Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 88
Filtrar
Más filtros

Banco de datos
Tipo del documento
Intervalo de año de publicación
1.
Cell ; 186(22): 4920-4935.e23, 2023 10 26.
Artículo en Inglés | MEDLINE | ID: mdl-37776859

RESUMEN

SpCas9 and AsCas12a are widely utilized as genome-editing tools in human cells. However, their relatively large size poses a limitation for delivery by cargo-size-limited adeno-associated virus (AAV) vectors. The type V-F Cas12f from Acidibacillus sulfuroxidans is exceptionally compact (422 amino acids) and has been harnessed as a compact genome-editing tool. Here, we developed an approach, combining deep mutational scanning and structure-informed design, to successfully generate two AsCas12f activity-enhanced (enAsCas12f) variants. Remarkably, the enAsCas12f variants exhibited genome-editing activities in human cells comparable with those of SpCas9 and AsCas12a. The cryoelectron microscopy (cryo-EM) structures revealed that the mutations stabilize the dimer formation and reinforce interactions with nucleic acids to enhance their DNA cleavage activities. Moreover, enAsCas12f packaged with partner genes in an all-in-one AAV vector exhibited efficient knock-in/knock-out activities and transcriptional activation in mice. Taken together, enAsCas12f variants could offer a minimal genome-editing platform for in vivo gene therapy.


Asunto(s)
Sistemas CRISPR-Cas , Edición Génica , Animales , Humanos , Ratones , Microscopía por Crioelectrón , Mutación , Terapia Genética
2.
J Orthop Sci ; 2024 May 16.
Artículo en Inglés | MEDLINE | ID: mdl-38760245

RESUMEN

BACKGROUND: Transcription factor protein IκBζ (encoded by the Nfkbiz gene) regulates nuclear factor-κB (NF-κB) and is involved in the pathophysiology of various inflammatory diseases. However, the role of IκBζ in secondary damage following spinal cord injury (SCI) remains to be determined. Here, we investigated the effect of IκBζ expressed in hematopoietic cells on the progression of secondary damage and functional recovery after SCI. METHODS: We used conditional IκBζ-knockout mice (Mx1-Cre;Nfkbizfl/f) to examine the role of IκBζ in hematopoietic cells after SCI. Contusion SCI was induced using a force of 60 kdyn. The recovery of locomotor performance was evaluated using the nine-point Basso Mouse Scale (BMS) until 42 days post-injury. Expression patterns of inflammatory cytokines and chemokines were examined by quantitative real-time PCR or proteome array analysis. Bone marrow transplantation (BMT) was performed to eliminate the effect of IκBζ deletion in non-hematopoietic cells. RESULTS: Mx1-Cre;Nfkbizfl/fl mice had significantly improved locomotor function compared with wild-type (WT) mice. The mRNA expression of Nfkbiz in WT mice peaked at 12 h after SCI and then decreased slowly in both the spinal cord and white blood cells. In situ hybridization showed that Nfkbiz mRNA was localized in cell nuclei, including macrophage-like cells, in the injured spinal cord of WT mice at 1 day after SCI. Compared with WT mice, Mx1-Cre;Nfkbizfl/fl mice had significantly increased mRNA expressions of interleukin (Il)-4 and Il-10 in the injured spinal cord. In addition, Mx1-Cre;Nfkbizfl/fl mice had significantly higher protein levels of granulocyte-macrophage colony-stimulating factor and C-C motif chemokine 11 compared with WT mice. BMT from Mx1-Cre;Nfkbizfl/fl mice into WT mice improved functional recovery after SCI compared with control mice (WT cells into WT mice). CONCLUSIONS: IκBζ deletion in hematopoietic cells improved functional recovery after SCI, possibly by shifting the inflammatory balance towards anti-inflammatory and pro-regenerative directions.

3.
J Gene Med ; 25(8): e3505, 2023 08.
Artículo en Inglés | MEDLINE | ID: mdl-36972408

RESUMEN

BACKGROUND: Intravenous administration of adeno-associated virus (AAV) vectors is a promising gene therapy approach for monogenic diseases. However, re-administration of the same AAV serotype is impossible because of the induction of anti-AAV neutralizing antibodies (NAbs). Here, we examined the feasibility of re-administrating AAV vector serotypes different from the initial AAV vector serotype. METHODS: Liver-targeting AAV3B, AAV5, and AAV8 vectors were intravenously injected in C57BL/6 mice, and the emergence of NAbs and the transduction efficacy following re-administration were evaluated. RESULTS: For all serotypes, re-administration of the same serotype was not possible. Although the highest neutralizing activity of NAb was induced by AAV5, anti-AAV5 NAbs did not react with other serotypes, resulting in successful re-administration with the other serotypes. AAV5 re-administration was also successful in all mice treated with AAV3B and AAV8. Effective secondary administration of AAV3B and AAV8 was observed in most mice initially administrated AAV8 and AAV3B, respectively. However, few mice developed NAbs cross-reacting with the other serotypes, especially those with close sequence homology. CONCLUSIONS: In summary, AAV vector administration induced NAbs relatively specific to the administrated serotype. Secondary administration of AAVs targeting liver transduction could be successfully achieved by switching AAV serotypes in mice.


Asunto(s)
Dependovirus , Vectores Genéticos , Animales , Ratones , Dependovirus/genética , Vectores Genéticos/genética , Ratones Endogámicos C57BL , Hígado , Anticuerpos Neutralizantes
4.
Rinsho Ketsueki ; 64(5): 377-388, 2023.
Artículo en Japonés | MEDLINE | ID: mdl-37271529

RESUMEN

Hemophiliacs are X-linked inherited bleeding disorders. The treatment of hemophilia is based on periodic replacement treatment with coagulation factor products to avoid the development of hemophilic arthropathy, as well as to control fatal bleeding. However, because of the coagulation factor's incredibly short half-life, the medication must be often given intravenously throughout one's life. Extended half-life coagulation factor preparations have lowered the burden of treatment by enhancing pharmacokinetics. However, the patients remain at risk of developing inhibitors to significantly interfere with the therapeutic effectiveness of coagulation factor concentrates. Recently, nonfactor replacement therapies have gained a lot of interest as a new class of therapeutic drugs preventing bleeding in patients with or without inhibitors. Moreover, gene therapy drugs for hemophilia have recently been approved in Europe and the United States. Because gene therapy can preserve coagulation factor levels in the blood for at least several years with a single treatment, it may become a curative treatment that removes the need for the therapy for a long time.


Asunto(s)
Enfermedades Hematológicas , Hemofilia A , Humanos , Hemofilia A/tratamiento farmacológico , Factores de Coagulación Sanguínea/uso terapéutico , Coagulación Sanguínea , Enfermedades Hematológicas/terapia , Terapia Genética , Factor VIII/uso terapéutico
5.
Haemophilia ; 28 Suppl 4: 61-67, 2022 May.
Artículo en Inglés | MEDLINE | ID: mdl-35521726

RESUMEN

INTRODUCTION: Gene therapy is emerging as a potential cure for haemophilia. Gene therapy is a one-time treatment that can elevate factor levels for many years and minimize or eliminate the need for clotting factor concentrate (CFC) replacement therapy. However, there is a paucity of reports on gene therapy efforts in countries outside of North America or Europe, especially in low-and-middle-income countries (LMIC). All indications are that gene therapy will be one of standard care treatments for haemophilia in the future. Still, it may not be accessible to many countries due to various barriers and challenges. At the same time, each country may formulate solutions that may be used globally. AIM: To summarize the approaches taken to establish haemophilia gene therapy in Japan, China, India, South Africa, and Brazil, and to describe the US-initiated multi-LMIC haemophilia gene therapy development program to include Peru, Vietnam, Thailand, Nepal, and Sri Lanka. METHODS: A review of related published information or as accessible by each country's author. RESULTS: Different starting conditions, differing input and level of support from the multitude of stakeholders, and strong leadership have led to various approaches for facilitating research and developing needed infrastructure and regulatory and financing models. Gene therapy programs are at various stages of development and include both adeno-associated viral and lentiviral vectors. CONCLUSION: Global partnerships and collaboration, exchange of knowledge and experience, and alignment of processes across borders will promote further progress towards global access to gene therapy for haemophilia.


Asunto(s)
Hemofilia A , Brasil , Dependovirus/genética , Países en Desarrollo , Europa (Continente) , Terapia Genética , Hemofilia A/genética , Hemofilia A/terapia , Humanos
6.
J Immunol ; 204(8): 2033-2042, 2020 04 15.
Artículo en Inglés | MEDLINE | ID: mdl-32144162

RESUMEN

IκBζ (encoded by the Nfkbiz) is a member of the nuclear IκB family, which is involved in the expression of secondary response genes based on signals from TLR or IL-1R. ST2L, an IL-33R, is a member of the IL-1R family and abundantly expressed in tissue-resident immune cells, such as mast cells and innate lymphoid cells; however, its downstream signaling pathway remains unelucidated. In this study, we examined the role of IκBζ in ST2L-mediated cytokine and chemokine production in mast cells. Murine bone marrow cells were differentiated ex vivo into bone marrow-derived mast cells (BMMCs). The treatment of BMMCs with IL-33 transiently induced robust IκBζ expression. Of the 40 cytokines and chemokines examined using a cytokine and chemokine array, the concentrations of IL-6, IL-13, CCL2, CCL3, and TNF-α in the supernatant were augmented by IL-33. The deletion of IκBζ in BMMCs resulted in a significant reduction of the production of these mediators and the expression of their mRNA. NF-κB p50 but not p65 translocated to the nucleus by IL-33 and was not affected by the deletion of IκBζ. However, induction of IκBζ and the resultant cytokine and chemokine productions were significantly inhibited by pretreatment with an NF-κB inhibitor. The deletion of IκBζ did not affect the phosphorylation of ERK, p38 MAPK, or JNK by IL-33, and the treatment with inhibitors of these mitogen-activated kinases failed to abolish the expression of Nfkbiz Our findings suggest that IκBζ augments IL-33-dependent cytokine and chemokine production in BMMCs through the action of NF-κB.


Asunto(s)
Citocinas/biosíntesis , Proteínas I-kappa B/metabolismo , Interleucina-33/inmunología , Mastocitos/metabolismo , Proteínas Adaptadoras Transductoras de Señales/deficiencia , Proteínas Adaptadoras Transductoras de Señales/inmunología , Animales , Citocinas/inmunología , Proteínas I-kappa B/deficiencia , Mastocitos/inmunología , Ratones , Ratones Noqueados , Ratones Transgénicos , FN-kappa B/metabolismo
7.
Rinsho Ketsueki ; 63(11): 1558-1565, 2022.
Artículo en Japonés | MEDLINE | ID: mdl-36476798

RESUMEN

Genome editing has been attracting increasing attention as a new treatment for several refractory diseases since the CRISPR-Cas discovery has facilitated easy modification of target chromosomal DNA. The concept of treating refractory diseases by genome editing has been achieved in various animal models, and genome editing has been applied to human clinical trials for ß-thalassemia, sickle cell disease, mucopolysaccharidosis, transthyretin amyloidosis, HIV infection, and CAR-T therapy. The genome editing technology targets the germline in industrial applications in animals and plants and is directed at the chromosomal DNA of the somatic cells in human therapeutic applications. Genome editing therapy for germline cells is currently forbidden due to ethical and safety concerns. Concerns regarding genome editing technology include safety (off-target effects) as well as technical aspects (low homologous recombination). Various technological innovations for genome editing are expected to expand its clinical application to various diseases in the future.


Asunto(s)
Edición Génica , Infecciones por VIH , Humanos , Terapia Genética , Tecnología , ADN
8.
Haemophilia ; 27 Suppl 3: 132-141, 2021 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-32638467

RESUMEN

Gene therapy is an opportunity for haemophilia patients to receive a one-time treatment and have lasting factor levels for years or decades instead of dependence on repeated administration within short intervals and on sustained supply of drug. Great strides have been made in the development of gene therapy for haemophilia in the last decade. Adeno-associated virus (AAV) vector-mediated gene transfer in haemophilia A and B has entered the phase III trial stage. Gene transfer by lentiviral vector or gene editing technologies using factor VIII (FVIII) or IX (FIX) genes are now entering clinical evaluation. It is expected that the first FVIII and FIX gene therapy products will soon be approved and distributed in major markets. Global access to gene therapy is a critical goal. This review presents new and ongoing efforts towards this goal in countries other than North America and Europe. In Japan, researchers, regulators and funders have established a promising gene therapy development platform for multiple diseases including haemophilia. Decades of scientific and clinical research in haemophilia gene therapy in China have led to a recently registered clinical trial of AAV-mediated gene therapy for haemophilia B. Other countries are in earlier phases of building gene therapy programmes or participate in international trials. A phase 2 feasibility trial of AAV-mediated FIX gene therapy in low- and middle-income countries aims to demonstrate that gene therapy could become available in resource-constrained socio-economic settings. The different strategies for establishing gene therapy provide opportunities for closing the global gap in haemophilia care.


Asunto(s)
Hemofilia A , Hemofilia B , China , Europa (Continente) , Factor IX/genética , Factor VIII/genética , Terapia Genética , Hemofilia A/genética , Hemofilia A/terapia , Hemofilia B/genética , Hemofilia B/terapia , Humanos
9.
J Orthop Sci ; 26(3): 487-493, 2021 May.
Artículo en Inglés | MEDLINE | ID: mdl-32402506

RESUMEN

BACKGROUND: Research has revealed the crucial roles of inflammasomes in various central nervous system disorders. However, the role of inflammasomes in secondary damage following spinal cord injury (SCI) remains incompletely understood. METHODS: Here, we investigated the role of apoptosis-associated speck-like protein (ASC), an adaptor protein for inflammasome formation, after contusion SCI in ASC homozygous knockout (ASC-/-) mice. Contusion SCI was induced using a force of 60 kdyn, and recovery of open-field locomotor performance was evaluated using the nine-point Basso Mouse Scale (BMS). Bone marrow transplantation (BMT) was performed to create mice chimeric for ASC expression in bone marrow cells. RESULTS: Western blot analysis revealed that protein expression of NLRP3, ASC, Caspase-1, and IL-ß were increased in injured spinal cords compared with sham-control spinal cords at 1 day post injury (dpi). Double immunostaining showed that ASC expression was co-localized to cellular constituents of the spinal cord, including NeuN+ neurons, CD11b+ microglia/macrophages, GFAP+ astrocytes, and MOG+ oligodendrocytes. ASC-/- mice had significantly better locomotor function assessed by BMS than wild-type (WT) mice. ASC-/- mice also had significantly reduced levels of Nlrp3, Casp1, IL1b, Il-6, Tnfa, Cxcl1, and Ly6g mRNA compared with WT mice. BMT (WT→ASC-/-) mice had significantly better BMS scores than BMT (WT→WT) mice. BMT (ASC-/-→WT) mice also had significantly better BMS scores than BMT (WT→WT) mice. However, the statistical significance was limited to time points between 7 and 21 dpi. CONCLUSIONS: These results suggest that ASC-dependent inflammasome formation, especially in resident cells of the spinal cord, plays a pivotal role in the progression of secondary damage following SCI.


Asunto(s)
Inflamasomas , Traumatismos de la Médula Espinal , Proteínas Adaptadoras Transductoras de Señales , Animales , Inflamasomas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Recuperación de la Función , Médula Espinal
10.
Gene Ther ; 27(9): 427-434, 2020 09.
Artículo en Inglés | MEDLINE | ID: mdl-32066928

RESUMEN

Adeno-associated virus (AAV) vectors can transduce hepatocytes efficiently in vivo in various animal species, including humans. Few reports, however, have examined the utility of pigs in gene therapy. Pigs are potentially useful in preclinical studies because of their anatomical and physiological similarity to humans. Here, we evaluated the utility of microminipigs for liver-targeted gene therapy. These pigs were intravenously inoculated with an AAV8 vector encoding the luciferase gene, and gene expression was assessed by an in vivo imaging system. Robust transgene expression was observed almost exclusively in the liver, even though the pig showed a low-titer of neutralizing antibody (NAb) against the AAV8 capsid. We assessed the action of NAbs against AAV, which interfere with AAV vector-mediated gene transfer by intravascular delivery. When a standard dose of vector was administered intravenously, transgene expression was observed in both NAb-negative and low-titer (14×)-positive subjects, whereas gene expression was not observed in animals with higher titers (56×). These results are compatible with our previous observations using nonhuman primates, indicating that pigs are useful in gene therapy experiments, and that the role of low-titer NAb in intravenous administration of the AAV vector shows similarities across species.


Asunto(s)
Anticuerpos Neutralizantes , Cápside , Animales , Dependovirus/genética , Expresión Génica , Técnicas de Transferencia de Gen , Vectores Genéticos/genética , Hígado , Porcinos
11.
Biol Blood Marrow Transplant ; 26(8): 1377-1385, 2020 08.
Artículo en Inglés | MEDLINE | ID: mdl-32311478

RESUMEN

Adult T cell leukemia/lymphoma (ATL) is an aggressive peripheral T cell neoplasm caused by infection with human T cell lymphotropic virus type-1 (HTLV-1). Its prognosis remains extremely poor. Tax, the most important regulatory protein for HTLV-1, is associated with the aggressive proliferation of host cells and is also a major target antigen for CD8+ cytotoxic T cells (CTLs). Based on our previous findings that Tax-specific CTLs with a T cell receptor (TCR) containing a unique amino-acid sequence motif exhibit strong HLA-A*24:02-restricted, Tax301-309-specific activity against HTLV-1, we aimed to develop a Tax-redirected T cell immunotherapy for ATL. TCR-ɑ/ß genes were cloned from a previously established CTL clone and transduced into peripheral blood mononuclear cells (PBMCs) of healthy volunteers using a retroviral siTCR vector. Then the cytotoxic efficacy against HTLV-1-infected T cells or primary ATL cells was assessed both in vitro and in vivo. The redirected CTLs (Tax-siCTLs) produced a large amount of cytokines and showed strong killing activity against ATL/HTLV-1-infected T cells in vitro, although they did not have universal activity against ATL cells. Next, in a xenograft mouse model using an HTLV-1-infected T cell line (MT-2), in all mice treated with Tax-siCTLs, the tumor rapidly diminished and finally disappeared without normal tissue damage, although all mice that were untreated or treated with non-gene-modified PBMCs died because of tumor progression. Our findings confirm that Tax-siCTLs can exert strong anti-ATL/HTLV-1 effects without a significant reaction against normal cells and have the potential to be a novel immunotherapy for ATL patients.


Asunto(s)
Virus Linfotrópico T Tipo 1 Humano , Leucemia-Linfoma de Células T del Adulto , Adulto , Animales , Productos del Gen tax/genética , Genes Codificadores de los Receptores de Linfocitos T , Humanos , Inmunoterapia , Leucemia-Linfoma de Células T del Adulto/terapia , Leucocitos Mononucleares , Ratones , Linfocitos T Citotóxicos
12.
Biochem Biophys Res Commun ; 531(2): 125-132, 2020 10 15.
Artículo en Inglés | MEDLINE | ID: mdl-32782151

RESUMEN

BACKGROUND: Platelets are critical mediators of vascular homeostasis and thrombosis, and also contribute to the development of inflammation. NLRP3 inflammasome is a cytosolic multi-protein complex that consists of NLRP3, ASC and caspase-1, and regulates IL-1ß-mediated inflammation. METHOD AND RESULTS: Using two mouse models of thrombosis (i.e., occlusion of the middle cerebral artery and inferior vena cava), we found that thrombus formation was significantly enhanced in ASC-deficient (ASC-/-) mice, compared to that in wild-type (WT) and IL-1ß-/- mice. ASC deficiency had no effects on blood coagulation parameters (i.e., prothrombin time [PT] and activated partial thromboplastin time [APTT]). Platelets from WT mice express ASC, but neither NLRP3 nor caspase-1. ASC deficiency significantly enhanced the expression of P-selectin and GPIIb/IIIa in response to a GPVI agonist (collagen-related peptide [CRP]), but not to thrombin, in platelets. CRP induced ASC speck formation in WT platelets. ASC deficiency also enhanced cytosolic Ca2+ elevation and phosphorylation of ERK1/2 and Akt in platelets. CONCLUSION: Our results demonstrate that ASC negatively regulates GPVI signaling in platelets and enhances thrombus formation, independent of NLRP3 inflammasome and IL-1ß, and provide novel insights into the link between inflammation and thrombosis.


Asunto(s)
Inflamasomas/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Activación Plaquetaria , Trombosis/metabolismo , Trombosis/patología , Animales , Proteínas Adaptadoras de Señalización CARD/deficiencia , Proteínas Adaptadoras de Señalización CARD/metabolismo , Calcio/metabolismo , Sistema de Señalización de MAP Quinasas , Ratones Endogámicos C57BL , Proteínas Proto-Oncogénicas c-akt/metabolismo
13.
Acta Haematol ; 143(3): 250-259, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-31461700

RESUMEN

BACKGROUND: Danaparoid sodium and synthetic protease inhibitors (SPIs) have been approved for the treatment of disseminated intravascular coagulation (DIC) in Japan. OBJECTIVES: To compare the clinical results of the treatment of DIC with danaparoid or SPIs. METHODS: We retrospectively examined 188 patients with hematological malignancy-related DIC. RESULTS: DIC resolution rate in the danaparoid group was higher than that in the SPIs group (61.5 vs. 42.6%; p = 0.031) on day 7. Multivariate analysis identified the response to chemotherapy as independent predictive factor for DIC resolution on day 7 (odds ratio, OR, 2.28; 95% confidence interval, CI, 1.21-4.31; p = 0.011). While there was no significant difference in the DIC resolution rate on day 14 (75.0 vs. 62.4%; p = 0.117), in a subgroup analysis of patients who did not show an improvement in the underlying disease, the danaparoid group showed a significantly better DIC resolution rate (OR 3.89; 95% CI 1.15-13.2; p = 0.030). There was no difference in the rate of cumulative mortality from bleeding within 28 days between the 2 groups (6.6 vs. 3.3%; p = 0.278). CONCLUSIONS: Danaparoid may be associated with more frequent resolution of DIC in patients with refractory underlying disease.


Asunto(s)
Sulfatos de Condroitina/uso terapéutico , Dermatán Sulfato/uso terapéutico , Neoplasias Hematológicas/sangre , Heparitina Sulfato/uso terapéutico , Inhibidores de Proteasas/uso terapéutico , Adulto , Anciano , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Transfusión de Componentes Sanguíneos , Sulfatos de Condroitina/efectos adversos , Dermatán Sulfato/efectos adversos , Coagulación Intravascular Diseminada/tratamiento farmacológico , Coagulación Intravascular Diseminada/terapia , Femenino , Productos de Degradación de Fibrina-Fibrinógeno/análisis , Fibrinógeno/análisis , Neoplasias Hematológicas/tratamiento farmacológico , Hemorragia/etiología , Hemorragia/mortalidad , Heparitina Sulfato/efectos adversos , Humanos , Masculino , Persona de Mediana Edad , Plasma , Inhibidores de Proteasas/efectos adversos , Tiempo de Protrombina , Estudios Retrospectivos , Resultado del Tratamiento
14.
Clin Exp Hypertens ; 42(4): 365-370, 2020 May 18.
Artículo en Inglés | MEDLINE | ID: mdl-31542950

RESUMEN

Background: Anticoagulant activity and blood pressure increase in the morning. The aim of this study was to evaluate changes of anticoagulant activity, blood pressure and target organ damage in patients with nonvalvular atrial fibrillation (AF) given combination treatment with Xa inhibitor and antihypertensive agent.Methods: We enrolled 72 patients with nonvalvular AF. Rivaroxaban (10-15 mg) was continuously administered once daily over 8 weeks (study period I). For subjects (n = 50) who exhibited uncontrolled morning hypertension (home systolic blood pressure [SBP]≥125 mmHg) at the end of study period I (at 8 weeks), nifedipine CR (20-40 mg) was added at bedtime, and rivaroxaban administration was continued an additional 8 weeks. We assessed prothrombin fragment 1 + 2 (optimal range: 69-229 pmol/L) and D-dimer (negative D-dimer measurement: <1.0 µg/mL).Results: The percentage of patients with optimal-range prothrombin fragment 1 + 2 was significantly increased at 4 weeks compared to baseline (70.8% vs. 86.1%, p = .033). In period II, office and home morning SBP were reduced at 12 compared to 8 weeks (office SBP: 135.2 ± 15.7 vs. 125.6 ± 18.4mmHg, p < .001; home morning SBP: 133.5 ± 10.5 vs. 119.9 ± 12.1mmHg, p<.001).The percentage of patients with negative D-dimer  was increased at 8 weeks compared to baseline (92% vs. 100%, p = .044), and remained at 100% at 16 weeks.Conclusions: Xa inhibitor therapy improved anticoagulant activity, and additional antihypertensive therapy maintained the anticoagulant activity in patients with nonvalvular AF.


Asunto(s)
Coagulación Sanguínea/efectos de los fármacos , Hipertensión , Nifedipino/administración & dosificación , Rivaroxabán/administración & dosificación , Accidente Cerebrovascular/prevención & control , Anciano , Antihipertensivos/administración & dosificación , Fibrilación Atrial/tratamiento farmacológico , Fibrilación Atrial/fisiopatología , Esquema de Medicación , Quimioterapia Combinada/métodos , Inhibidores del Factor Xa/administración & dosificación , Femenino , Humanos , Hipertensión/tratamiento farmacológico , Hipertensión/fisiopatología , Masculino , Accidente Cerebrovascular/sangre , Resultado del Tratamiento
15.
Thromb J ; 16: 5, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-29568240

RESUMEN

BACKGROUND: Although prasugrel exerts stronger antiplatelet effects compared with clopidogrel, the factors affecting platelet reactivity under prasugrel have not been fully determined. This study aimed to find the novel mechanistic differences between two thienopyridines and identify the factor that influence platelet reactivity to each drug. METHODS: Forty patients with stable angina who underwent elective percutaneous coronary intervention were randomly assigned to receive either prasugrel (20 mg) or clopidogrel (300 mg) as a loading dose. Platelet function (light transmission, laser light scattering, and vasodilator-stimulated phosphoprotein phosphorylation) and plasma active metabolite levels were measured after the loading dose. RESULTS: Prasugrel consistently inhibited adenosine diphosphate receptor P2Y12 signalling to abolish amplification of platelet aggregation. Prasugrel abolished even small platelet aggregates composed of less than 100 platelets. On the other hand, clopidogrel inhibited large aggregates but increased small and medium platelet aggregates. Diabetes was the only independent variable for determining antiplatelet effects and active metabolite concentration of prasugrel, but not clopidogrel. Sleep-disordered breathing was significantly correlated with platelet reactivity in patients who had clopidogrel. CONCLUSIONS: Prasugrel efficiently abolishes residual P2Y12 signalling that causes small platelet aggregates, but these small aggregates are not inhibited by clopidogrel. Considering the differential effect of diabetes on antiplatelet effects between these two drugs, the pharmacokinetics of prasugrel, other than cytochrome P450 metabolism, might be affected by diabetes. TRIAL REGISTRATION: UMIN-CTR UMIN000017624, retrospectively registered 21 May 2015.

16.
Rinsho Ketsueki ; 59(10): 2238-2246, 2018.
Artículo en Japonés | MEDLINE | ID: mdl-30305531

RESUMEN

Hemophilia is congenital hemorrhagic disease due to genetic abnormality of blood coagulation factor VIII or factor IX. Hemophilia appears suitable for gene therapy because it is caused by a single gene abnormality, and therapeutic coagulation factor levels vary across a broad range. Since the success of gene therapy eliminates the need for regular administration of factor concentrates, the development has been met with great expectation from patients and their families. In fact, several clinical trials using adeno-associated virus (AAV) vectors have been started in Western countries, and long-term therapeutic effect could be obtained by single injection. More recently, the modifications of therapeutic gene and AAV serotypes have further improved therapeutic effects, and it appears to have reached the level of "cure". However, gene therapy using AAV vectors presents difficulties such as the existence of anti AAV capsid neutralizing antibody and hepatic injury due to cellular immunity with CD8+ T cells. Thus, it is essential to develop technologies adapted to a wide range of hemophilia patients, and to observe long-term safety and therapeutic effect after the gene therapy.


Asunto(s)
Terapia Genética , Hemofilia A/terapia , Hemofilia B/terapia , Dependovirus , Factor IX , Factor VIII , Vectores Genéticos , Humanos
17.
J Med Virol ; 86(11): 1990-7, 2014 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-24136735

RESUMEN

Pre-existing antibodies against adeno-associated virus (AAV), caused by natural AAV infections, interfere with recombinant AAV vector-mediated gene transfer. We studied the prevalence of neutralizing antibodies against AAV serotypes 1, 2, 5, 8, and 9 in healthy subjects (n = 85) and hemophilia patients (n = 59) in a Japanese population. For healthy subjects, the prevalence of neutralizing antibodies against AAV serotypes 1, 2, 5, 8, and 9 was 36.5%, 35.3%, 37.6%, 32.9%, and 36.5%, respectively, while that in hemophilia patients was 39.7%, 28.8%, 35.6%, 32.9%, and 27.4%, respectively. There was no difference in the prevalence of neutralizing antibody against each AAV serotype between the healthy subjects and the hemophilia patients. The prevalence of neutralizing antibodies against all AAV serotypes increased with age in both healthy subjects and hemophilia patients. High titers of neutralizing antibodies against AAV2 (≥1:224) and AAV8 (≥1:224) were more evident in older individuals (≥42 years old). Approximately 50% of all screened individuals were seronegative for neutralizing antibodies against each AAV tested, while approximately 25% of individuals were seropositive for each AAV serotype tested. The prevalence of seronegativity for all AAV serotypes was 67.0% (healthy subjects, 68.6%; hemophilia patients, 65.0%) and 18.6% (healthy subjects, 20.5%; hemophilia patients, 15.7%) in young (<42 years old) and older subjects (≥42 years old), respectively. The findings from this study suggested that young subjects are more likely to be eligible for gene therapy based on AAV vectors delivered via an intravascular route because of the low prevalence of antibodies to AAV capsids.


Asunto(s)
Anticuerpos Neutralizantes/sangre , Anticuerpos Antivirales/sangre , Cápside/inmunología , Dependovirus/inmunología , Infecciones por Parvoviridae/epidemiología , Adulto , Factores de Edad , Pueblo Asiatico , Humanos , Japón/epidemiología , Persona de Mediana Edad , Infecciones por Parvoviridae/virología , Estudios Seroepidemiológicos
18.
Thromb J ; 12(1): 1, 2014 Jan 02.
Artículo en Inglés | MEDLINE | ID: mdl-24383745

RESUMEN

BACKGROUND: Paxillin is a LIM domain protein localized at integrin-mediated focal adhesions. Although paxillin is thought to modulate the functions of integrins, little is known about the contribution of paxillin to signaling pathways in platelets. Here, we studied the role of paxillin in platelet activation in vitro and in vivo. METHODS AND RESULTS: We generated paxillin knockdown (Pxn-KD) platelets in mice by transplanting bone marrow cells transduced with a lentiviral vector carrying a short hairpin RNA sequence, and confirmed that paxillin expression was significantly reduced in platelets derived from the transduced cells. Pxn-KD platelets showed a slight increased in size and augmented integrin αIIbß3 activation following stimulation of multiple receptors including glycoprotein VI and G protein-coupled receptors. Thromboxane A2 biosynthesis and the release of α-granules and dense granules in response to agonist stimulation were also enhanced in Pxn-KD platelets. However, Pxn-KD did not increase tyrosine phosphorylation or intracellular calcium mobilization. Intravital imaging confirmed that Pxn-KD enhanced thrombus formation in vivo. CONCLUSIONS: Our findings suggest that paxillin negatively regulates several common platelet signaling pathways, resulting in the activation of integrin αIIbß3 and release reactions.

19.
Crit Care ; 18(1): R13, 2014 Jan 13.
Artículo en Inglés | MEDLINE | ID: mdl-24410881

RESUMEN

INTRODUCTION: Current criteria for early diagnosis of coagulopathy in sepsis are limited. We postulated that coagulopathy is already complicated with sepsis in the initial phase, and severe coagulopathy or disseminated intravascular coagulation (DIC) becomes overt after progressive consumption of platelet and coagulation factors. To determine early diagnostic markers for severe coagulopathy, we evaluated plasma biomarkers for association with subsequent development of overt DIC in patients with sepsis. METHODS: A single-center, prospective observational study was conducted in an adult ICU at a university hospital. Plasma samples were obtained from patients with sepsis at ICU admission. Fourteen biomarkers including global markers (platelet count, prothrombin time, activated partial thromboplastin time, fibrinogen and fibrin degradation product (FDP)); markers of thrombin generation (thrombin-antithrombin complex (TAT) and soluble fibrin); markers of anticoagulants (protein C (PC) and antithrombin); markers of fibrinolysis (plasminogen, α2-plasmin inhibitor (PI), plasmin-α2-PI complex, and plasminogen activator inhibitor (PAI)-1); and a marker of endothelial activation (soluble E-selectin) were assayed. Patients who had overt DIC at baseline were excluded, and the remaining patients were followed for development of overt DIC in 5 days, and for mortality in 28 days. RESULTS: A total of 77 patients were enrolled, and 37 developed overt DIC within the following 5 days. Most patients demonstrated hemostatic abnormalities at baseline with 98.7% TAT, 97.4% FDP and 88.3% PC. Most hemostatic biomarkers at baseline were significantly associated with subsequent development of overt DIC. Notably, TAT, PAI-1 and PC discriminated well between patients with and without developing overt DIC (area under the receiver operating characteristic curve (AUROC), 0.77 (95% confidence interval, 0.64 to 0.86); 0.87 (0.78 to 0.92); 0.85 (0.76 to 0.91), respectively), and using the three together, significantly improved the AUROC up to 0.95 (vs. TAT, PAI-1, and PC). Among the significant diagnostic markers for overt DIC, TAT and PAI-1 were also good predictors of 28-day mortality (AUROC, 0.77 and 0.81, respectively). CONCLUSIONS: Severe coagulation and fibrinolytic abnormalities on ICU admission were associated with subsequent development of overt DIC. A single measurement of TAT, PAI-1, and PC activity could identify patients with ongoing severe coagulopathy, early in the course of sepsis.


Asunto(s)
Trastornos de la Coagulación Sanguínea/sangre , Péptido Hidrolasas/sangre , Inhibidor 1 de Activador Plasminogénico/sangre , Proteína C/metabolismo , Sepsis/sangre , Índice de Severidad de la Enfermedad , Anciano , Anciano de 80 o más Años , Antitrombina III , Biomarcadores/sangre , Coagulación Sanguínea/fisiología , Trastornos de la Coagulación Sanguínea/diagnóstico , Trastornos de la Coagulación Sanguínea/epidemiología , Diagnóstico Precoz , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Sepsis/diagnóstico , Sepsis/epidemiología
20.
Mol Ther ; 21(2): 318-23, 2013 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-23247100

RESUMEN

Neutralizing antibodies (NAbs) against adeno-associated viruses (AAVs) are known to interfere with AAV vector-mediated gene transfer by intravascular delivery. Evading the inhibitory effects of antibodies against AAV vectors is necessary for efficient transfer of therapeutic genes clinically. For this purpose, we tested the efficacy of saline flushing in order to avoid contact of vectors with NAbs present in blood. Direct injection of the AAV8 vector carrying the factor IX (FIX) gene into the portal vein of macaques using saline flushing achieved transgene-derived FIX expression (4.7 ± 2.10-10.1 ± 5.45% of normal human FIX concentration) in the presence of NAbs. Expression was as efficient as that (5.43 ± 2.59-12.68 ± 4.83%) in macaques lacking NAbs. We next tested the efficacy of saline flushing using less invasive balloon catheter-guided injection. This approach also resulted in efficient expression of transgene-derived FIX (2.5 ± 1.06-9.0 ± 2.37%) in the presence of NAbs (14-56× dilutions). NAbs at this range of titers reduced the efficiency of transduction in the macaque liver by 100-fold when the same vector was injected into mesenteric veins without balloon catheters. Our results suggest that portal vein-directed vector delivery strategies with flushing to remove blood are efficacious for minimizing the inhibitory effect of anti-AAV antibodies.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Dependovirus/inmunología , Expresión Génica , Técnicas de Transferencia de Gen , Hígado/metabolismo , Animales , Catéteres , Dependovirus/genética , Factor IX/genética , Terapia Genética , Vectores Genéticos , Humanos , Macaca , Mutación Missense , Vena Porta , Transgenes
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA