RESUMO
Nanoparticle-based thrombolysis is a potential new treatment for stroke. The aim of this study was to investigate the efficacy of targeted thrombolysis using recombinant tissue plasminogen activator (rtPA). The rtPA was covalently bound to magnetic nanoparticles (MNP) and maintained at the target site using an external magnet. Polyacrylic acid (PAA)-coated MNP were synthesized and rtPA was then bound to the resultant PAA-MNP via carbodiimide-mediated amide bonds. For the in vitro tests, blood clots were formed in plastic centrifuge tubes with anti-coagulated plasma, thrombin and calcium chloride. For the in vivo tests, mice with ferric chloride-induced distal middle cerebral artery occlusion were treated with phosphate-buffered saline (PBS), MNP, rtPA, or MNP-rtPA (n = 6 mice per group). The binding efficacy was 80.7 ± 1.5 µg rtPA bound to 1 mg PAA-MNP. In the in vitro tests, the mean lysis percentage dramatically increased from 1.28% in the MNP group without rotation to 77.40% in the rtPA + MNP group with rotating magnetic field. The lysis efficiency of MNP-rtPA was 27.3 ± 1.3%, and it increased to 42.8 ± 2.8% with magnetic field rotation. The mean sizes of the infarct areas of the PBS, MNP, rtPA, and MNP-rtPA mouse groups were 20.09 ± 6.07, 18.28 ± 2.69, 8.65 ± 3.63 and 4.40 ± 2.46 mm3, respectively. Thus, targeted MNP-rtPA accelerated thrombolysis and reduced the infarct area in a mouse model of cerebral embolism. This approach may serve as a feasible and effective treatment for embolic cerebral ischemia.
Assuntos
Resinas Acrílicas/química , Isquemia Encefálica/tratamento farmacológico , Materiais Revestidos Biocompatíveis/química , Nanopartículas de Magnetita/química , Proteínas Recombinantes/uso terapêutico , Acidente Vascular Cerebral/tratamento farmacológico , Ativador de Plasminogênio Tecidual/uso terapêutico , Animais , Infarto Encefálico/patologia , Isquemia Encefálica/complicações , Modelos Animais de Doenças , Embolia/complicações , Embolia/tratamento farmacológico , Embolia/patologia , Infarto da Artéria Cerebral Média/complicações , Infarto da Artéria Cerebral Média/tratamento farmacológico , Masculino , Camundongos Endogâmicos C57BL , Ratos Sprague-Dawley , Acidente Vascular Cerebral/complicações , Trombose/complicações , Trombose/patologia , Fatores de TempoRESUMO
Experimental modeling of traumatic brain injury (TBI) in animals has identified several potential means and interventions that might have beneficial applications for treating traumatic brain injury clinically. Several of these interventions have been applied and tried with humans that are at different phases of testing (completed, prematurely terminated and others in progress). The promising results achieved in the laboratory with animal models have not been replicated with human trails as expected. This review will highlight some insights and significance attained via laboratory animal modeling of TBI as well as factors that require incorporation into the experimental studies that could help in translating results from laboratory to the bedside. Major progress has been made due to laboratory studies; in explaining the mechanisms as well as pathophysiological features of brain damage after TBI. Attempts to intervene in the cascade of events occurring after TBI all rely heavily on the knowledge from basic laboratory investigations. In looking to discover treatment, this review will endeavor to sight and state some central discrepancies between laboratory models and clinical scenarios.
Assuntos
Lesões Encefálicas Traumáticas/fisiopatologia , Lesões Encefálicas Traumáticas/terapia , Modelos Animais de Doenças , Animais , Lesões Encefálicas Traumáticas/genética , HumanosRESUMO
Abstract Pyroptosis is considered one of a critical factor in the recovery of neurological function following traumatic brain injury. Brain injury activates a molecular signaling cascade associated with pyroptosis and inflammation, including NLRP3, inflammatory cytokines, caspase-1, gasdermin D (GSDMD), and other pyroptosis-related proteins. In this study, we explored the neuroprotective effects of LDC7559, a GSDMD inhibitor. Briefly, LDC7559, siRNA-GSDMD (si-GSDMD), or equal solvent was administrated to mice with a lipopolysaccharide + nigericin (LPS + Nig) model in vitro or with controlled cortical impact brain injury. The findings revealed that inflammation and pyroptosis levels were decreased by LDC7559 or si-GSDMD treatment both in vitro and in vivo. Immunofluorescence staining, brain water content, hematoxylin and eosin staining, and behavioral investigations suggested that LDC7559 or si-GSDMD inhibited microglial proliferation, ameliorated cerebral edema, reduced brain tissue loss, and promoted brain function recovery. Taken together, LDC7559 may inhibit pyroptosis and reduce inflammation by inhibiting GSDMD, thereby promoting the recovery of neurological function.
Assuntos
Lesões Encefálicas Traumáticas , Lesões Encefálicas , Fármacos Neuroprotetores , Camundongos , Animais , Microglia/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/farmacologia , Fármacos Neuroprotetores/farmacologia , Piroptose , Inflamação/metabolismo , Lesões Encefálicas Traumáticas/metabolismo , Lesões Encefálicas/metabolismoRESUMO
BACKGROUND: Ferric chloride is widely utilized in inducing thrombosis in small vessels of experimental animals. However, the lack of its application in large blood vessels of experimental animals and inconsistent concentration has limited its application. Therefore, we systematically tested the most suitable concentration and reliable induction time in the experiment of using ferric chloride to induce rat carotid artery thrombosis. METHODS: In this study, we selected the common carotid artery of 59 Sprague-Dawley rats as the target vessel. The exploration process was divided into three stages. First, to determine the optimum induction concentration, we compared the effects of 30-60% ferric chloride on thrombus formation within 24 h. Second, to confirm the handling time, we tested different induction times from 3 min to 10 min. Lastly, we used the thrombolytic drug rt-PA to detect whether the formed thrombus can be lysed. Doppler blood flow imaging and H-E staining were employed to estimate the blood flow and thrombus. The ATP levels in the brain were measured using a bioluminescence ATP assay kit. RESULTS: We found that the application of 50% ferric chloride for 10 min was enough to successfully induce thrombosis in the rat carotid artery and without spontaneous thrombolysis after 24 h. It is better than other concentrations and will lead to the decline of the ATP content in the ischemic hemisphere. CONCLUSIONS: Our results indicate that the rat carotid artery thrombosis model induced by 50% ferric chloride for 10 min is stable and reliable.
RESUMO
Bone marrow mesenchymal stem cells (BMMSCs)-based therapy has emerged as a promising novel therapy for Traumatic Brain Injury (TBI). However, the therapeutic quantity of viable implanted BMMSCs necessary to initiate efficacy is still undetermined. Increased oxidative stress following TBI, which leads to the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase signaling pathway, has been implicated in accounting for the diminished graft survival and therapeutic effect. To prove this assertion, we silenced the expression of NADPH subunits (p22-phox, p47-phox, and p67-phox) and small GTPase Rac1 in BMMSCs using shRNA. Our results showed that silencing these proteins significantly reduced oxidative stress and cell death/apoptosis, and promoted implanted BMMSCs proliferation after TBI. The most significant result was however seen with Rac1 silencing, which demonstrated decreased expression of apoptotic proteins, enhanced in vitro survival ratio, reduction in TBI lesional volume and significant improvement in neurological function post shRac1-BMMSCs transplantation. Additionally, two RNA-seq hub genes (VEGFA and MMP-2) were identified to play critical roles in shRac1-mediated cell survival. In summary, we propose that knockdown of Rac1 gene could significantly boost cell survival and promote the recovery of neurological functions after BMMSCs transplantation in TBI mice.
Assuntos
Lesões Encefálicas Traumáticas/terapia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/metabolismo , NADPH Oxidases/genética , Recuperação de Função Fisiológica , Proteínas rac1 de Ligação ao GTP/genética , Animais , Apoptose/genética , Comportamento Animal , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Transplante de Medula Óssea , Lesões Encefálicas Traumáticas/fisiopatologia , Proliferação de Células/genética , Sobrevivência Celular/genética , Técnicas de Silenciamento de Genes , Inativação Gênica , Técnicas In Vitro , Metaloproteinase 2 da Matriz/genética , Células-Tronco Mesenquimais/citologia , Camundongos , NADH NADPH Oxirredutases/genética , Neurônios/citologia , Estresse Oxidativo/genética , RNA Interferente Pequeno , Ratos , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
Traumatic brain injury (TBI) is a leading cause of mortality and disability worldwide. Although treatment guidelines have been developed, no best treatment option or medicine for this condition exists. Recently, mesenchymal stem cells (MSCs)-derived exosomes have shown lots of promise for the treatment of brain disorders, with some results highlighting the neuroprotective effects through neurogenesis and angiogenesis after TBI. However, studies focusing on the role of exosomes in the early stages of neuroinflammation post-TBI are not sufficient. In this study, we investigated the role of bone mesenchymal stem cells (BMSCs)-exosomes in attenuating neuroinflammation at an early stage post-TBI and explored the potential regulatory neuroprotective mechanism. We administered 30 µg protein of BMSCs-exosomes or an equal volume of phosphate-buffered saline (PBS) via the retro-orbital route into C57BL/6 male mice 15 min after controlled cortical impact (CCI)-induced TBI. The results showed that the administration of BMSCs-exosomes reduced the lesion size and improved the neurobehavioral performance assessed by modified Neurological Severity Score (mNSS) and rotarod test. In addition, BMSCs-exosomes inhibited the expression of proapoptosis protein Bcl-2-associated X protein (BAX) and proinflammation cytokines, tumor necrosis factor-α (TNF-α) and interleukin (IL)-1ß, while enhancing the expression of the anti-apoptosis protein B-cell lymphoma 2 (BCL-2). Furthermore, BMSCs-exosomes modulated microglia/macrophage polarization by downregulating the expression of inducible nitric oxide synthase (INOS) and upregulating the expression of clusters of differentiation 206 (CD206) and arginase-1 (Arg1). In summary, our result shows that BMSCs-exosomes serve a neuroprotective function by inhibiting early neuroinflammation in TBI mice through modulating the polarization of microglia/macrophages. Further research into this may serve as a potential therapeutic strategy for the future treatment of TBI.
RESUMO
Intracerebral hemorrhage (ICH) refers to bleeding in the brain and is associated with the release of large amount of inflammasomes, and the activation of different cell death pathways. These cell death pathways lead to removal of inactivated and damaged cells and also result in neuronal cell damage. Pyroptosis is a newly discovered cell death pathway that has gained attention in recent years. This pathway mainly depends on activation of caspase-1-mediated cascades to cause cell death. We tested a well-known selective inhibitor of caspase-1, AC-YVAD-CMK, which has previously been found to have neuroprotective effects in ICH mice model, to ascertain its effects on the activation of inflammasomes mediated pyroptosis. Our results showed that AC-YVAD-CMK could reduce caspase-1 activation and inhibit IL-1ß production and maturation, but has no effect on NLRP3 expression, an upstream inflammatory complex. AC-YVAD-CMK administration also resulted in reduction in M1-type microglia polarization around the hematoma, while increasing the number of M2-type cells. Furthermore, AC-YVAD-CMK treated mice showed some recovery of neurological function after hemorrhage especially at the hyperacute and subacute stage resulting in some degree of limb movement. In conclusion, we are of the view that AC-YVAD-CMK could inhibit pyroptosis, decrease the secretion or activation of inflammatory factors, and affect the polarization of microglia resulting in improvement of neurological function after ICH.
Assuntos
Clorometilcetonas de Aminoácidos/farmacologia , Hemorragia Cerebral/metabolismo , Fármacos Neuroprotetores/farmacologia , Piroptose/efeitos dos fármacos , Animais , Comportamento Animal/efeitos dos fármacos , Caspase 1/metabolismo , Inibidores de Caspase , Inflamassomos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismoRESUMO
The effectiveness of noninvasive treatment for neurological disease is generally limited by the poor entry of therapeutic agents into the central nervous system (CNS). Most CNS drugs cannot permeate into the brain parenchyma because of the blood-brain barrier thus, overcoming this problem has become one of the most significant challenges in the development of neurological therapeutics. Nanotechnology has emerged as an innovative alternative for treating neurological diseases. In fact, rapid advances in nanotechnology have provided promising solutions to this challenge. This review highlights the applications of nanomaterials in the developing neurological field and discusses the evidence for their efficacies.
Assuntos
Fármacos do Sistema Nervoso Central/administração & dosagem , Sistemas de Liberação de Medicamentos , Nanoestruturas , Doenças do Sistema Nervoso/tratamento farmacológico , Animais , Sistemas de Liberação de Medicamentos/efeitos adversos , Humanos , Nanoestruturas/efeitos adversosRESUMO
Intracerebral hemorrhage (ICH) which refers to bleeding in the brain is a very deleterious condition with high mortality and disability rate. Surgery or conservative therapy remains the treatment option. Various studies have divided the disease process of ICH into primary and secondary injury, for which knowledge into these processes has yielded many preclinical and clinical treatment options. The aim of this review is to highlight some of the new experimental drugs as well as other treatment options like stem cell therapy, rehabilitation, and nanomedicine and mention some translational clinical applications that have been done with these treatment options.