ABSTRACT
Mesenchymal stem cell (MSC) therapy shows promise in regenerative medicine. For osteoarthritis (OA), MSCs delivered to the joint have a temporal window in which they can secrete growth factors and extracellular matrix molecules, contributing to cartilage regeneration and cell proliferation. However, upon injection in the non-vascularized joint, MSCs lacking energy supply, starve and die too quickly to efficiently deliver enough of these factors. To feed injected MSCs, we developed a hyaluronic acid (HA) derivative, where glucose is covalently bound to hyaluronic acid. To achieve this, the glucose moiety in 4-aminophenyl-ß-D-glucopyranoside was linked to the HA backbone through amidation. The hydrogel was able to deliver glucose in a controlled manner using a trigger system based on hydrolysis catalyzed by endogenous ß-glucosidase. This led to glucose release from the hyaluronic acid backbone inside the cell. Indeed, our hydrogel proved to rescue starvation and cell mortality in a glucose-free medium. Our approach of adding a nutrient to the polymer backbone in hydrogels opens new avenues to deliver stem cells in poorly vascularized, nutrient-deficient environments, such as osteoarthritic joints, and for other regenerative therapies.
ABSTRACT
Osteoarthritis is the most common chronic joint disease and a major health care concern due to the lack of efficient treatments. This is mainly related to the local and degenerative nature of this disease. Kartogenin was recently reported as a disease-modifying osteoarthritis drug that promotes cartilage repair, but its therapeutic effect is impeded by its very low solubility. Therefore, we designed a unique nanocrystal-chitosan particle intra-articular delivery system for osteoarthritis treatment that merges the following formulation techniques: nanosize reduction of a drug by wet milling and spray drying. The intermediate formulation (kartogenin nanocrystals) increased the solubility and dissolution rates of kartogenin. The final drug delivery system consisted of an easily resuspendable and ready-to-use microsphere powder for intra-articular injection. Positively charged chitosan microspheres with a median size of approximately 10 µm acted as a mothership drug delivery system for kartogenin nanocrystals in a simulated intra-articular injection. The microspheres showed suitable stability and a controlled release profile in synovial fluid and were nontoxic in human synoviocytes. The cartilage retention skills of the microspheres were also explored ex vivo using cartilage. This drug delivery system shows promise for advancement to preclinical stages in osteoarthritis therapy and scale-up production.
Subject(s)
Anilides , Chitosan , Nanoparticles , Osteoarthritis , Phthalic Acids , Humans , Chitosan/chemistry , Pharmaceutical Preparations , Osteoarthritis/drug therapy , Drug Delivery Systems , Nanoparticles/chemistry , Injections, Intra-Articular , MicrospheresABSTRACT
The Rad5/HLTF protein has a central role in the tolerance to DNA damage by mediating an error-free mode of bypassing unrepaired DNA lesions, and is therefore critical for the maintenance of genome stability. We show in this work that, following cellular stress, Rad5 is regulated by relocalization into two types of nuclear foci that coexist within the same cell, which we termed 'S' and 'I'. Rad5 S-foci form in response to genotoxic stress and are associated with Rad5's function in maintaining genome stability, whereas I-foci form in the presence of proteotoxic stress and are related to Rad5's own proteostasis. Rad5 accumulates into S-foci at DNA damage tolerance sites by liquid-liquid phase separation, while I-foci constitute sites of chaperone-mediated sequestration of Rad5 at the intranuclear quality control compartment (INQ). Relocalization of Rad5 into each type of foci involves different pathways and recruitment mechanisms, but in both cases is driven by the evolutionarily conserved E2 ubiquitin-conjugating enzyme Rad6. This coordinated differential relocalization of Rad5 interconnects DNA damage response and proteostasis networks, highlighting the importance of studying these homeostasis mechanisms in tandem. Spatial regulation of Rad5 under cellular stress conditions thus provides a useful biological model to study cellular homeostasis as a whole.
Subject(s)
DNA Helicases , Saccharomyces cerevisiae Proteins , Saccharomyces cerevisiae , Humans , DNA Damage , DNA Damage Tolerance , DNA Helicases/genetics , DNA Repair , DNA Replication , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Genomic Instability , Proteostasis/genetics , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/metabolism , Transcription Factors/metabolism , Ubiquitin-Conjugating Enzymes/genetics , Ubiquitin-Conjugating Enzymes/metabolismABSTRACT
Thermo-responsive hyaluronan-based hydrogels and FE002 human primary chondroprogenitor cell sources have both been previously proposed as modern therapeutic options for the management of osteoarthritis (OA). For the translational development of a potential orthopedic combination product based on both technologies, respective technical aspects required further optimization phases (e.g., hydrogel synthesis upscaling and sterilization, FE002 cytotherapeutic material stabilization). The first aim of the present study was to perform multi-step in vitro characterization of several combination product formulas throughout the established and the optimized manufacturing workflows, with a strong focus set on critical functional parameters. The second aim of the present study was to assess the applicability and the efficacy of the considered combination product prototypes in a rodent model of knee OA. Specific characterization results (i.e., spectral analysis, rheology, tribology, injectability, degradation assays, in vitro biocompatibility) of hyaluronan-based hydrogels modified with sulfo-dibenzocyclooctyne-PEG4-amine linkers and poly(N-isopropylacrylamide) (HA-L-PNIPAM) containing lyophilized FE002 human chondroprogenitors confirmed the suitability of the considered combination product components. Specifically, significantly enhanced resistance toward oxidative and enzymatic degradation was shown in vitro for the studied injectable combination product prototypes. Furthermore, extensive multi-parametric (i.e., tomography, histology, scoring) in vivo investigation of the effects of FE002 cell-laden HA-L-PNIPAM hydrogels in a rodent model revealed no general or local iatrogenic adverse effects, whereas it did reveal some beneficial trends against the development of knee OA. Overall, the present study addressed key aspects of the preclinical development process for novel biologically-based orthopedic combination products and shall serve as a robust methodological basis for further translational investigation and clinical work.
ABSTRACT
Hyaluronic acid (HA) constitutes a versatile chemical framework for the development of osteoarthritis pain treatment by means of injection in the joints, so-called viscosupplementation. Without appropriate physico-chemical tuning, such preparations are inherently hindered by prompt in vivo degradation, mediated by hyaluronidases and oxidative stress. To prolong hydrogel residence time and confer optimized product functionality, novel thermoresponsive nanoforming HA derivatives were proposed and characterized. Combined use of sulfo-dibenzocyclooctyne-PEG4-amine linkers and poly(N-isopropylacrylamide) in green chemistry process enabled the synthesis of HA-based polymers, with in situ obtention of appropriate viscoelastic properties. Spontaneous and reversible thermoformation of nanoparticles above 30 °C was experimentally confirmed. Lead formulations were compared to a commercially available HA-based product and shown significantly better in vitro resistance to enzymatic and oxidative degradation, required half the injection force with optimal viscoelastic hydrogel properties in equine synovial fluids. Results highlighted the vast potential of appropriately engineered HA-based systems as next-generation long-acting viscosupplementation products for osteoarthritic patients.
ABSTRACT
Osteoarthritis (OA) is a chronic and inflammatory disease with no effective regenerative treatments to date. The therapeutic potential of mesenchymal stem cells (MSCs) remains to be fully explored. Intra-articular injection of these cells promotes cartilage protection and regeneration by paracrine signaling and differentiation into chondrocytes. However, joints display a harsh avascular environment for these cells upon injection. This phenomenon prompted researchers to develop suitable injectable materials or systems for MSCs to enhance their function and survival. Among them, hydrogels can absorb a large amount of water and maintain their 3D structure but also allow incorporation of bioactive agents or small molecules in their matrix that maximize the action of MSCs. These materials possess advantageous cartilage-like features such as collagen or hyaluronic acid moieties that interact with MSC receptors, thereby promoting cell adhesion. This review provides an up-to-date overview of the progress and opportunities of MSCs entrapped into hydrogels, combined with bioactive/small molecules to improve the therapeutic effects in OA treatment.
Subject(s)
Cartilage, Articular , Mesenchymal Stem Cells , Osteoarthritis , Cartilage, Articular/metabolism , Chondrocytes , Humans , Hydrogels/chemistry , Mesenchymal Stem Cells/metabolism , Osteoarthritis/drug therapyABSTRACT
Se realizó una investigación en sistemas y servicios de salud para evaluar la calidad del proceso de ejecución del Programa de Riesgo Preconcepcional a través de la competencia profesional de 45 médicos y enfermeras del Policlínico Ramón López Peña de Santiago de Cuba, desde abril hasta noviembre del 2016. Un comité de expertos preestableció criterios, indicadores y estándares de evaluación. Hubo dificultades en el conocimiento sobre criterios de control, salida y fallo del programa, en el seguimiento de las parejas, así como en lo concerniente a las historias clínicas y la interconsulta con obstetra u otros especialistas afines al factor de riesgo. La calidad en la ejecución del programa no fue la mejor; las dificultades detectadas pudieran incidir en los resultados negativos del Programa de Atención Materno-Infantil en el área de salud. Resulta necesario evaluar otras dimensiones de la calidad y capacitar a los profesionales en los aspectos con dificultades(AU)
An investigation in health systems and services was carried out to evaluate the process quality in the implementation of the Preconceptional Risk Program by means of the professional competence of 45 doctors and nurses of Ramón López Peña Polyclinic in Santiago de Cuba, from April to November, 2016. An experts committee prestablished evaluation approaches, indicators and standards. There were difficulties in the knowledge about control, emission and failure of the program approaches, in the follow up of couples, as well as in everything concerning the medical records and the consultation with the obstetrician or other specialists related to the risk factor. The quality in the implementation of the program was not the best; the difficulties detected could impact in the negative results of the Maternal and Child Care Program in the health area. It is necessary to evaluate other dimensions of the quality and qualify the professionals in the aspects with difficulties(AU)
Subject(s)
Humans , Male , Female , Reproductive Behavior , Reproductive Health , Preconception Care , Professional Competence , Risk FactorsABSTRACT
Se realizó una investigación en sistemas y servicios de salud para evaluar la calidad del proceso de ejecución del Programa de Riesgo Preconcepcional a través de la competencia profesional de 45 médicos y enfermeras del Policlínico Ramón López Peña de Santiago de Cuba, desde abril hasta noviembre del 2016. Un comité de expertos preestableció criterios, indicadores y estándares de evaluación. Hubo dificultades en el conocimiento sobre criterios de control, salida y fallo del programa, en el seguimiento de las parejas, así como en lo concerniente a las historias clínicas y la interconsulta con obstetra u otros especialistas afines al factor de riesgo. La calidad en la ejecución del programa no fue la mejor; las dificultades detectadas pudieran incidir en los resultados negativos del Programa de Atención Materno-Infantil en el área de salud. Resulta necesario evaluar otras dimensiones de la calidad y capacitar a los profesionales en los aspectos con dificultades
An investigation in health systems and services was carried out to evaluate the process quality in the implementation of the Preconceptional Risk Program by means of the professional competence of 45 doctors and nurses of Ramón López Peña Polyclinic in Santiago de Cuba, from April to November, 2016. An experts committee prestablished evaluation approaches, indicators and standards. There were difficulties in the knowledge about control, emission and failure of the program approaches, in the follow up of couples, as well as in everything concerning the medical records and the consultation with the obstetrician or other specialists related to the risk factor. The quality in the implementation of the program was not the best; the difficulties detected could impact in the negative results of the Maternal and Child Care Program in the health area. It is necessary to evaluate other dimensions of the quality and qualify the professionals in the aspects with difficulties
