RESUMO
Schizophyllan, a triple helical polysaccharide, exhibits cooperative order-disorder transition (CODT) in aqueous solutions. The transition transforms the ordered structure (triple helix I) formed between the branched side chains and solvent molecules into the disordered structure (triple helix II) without dissociation of the triple helix. The CODT behaviors in H2O-imidazole mixtures containing HCl with different molar ratios of imidazole/HCl were investigated by adiabatic calorimetry and differential scanning calorimetry on two schizophyllan solutions with different molar masses. The transition temperature (Tr) and the transition enthalpy (ΔHr) significantly depended on both of the mole fractions of imidazole and imidazole/HCl. The composition dependences of Tr and ΔHr in H2O-imidazole mixtures were analyzed with linear cooperative transition theory for the solvent-stabilizing effect in the mixture with active compounds. Theoretical analyses confirmed that both imidazole and imidazolium ions in the solutions competitively interact with the side chain of the triple helix.
Assuntos
Sizofirano , Varredura Diferencial de Calorimetria , Configuração de Carboidratos , Imidazóis , Sizofirano/química , Soluções , Água/químicaRESUMO
Alginate is a linear polysaccharide from brown algae consisting of 1,4-linked ß-d-mannuronic acid (M) and α-l-guluronic acid (G) arranged in M, G, and mixed MG blocks. Alginate was assumed to be indigestible in humans, but bacteria isolated from fecal samples can utilize alginate. Moreover, genomes of some human gut microbiome-associated bacteria encode putative alginate-degrading enzymes. Here, we genome-mined a polysaccharide lyase family 6 alginate lyase from the gut bacterium Bacteroides cellulosilyticus (BcelPL6). The structure of recombinant BcelPL6 was solved by X-ray crystallography to 1.3 Å resolution, revealing a single-domain, monomeric parallel ß-helix containing a 10-step asparagine ladder characteristic of alginate-converting parallel ß-helix enzymes. Substitutions of the conserved catalytic site residues Lys-249, Arg-270, and His-271 resulted in activity loss. However, imidazole restored the activity of BcelPL6-H271N to 2.5% that of the native enzyme. Molecular docking oriented tetra-mannuronic acid for syn attack correlated with M specificity. Using biochemical analyses, we found that BcelPL6 initially releases unsaturated oligosaccharides of a degree of polymerization of 2-7 from alginate and polyM, which were further degraded to di- and trisaccharides. Unlike other PL6 members, BcelPL6 had low activity on polyMG and none on polyG. Surprisingly, polyG increased BcelPL6 activity on alginate 7-fold. LC-electrospray ionization-MS quantification of products and lack of activity on NaBH4-reduced octa-mannuronic acid indicated that BcelPL6 is an endolyase that further degrades the oligosaccharide products with an intact reducing end. We anticipate that our results advance predictions of the specificity and mode of action of PL6 enzymes.
Assuntos
Bacteroides/enzimologia , Microbioma Gastrointestinal , Ácidos Hexurônicos/metabolismo , Polissacarídeo-Liases/química , Polissacarídeo-Liases/metabolismo , Alginatos/química , Alginatos/metabolismo , Bacteroides/genética , Genoma Bacteriano , Humanos , Cinética , Simulação de Acoplamento Molecular , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Mutação/genética , Estrutura Secundária de Proteína , Eletricidade Estática , Homologia Estrutural de Proteína , Relação Estrutura-Atividade , Especificidade por SubstratoRESUMO
Diblock oligosaccharides based on renewable resources allow for a range of new but, so far, little explored biomaterials. Coupling of blocks through their reducing ends ensures retention of many of their intrinsic properties that otherwise are perturbed in classical lateral modifications. Chitin is an abundant, biodegradable, bioactive, and self-assembling polysaccharide. However, most coupling protocols relevant for chitin blocks have shortcomings. Here we exploit the highly reactive 2,5-anhydro-d-mannose residue at the reducing end of chitin oligomers obtained by nitrous acid depolymerization. Subsequent activation by dihydrazides or dioxyamines provides precursors for chitin-based diblock oligosaccharides. These reactions are much faster than for other carbohydrates, and only acyclic imines (hydrazones or oximes) are formed (no cyclic N-glycosides). α-Picoline borane and cyanoborohydride are effective reductants of imines, but in contrast to most other carbohydrates, they are not selective for the imines in the present case. This could be circumvented by a simple two-step procedure. Attachment of a second block to hydrazide- or aminooxy-functionalized chitin oligomers turned out to be even faster than the attachment of the first block. The study provides simple protocols for the preparation of chitin-b-chitin and chitin-b-dextran diblock oligosaccharides without involving protection/deprotection strategies.
Assuntos
Quitina , Manose , OligossacarídeosRESUMO
BACKGROUND: Bone marrow stimulation (BMS) remains a dominant treatment strategy for symptomatic full thickness articular cartilage defects. Autologous platelet-rich plasma (PRP), may improve biological cartilage repair as an adjunct to BMS. OBJECTIVES: To assess the histological quality of cartilage repair after BMS with and without repeated local injections of PRP for the treatment of full-thickness focal chondral defects of the knee. METHODS: Two full-thickness chondral defects (Ø = 6 mm) were surgically performed in the medial and lateral trochlea of each knee in skeletally mature Göttingen minipigs. The two treatment groups with 12 defect for each groups were (1) BMS with one weekly PRP injection for 4 weeks, and (2) BMS alone. The animals were euthanized after 6 months. Samples of both whole blood and PRP were analysed with an automated hematology analyzer to determine the concentrations of platelets and nucleated cells. The composition of cartilage repair tissue was assessed using gross appearance assessment, histomorphometry and semi-quantitative scoring (ICRS II). RESULTS: The average fold increase in platelets was 10.2 ± 2.2. Leukocyte concentration increased in PRP samples by an average fold change of 7.2 ± 1.3. Our macroscopic findings showed that the defects in the BMS + PRP-treated group, were filled with an irregular, partially rough tissue similar to the BMS-treated group. No significant difference in amount of hyalin cartilage, fibrocartilage or fibrous tissue content and ICRS II scores was found between the groups. CONCLUSIONS: Four repeated local injections of leukocyte-rich PRP after BMS in the treatment of full-thickness cartilage injuries demonstrated no beneficial effects in terms of macroscopic and histological cartilage repair tissue quality.
Assuntos
Artroscopia/métodos , Células da Medula Óssea , Doenças das Cartilagens/terapia , Plasma Rico em Plaquetas , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/fisiologia , Modelos Animais de Doenças , Suínos , Porco MiniaturaRESUMO
ß-1,3-d-glucan with different degrees of branching were obtained by selectively and gradually removing side chains from schizophyllan, a water-soluble triple helical polysaccharide, using the Smith degradation. Size exclusion chromatography combined with a multi-angle light scattering detection was performed in aqueous 0.1 M NaCl. The degree of branching decreased after the Smith degradation, while the molar mass distributions were almost unchanged. The molecular conformation of the Smith-degraded ß-1,3-d-glucan was analyzed on the basis of the molar mass dependency of the radius gyration, and found to be comparable to the original triple helix of schizophyllan. Differential scanning calorimetry in deuterium oxide-hexadeuterodimethylsulfoxide mixtures was performed to investigate the effects of the degree of branching on the cooperative order-disorder transition. Removal of side chains affects both the transition temperature and transition enthalpy. The ordered structure is formed by the residual side chains in the triplex unit, so that the linear cooperative system of the triplex is maintained after the Smith degradation.
Assuntos
Sizofirano/química , beta-Glucanas/química , Varredura Diferencial de Calorimetria , Configuração de Carboidratos , Cromatografia em Gel , Difusão Dinâmica da Luz , Glucana 1,3-beta-Glucosidase/química , Glucana 1,3-beta-Glucosidase/metabolismo , Peso Molecular , Proteoglicanas , Cloreto de Sódio , Soluções/química , Termodinâmica , Água/químicaRESUMO
INTRODUCTION: Treatment of osteochondral injuries is challenging, and no gold standard has been established. Layered cell-free scaffolds are a new treatment option for these defects. The aim of this study was to evaluate the osteochondral repair in patients treated with the MaioRegen(®) scaffold, a cell-free biomimetic scaffold consisting of type I collagen and hydroxyapatite. Treatment using this scaffold has previously shown promising clinical results. METHODS: Ten patients with osteochondral lesions in the knee (n = 6) or in the talus (n = 4) were enrolled. The patients underwent pre-operative MRI and CT scans and were assessed at 1- and 2.5-year timescales post-operatively. The cartilage and bone formations were evaluated semi-quantitatively using the MOCART score. Knee patients were clinically evaluated using KOOS, subjective IKDC and Tegner scores, whereas ankle patients were evaluated using AOFAS Hindfoot and Tegner scores. RESULTS: Two patients were re-operated and excluded from further follow-up due to treatment failure. None of the patients had complete regeneration of the subchondral bone evaluated using CT. At 2.5 years, 6/8 patients had no or very limited (<10 %) bone formation in the defects and 2/8 had 50-75 % bone formation in the treated defect. MRI showed no improvement in the MOCART score at any time point. The IKDC score improved from 41.3 to 80.7, and the KOOS pain subscale improved from 63.8 to 90.8 at 2.5-year follow-up. No improvement was found with the remaining KOOS subscales, the Tegner or AOFAS Ankle-Hindfoot score. CONCLUSION: Treatment of osteochondral defects in the ankle and knee joint with a biomimetic scaffold resulted in incomplete cartilage repair and poor subchondral bone repair at 1- and 2.5-year follow-up. Clinical significant improvements were observed. These results raise serious concerns about the biological repair potential of the MaioRegen(®) scaffold, and we advise to use the MaioRegen(®) scaffold with caution. LEVEL OF EVIDENCE: Prospective therapeutic study, Level IV.
Assuntos
Articulação do Tornozelo/cirurgia , Materiais Biomiméticos , Cartilagem Articular/cirurgia , Colágeno Tipo I , Durapatita , Articulação do Joelho/cirurgia , Alicerces Teciduais , Adulto , Traumatismos do Tornozelo/diagnóstico por imagem , Traumatismos do Tornozelo/cirurgia , Articulação do Tornozelo/diagnóstico por imagem , Cartilagem/cirurgia , Cartilagem Articular/diagnóstico por imagem , Feminino , Seguimentos , Humanos , Traumatismos do Joelho/diagnóstico por imagem , Traumatismos do Joelho/cirurgia , Articulação do Joelho/diagnóstico por imagem , Imageamento por Ressonância Magnética/métodos , Masculino , Osteocondrite Dissecante/diagnóstico por imagem , Osteocondrite Dissecante/cirurgia , Osteogênese , Estudos Prospectivos , Radiografia , Procedimentos de Cirurgia Plástica , Regeneração , Tálus/cirurgia , Tomografia Computadorizada por Raios X , Adulto JovemRESUMO
The combination of modern interventional and preventive medicine has led to an epidemic of ageing. While this phenomenon is a positive consequence of an improved lifestyle and achievements in a society, the longer life expectancy is often accompanied by decline in quality of life due to musculoskeletal pain and disability. The Aarhus Regenerative Orthopaedics Symposium (AROS) 2015 was motivated by the need to address regenerative challenges in an ageing population by engaging clinicians, basic scientists, and engineers. In this position paper, we review our contemporary understanding of societal, patient-related, and basic science-related challenges in order to provide a reasoned roadmap for the future to deal with this compelling and urgent healthcare problem.
Assuntos
Envelhecimento/fisiologia , Sistema Musculoesquelético/fisiopatologia , Medicina Regenerativa/métodos , Animais , Comorbidade , Modelos Animais de Doenças , Humanos , Regeneração/fisiologiaRESUMO
Alginate-based biomaterials and medical devices are commonly subjected to γ-irradiation as a means of sterilization, either in the dry state or the gel (hydrated) state. In this process the alginate chains degrade randomly in a dose-dependent manner, altering alginates' material properties. The addition of free radical scavenging amino acids such as histidine and phenylalanine protects the alginate significantly against degradation, as shown by monitoring changes in the molecular weight distributions using SEC-MALLS and determining the pseudo first order rate constants of degradation. Tris buffer (0.5 M), but not acetate, citrate, or phosphate buffers had a similar effect on the degradation rate. Changes in pH itself had only marginal effects on the rate of alginate degradation and on the protective effect of amino acids. Contrary to previous reports, the chemical composition (M/G profile) of the alginates, including homopolymeric mannuronan, was unaltered following irradiation up to 10 kGy.
Assuntos
Alginatos/química , Aminoácidos/química , Raios gama , Alginatos/efeitos da radiação , Materiais Biocompatíveis , Soluções Tampão , Ácido Glucurônico/química , Ácido Glucurônico/efeitos da radiação , Ácidos Hexurônicos/química , Ácidos Hexurônicos/efeitos da radiação , Concentração de Íons de Hidrogênio , Peso Molecular , EsterilizaçãoRESUMO
PURPOSE: This study investigates the effect of cell seeding density on cartilage repair in matrix-assisted chondrocyte implantation in vitro and in vivo. METHODS: In vitro: Four different cell seeding densities of human chondrocytes were seeded onto a porous methoxy-polyethylene glycol-polylactic-co-glycolic acid scaffold (MPEG-PLGA) polymer scaffold ASEED™ (1.2 × 10(6), 4.0 × 10(6), 1.2 × 10(7) and 2.0 × 10(7) cells/cm(3)). The cartilage repair response was evaluated by relative gene expression of the chondrogenic markers sox9, collagen types I, II and X, and aggrecan, total DNA content and sulphated glycosaminoglycan synthesis. In vivo: Using a New Zealand white rabbit intercondylar osteochondral defect model, three different cell seeding densities (1.2 × 10(6), 4.0 × 10(6) and 1.2 × 10(7) cells/cm(3)) were tested with an empty scaffold as control. The cartilage repair response was evaluated using O'Driscoll score. RESULTS: In vitro: A significant difference (p < 0.05) in total DNA content was found at day 2 but not at day 7. The low cell seeding densities yielded the highest GAG content (p < 0.001) at day 7. Collagen type I was highest (p < 0.01) at the lowest density at day 7. In vivo: No significant difference was found between the 4 groups. CONCLUSIONS: No positive effect on cartilage repair was found using increased cell seeding density. LEVEL OF EVIDENCE: Controlled experimental study, Level II.
Assuntos
Cartilagem Articular/fisiologia , Condrócitos/transplante , Alicerces Teciduais , Cicatrização/fisiologia , Animais , Materiais Biocompatíveis , Cartilagem Articular/lesões , Contagem de Células , Técnicas de Cultura de Células , Transplante de Células , Perfilação da Expressão Gênica , Humanos , Articulação do Joelho/fisiologia , Poliésteres , Polietilenoglicóis , Coelhos , Transplante AutólogoRESUMO
Alginates are a broad family of linear (unbranched) polysaccharides derived from brown seaweeds and some bacteria. Despite having only two monomers, i.e. ß-d-mannuronate (M) and its C5 epimer α-l-guluronate (G), their blockwise arrangement in oligomannuronate (..MMM..), oligoguluronate (..GGG..), and polyalternating (..MGMG..) blocks endows it with a rather complex interaction pattern with specific counterions and salts. Classic polyelectrolyte theories well apply to alginate as polyanion in the interaction with monovalent and non-gelling divalent cations. The use of divalent gelling ions, such as Ca2+, Ba2+ or Sr2+, provides thermostable homogeneous or heterogeneous hydrogels where the block composition affects both macroscopic and microscopic properties. The mechanism of alginate gelation is still explained in terms of the original egg-box model, although over the years some novel insights have been proposed. In this review we summarize several decades of research related to structure-functionships in alginates in the presence of non-gelling and gelling cations and present some novel applications in the field of self-assembling nanoparticles and use of radionuclides.
Assuntos
Alginatos , Hidrogéis , Cátions , Substâncias Macromoleculares , MetaisRESUMO
Alginates in the format of scaffolds provide important functions as materials for cell encapsulation, drug delivery, tissue engineering and wound healing among others. The method for preparation of alginate-based foams presented here is based on homogeneous, ionotropic gelation of aerated alginate solutions, followed by air drying. The method allows higher flexibility and better control of the pore structure, hydration properties and mechanical integrity compared to foams prepared by other techniques. The main variables for tailoring hydrogel properties include operational parameters such as degree of aeration and mixing times and concentration of alginate, as well as macromolecular properties such as the type of alginate (chemical composition and molecular weight distribution). Exposure of foams to γ-irradiation resulted in a dose-dependent (0-30 kGy) reduction in molecular weight of the alginate and a corresponding reduction in tensile strength of the foams.
Assuntos
Alginatos/química , Portadores de Fármacos , Géis/química , Engenharia Tecidual , Alicerces Teciduais , Materiais Biocompatíveis/química , Biopolímeros/química , Hidrogel de Polietilenoglicol-Dimetacrilato/química , Peso Molecular , Resistência à Tração , CicatrizaçãoRESUMO
BACKGROUND: Excess peroxisome proliferator-activated receptor (PPAR) stimulation has been associated with detrimental health effects including impaired myocardial function. Recently, supplementation with n-3 polyunsaturated fatty acids (PUFA) has been associated with improved left ventricular function and functional capacity in patients with dilated cardiomyopathy. We investigated the long-term effects of the pan-PPAR agonist tetradecylthioacetic acid (TTA) and/or high-dose fish oil (FO) on cardiac fatty acid (FA) composition and lipid metabolism. Male Wistar rats were given one out of four different 25% (w/v) fat diets: control diet; TTA diet; FO diet; or diet containing both TTA and FO. RESULTS: After 50 weeks n-3 PUFA levels were increased by TTA and FO in the heart, whereas liver levels were reduced following TTA administration. TTA was associated with a decrease in arachidonic acid, increased activities of carnitine palmitoyltransferase II, fatty acyl-CoA oxidase, glycerol-3-phosphate acyltransferase, and fatty acid synthase in the heart. Furthermore, cardiac Ucp3 and Cact mRNA was upregulated. CONCLUSIONS: Long-term treatment with the pan-PPAR agonist TTA or high-dose FO induced marked changes in PUFA composition and enzymatic activity involved in FA metabolism in the heart, different from liver. Changes included increased FA oxidation and a selective increase in cardiac n-3 PUFA.
Assuntos
Ácidos Graxos Ômega-3/metabolismo , Óleos de Peixe/administração & dosagem , Miocárdio/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/agonistas , Sulfetos/administração & dosagem , Acil-CoA Oxidase/metabolismo , Animais , Carnitina O-Palmitoiltransferase/genética , Carnitina O-Palmitoiltransferase/metabolismo , Óleos de Peixe/farmacologia , Expressão Gênica , Glicerol-3-Fosfato O-Aciltransferase/metabolismo , Canais Iônicos/genética , Canais Iônicos/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/metabolismo , Masculino , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Miocárdio/enzimologia , Especificidade de Órgãos , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Ratos , Ratos Wistar , Sulfetos/farmacologia , Fatores de Tempo , Proteína Desacopladora 3Assuntos
Tempestades Ciclônicas , Desastres , Meteorologia , Terminologia como Assunto , Feminino , Humanos , MasculinoRESUMO
PURPOSE: To develop a nano-structured porous polycaprolactone (NSP-PCL) scaffold and compare the articular cartilage repair potential with that of a commercially available collagen type I/III (Chondro-Gide) scaffold. METHODS: By combining rapid prototyping and thermally induced phase separation, the NSP-PCL scaffold was produced for matrix-assisted autologous chondrocyte implantation. Lyophilizing a water-dioxane-PCL solution created micro and nano-pores. In vitro: The scaffolds were seeded with rabbit chondrocytes and cultured in hypoxia for 6 days. qRT-PCR was performed using primers for sox9, aggrecan, collagen type 1 and 2. In vivo: 15 New Zealand White Rabbits received bilateral osteochondral defects in the femoral intercondylar grooves. Autologous chondrocytes were harvested 4 weeks prior to surgery. There were 3 treatment groups: (1) NSP-PCL scaffold without cells. (2) The Chondro-Gide scaffold with autologous chondrocytes and (3) NSP-PCL scaffold with autologous chondrocytes. Observation period was 13 weeks. Histological evaluation was made using the O'Driscoll score. RESULTS: In vitro: The expressions of sox9 and aggrecan were higher in the NSP-PCL scaffold, while expression of collagen 1 was lower compared to the Chondro-Gide scaffold. In vivo: Both NSP-PCL scaffolds with and without cells scored significantly higher than the Chondro-Gide scaffold when looking at the structural integrity and the surface regularity of the repair tissue. No differences were found between the NSP-PCL scaffold with and without cells. CONCLUSION: The NSP-PCL scaffold demonstrated higher in vitro expression of chondrogenic markers and had higher in vivo histological scores compared to the Chondro-Gide scaffold. The improved chondrocytic differentiation can potentially produce more hyaline cartilage during clinical cartilage repair. It appears to be a suitable cell-free implant for hyaline cartilage repair and could provide a less costly and more effective treatment option than the Chondro-Gide scaffold with cells.
Assuntos
Cartilagem Articular/fisiologia , Colágeno Tipo III , Colágeno Tipo I , Regeneração Tecidual Guiada/instrumentação , Nanoestruturas , Poliésteres , Alicerces Teciduais , Agrecanas/metabolismo , Animais , Biomarcadores/metabolismo , Cartilagem Articular/citologia , Cartilagem Articular/lesões , Condrócitos/transplante , Colágeno Tipo I/metabolismo , Colágeno Tipo III/metabolismo , Masculino , Modelos Animais , Coelhos , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição SOX9/metabolismo , CicatrizaçãoRESUMO
Sequential periodate-chlorite oxidation of sodium hyaluronate to 2,3-dicarboxylated hyaluronate (DCH), a novel biocompatible and highly functionalized derivative bearing additional pair of COOH groups at C2 and C3 carbons of oxidized á´ -glucuronic acid units, is investigated. The impact of various reaction parameters (time, oxidizer concentration, and molar amount) on DCH's composition, molecular weight, degree of oxidation, and cytotoxicity are investigated to guide the synthesis of DCH derivatives of desired properties. Subsequently, fully (99%) and partially (70%) oxidized DCH derivatives were compared to untreated sodium hyaluronate in terms of anticancer drug cisplatin loading efficacy, carrier capacity, drug release rates, and cytotoxicity towards healthy and cancerous cell lines. DCH derivatives were found to be superior in every aspect, having nearly twice the carrier capacity, significantly slower release rates, and higher efficacy. DCH is thus a highly interesting hyaluronate derivative with an adjustable degree of oxidation, molecular weight, and great potential for further modifications.
Assuntos
Glicosaminoglicanos , Ácido Hialurônico , Liberação Controlada de Fármacos , Peso Molecular , OxirreduçãoRESUMO
PURPOSE: To investigate the effect of bone-marrow stimulation (BMS) on subchondral bone plate morphology and remodeling compared to untreated subchondral bone in a validated minipig model. METHODS: Three Göttingen minipigs received BMS with drilling as treatment for two chondral defects in each knee. The animals were euthanized after six months. Follow-up consisted of a histological semiquantitative evaluation using a novel subchondral bone scoring system and micro computed tomography (µCT) of the BMS subchondral bone. The histological and microstructural properties of the BMS-treated subchondral bone were compared to that of the adjacent healthy subchondral bone. RESULTS: The µCT analysis showed that subchondral bone treated with BMS had significantly higher connectivity density compared to adjacent untreated subchondral bone (26 1/mm3 vs. 21 1/mm3, P = 0.048). This was the only microstructural parameter showing a significant difference. The histological semiquantitative score differed significantly between the subchondral bone treated with BMS and the adjacent untreated subchondral (8.0 vs. 10 P = < 0.001). Surface irregularities were seen in 43% and bone overgrowth in 27% of the histological sections. Only sparse formation of bone cysts was detected (1%). CONCLUSIONS: BMS with drilling does not cause extensive changes to the subchondral bone microarchitecture. Furthermore, the morphology of BMS subchondral bone resembled that of untreated subchondral bone with almost no formation of bone cyst, but some surface irregularities and bone overgrowth.
Assuntos
Medula Óssea , Cartilagem Articular , Animais , Placas Ósseas , Cartilagem Articular/fisiologia , Cartilagem Articular/cirurgia , Suínos , Porco Miniatura , Microtomografia por Raio-XRESUMO
Engineered block polysaccharides is a relatively new class of biomacromolecules consisting of chemical assembly of separate block structures at the chain termini. In contrast to conventional, laterally substituted polysaccharide derivatives, the block arrangement allows for much higher preservation of inherent chain properties such as biodegradability and stimuli-responsive self-assembly, while at the same time inducing new macromolecular properties. Abundant, carbon neutral, and even recalcitrant biomass is an excellent source of blocks, opening for numerous new uses of biomass for a wide range of novel biomaterials. Among a limited range of methodologies available for block conjugation, bifunctional linkers allowing for oxyamine and hydrazide 'click' reactions have recently proven useful additions to the repertoire. This article focuses the chemistry and kinetics of these reactions. It also presents some new data with the aim to provide useful protocols and methods for general use towards new block polysaccharides.
Assuntos
Aminas/farmacologia , Hidrazonas/farmacologia , Polissacarídeos/antagonistas & inibidores , Aminas/química , Configuração de Carboidratos , Química Click , Hidrazonas/químicaRESUMO
Animals and higher plants express endogenous peptide antibiotics called defensins. These small cysteine-rich peptides are active against bacteria, fungi and viruses. Here we describe plectasin-the first defensin to be isolated from a fungus, the saprophytic ascomycete Pseudoplectania nigrella. Plectasin has primary, secondary and tertiary structures that closely resemble those of defensins found in spiders, scorpions, dragonflies and mussels. Recombinant plectasin was produced at a very high, and commercially viable, yield and purity. In vitro, the recombinant peptide was especially active against Streptococcus pneumoniae, including strains resistant to conventional antibiotics. Plectasin showed extremely low toxicity in mice, and cured them of experimental peritonitis and pneumonia caused by S. pneumoniae as efficaciously as vancomycin and penicillin. These findings identify fungi as a novel source of antimicrobial defensins, and show the therapeutic potential of plectasin. They also suggest that the defensins of insects, molluscs and fungi arose from a common ancestral gene.
Assuntos
Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Fungos/química , Sequência de Aminoácidos , Animais , Antibacterianos/química , Clonagem Molecular , DNA Complementar/genética , Defensinas/química , Modelos Animais de Doenças , Fungos/genética , Bactérias Gram-Positivas/efeitos dos fármacos , Bactérias Gram-Positivas/fisiologia , Humanos , Camundongos , Dados de Sequência Molecular , Peptídeos , Infecções Pneumocócicas/tratamento farmacológico , Infecções Pneumocócicas/microbiologia , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/farmacologiaRESUMO
Most polysaccharides used in polysaccharide-based block copolymers are attached to the second block through the reducing end, due to the few and highly polysaccharide specific non-reducing end (NRE) functionalisation methods available. Chitin oligomers, prepared by nitrous acid degradation of chitosan (AnM) can, however, be selectively oxidised by periodate since they only possess a single vicinal diol in the NRE residue. Here, we show that both aldehydes formed after oxidation are highly reactive towards bifunctional oxyamines and hydrazide linkers. Sub-stochiometric amounts of linkers resulted in conjugation of AnM oligomers through both chain termini to yield a discrete distribution of 'polymerised' oligomers. Such chitin-based block polymers were, in contrast to chitins of the same chain lengths, water-soluble. Oxidised AnM oligomers, functionalised at both termini can also enable the preparation of more complex block polysaccharides such as ABA- or ABC-type.
Assuntos
Quitina/química , Ácido Periódico/química , Água/química , Adipatos/química , Aldeídos/síntese química , Aldeídos/química , Sequência de Carboidratos , Quitina/síntese química , Hidroxilaminas/química , Manose/análogos & derivados , Manose/química , Oxirredução , SolubilidadeRESUMO
OBJECTIVE: Focal cartilage injuries are debilitating and difficult to treat. Biological cartilage repair procedures are used for patients younger than 40 years, and knee arthroplasties are generally reserved for patients older than 60 years. Resurfacing implants are well suited for patients in this treatment gap. The objective was to investigate the 10-year survival of resurfacing implants in the Danish Knee Arthroplasty Registry. DESIGN: In this retrospective cohort study, patients treated with resurfacing implants were followed longitudinally in the Danish Knee Arthroplasty Registry from 1997 to 2020. The primary endpoint was revision surgery. The survival of the resurfacing implants was analyzed by Kaplan-Meier method. RESULTS: A total of 379 resurfacing implant procedures were retrieved from the Danish Knee Arthroplasty Registry. The mean age and weight of patients were 50 years (SD = 11) and 84 kg (SD = 17), respectively. The indications for surgery were as follows: secondary osteoarthritis (42%), primary osteoarthritis (32%), and osteochondral lesions (20%). Within the follow-up period, 70 (19%) of the implants were revised to arthroplasties. The 1-, 5-, and 10-year revision-free survival estimation was 0.95 (95% CI 0.93-0.97), 0.84 (95% CI 0.80-0.88), and 0.80 (95% CI 0.75-0.84), respectively. The median time to revision was 2 years. CONCLUSION: The 10-year revision-free survival rate for resurfacing implants was 80%. Based on the revision rates, this treatment offers a viable alternative to biological cartilage repair methods in patients aged 40 to 60 years with focal cartilage pathology. Improved patient selection could further improve the implant survival rate. Further studies are needed to investigate this treatment method.