RESUMEN
The best tissue-engineered spinal cord grafts not only match the structural characteristics of the spinal cord but also allow the seed cells to grow and function in situ. Platelet-derived growth factor (PDGF) has been shown to promote the migration of bone marrow stromal cells; however, cytokines need to be released at a steady rate to maintain a stable concentration in vivo. Therefore, new methods are needed to maintain an optimal concentration of cytokines over an extended period of time to effectively promote seed cell localization, proliferation and differentiation. In the present study, a partition-type tubular scaffold matching the anatomical features of the thoracic 8-10 spinal cord of the rat was fabricated using chitosan and then subsequently loaded with chitosan-encapsulated PDGF-BB microspheres (PDGF-MSs). The PDGF-MS-containing scaffold was then examined in vitro for sustained-release capacity, biocompatibility, and its effect on neural progenitor cells differentiated in vitro from multilineage-differentiating stress-enduring cells (MUSE-NPCs). We found that pre-freezing for 2 hours at -20°C significantly increased the yield of partition-type tubular scaffolds, and 30 µL of 25% glutaraldehyde ensured optimal crosslinking of PDGF-MSs. The resulting PDGF-MSs cumulatively released 52% of the PDGF-BB at 4 weeks in vitro without burst release. The PDGF-MS-containing tubular scaffold showed suitable biocompatibility towards MUSE-NPCs and could promote the directional migration and growth of these cells. These findings indicate that the combination of a partition-type tubular scaffold, PDGF-MSs and MUSE-NPCs may be a promising model for the fabrication of tissue-engineered spinal cord grafts.
RESUMEN
Tendon injuries are a common injury of musculocutaneous system. Due to the lack of sufficient cellularity and low growth factor activity, healing of disrupted digital flexor tendon is troublesome and the process is lengthy and ineffective. bFGF and VEGFA gene were proved to be responsible and critical for promoting tendon healing. How to continuously enhance expression of these genes is a challenge. In this study, we developed a combination therapeutic approach that corrects the fundamental problem underlying intrasynovial tendon healing with introduction of growth factor genes via non-viral vector nanoparticle. PLGA nanoparticles as vehicle were used to delivery bFGF+VEGFA genes into injured tendon tissues. The expression of bFGF and VEGFA was upregulated in the tenocytes after transfection. We injected nanoparticle/bFGF+VEGFA gene complexes into injured tendons producing sufficient amounts of these factors required during early tendon healing period. After treatment, the ultimate strength of repaired tendons treated with nanoparticle/bFGF+VEGFA plasmid complexes was significantly increased, and combination therapy could also enhance flexor tendon gliding function. Therefore, combination gene therapy via nanoparticles may be an effective biological strategy for tendon repair.
Asunto(s)
Factor 2 de Crecimiento de Fibroblastos/genética , Técnicas de Transferencia de Gen , Nanopartículas/química , Tendones/patología , Factor A de Crecimiento Endotelial Vascular/genética , Cicatrización de Heridas , Animales , Fenómenos Biomecánicos , Supervivencia Celular , Pollos , Factor 2 de Crecimiento de Fibroblastos/administración & dosificación , Proteínas Fluorescentes Verdes/metabolismo , Ácido Láctico/química , Plásmidos/metabolismo , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Tenocitos/metabolismo , Adherencias Tisulares/patología , Adherencias Tisulares/terapia , Transfección , Factor A de Crecimiento Endotelial Vascular/administración & dosificaciónRESUMEN
Since neurotrophic factor is easy to degrade and aggregate, it usually has a short half-life in vitro. To overcome this shortage, neurotrophic factor has been combined with the silk fibroin (SF) membrane to realize less degradation, optimal loading efficiency, sustained release, and good adsorption. By optimizing its binding conditions, main parameters were investigated and its optimal loading efficiency was obtained. bFGF was combined to SF membrane by layer by layer (LbL) static adsorption technique. The natural and nontoxic chondroitin sulfate (CS) was used as a crosslinking agent. Optimization was carried out in three aspects: the concentration of bFGF, the concentration of CS, and the reaction time. This experiment provides a better environment for the growth of cells and offers a new kind material of absorbing neurotrophic factor to meet increasing demand for biological materials.
Asunto(s)
Factor 2 de Crecimiento de Fibroblastos/química , Fibroínas/química , Animales , Técnicas de Cultivo de Célula , Células PC12 , RatasRESUMEN
Mulberroside A is a natural polyhydroxylated stilbene compound present at relatively high abundance in the roots and twigs of Morus alba L. It is known for its nephroprotective, hypoglycemic, and antidiabetic effects. Because its metabolite, oxyresveratrol, possessed purported anti-inflammatory and neuroprotective effects, we proposed that mulberroside A may elicit neuroprotective effects that can be used in the treatment of brain ischemic injury. Therefore, we decided to investigate the pharmacological properties of mulberroside A in primary culture of rat cortical neurons after oxygen-glucose deprivation followed by reperfusion (OGD/R), evaluating its ability to counteract the hypoxia-ischemia impairment. The results showed that mulberroside A elicited neuroprotective effects comparable to nimodipine. The mechanistic studies showed that mulberroside A decreased the expressions of tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß, and IL-6 and inhibited the activation of NALP3, caspase-1, and nuclear factor-κB and the phosphorylation of extracellular signal-regulated protein kinases, the c-Jun N-terminal kinase, and p38, exhibiting anti-inflammatory antiapoptotic effects. Our results also further demonstrate that the proinflammatory cytokines of IL-1ß, IL-6, and TNF-α are promising targets for treatment of cerebral ischemic injury. Although further investigation is required for its development, all of these findings led us to speculate that mulberroside A is a candidate for the treatment of ischemic stroke, which would act as a multifactorial neuroprotectant.
Asunto(s)
Corteza Cerebral/citología , Disacáridos/farmacología , Glucosa/deficiencia , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Estilbenos/farmacología , Animales , Apoptosis/efectos de los fármacos , Proteínas Portadoras , Caspasa 1/metabolismo , Hipoxia de la Célula , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Relación Dosis-Respuesta a Droga , Embrión de Mamíferos , FN-kappa B/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR , Ratas , Ratas Sprague-Dawley , Receptores Citoplasmáticos y Nucleares/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
In the present study, oxygen-glucose deprivation followed by reperfusion (OGD/R), an in vitro model of ischemia, was used to evaluate the neuroprotective effect of isoquercetin in primary culture of rat cortical neuronal cells. It was found that isoquercetin administered prior to the insult could prevent OGD/R-induced intracellular calcium concentrations ([Ca(2+)]i) increase, lactate dehydrogenase (LDH) release and cell viability decrease. For the first time, isoquercetin is described as a neuroprotective agent that potentially explains the alleviation and prevention from OGD/R-induced injury in neurons. Mechanistic studies showed that the neuroprotective effect of isoquercetin was carried out by anti-inflammatory signaling pathway of inhibiting protein expression of toll-like receptor 4 (TLR4) and nuclear factor-kappa B (NF-κB), and mRNA expression of TNF-α and IL-6, accompanied by the anti-apoptotic signaling pathway of deactivation of extracellular-regulated kinase (ERK), Jun kinase (JNK) and p38, and inhibition of activity of caspase-3. Therefore, these studies highlighted the confirmation of isoquercetin, a flavonoid compound, as an anti-inflammation and anti-apoptosis factor which might be used as a therapeutic strategy for the ischemia/reperfusion (I/R) brain injury and related diseases.