal stem cell proliferation based on the structure of their HS GAG component.
Asunto(s)
Matriz Extracelular/metabolismo , Factor 2 de Crecimiento de Fibroblastos/química , Heparinoides/química , Células Madre/metabolismo , Animales , Glicosaminoglicanos/químicaRESUMEN
Radiation-induced proctitis (RIP) is the most common clinical adverse effect for patients receiving radiotherapy as part of the standard course of treatment for ovarian, prostate, colon, and bladder cancers. RIP limits radiation dosage, interrupts treatment, and lowers patients' quality of life. A prophylactic treatment that protects the gastrointestinal tract from deleterious effects of radiotherapy will significantly improve patient quality of life and may allow for higher and more regular doses of radiation therapy. Semi-synthetic glycosaminoglycan (GAG), generated from the sulfation of hyaluronic acid, are anti-inflammatory but have difficulty achieving therapeutic levels in many tissues. To enhance the delivery of GAG, we created an in situ gelling rectal delivery system using silk-elastinlike protein polymers (SELPs). Using solutions of SELP 815K (which contains 6 repeats of blocks comprised of 8 silk-like units, 15 elastin-like units, and 1 lysine-substituted elastin-like unit) with GAG GM-0111, we created an injectable delivery platform that transitioned in <5min from a liquid at room temperature to a hydrogel at body temperature. The hydrogels released 50% of their payload within 30min and enhanced the accumulation of GAG in the rectum compared to traditional enema-based delivery. Using a murine model of radiation-induced proctitis, the prophylactic delivery of a single dose of GAG from a SELP matrix administered prior to irradiation significantly reduced radiation-induced pain after 3, 7, and 21days by 53±4%, 47±10%, and 12±6%, respectively. Matrix-mediated delivery of GAG by SELP represents an innovative method for more effective treatment of RIP and promises to improve quality of life of cancer patients by allowing higher radiotherapy doses with improved safety.
Asunto(s)
Glicosaminoglicanos/administración & dosificación , Hidrogeles/administración & dosificación , Dolor/tratamiento farmacológico , Proctitis/tratamiento farmacológico , Proteínas/administración & dosificación , Traumatismos Experimentales por Radiación/tratamiento farmacológico , Animales , Conducta Animal/efectos de los fármacos , Liberación de Fármacos , Enema , Femenino , Glicosaminoglicanos/química , Glicosaminoglicanos/farmacocinética , Glicosaminoglicanos/uso terapéutico , Hidrogeles/química , Hidrogeles/farmacocinética , Hidrogeles/uso terapéutico , Ratones , Dolor/etiología , Dolor/metabolismo , Dolor/prevención & control , Proctitis/etiología , Proctitis/metabolismo , Proctitis/prevención & control , Proteínas/química , Proteínas/farmacocinética , Proteínas/uso terapéutico , Traumatismos Experimentales por Radiación/etiología , Traumatismos Experimentales por Radiación/metabolismo , Traumatismos Experimentales por Radiación/prevención & control , Recto/metabolismo , Reología , Rayos X/efectos adversosRESUMEN
The importance of extracellular matrix (ECM) integrity in maintaining normal tissue function is highlighted by numerous pathologies and situations of acute and chronic injury associated with dysregulation or destruction of ECM components. Heparan sulfate (HS) is a key component of the ECM, where it fulfils important functions associated with tissue homeostasis. Its degradation following tissue injury disrupts this delicate equilibrium and may impair the wound healing process. ReGeneraTing Agents (RGTA®s) are polysaccharides specifically designed to replace degraded HS in injured tissues. The unique properties of RGTA® (resistance to degradation, binding and protection of ECM structural and signaling proteins, like HS) permit the reconstruction of the ECM, restoring both structural and biochemical functions to this essential substrate, and facilitating the processes of tissue repair and regeneration. Here, we review 25 years of research surrounding this HS mimic, supporting the mode of action, pre-clinical studies and therapeutic efficacy of RGTA® in the clinic, and discuss the potential of RGTA® in new branches of regenerative medicine.
Asunto(s)
Materiales Biomiméticos/farmacología , Lesiones de la Cornea/tratamiento farmacológico , Glicosaminoglicanos/farmacología , Sustancias Protectoras/farmacología , Piel/efectos de los fármacos , Cicatrización de Heridas/efectos de los fármacos , Animales , Materiales Biomiméticos/química , Huesos/efectos de los fármacos , Huesos/lesiones , Ensayos Clínicos como Asunto , Lesiones de la Cornea/rehabilitación , Evaluación Preclínica de Medicamentos , Matriz Extracelular/química , Tracto Gastrointestinal/efectos de los fármacos , Tracto Gastrointestinal/lesiones , Glicosaminoglicanos/química , Heparitina Sulfato/química , Heparitina Sulfato/farmacología , Humanos , Músculos/efectos de los fármacos , Músculos/lesiones , Sustancias Protectoras/química , Medicina Regenerativa/métodos , Piel/lesiones , Andamios del TejidoRESUMEN
This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article has been retracted at the request of authors. The authors have recently found a serious mistake in Table 1 of the article, where the molecular ratio of different monosaccharides is inconsistent with their previously published work. This error flaws the paper and so the authors wish it to be retracted to avoid misunderstanding and misinterpretation of their research work. The authors apologise for any concern or confusion that might have resulted in publishing this article.
Asunto(s)
Antitrombinas/química , Glicosaminoglicanos/química , Trombosis/tratamiento farmacológico , Animales , Antitrombinas/farmacología , Evaluación Preclínica de Medicamentos , Glicosaminoglicanos/farmacología , Holothuria/química , Masculino , Ratones Endogámicos BALB C , Peso Molecular , Polimerizacion , Ratas WistarRESUMEN
We report on the build-up and the intrinsic properties of polyelectrolyte multilayer films from poly-l-lysine and glycosaminoglycans (GAGs) with different sulfation degree, i.e. different charge. We used three complementary techniques, namely electrokinetic analysis (EKA), quartz-crystal microbalance with dissipation (QCM-D) and surface plasmon resonance (SPR), to characterize the assembly process and to assess the properties of the obtained films. EKA elucidated the contribution of the polymers charged groups to the net surface charge of the films and suggested that the assembly process is not solely driven by electrostatic interactions. The combined analysis of QCM-D and SPR data demonstrated that the mechanical properties of the films are dependent on the polymer charge: sulfated GAGs (heparin and chondroitin sulfate) form elastic films while hyaluronan (no sulfation) assembles into multilayer constructs with viscous behavior. The contribution of the water content to these distinct regimes is also discussed. Finally, we show that rather complete characterization of the film properties is possible by SPR employing the two-wavelength and two-media approach: thickness, adsorbed mass, refractive index, and interaction kinetics of the assembly process can be studied by SPR alone.
Asunto(s)
Glicosaminoglicanos/química , Polilisina/química , Sulfatos/química , Ácido Hialurónico/química , Concentración de Iones de Hidrógeno , Tecnicas de Microbalanza del Cristal de Cuarzo , Refractometría , Electricidad Estática , Resonancia por Plasmón de SuperficieRESUMEN
Oligophenylenevinylene (OPV)-based fluorescent (FL) chemosensors exhibiting linear FL responses toward polyanions were designed. Their application to FL sensing of glycosaminoglycans (heparin: HEP, chondroitin 4-sulfate: ChS, and hyaluronic acid: HA) revealed that the charge density encoded as the unit structure directs the mode of OPV self-assembly: H-type aggregate for HEP with 16-times FL increase and J-type aggregate for HA with 93-times FL increase, thus unexpectedly achieving the preferential selectivity for HA in contrast to the conventional HEP selective systems. We have found that the integral magnitude of three factors consisting of binding mechanism, self-assembly, and FL response can amplify the structural information on the target input into the characteristic FL output. This emergent property has been used for a novel molecular recognition system that realizes unconventional FL sensing of HA, potentially applicable to the clinical diagnosis of cancer-related diseases.
Asunto(s)
Colorantes Fluorescentes/química , Glicosaminoglicanos/química , Ácido Hialurónico/química , Espectrometría de Fluorescencia , Sulfatos de Condroitina/química , Heparina/química , Concentración de Iones de HidrógenoRESUMEN
Phytochemicals can exert their bioactivity without reaching the systemic circulation; scarcely absorbed antioxidants might reach the large bowel contributing to protection from oxidative damage-induced gastrointestinal diseases. In the present work, we aimed to study the relationship between potential activity of polyphenol-rich extracts from Cichorium intybus L. and changes in morphological characteristics on Caco-2 cells. Phytochemicals content (carotenoids and flavonoids) and total antioxidant activity of Red Chicory of Treviso and Variegated Chicory of Castelfranco were evaluated. The bioactivity of polyphenol-rich extracts from chicories was studied in in vitro Caco-2 cell monolayers model. Morphological characteristics changes to test the antioxidant and/or prooxidant effect were verified by histological analysis and observed by Electronic Scansion Microscopy (SEM). On Caco-2 cell model, the polyphenols fractions from chicories have indicated a moderate antioxidant behavior until 17 µM concentration, while 70 µM and 34 µM exert cytotoxic effects for Treviso's and Castelfranco's Chicory, respectively, highlighted by TEER decreasing, increased permeability, and alteration of epithelium. Our findings support the beneficial effects of these products in counteracting the oxidative stress and cellular damage, induced in vitro on Caco-2 cell model, through interaction with the mucopolysaccharide complexes in the glycocalyx, maintaining in vivo a healthy and effective intestinal barrier.
Asunto(s)
Cichorium intybus/química , Extractos Vegetales/química , Polifenoles/química , Antioxidantes/química , Células CACO-2/efectos de los fármacos , Impedancia Eléctrica , Flavonoides/química , Glicocálix/química , Glicosaminoglicanos/química , Humanos , Microscopía Electrónica de Rastreo , Oxidación-Reducción , Estrés Oxidativo , Permeabilidad , Fitoquímicos/química , Especies Reactivas de Oxígeno/metabolismo , Uniones Estrechas/metabolismoRESUMEN
Interstitial cystitis (IC), often referred to in combination with painful bladder syndrome, is a chronic inflammatory disease of the bladder. Current therapies primarily focus on replenishing urothelial glycosaminoglycan (GAG) layer using GAG analogs and managing pain with supportive therapies. However, the elusive etiology of IC and the lack of animal models to study the disease have been major hurdles developing more effective therapeutics. Previously, we showed an increased urinary concentration of antimicrobial peptide LL-37 in spina bifida patients and used LL-37 to develop a mouse model of cystitis that mimics important clinical findings of IC. Here we investigate (1) the molecular mechanism of LL-37 induced cystitis in cultured human urothelial cells and in mice, (2) the protective effects of GM-0111, a modified GAG, within the context of this mechanism, (3) the physiological and molecular markers that correlate with the severity of the inflammation, and (4) the protective effects of several GAGs using these biomarkers in our LL-37 induced cystitis model. We find that LL-37 quickly induces release of ATP and apoptosis in the urothelium. These changes can be inhibited by a chemically-modified GAG, GM-0111. Furthermore, we also find that GAG analogs provide varying degrees of protection against LL-37 challenge in mice. These findings suggest that GM-0111 and possibly GAG molecules prevent the development of cystitis by blocking the apoptosis and the concurrent release of ATP from the urothelium.
Asunto(s)
Adenosina Trifosfato/metabolismo , Apoptosis/efectos de los fármacos , Catelicidinas/toxicidad , Cistitis/tratamiento farmacológico , Modelos Animales de Enfermedad , Glicosaminoglicanos/uso terapéutico , Inflamación/prevención & control , Vejiga Urinaria/efectos de los fármacos , Adyuvantes Inmunológicos/toxicidad , Animales , Péptidos Catiónicos Antimicrobianos , Cistitis/etiología , Cistitis/metabolismo , Cistitis/patología , Femenino , Glicosaminoglicanos/química , Humanos , Técnicas para Inmunoenzimas , Inflamación/etiología , Inflamación/metabolismo , Ratones , Ratones Endogámicos C57BL , Vejiga Urinaria/metabolismo , Vejiga Urinaria/patologíaRESUMEN
The antihypertensive effects of both extracts and glycosaminoglycan derived from Isaria sinclairii (IS) were investigated in a spontaneously hypertensive rat (SHR) model. Groups of rats were treated orally with 30 mg/kg each of: (1) saline control or extracts of (2) water-IS (3) methanol-IS, (4) butanol-IS, (5) ethyl acetate-IS, or (6) captopril as positive control. The 30-mg/kg dose was administered with a standard diet every day for a period of 2 wk. The antihypertensive effects of the individual extracts were in the following order: methanol > water > ethyl acetate > butanol. Glycosaminoglycan (GAG) obtained from IS as a water-soluble alcohol precipitation fraction produced an antihypertensive effect. One month following administration of GAG derived from IS to SHR animals there was a marked decrease in systolic blood pressure from 183 to 105 mm Hg and reduced diastolic blood pressure from 148 to 80 mm Hg compared to untreated control SHR rats. It was found that GAG produced an antihypertensive effect, which was more effective than the positive control captopril. In the SHR animal model a fall of 19% in body weight was observed in the group that received GAG. Data thus indicate that GAG derived from I. sinclairii may be a potent, naturally occurring antihypertensive agent.
Asunto(s)
Antihipertensivos/farmacología , Presión Sanguínea/efectos de los fármacos , Glicosaminoglicanos/farmacología , Hipertensión/tratamiento farmacológico , Hypocreales/química , Animales , Coagulación Sanguínea/efectos de los fármacos , Glicosaminoglicanos/química , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/enzimología , Humanos , Hipertensión/genética , Hipertensión/fisiopatología , Masculino , Óxido Nítrico Sintasa de Tipo III/análisis , Extractos Vegetales/farmacología , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Aumento de Peso/efectos de los fármacos , Tiempo de Coagulación de la Sangre TotalRESUMEN
Recently, a complete characterization and detailed evaluation of the glycosaminoglycans of human milk were performed. The total glycosaminoglycans content in milk from healthy mothers having delivered term or preterm newborns showed a constant pattern which was essentially composed of two main polysaccharides: chondroitin sulfate (60-70%) and heparin (30-40%). Moreover, considerable variations of glycosaminoglycans concentration were found during the first month of lactation, the highest values being present in colostrum compared to mature milk. Metabolism and potential biological functions of human milk glycosaminoglycans are hypothesized and future studies are encouraged.
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Calostro/química , Glicosaminoglicanos/química , Leche Humana/química , Leche/química , Animales , Sulfatos de Condroitina/química , Heparina/química , HumanosRESUMEN
The design of sulfated, small, nonsaccharide molecules as modulators of proteins is still in its infancy as standard drug discovery tools such as library of diverse sulfated molecules and in silico docking and scoring protocol have not been firmly established. Databases, such as ZINC, contain too few sulfate-containing nonsaccharide molecules, which severely limits the identification of new hits. Lack of a generally applicable protocol for scaffold hopping limits the development of sulfated small molecules as synthetic mimetics of the highly sulfated glycosaminoglycans. We explored a sequential ligand-based (LBVS) and structure-based virtual screening (SBVS) approach starting from our initial discovery of monosulfated benzofurans to discover alternative scaffolds as allosteric modulators of thrombin, a key coagulation enzyme. Screening the ZINC database containing nearly 1 million nonsulfated small molecules using a pharmacophore developed from the parent sulfated benzofurans followed by a genetic algorithm-based dual-filter docking and scoring screening identified a group of 10 promising hits, of which three top-scoring hits were synthesized. Each was found to selectively inhibit human alpha-thrombin suggesting the possibility of this approach for scaffold hopping. Michaelis-Menten kinetics showed allosteric inhibition mechanism for the best molecule and human plasma studies confirmed good anticoagulation potential as expected. Our simple sequential LBVS and SBVS approach is likely to be useful as a general strategy for identification of sulfated small molecules hits as modulators of glycosaminoglycan-protein interactions.