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[Purpose] In stroke rehabilitation, gait assessment measures the maximal walking speed and six-minute walking distance, both of which have not been thoroughly investigated as determinants of walking ability. Here, we assessed the methods for evaluating these gait parameters using clinical data on hybrid assistive limb treatment compared with conventional training. [Participants and Methods] In total, 20 stroke patients (hybrid assistive limb group, n=9; conventional group, n=11) participated in this randomized controlled trial. For 12 sessions (three times per week in 4 weeks), the hybrid assistive limb and conventional groups performed gait treatment with hybrid assistive limb and conventional gait training, respectively. Short physical performance battery and walking ability (maximal walking speed and six-minute walking distance) were evaluated pre- and post-intervention. Subsequently, the patients were divided further into two groups: low- and high-balance score groups. [Results] Maximum walking speed and six-minute walking distance were significantly associated, with a positive relationship observed post-intervention. The high-balance score group showed a significant improvement in the six-minute walking distance compared to the low-balance score group. However, no significant improvement in maximum walking speed was observed between both groups. [Conclusion] Due to its sensitivity in detecting differences in balance, six-minute walking distance may be a useful assessment parameter for stroke rehabilitation, particularly in the recovery of physiological walking ability.
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BACKGROUND AND PURPOSE: Reperfusion therapy by mechanical thrombectomy is used to treat acute ischemic stroke. However, reactive oxygen species generation after reperfusion therapy causes cerebral ischemia-reperfusion injury, which aggravates cerebral infarction. There is limited evidence for clinical efficacy in stroke for antioxidants. Here, we developed a novel core-shell type nanoparticle containing 4-amino-4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (nitroxide radical-containing nanoparticles [RNPs]) and investigated its ability to scavenge reactive oxygen species and confer neuroprotection. METHODS: C57BL/6J mice underwent transient middle cerebral artery occlusion and then received RNPs (9 mg/kg) through the common carotid artery. Infarction size, neurological scale, and blood-brain barrier damage were visualized by Evans blue extravasation 24 hours after reperfusion. RNP distribution was detected by rhodamine labeling. Blood-brain barrier damage, neuronal apoptosis, and oxidative neuronal cell damage were evaluated in ischemic brains. Multiple free radical-scavenging capacities were analyzed by an electron paramagnetic resonance-based method. RESULTS: RNPs were detected in endothelial cells and around neuronal cells in the ischemic lesion. Infarction size, neurological scale, and Evans blue extravasation were significantly lower after RNP treatment. RNP treatment preserved the endothelium and endothelial tight junctions in the ischemic brain; neuronal apoptosis, O2- production, and gene oxidation were significantly suppressed. Reactive oxygen species scavenging capacities against OH, ROO, and O2- improved by RNP treatment. CONCLUSIONS: An intra-arterial RNP injection after cerebral ischemia-reperfusion injury reduced blood-brain barrier damage and infarction volume by improving multiple reactive oxygen species scavenging capacities. Therefore, RNPs can provide neurovascular unit protection.
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Isquemia Encefálica/metabolismo , Radicais Livres/administração & dosagem , Nanopartículas/administração & dosagem , Fármacos Neuroprotetores/administração & dosagem , Traumatismo por Reperfusão/metabolismo , Animais , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Isquemia Encefálica/patologia , Isquemia Encefálica/prevenção & controle , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio/metabolismo , Traumatismo por Reperfusão/patologia , Traumatismo por Reperfusão/prevenção & controleRESUMO
Surface-mediated gene transfer systems using biocompatible calcium phosphate (CaP)-based composite layers have attracted attention as a tool for controlling cell behaviors. In the present study we aimed to demonstrate the potential of CaP-based composite layers to mediate area-specific dual gene transfer and to stimulate cells on an area-by-area basis in the same well. For this purpose we prepared two pairs of DNA-fibronectin-apatite composite (DF-Ap) layers using a pair of reporter genes and pair of differentiation factor genes. The results of the area-specific dual gene transfer successfully demonstrated that the cells cultured on a pair of DF-Ap layers that were adjacently placed in the same well showed specific gene expression patterns depending on the gene that was immobilized in the underlying layer. Moreover, preliminary real-time PCR results indicated that multipotential C3H10T1/2 cells may have a potential to change into different types of cells depending on the differentiation factor gene that was immobilized in the underlying layer, even in the same well. Because DF-Ap layers have a potential to mediate area-specific cell stimulation on their surfaces, they could be useful in tissue engineering applications.
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Apatitas/química , Transfecção/métodos , Animais , Células CHO , Diferenciação Celular , Cricetinae , Cricetulus , DNA/química , Fibronectinas/química , Genes Reporter , Luciferases de Vaga-Lume/biossíntese , Luciferases de Vaga-Lume/genética , Luciferases de Renilla/biossíntese , Luciferases de Renilla/genética , Camundongos , Propriedades de SuperfícieRESUMO
Although fibroblast growth factor 2 (FGF2) is a promising agent for treating brain infarction, current methods of FGF2 administration are associated with a short circulating half-life. An FGF2 apatite coating was developed as a slow-releasing drug delivery system (DDS) by forming an FGF2/calcium phosphate composite layer. Hydroxyapatite was coated with high or low doses of FGF2, denoted as FGF-high and FGF-low. This study investigated the efficacy of the coating as angiogenesis therapy for brain infarction. Rats were subjected to permanent occlusion of the middle cerebral artery, an FGF2 apatite-coated implant was inserted, and the rat brains were removed 2 weeks after implantation. Rats in groups treated with FGF-high had significantly smaller areas of brain infarction, particularly in the external capsule and the lateral side of the putamen, and better capillary density than rats in groups treated with non-FGF2 apatite-coated implants. Histologic analysis indicated that the new vessels were larger and had thicker walls in the FGF2 apatite-coated groups than in the non-FGF2 groups. Fluorescence immunohistochemistry of the peri-infarction region showed that FGF2 released from FGF2 apatite-coated implants might have biological activity. Moreover, fluorescence immunohistochemistry showed that released FGF2 influenced microglia cells. This new FGF2 DDS involving an FGF2 apatite coating can prevent infarction of the penumbra through the multipotential effects of FGF2.
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Indutores da Angiogênese/administração & dosagem , Infarto Encefálico/tratamento farmacológico , Encéfalo/irrigação sanguínea , Encéfalo/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/administração & dosagem , Animais , Apatitas/química , Encéfalo/patologia , Infarto Encefálico/patologia , Capilares/patologia , Preparações de Ação Retardada/administração & dosagem , Preparações de Ação Retardada/química , Fator 2 de Crescimento de Fibroblastos/química , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Basic fibroblast growth factor (bFGF) is a therapeutic protein that can enhance angiogenesis, wound healing, and tissue regeneration; however, it is extremely unstable even under a normal physiological environment. Biocompatible calcium phosphate (CaP) nanoparticles (NPs) co-immobilizing bFGF, heparin, and ferucarbotran would be useful as a multifunctional delivery carrier of bFGF. In this study, such NPs were successfully fabricated by a coprecipitation process, using a labile supersaturated CaP solution containing bFGF, heparin, and ferucarbotran. The NPs showed relatively high negative zeta potential (-12 mV) because of the negatively charged heparin, which enabled their stable dispersion in water. The hydrodynamic diameter of the NPs was around 200 nm. Immunoreactive bFGF was released from the NPs in an acellular medium dose-dependently. The NPs promoted proliferation of baby hamster kidney fibroblasts (BHK-21 cells) and mouse osteoblastic MC3T3-E1 cells at a certain dose range, although they inhibited proliferation of rat pheochromocytoma (PC-12) cells. These results demonstrated that the effect of the NPs on cell proliferation was dependent on the cell type and dose, the details of which should be investigated in a future study.
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Fator 2 de Crescimento de Fibroblastos , Heparina , Ratos , Camundongos , Animais , Fator 2 de Crescimento de Fibroblastos/farmacologia , Proliferação de Células , Heparina/farmacologia , Fibroblastos , Fosfatos de Cálcio/farmacologiaRESUMO
Previous studies have examined the therapeutic effect of endothelial progenitor cells (EPCs) during the chronic phase of cerebral infarction in rats; however, few studies have investigated the effects of EPCs during the acute phase of infarction. In this study, we evaluated the therapeutic effect of EPCs with low aldehyde dehydrogenase activity (Alde-Low EPCs) in rats with acute cerebral infarction, and our results provide insight that may help to identify a therapeutic mechanism of EPCs for acute cerebral infarction. The administration of Alde-Low EPCs into rats with acute cerebral infarction results in the accumulation and migration of the Alde-Low EPCs into the infarct area and the subsequent decrease of infarct volume. Moreover, we found that the stromal cell-derived factor-1 (SDF-1) and CXC chemokine receptor 4 (CXCR4) signaling pathway may regulate the accumulation of Alde-Low EPCs. The transplantation of Alde-Low EPCs may represent a potential treatment strategy for acute cerebral infarction.
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Aldeído Desidrogenase/biossíntese , Isquemia Encefálica/cirurgia , Células Endoteliais da Veia Umbilical Humana/enzimologia , Células Endoteliais da Veia Umbilical Humana/transplante , Traumatismo por Reperfusão/cirurgia , Transplante de Células-Tronco/métodos , Células-Tronco/enzimologia , Animais , Infarto Encefálico/patologia , Infarto Encefálico/cirurgia , Isquemia Encefálica/patologia , Modelos Animais de Doenças , Humanos , Masculino , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/patologia , Resultado do TratamentoRESUMO
BACKGROUND: An inflammatory reaction in vascular tissue is a potential factor linking restenosis after angioplasty. Although cilostazol, a selective phosphodiesterase type 3 inhibitor that is a unique antiplatelet drug and vasodilator, has been reported to be anti-inflammatory, its effect on the inflammatory action of mononuclear cells homing to endothelial cells is not clearly understood. In this study, whether cilostazol inhibits neointimal formation and improves inflammatory actions by inhibiting sialyl Lewis X (SLX) expression on mononuclear cells and E-selectin expression on endothelial cells was evaluated. METHODS: The effect of cilostazol (1, 3, 10, 30 µM) on expression of E-selectin in human umbilical vein endothelial cells and SLX in rat mononuclear cells stimulated with lipopolysaccharide by immunofluorescence and real-time polymerase chain reaction (n = 3) was studied. Additionally, a double-balloon injury model was used on rat carotid arteries to evaluate vascular intimal hyperplasia. 0.1% cilostazol was administered 3 days before the first balloon injury, and the second balloon injury was performed 7 days after the first injury. Cilostazol administration was continued until rats were sacrificed 14 days after the second angioplasty. The expression of SLX on mononuclear cells and E-selectin on endothelial cells by immunofluorescence (n = 10) and real-time polymerase chain reaction (n = 5) were studied. RESULTS: Cilostazol effectively inhibited the expression of SLX on mononuclear cells and E-selectin on endothelial cells. Cilostazol inhibited the migration of mononuclear cells in neointimal regions and neointimal hyperplasia after balloon injury. The numbers of macrophages and T-lymphocytes and the hyperplasia area in neointimal regions decreased from 71.06 ± 20.04, 1121 ± 244.4 cells per section, 206,400 ± 96,150 mm(2) to 29.65 ± 16.73, 374.2 ± 124.5 cells per section, and 101,900 ± 16,150 mm(2) due to the administration of cilostazol. CONCLUSIONS: These results demonstrate that the protective effect of cilostazol against neointimal hyperplasia may be mediated by its anti-inflammatory actions of mononuclear cells homing to endothelial cells by decreasing SLX and E-selectin expression. CLINICAL RELEVANCE: It is reported that cilostazol inhibits neointimal hyperplasia by decreasing the expression of some cell-adhesion molecules. We evaluated the effects of cilostazol for the expression of sialyl Lewis X (SLX) on mononuclear cells and E-selectin on endothelial cells, which interaction is the first step of inflammation action. Cilostazol was thought to show the anti-inflammatory actions by decreasing SLX and E-selectin expression in addition to decreasing the expression of some cell-adhesion molecules.
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Anti-Inflamatórios/farmacologia , Artérias Carótidas/efeitos dos fármacos , Lesões das Artérias Carótidas/tratamento farmacológico , Proliferação de Células/efeitos dos fármacos , Selectina E/metabolismo , Células Endoteliais/efeitos dos fármacos , Leucócitos Mononucleares/efeitos dos fármacos , Oligossacarídeos/metabolismo , Inibidores da Fosfodiesterase 3/farmacologia , Tetrazóis/farmacologia , Túnica Íntima/efeitos dos fármacos , Angioplastia com Balão , Animais , Artérias Carótidas/metabolismo , Artérias Carótidas/patologia , Lesões das Artérias Carótidas/metabolismo , Lesões das Artérias Carótidas/patologia , Células Cultivadas , Cilostazol , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Regulação para Baixo , Selectina E/genética , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Imunofluorescência , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Hiperplasia , Leucócitos Mononucleares/metabolismo , Masculino , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Antígeno Sialil Lewis X , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Fatores de Tempo , Túnica Íntima/metabolismo , Túnica Íntima/patologiaRESUMO
A surface-mediated gene transfer system using biocompatible apatite-based composite layers has great potential for tissue engineering. Among the apatite-based composite layers developed to date, we focused on a DNA-lipid-apatite composite layer (DLp-Ap layer), which has the advantage of relatively high efficiency as a non-viral system. In this study, various lipid transfection reagents, including a newly developed reagent, polyamidoamine dendron-bearing lipid (PD), were employed to prepare the DLp-Ap layer, and the preparation condition was optimized in terms of efficiency of gene transfer to epithelial-like CHO-K1 cells in the presence of serum. The optimized DLp-Ap layer derived from PD had the highest gene transfer efficiency among all the apatite-based composite layers prepared in this study. In addition, the optimized DLp-Ap layer demonstrated higher gene transfer efficiency in the presence of serum than the conventional particle-mediated systems using commercially available lipid transfection reagents. It was also shown that the optimized DLp-Ap layer mediated the area-specific gene transfer on its surface, i.e., DNA was preferentially transferred to the cells adhering to the surface of the layer. The present gene transfer system using the PD-derived DLp-Ap layer, with the advantages of high efficiency in the presence of serum and area-specificity, would be useful in tissue engineering.
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Apatitas/química , DNA/química , Técnicas de Transferência de Genes , Lipídeos/química , Animais , Células CHO , Cricetinae , Microscopia Eletrônica de Varredura , Espectrofotometria Ultravioleta , Difração de Raios XRESUMO
Surface-mediated gene transfer systems using apatite (Ap)-based composite layers have received increased attention in tissue engineering applications owing to their safety, biocompatibility and relatively high efficiency. In this study, DNA-antibody-apatite composite layers (DA-Ap layers), in which DNA and antibody molecules are immobilized within a matrix of apatite nanocrystals, were fabricated using a biomimetic coating process. They were then assayed for their gene transfer capability for application in a specific cell-targeted gene transfer. A DA-Ap layer that was fabricated with an anti-CD49f antibody showed a higher gene transfer capability to the CD49f-positive CHO-K1 cells than a DNA-apatite composite layer (D-Ap layer). The antibody facilitated the gene transfer capability of the DA-Ap layer only to the specific cells that were expressing corresponding antigens. When the DA-Ap layer was fabricated with an anti-N-cadherin antibody, a higher gene transfer capability compared with the D-Ap layer was found in the N-cadherin-positive P19CL6 cells, but not in the N-cadherin-negative UVâ2 cells or in the P19CL6 cells that were pre-blocked with anti-N-cadherin. Therefore, the antigen-antibody binding that takes place at the cell-layer interface should be responsible for the higher gene transfer capability of the DA-Ap than D-Ap layer. These results suggest that the DA-Ap layer works as a mediator in a specific cell-targeted gene transfer system.
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Electronic health records should efficiently store the information required for clinical decision-making and contain progress notes that reference this information. However, beyond the inclusion of subjective data, objective data, assessment, and plan framework, the content required to make progress notes useful for readers with diverse specialties has not been clarified. Moreover, the documentation burden that including additional content places on medical doctors (MDs) has not been determined. We conducted a questionnaire with 74 MDs, nurses, and other clinical professionals to determine whether they found progress notes with varying specific contents useful. In addition, the degree of the burden of writing progress notes that contain specific content was measured when 25 MDs were instructed to add specific content. Our results reveal that progress notes are more useful for clinical reasoning for readers other than MDs when more specific information is included; this can be achieved without increasing the documentation burden.
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Documentação , Médicos , Comunicação , Registros Eletrônicos de Saúde , Humanos , RedaçãoRESUMO
Edema is a common symptom of kidney disease, and quantitative measurement of edema is desired. This paper presents a method to estimate the degree of edema from facial images taken before and after dialysis of renal failure patients. As tasks to estimate the degree of edema, we perform pre- and post-dialysis classification and body weight prediction. We develop a multi-patient pre-training framework for acquiring knowledge of edema and transfer the pre-trained model to a model for each patient. For effective pre-training, we propose a novel contrastive representation learning, called weight-aware supervised momentum contrast (WeightSupMoCo). WeightSupMoCo aims to make feature representations of facial images closer in similarity of patient weight when the pre- and post-dialysis labels are the same. Experimental results show that our pre-training approach improves the accuracy of pre- and post-dialysis classification by 15.1% and reduces the mean absolute error of weight prediction by 0.243 kg compared with training from scratch. The proposed method accurately estimate the degree of edema from facial images; our edema estimation system could thus be beneficial to dialysis patients.
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Biodegradable sheets loaded with basic fibroblast growth factor (bFGF) are prepared as novel bFGF-releasing systems from polyglycolic acid nonwoven fabric by oxygen plasma treatment followed by bFGF adsorption. In the present study, we investigated the therapeutic effects of this system on a focal cerebral infarction model (CB-17 mouse). A preliminary in vitro study showed that this system released bFGF in an acellular culture medium, thereby keeping the bFGF concentration in the medium at ≥5 ng/ml for a prolonged period of 7 days. The released bFGF from this system retained its biological activity to enhance endothelial tube formation in vitro. In a mouse model of subacute focal cerebral infarction, this system increased the expression of endogenous vascular endothelial growth factor in the peri-infarct cortex and subventricular zone, promoted angiogenesis in the striatum, and increased neural progenitor cells in the peri-infarct cortex. Thus, this bFGF-releasing system has the potential to be a novel therapeutic approach for cerebral infarction.
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Células-Tronco Neurais , Ácido Poliglicólico , Animais , Infarto Cerebral/terapia , Fator 2 de Crescimento de Fibroblastos/farmacologia , Camundongos , Células-Tronco Neurais/metabolismo , Oxigênio , Fator A de Crescimento do Endotélio Vascular/metabolismo , Fatores de Crescimento do Endotélio VascularRESUMO
Hybrid Assistive Limb (HAL) is a wearable human assistant cyborg-type robot that helps lower-leg movement based on bioelectrical signals detected from the voluntary movement of the person wearing it. In this study, we developed a novel staged HAL treatment protocol for patients with acute stroke. The Regain Program for Gait with HAL (RPG-HAL) was formulated in four steps, based on the severity of limb paralysis. Twenty-one patients with acute stroke received a combination treatment of RPG-HAL and conventional rehabilitation. The feasibility and safety of RPG-HAL were evaluated based on changes in physical function and activities of daily living (ADL). RPG-HAL yielded improvement in gait speed, cadence, step length, and functional ambulation category (FAC). The effect size was >0.8 in all measurements. FAC (1.90) and Barthel Index (BI) (1.92) exhibited the highest scores. Twelve out of 14 patients with FAC 0 before RPG-HAL reached the upper FAC. Thus, earlier intervention using RPG-HAL as improving physical function, ADL, and gait ability in patients with stroke.
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Reabilitação do Acidente Vascular Cerebral , Acidente Vascular Cerebral , Atividades Cotidianas , Protocolos Clínicos , Marcha , Humanos , Acidente Vascular Cerebral/complicações , Acidente Vascular Cerebral/terapia , Reabilitação do Acidente Vascular Cerebral/métodosRESUMO
BACKGROUND: Safe and efficient gene transfer systems are needed for tissue engineering. We have developed an apatite composite layer including the bone morphogenetic protein-2 (BMP-2) gene and fibronectin (FB), and we evaluated its ability to induce bone formation. METHODS: An apatite composite layer was evaluated to determine the efficiency of gene transfer to cells cultured on it. Cells were cultured on a composite layer including the BMP-2 gene and FB, and BMP-2 gene expression, BMP-2 protein concentrations, alkaline phosphatase (ALP) activity, and osteocalcin (OC) concentrations were measured. A bone defect on the cranium of rats was treated with hydroxyapatite (HAP)-coated ceramic buttons with the apatite composite layer including the BMP-2 gene and FB (HAP-BMP-FB). The tissue concentration of BMP-2, bone formation, and the expression levels of the BMP-2, ALP, and OC genes were all quantified. RESULTS: The apatite composite layer provided more efficient gene transfer for the cultured cells than an apatite composite layer without FB. The BMP-2 concentration was approximately 100-600 pg/mL in the cell-culture medium. Culturing the cells on the apatite composite layer for 27 days increased ALP activity and OC concentrations. In animal experiments, the tissue concentrations of BMP-2 were over 100 pg/mg in the HAP-BMP-FB group and approximately 50 pg/mg in the control groups. Eight weeks later, bone formation was more enhanced in the HAP-BMP-FB group than in the control groups. In the tissues surrounding the HAP button, the gene expression levels of ALP and OC increased. CONCLUSION: The BMP-2 gene-FB-apatite composite layer might be useful for bone engineering.
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Proteína Morfogenética Óssea 2/fisiologia , Durapatita/metabolismo , Fibronectinas/fisiologia , Técnicas de Transferência de Genes , Osteogênese/fisiologia , Engenharia Tecidual/métodos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Proteína Morfogenética Óssea 2/genética , Proteína Morfogenética Óssea 2/metabolismo , Fosfatos de Cálcio/metabolismo , Células Cultivadas , Primers do DNA/genética , Ensaio de Imunoadsorção Enzimática , Células HeLa , Humanos , Proteínas de Membrana/metabolismo , Camundongos , Microscopia Eletrônica de Varredura , Osteocalcina/metabolismo , Osteogênese/genética , Plasmídeos/genética , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Crânio/lesões , Crânio/cirurgiaRESUMO
We hypothesized that gait treatment with a wearable cyborg Hybrid Assistive Limb (HAL) would improve the walking ability of patients with hemiparesis after stroke. This study aims to evaluate the efficacy and safety of gait treatment using HAL versus conventional gait training (CGT) in hemiplegic patients with acute stroke and establish a protocol for doctor-initiated clinical trials for acute stroke. We will enroll patients with acute stroke at the University of Tsukuba Hospital. This study is a single-center, randomized, parallel-group, controlled trial (HAL group, n = 20; control group, n = 20) that will include three phases: (1) pre-observation phase (patient enrollment, baseline assessment, and randomization); (2) treatment phase (nine sessions, twice or thrice per week over 3-4 weeks; the HAL and control groups will perform gait treatment using HAL or CGT, respectively, and finally (3) post-treatment evaluation phase. The Functional Ambulation Category score will be the primary outcome measure, and the following secondary outcome measures will be assessed: Mini-Mental State Examination, Brunnstrom recovery stage of lower limbs, Fugl-Meyer assessment of lower limbs, 6-min walking distance, comfortable gait speed, step length, cadence, Barthel Index, Functional Independence Measure, gait posture, motion analysis (muscle activity), amount of activity (evaluated using an activity meter), stroke-specific QOL, and modified Rankin Scale score. The baseline assessment, post-treatment evaluation, and follow-up assessment will evaluate the overall outcome measures; for other evaluations, physical function evaluation centered on walking will be performed exclusively, excluding ADL and QOL scores. This study is a randomized controlled trial that aims to clarify the efficacy and safety of gait treatment using HAL compared with CGT in hemiplegic patients with acute stroke. In addition, we aim to establish a protocol for doctor-initiated clinical trials for acute stroke based on the study results. If our results demonstrate the effectiveness of the proposed treatment regarding outcomes of patients with hemiplegic acute stroke, this study will promote the treatment of these patients using the HAL system as an effective tool in future stroke rehabilitation programs. The study protocol was registered with the Japan Registry of Clinical Trials on October 14, 2020 (jRCTs032200151).
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BACKGROUND: We have recently developed a safe and efficient gene transfer system using a laminin-DNA-apatite composite layer. The objectives of the present study were to fully characterize and optimize the laminin-DNA-apatite composite layer in relation to the efficiency of gene transfer and to demonstrate the feasibility of the composite layer in the induction of cell differentiation. METHODS: The laminin-DNA-apatite composite layer was prepared under various conditions. The efficiency of gene transfer on the resulting composite layer was evaluated using luciferase and ss-galactosidase gene expression assay systems. A laminin-DNA-apatite composite layer, prepared under the optimized condition using a plasmid including cDNA of nerve growth factor (NGF), was then applied to the neuron-like differentiation of PC12 cells. RESULTS: The laminin content of the laminin-DNA-apatite composite layer was found to be a dominant factor improving the efficiency of gene transfer rather than the DNA content. The cell adhesion property of laminin in the composite layer should be responsible for the improvement in efficiency of gene transfer because the immobilization of albumin without the cell adhesion property in a DNA-apatite composite layer had no effect on the efficiency of gene transfer. A laminin-DNA-apatite composite layer, prepared under the optimized condition using a plasmid including cDNA of NGF, successfully induced the neuron-like differentiation of PC12 cells. CONCLUSIONS: The present gene transfer system, with the potential to control cell differentiation and having features of safety and relatively high and controllable efficiency, would be a useful tool for tissue engineering applications and the production of transfection microarrays.
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Apatitas/metabolismo , DNA/metabolismo , Técnicas de Transferência de Genes , Laminina/metabolismo , Animais , Células CHO , Adesão Celular , Diferenciação Celular , Cricetinae , Cricetulus , Proteínas Imobilizadas/metabolismo , Cinética , Luciferases/metabolismo , Camundongos , Microscopia Eletrônica de Varredura , Células PC12 , Ratos , Soluções , Propriedades de Superfície , Fatores de Tempo , Difração de Raios XRESUMO
This study investigated whether nitroxide radical (4-amino-TEMPOL)-containing nanoparticles (RNPs; antioxidant nanomedicine) can prevent neurovascular unit impairment caused by reactive oxygen species (ROS) after cerebral ischemia-reperfusion. C57BL/6J mice underwent transient middle cerebral artery occlusion (tMCAO). The mice were randomly divided and administered intra-arterial RNPs injection (9 mg/kg, 7 µM/kg), edaravone (3 mg/kg, 17 µM/kg), or phosphate-buffered saline (control group). Survival rate and neurological score were evaluated 24 h post-injection. RNPs distribution was determined using immunofluorescence staining and blood-brain barrier (BBB) disruption using Evans blue extravasation assay. Effect of RNPs and edaravone on microglia polarization into microglia M1 and M2 was evaluated. We also determined multiple ROS-scavenging activities in brain homogenates of RNPs- and edaravone-treated animals using an electron spin resonance-based spin-trapping method. Compared with edaravone, RNPs significantly improved the survival rate and neurological deficit, inhibited BBB disruption and supported polarization of microglia into M2 microglia. RNPs were localized in endothelial cells, the perivascular space, neuronal cell cytoplasm, astrocytes, and microglia. Scavenging capacities of hydroxyl, alkoxyl, and peroxyl radicals were significantly higher in the RNPs-treated group. RNPs show promising results as a future neuroprotective nanomedicine approach for cerebral ischemia-reperfusion injury.
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Antioxidantes/farmacologia , Isquemia Encefálica/patologia , Encéfalo/efeitos dos fármacos , Nanopartículas , Traumatismo por Reperfusão/patologia , Animais , Óxidos N-Cíclicos/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Nanomedicina/métodos , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Espécies Reativas de Oxigênio , Marcadores de SpinRESUMO
BACKGROUND: Gait disturbance often occurs in stroke survivors. Recovery of walking function is challenging, as some gait disturbance due to hemiparesis often remains even after rehabilitation therapy, presenting a major obstacle towards regaining activities-of-daily-living performance and achieving social reintegration. OBJECTIVE: This study aims to clarify the effectiveness of a walking program involving the wearable Hybrid Assistive Limb (HAL-TS01) robotic exoskeleton for improving walking ability in stroke patients with hemiparesis and stagnant recovery despite ongoing rehabilitation. METHODS: This is a multicenter, randomized, parallel-group, controlled study (HAL group, n=27; control group, n=27). The study period includes preintervention observation (until stagnant recovery), intervention (HAL-based walking therapy or conventional rehabilitation; 5 weeks), and postintervention observation (2 weeks). Following provision of informed consent and primary registration, the patients undergo conventional rehabilitation for preintervention observation, during which the recovery of walking ability is monitored to identify patients with stagnant recovery (based on weekly assessments using the 10-meter maximum walking speed [MWS] test). Patients with an MWS of 30-60 m/minute and insufficient weekly improvement in MWS undergo secondary registration and are randomly assigned to undergo HAL-based walking therapy (HAL group) or conventional rehabilitation (control group). The primary outcome is the change in MWS from baseline to the end of the 5-week intervention. RESULTS: This study began in November 2016 and is being conducted at 15 participating facilities in Japan. CONCLUSIONS: Assessments of walking ability vary greatly and it is difficult to define the threshold for significant differences. To reduce such variability, our study involves conducting conventional rehabilitation to the point of saturation before starting the intervention. Stagnation in the recovery of walking ability despite conventional rehabilitation highlights the limits of current medical care. The present study may bring evidence that HAL-based therapy can overcome such limitations and induce added recovery of walking ability, which would promote the use of HAL technology in the clinical setting. TRIAL REGISTRATION: UMIN Clinical Trials Registry UMIN000024805; https://upload.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000028545.
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We hypothesized that a single-leg version of the Hybrid Assistive Limb (HAL) system could improve the gait and physical function of patients with hemiparesis following a stroke. In this pilot study, we therefore compared the efficacy of HAL-based gait training with that of conventional gait training (CGT) in patients with acute stroke. Patients admitted to the participating university hospital were assigned to the HAL group, whereas those admitted to outside teaching hospitals under the same rehabilitation program who did not use the HAL were assigned to the control group. Over 3 weeks, all participants completed nine 20 min sessions of gait training, using either HAL (i.e., the single-leg version of HAL on the paretic side) or conventional methods (i.e., walking aids and gait orthoses). Outcome measures were evaluated before and after the nine training sessions. The Functional Ambulation Category (FAC) was the primary outcome measure, but the following secondary outcome measures were also assessed: National Institutes of Health Stroke Scale, Fugl-Meyer Assessment (Lower Extremity), comfortable walking speed, step length, cadence, 6-min walk distance, Barthel Index, and Functional Independence Measure. In total, 22 post-stroke participants completed the clinical trial: 12 in the HAL group and 10 in the CGT group. No serious adverse events occurred in either group. The HAL group showed significant improvement in FAC after nine sessions when compared with the CGT group (P = 0.014). However, secondary outcomes did not differ significantly between the groups. Our results demonstrate that HAL-based gait therapy may improve independent walking in patients with acute stroke hemiplegia who are dependent on ambulatory assistance. A larger-scale randomized controlled trial is needed to clarify the effectiveness of single-leg HAL therapy. Clinical Trial Registration: UMIN Clinical Trials Registry, identifier UMIN000022410.
RESUMO
PURPOSE: Patients with malignant gliomas have a poor prognosis. To explore a novel and more effective approach for the treatment of patients with malignant gliomas, we designed a strategy that combines caspase-8 (CSP8) gene therapy and radiation treatment (RT). In addition, the specificity of the combined therapy was investigated to decrease the unpleasant effects experienced by the surrounding normal tissue. METHODS AND MATERIALS: We constructed the plasmid pEGR-green fluorescence protein that included the radiation-inducible early growth response gene-1 (Egr-1) promoter and evaluated its characteristics. The pEGR-CSP8 was constructed and included the Egr-1 promoter and CSP8 complementary DNA. Assays that evaluated the apoptosis inducibility and cytotoxicity caused by CSP8 gene therapy combined with RT were performed using U251 and U87 glioma cells. The pEGR-CSP8 was transfected into the subcutaneous U251 glioma cells of nude mice by means of in vivo electroporation. The in vivo effects of CSP8 gene therapy combined with RT were evaluated. RESULTS: The Egr-1 promoter yielded a better response with fractionated RT than with single-dose RT. In the assay of apoptosis inducibility and cytotoxicity, pEGR-CSP8 showed response for RT. The pEGR-CSP8 combined with RT is capable of inducing cell death effectively. In mice treated with pEGR-CSP8 and RT, apoptotic cells were detected in pathologic sections, and a significant difference was observed in tumor volumes. CONCLUSIONS: Our results indicate that radiation-inducible gene therapy may have great potential because this can be spatially or temporally controlled by exogenous RT and is safe and specific.