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OBJECTIVE: The use of intrawound vancomycin powder in spine surgery has been shown to decrease the rate of surgical site infections; however, the optimal dose is unknown. High-dose vancomycin inhibits osteoblast proliferation in vitro and may decrease the rate of solid arthrodesis. Bone marrow-derived mesenchymal stem cells (BMSCs) are multipotent cells that are a source of osteogenesis in spine fusions. The purpose of this study was to determine the effects of vancomycin on rat BMSC viability and differentiation in vitro. METHODS: BMSCs were isolated from the femurs of immature female rats, cultured, and then split into two equal groups; half were treated to stimulate osteoblastic differentiation and half were not. Osteogenesis was stimulated by the addition of 50 µg/mL l-ascorbic acid, 10 mM ß-glycerol phosphate, and 0.1 µM dexamethasone. Vancomycin was added to cell culture medium at concentrations of 0, 0.04, 0.4, or 4 mg/mL. Early differentiation was determined by alkaline phosphatase activity (4 days posttreatment) and late differentiation by alizarin red staining for mineralization (9 days posttreatment). Cell viability was determined at both the early and late time points by measurement of formazan colorimetric product. RESULTS: Viability within the first 4 days decreased with high-dose vancomycin treatment, with cells receiving 4 mg/mL vancomycin having 40%-60% viability compared to the control. A gradual decrease in alizarin red staining and nodule formation was observed with increasing vancomycin doses. In the presence of the osteogenic factors, vancomycin did not have deleterious effects on alkaline phosphatase activity, whereas a trend toward reduced activity was seen in the absence of osteogenic factors when compared to osteogenically treated cells. CONCLUSIONS: Vancomycin reduced BMSC viability and impaired late osteogenic differentiation with high-dose treatment. Therefore, the inhibitory effects of high-dose vancomycin on spinal fusion may result from both reduced BMSC viability and some impairment of osteogenic differentiation.
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Phosphorene, also known as black phosphorus (BP), is a two-dimensional (2D) material that has gained significant attention in several areas of current research. Its unique properties such as outstanding surface activity, an adjustable bandgap width, favorable on/off current ratios, infrared-light responsiveness, good biocompatibility, and fast biodegradation differentiate this material from other two-dimensional materials. The application of BP in the biomedical field has been rapidly emerging over the past few years. This article aimed to provide a comprehensive review of the recent progress on the unique properties and extensive medical applications for BP in bone, nerve, skin, kidney, cancer, and biosensing related treatment. The details of applications of BP in these fields were summarized and discussed.
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Nanotubos de Carbono , Neoplasias , Pontos Quânticos , Osso e Ossos , Humanos , FósforoRESUMO
Trauma, surgery, and other inflammatory conditions can lead to debilitating joint contractures. Adjunct pharmacologic modalities may permit clinical prevention and treatment of recalcitrant joint contractures. We investigated the therapeutic potential of rosiglitazone by intra-articular delivery via oligo[poly(ethylene glycol)fumarate] (OPF) hydrogels in an established rabbit model of arthrofibrosis. OPF hydrogels loaded with rosiglitazone were characterized for drug elution properties upon soaking in minimum essential media (MEM) with 10% fetal bovine serum and measurements of drug concentrations via High Performance Liquid Chromatography (HPLC). Drug-loaded scaffolds were surgically implanted into 24 skeletally mature female New Zealand White rabbits that were divided into equal groups receiving OPF hydrogels loaded with rosiglitazone (1.67 mg), or vehicle control (10 µl DMSO). After 8 weeks of joint immobilization, rabbits were allowed unrestricted cage activity for 16 weeks. Contracture angles of rabbit limbs treated with rosiglitazone showed statistically significant improvements in flexion compared to control animals (mean angles, respectively, 64.4° vs. 53.3°, p < 0.03). At time of sacrifice (week 24), animals in the rosiglitazone group continued to exhibit less joint contracture than controls (119.0° vs. 99.5°, p = 0.014). The intra-articular delivery of rosiglitazone using implanted OPF hydrogels decreases flexion contractures in a rabbit model of arthrofibrosis without causing adverse effects (e.g., gross inflammation or arthritis). Statement of Clinical Significance: Post-traumatic joint contractures are common and debilitating, with limited available treatment options. Pharmacologic interventions can potentially prevent and treat such contractures. This study is translational in that a commercially approved medication has been repurposed through a novel delivery device. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2949-2955, 2018.
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Contratura/tratamento farmacológico , Hipoglicemiantes/administração & dosagem , Rosiglitazona/administração & dosagem , Alicerces Teciduais , Animais , Células Cultivadas , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Feminino , Fibroblastos/efeitos dos fármacos , Fibrose , Humanos , Poliésteres , Polietilenoglicóis , CoelhosRESUMO
The purpose of this study was to develop and validate a screening method based on scintillation probes for the simultaneous evaluation of in vivo growth factor release profiles of multiple implants in the same animal. First, we characterized the scintillation probes in a series of in vitro experiments to optimize the accuracy of the measurement setup. The scintillation probes were found to have a strong geometric dependence and experience saturation effects at high activities. In vitro simulation of 4 subcutaneous limb implants in a rat showed minimal interference of surrounding implants on local measurements at close to parallel positioning of the probes. These characteristics were taken into consideration for the design of the probe setup and in vivo experiment. The measurement setup was then validated in a rat subcutaneous implantation model using 4 different sustained release carriers loaded with (125)I-BMP-2 per animal. The implants were removed after 42 or 84 days of implantation, for comparison of the non-invasive method to ex vivo radioisotope counting. The non-invasive method demonstrated a good correlation with the ex vivo counting method at both time-points of all 4 carriers. Overall, this study showed that scintillation probes could be successfully used for paired measurement of 4 release profiles with minimal interference of the surrounding implants, and may find use as non-invasive screening tools for various drug delivery applications.
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Implantes Absorvíveis , Proteínas Morfogenéticas Ósseas , Sistemas de Liberação de Medicamentos/métodos , Fêmur , Fator de Crescimento Transformador beta , Animais , Materiais Biocompatíveis/química , Proteína Morfogenética Óssea 2 , Proteínas Morfogenéticas Ósseas/química , Proteínas Morfogenéticas Ósseas/farmacocinética , Proteínas Morfogenéticas Ósseas/farmacologia , Preparações de Ação Retardada , Avaliação Pré-Clínica de Medicamentos/instrumentação , Avaliação Pré-Clínica de Medicamentos/métodos , Fêmur/efeitos dos fármacos , Fêmur/metabolismo , Câmaras gama , Humanos , Radioisótopos do Iodo , Masculino , Ratos , Ratos Endogâmicos , Proteínas Recombinantes/química , Proteínas Recombinantes/farmacocinética , Proteínas Recombinantes/farmacologia , Distribuição Tecidual , Fator de Crescimento Transformador beta/química , Fator de Crescimento Transformador beta/farmacocinética , Fator de Crescimento Transformador beta/farmacologiaRESUMO
BACKGROUND: Others have reported significant changes in red blood cell (RBC) transfusion practice during the past two decades during knee, hip, prostate, and carotid surgery. Similar data for patients undergoing major spine surgery, however, are not available. STUDY DESIGN AND METHODS: After institutional review board approval, adult patients undergoing elective major spine surgery were stratified into one of two transfusion-related groups: 1) 1980 to 1985 (i.e., before human immunodeficiency virus screening, early practice group; n = 699) or 2) 1995 to 2000 (i.e., late practice group; n = 610). RESULTS: Patients in the late practice group were older, had greater numbers of preoperative coexisting diseases (e.g., hypertension, cardiac dysrhythmias, coronary artery disease, prior myocardial infarction, diabetes mellitus, renal disease, cerebrovascular disease, and asthma), and were exposed to longer operations (p < 0.01 for each variable). Over time, allogeneic RBC administration significantly decreased, whereas autologous and intraoperative autotransfusion significantly increased. Compared to the early practice group, all perioperative Hb concentrations were significantly lower than the late practice group, yet no significant difference in major morbidity or mortality was observed between groups. CONCLUSION: In this retrospective analysis, significantly lower acceptable perioperative Hb concentrations were observed in older patients having substantially worse baseline comorbidity and exposed to longer major spine operations, without significant change in the incidence of perioperative morbidity or mortality.
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Transfusão de Sangue Autóloga , Bases de Dados como Assunto , Procedimentos Cirúrgicos Eletivos , Transfusão de Eritrócitos , Procedimentos Neurocirúrgicos , Coluna Vertebral/cirurgia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Transfusão de Sangue Autóloga/tendências , Procedimentos Cirúrgicos Eletivos/mortalidade , Procedimentos Cirúrgicos Eletivos/tendências , Transfusão de Eritrócitos/mortalidade , Transfusão de Eritrócitos/tendências , Feminino , Humanos , Prática Institucional/tendências , Masculino , Pessoa de Meia-Idade , Procedimentos Neurocirúrgicos/mortalidade , Procedimentos Neurocirúrgicos/tendências , Estudos RetrospectivosRESUMO
In this study, we investigated the effect of signaling peptides incorporated into oligo(poly(ethylene glycol) fumarate) (OPF) hydrogels on in vitro differentiation and mineralization of marrow stromal cells (MSCs) cultured in media without soluble osteogenic supplements (dexamethasone and beta-glycerol phosphate). When MSCs were cultured for 16 days on OPF hydrogels modified with Arg-Gly-Asp (RGD) containing peptides, the normalized cell number was dependent on the peptide concentration between days 0 and 5 and reached comparable values at day 10 regardless of the concentration. The alkaline phosphatase (ALP) activity of MSCs on the peptide-modified OPF hydrogels was also concentration-dependent: ALP activity showed peaks on day 10 or day 13 on OPF hydrogels modified with 2.0 and 1.0 micromol peptide/g, which were significantly greater than those on the OPF hydrogels modified with 0.1 micromol peptides/g or no peptide. A characteristic marker of osteoblastic differentiation, osteopontin (OPN), was detected for all the test groups. However, OPN secretion between days 0 and 10 was significantly higher on the peptide modified hydrogels compared to that on tissue culture-treated polystyrene. Taken together, the results indicate that the presence of signaling peptide allows for a favorable microenvironment for MSCs to differentiate into osteoblasts and produce mineralized matrix, although the soluble factors may further enhance calcium deposition. These findings further support the usefulness of OPF hydrogels as scaffolds for guided bone regeneration, and represent an initial step in exploring the complex relationship between soluble and insoluble factors in osteogenic differentiation on biodegradable materials.
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Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Oligopeptídeos/administração & dosagem , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Engenharia Tecidual/métodos , Implantes Absorvíveis , Adsorção , Animais , Células da Medula Óssea/fisiologia , Calcificação Fisiológica/efeitos dos fármacos , Calcificação Fisiológica/fisiologia , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/fisiologia , Células Cultivadas , Materiais Revestidos Biocompatíveis/administração & dosagem , Materiais Revestidos Biocompatíveis/química , Dexametasona/administração & dosagem , Glicerofosfatos/administração & dosagem , Hidrogéis/química , Masculino , Teste de Materiais , Osteoblastos/fisiologia , Osteogênese/fisiologia , Ligação Proteica , Ratos , Ratos Wistar , Células Estromais/citologia , Células Estromais/efeitos dos fármacos , Células Estromais/fisiologiaRESUMO
We synthesized biomimetic hydrogels modified with an osteopontin-derived peptide (ODP) and used them as a substrate for in vitro culture of marrow stromal cells (MSCs) to investigate the effect of the biomimetic surface on differentiation of MSCs into osteoblasts. Proliferation and biological assays for 16 days proved that MSCs became differentiated into osteoblasts secreting osteogenic phenotypic markers such as alkaline phosphatase (ALP), osteopontin, and mineralized calcium. In addition, there was an additive effect of the cell-binding peptide on differentiation and mineralization of MSCs cultured in the presence of soluble osteogenic supplements in cell culture media. For example, calcium content at day 16 on peptide-modified hydrogels was significantly higher than on tissue culture polystyrene. Two general trends were observed: (1) proliferation of MSCs decreased as the amount of differentiation markers increased, and (2) higher peptide concentrations accelerated the differentiation of MSCs. On the hydrogel modified with ODP, ALP activity exhibited a maximum value of 36.7 +/- 4.2 pmol/cell/h at day 10 for the concentration of 2 micromol/g while the culture time needed for maximum ALP activity occurred on day 13 for the lower concentrations. On the same hydrogel, the calcium content at day 10 was 21.4 +/- 2.3 ng/cell for the peptide concentration of 2 micromol/g and 1.0 +/- 0.3 ng/cell for 1.0 micromol/g. We used Gly-Arg-Gly-Asp-Ser (GRGDS) for modification of the hydrogel as a comparison to the results with ODP. However, osteoblast development was not significantly affected by the nature of the binding peptide sequences. These results suggest that MSC function can be modulated by variation of the peptide concentration in biomimetic hydrogels used for scaffold-based bone tissue engineering.
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Materiais Biomiméticos/metabolismo , Células da Medula Óssea/fisiologia , Calcificação Fisiológica/fisiologia , Diferenciação Celular/fisiologia , Hidrogéis/metabolismo , Osteoblastos/fisiologia , Peptídeos/metabolismo , Células Estromais/fisiologia , Fosfatase Alcalina/metabolismo , Animais , Materiais Biomiméticos/química , Células da Medula Óssea/citologia , Cálcio/metabolismo , Técnicas de Cultura de Células/métodos , Células Cultivadas , Hidrogéis/química , Masculino , Teste de Materiais , Osteoblastos/citologia , Osteopontina , Peptídeos/química , Peptídeos/genética , Polímeros/química , Polímeros/metabolismo , Ratos , Ratos Wistar , Sialoglicoproteínas/química , Sialoglicoproteínas/metabolismo , Células Estromais/citologiaRESUMO
This study investigated the temporary encapsulation of rat marrow stromal osteoblasts in surface crosslinked gelatin microparticles. Cells were encapsulated in uncrosslinked gelatin microparticles of average diameter of 630 microm containing approximately 53 cells. Gelatin microparticles were crosslinked to shell thicknesses of approximately 75 microm via exposure to 1 mM dithiobis(succinimidylpropionate) (DSP) solution for 15 min or 5 mm DSP solution for 5 min for the production of microparticles dispersing approximately 60 min after placement into a physiologic fluid at 37 degrees C. Formed microparticles were placed into culture wells at a cell seeding density of 5.3 x 10(4) cells/cm2 and, following the degradation and/or dissolution of gelatin, the cells were cultured in the presence of osteogenic supplements for 28 days. Samples were taken at specified time points and analyzed by a DNA assay for cell number and a 3H-thymidine incorporation assay for proliferative potential. Samples were also obtained and analyzed at several time points by alkaline phosphatase, osteocalcin, and mineralization assays for early and late phenotypic expression markers of osteoblastic differentiation. The measurements from the different assays for encapsulated cells (EC) in uncrosslinked and crosslinked gelatin microparticles were normalized with the cell numbers from the DNA assay and compared with those for nonencapsulated control cells. The results demonstrated that the marrow stromal cells survived the encapsulation procedure in uncrosslinked gelatin microparticles and also retained their proliferative potential and osteoblastic phenotype over a 28 day period, although at a slightly lower level than the nonencapsulated cells. The results further showed that the marrow stromal cells survived the encapsulation in crosslinked gelatin microparticles prepared via exposure to 5mm DSP for 5 min and also retained their proliferative potential and osteoblastic phenotype over a 28 day period, but at a slightly lower level than the EC in uncrosslinked gelatin microparticles. In contrast, exposure to 1 mM DSP for 15 min led to severely limited cell viability and phenotypic expression probably due to the increased crosslinking time. These results suggest that temporary encapsulation of cells in gelatin microparticles may protect cells from short-term environmental effects such as those associated with the crosslinking of an injectable polymeric carrier for bone tissue engineering.