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Purpose: Exosomes are membrane vesicles secreted by various cells and play a crucial role in intercellular communication. They can be excellent delivery vehicles for oligonucleotide drugs, such as microRNAs, due to their high biocompatibility. MicroRNAs have been shown to be more stable when incorporated into exosomes; however, the lack of targeting and immune evasion is still the obstacle to the use of these microRNA-containing nanocarriers in clinical settings. Our goal was to produce functional exosomes loaded with target ligands, immune evasion ligand, and oligonucleotide drug through genetic engineering in order to achieve more precise medical effects. Methods: To address the problem, we designed engineered exosomes with exogenous cholecystokinin (CCK) or somatostatin (SST) as the targeting ligand to direct the exosomes to the brain, as well as transduced CD47 proteins to reduce the elimination or phagocytosis of the targeted exosomes. MicroRNA-29b-2 was the tested oligonucleotide drug for delivery because our previous research showed that this type of microRNA was capable of reducing presenilin 1 (PSEN1) gene expression and decreasing the ß-amyloid accumulation for Alzheimer's disease (AD) in vitro and in vivo. Results: The engineered exosomes, containing miR29b-2 and expressing SST and CD47, were produced by gene-modified dendritic cells and used in the subsequent experiments. In comparison with CD47-CCK exosomes, CD47-SST exosomes showed a more significant increase in delivery efficiency. In addition, CD47-SST exosomes led to a higher delivery level of exosomes to the brains of nude mice when administered intravenously. Moreover, it was found that the miR29b-2-loaded CD47-SST exosomes could effectively reduce PSEN1 in translational levels, which resulted in an inhibition of beta-amyloid oligomers production both in the cell model and in the 3xTg-AD animal model. Conclusion: Our results demonstrated the feasibility of the designed engineered exosomes. The application of this exosomal nanocarrier platform can be extended to the delivery of other oligonucleotide drugs to specific tissues for the treatment of diseases while evading the immune system.
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Enfermedad de Alzheimer , Péptidos beta-Amiloides , Encéfalo , Antígeno CD47 , Exosomas , MicroARNs , Presenilina-1 , Receptores de Somatostatina , Animales , Exosomas/metabolismo , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/genética , MicroARNs/genética , MicroARNs/administración & dosificación , Presenilina-1/genética , Encéfalo/metabolismo , Receptores de Somatostatina/genética , Receptores de Somatostatina/metabolismo , Péptidos beta-Amiloides/metabolismo , Ratones , Antígeno CD47/genética , Antígeno CD47/metabolismo , Somatostatina , Humanos , Modelos Animales de EnfermedadRESUMEN
A patient-friendly and efficient treatment method for patients with spinocerebellar ataxia type 3 (SCA3) was provided through a nose-to-brain liposomal system. Initially, PGK1 was overexpressed in HEK 293-84Q-GFP diseased cells (HEK 293-84Q-GFP-PGK1 cells) to confirm its effect on the diseased protein polyQ. A decrease in polyQ expression was demonstrated in HEK 293-84Q-GFP-PGK1 cells compared to HEK 293-84Q-GFP parental cells. Subsequently, PGK1 was encapsulated in a liposomal system to evaluate its therapeutic efficiency in SCA3. The optimized liposomes exhibited a significantly enhanced positive charge, facilitating efficient intracellular protein delivery to the cells. The proteins were encapsulated within the liposomes using an optimized method involving a combination of heat shock and sonication. The liposomal system was further demonstrated to be deliverable to the brain via intranasal administration. PGK1/liposomes were intranasally delivered to SCA3 mice, which subsequently exhibited an amelioration of motor impairment, as assessed via the accelerated rotarod test. Additionally, fewer shrunken morphology Purkinje cells and a reduction in polyQ expression were observed in SCA3 mice that received PGK1/liposomes but not in the untreated, liposome-only, or PGK1-only groups. This study provides a non-invasive route for protein delivery and greater delivery efficiency via the liposomal system for treating neurodegenerative diseases.
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Administración Intranasal , Encéfalo , Liposomas , Enfermedad de Machado-Joseph , Fosfoglicerato Quinasa , Animales , Humanos , Fosfoglicerato Quinasa/genética , Encéfalo/metabolismo , Células HEK293 , Enfermedad de Machado-Joseph/tratamiento farmacológico , Enfermedad de Machado-Joseph/genética , Ratones , Péptidos/administración & dosificación , Péptidos/químicaRESUMEN
Exosome therapy is a novel trend in regeneration medicine. However, identifying a suitable biomarker that can associate the therapeutic efficacy of exosomes with SCA3/MJD is essential. In this study, parental cells were preconditioned with butylidenephthalide (Bdph) for exosome preparation to evaluate the therapeutic effect of SCA3/MJD. The therapeutic agent hsa-miRNA-6780-5p was enriched up to 98-fold in exosomes derived from butylidenephthalide (Bdph)-preconditioned human olfactory ensheathing cells (hOECs) compared with that in naïve hOECs exosomes. The particle sizes of exosomes derived from naïve hOECs and those derived from hOECs preconditioned with Bdph were approximately 113.0 ± 3.5 nm and 128.9 ± 0.7 nm, respectively. A liposome system was used to demonstrate the role of hsa-miRNA-6780-5p, wherein hsa-miRNA-6780-5p was found to enhance autophagy and inhibit the expression of spinocerebellar ataxia type 3 (SCA3) disease proteins with the polyglutamine (polyQ) tract. Exosomes with enriched hsa-miRNA-6780-5p were further applied to HEK-293-84Q cells, leading to decreased expression of polyQ and increased autophagy. The results were reversed when 3MA, an autophagy inhibitor, was added to the cells treated with hsa-miRNA-6780-5p-enriched exosomes, indicating that the decreased polyQ expression was modulated via autophagy. SCA3 mice showed improved motor coordination behavior when they intracranially received exosomes enriched with hsa-miRNA-6780-5p. SCA3 mouse cerebellar tissues treated with hsa-miRNA-6780-5p-enriched exosomes showed decreased expression of polyQ and increased expression of LC3II/I, an autophagy marker. In conclusion, our findings can serve as a basis for developing an alternative therapeutic strategy for SCA3 disease treatment using miRNA-enriched exosomes derived from chemically preconditioned cells.
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Exosomas , Enfermedad de Machado-Joseph , MicroARNs , Humanos , Ratones , Animales , Enfermedad de Machado-Joseph/tratamiento farmacológico , Enfermedad de Machado-Joseph/metabolismo , Exosomas/metabolismo , Células HEK293 , MicroARNs/metabolismoRESUMEN
Human pluripotent stem cell (hPSC)-derived motor neurons (MNs) act as models for motor neuron diseases (MNDs), such as amyotrophic lateral sclerosis (ALS) or spinal muscular atrophy. However, the MN differentiation efficiency and viability following cryopreservation require further development for application in large-scale studies and drug screening. Here, we developed a robust protocol to convert hPSCs into MN cryopreservation stocks (hPSCs were converted into >92% motor neural progenitors and >91% MNs). Near-mature MNs were cryopreserved at a high thawing survival rate and 89% MN marker expression on day 32. Moreover, these MNs exhibited classical electrophysiological properties and neuromuscular junction (NMJ) formation ability within only 4−6 days after thawing. To apply this platform as an MND model, MN stocks were generated from SOD1G85R, SOD1G85G isogenic control, and sporadic ALS hPSC lines. The thawed ALS MNs expressed ALS-specific cytopathies, including SOD1 protein aggregation and TDP-43 redistribution. Thus, a stable and robust protocol was developed to generate ready-to-use cryopreserved MNs without further neuronal maturation processes for application in MND mechanistic studies, NMJ model establishment, and large-scale drug screening.
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Esclerosis Amiotrófica Lateral , Células Madre Pluripotentes Inducidas , Células Madre Pluripotentes , Humanos , Esclerosis Amiotrófica Lateral/metabolismo , Superóxido Dismutasa-1/metabolismo , Neuronas Motoras/metabolismo , Células Madre Pluripotentes/metabolismo , CriopreservaciónRESUMEN
Retinal pigmented epithelial (RPE) cells possess high mitochondria content for energy production, which is required for phagocytosis and vision cycle metabolism. The mitochondrial integrity in RPE cells helps the homeostasis of photoreceptor turnover and prevents retina aging and degeneration. Mitochondrial transplantation benefits the recovery of several acute inflammatory diseases, leading us to investigate the effects of mitochondrial transplantation on retina degeneration. Allogeneic mitochondria were isolated and delivered into the vitreous chamber in the Royal College of Surgeons (RCS) rats, which exhibit inherited and early-onset retina degeneration. The progress of retina degeneration was examined with optical coherence tomography (OCT) and visual evoked potential (VEP) to determine the retina thickness and integrity of afferent electrical signals from affected eyes, respectively. We found that mitochondria engraftment moderately attenuated the degeneration of retinal layers in RCS rats by histological examination. This result was consistent with the OCT measurement of retina thickness around the optic disc. The VEP analysis revealed that the peak one (N1) latency, representing the arriving time of electrical impulse from the retina to cortex, was substantially maintained as the normal value after the mitochondrial transplantation. This result suggests that the intra-vitreous transplanted mitochondria ameliorate the degeneration of photoreceptors in RCS rats and might be potential for clinical application.
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Presenilin-1 (PSEN1) is a crucial subunit within the γ-secretase complex and regulates ß-amyloid (Aß) production. Accumulated evidence indicates that n-butylidenephthalide (BP) acts effectively to reduce Aß levels in neuronal cells that are derived from trisomy 21 (Ts21) induced pluripotent stem cells (iPSCs). However, the mechanism underlying this effect remains unclear. This article aims to investigate the possible mechanisms through which BP ameliorates the development of Alzheimer's disease (AD) and verify the effectiveness of BP through animal experiments. Results from RNA microarray analysis showed that BP treatment in Ts21 iPSC-derived neuronal cells reduced long noncoding RNA (lncRNA) CYP3A43-2 levels and increased microRNA (miR)-29b-2-5p levels. Bioinformatics tool prediction analysis, biotin-labeled miR-29b-2-5p pull-down assay, and dual-luciferase reporter assay confirmed a direct negative regulatory effect for miRNA29b-2-5p on lnc-RNA-CYP3A43-2 and PSEN1. Moreover, BP administration improved short-term memory and significantly reduced Aß accumulation in the hippocampus and cortex of 3xTg-AD mice but failed in miR-29b-2-5p mutant mice generated by CRISP/Cas9 technology. In addition, analysis of brain samples from patients with AD showed a decrease in microRNA-29b-2-5p expression in the frontal cortex region. Our results provide evidence that the LncCYP3A43-2/miR29-2-5p/PSEN1 network might be involved in the molecular mechanisms underlying BP-induced Aß reduction.
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Enfermedad de Alzheimer , MicroARNs , ARN Largo no Codificante , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/metabolismo , Secretasas de la Proteína Precursora del Amiloide/genética , Péptidos beta-Amiloides/genética , Péptidos beta-Amiloides/metabolismo , Animales , Biotina , Cognición , Ratones , MicroARNs/metabolismo , Placa Amiloide , Presenilina-1/genética , ARN Largo no Codificante/genéticaRESUMEN
Spinocerebellar ataxia type 3 (SCA3) is characterized by the over-repetitive CAG codon in the ataxin-3 gene (ATXN3), which encodes the mutant ATXN3 protein. The pathological defects of SCA3 such as the impaired aggresomes, autophagy, and the proteasome have been reported previously. To date, no effective treatment is available for SCA3 disease. This study aimed to study anti-excitotoxic effects of n-butylidenephthalide by chemically insulted Purkinje progenitor cells derived from SCA3 iPSCs. We successfully generated Purkinje progenitor cells (PPs) from SCA3 patient-derived iPSCs. The PPs, expressing both neural and Purkinje progenitor's markers, were acquired after 35 days of differentiation. In comparison with the PPs derived from control iPSCs, SCA3 iPSCs-derived PPs were more sensitive to the excitotoxicity induced by quinolinic acid (QA). The observations of QA-treated SCA3 PPs showing neural degeneration including neurite shrinkage and cell number decrease could be used to quickly and efficiently identify drug candidates. Given that the QA-induced neural cell death of SCA3 PPs was established, the activity of calpain in SCA3 PPs was revealed. Furthermore, the expression of cleaved poly (ADP-ribose) polymerase 1 (PARP1), a marker of apoptotic pathway, and the accumulation of ATXN3 proteolytic fragments were observed. When SCA3 PPs were treated with n-butylidenephthalide (n-BP), upregulated expression of calpain 2 and concurrent decreased level of calpastatin could be reversed, and the overall calpain activity was accordingly suppressed. Such findings reveal that n-BP could not only inhibit the cleavage of ATXN3 but also protect the QA-induced excitotoxicity from the Purkinje progenitor loss.
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Ataxina-3/metabolismo , Anhídridos Ftálicos/farmacología , Células de Purkinje/efectos de los fármacos , Proteínas Represoras/metabolismo , Animales , Autofagia/efectos de los fármacos , Calpaína/metabolismo , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Enfermedad de Machado-Joseph/metabolismo , Masculino , Complejo de la Endopetidasa Proteasomal/metabolismo , Células de Purkinje/metabolismoRESUMEN
In recurrent glioblastoma, Gliadel wafer implantation after surgery has been shown to result in incomplete chemical removal of residual tumor and development of brain edema. Furthermore, temozolomide (TMZ) resistance caused by O6-methylguanine-DNA-methyltransferase (MGMT) activation and programmed cell death-ligand 1 (PD-L1) expression leads to immune-cold lesions that result in poorer prognosis. Cerebraca wafer, a biodegradable polymer containing (Z)-n-butylidenephthalide (BP), is designed to eliminate residual tumor after glioma resection. An open-label, one-arm study with four dose cohorts, involving a traditional 3 + 3 dose escalation clinical trial, of the Cerebraca wafer combined with TMZ on patients with recurrent high-grade glioma, was conducted. Of the 12 patients who receive implantation of Cerebraca wafer, there were no drug-related adverse events (AEs) or serious AEs (SAEs). The median overall survival (OS) of patients receiving low-dose Cerebraca wafer was 12 months in the group with >25% wafer coverage of the resected tumor, which is longer than OS duration in previously published studies (Gliadel wafer, 6.4 months). Patients who received high-dose Cerebraca wafer treatment had not yet died at the data cut-off date; a 100% progression-free survival (PFS) rate at six month was achieved, indicating the median OS of cohort IV was more than 17.4 months. In vitro study of the primary cells collected from the patients revealed that the IC50 of BP against tumor stem cells was four times lower than that of bis-chloroethylnitrosourea (BCNU). A synergistic effect between BP and TMZ was demonstrated by a reduction in MGMT expression. Furthermore, BP inhibited PD-L1 expression, thereby activating T-cell cytotoxicity and increasing interferon-gamma (IFN-γ) secretion. The better therapeutic effect of Cerebraca wafer on recurrent high-grade glioma could occur through re-sensitization of TMZ and reduction of PD-L1.
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Cognitive impairment is a serious side effect of post-myocardial infarction (MI) course. We have recently demonstrated that human adipose-derived stem cells (hADSCs) ameliorated myocardial injury after MI by attenuating reactive oxygen species (ROS) levels. Here, we studied whether the beneficial effects of intramyocardial hADSC transplantation can extend to the brain and how they may attenuate cognitive dysfunction via modulating ROS after MI. After coronary ligation, male Wistar rats were randomized via an intramyocardial route to receive either vehicle, hADSC transplantation (1 × 106 cells), or the combination of hADSCs and 3-Morpholinosydnonimine (SIN-1, a peroxynitrite donor). Whether hADSCs migrated into the hippocampus was assessed by using human-specific primers in qPCR reactions. Passive avoidance test was used to assess cognitive performance. Postinfarction was associated with increased oxidative stress in the myocardium, circulation, and hippocampus. This was coupled with decreased numbers of dendritic spines as well as a significant downregulation of synaptic plasticity consisting of synaptophysin and PSD95. Step-through latency during passive avoidance test was impaired in vehicle-treated rats after MI. Intramyocardial hADSC injection exerted therapeutic benefits in improving cardiac function and cognitive impairment. None of hADSCs was detected in rat's hippocampus at the 3rd day after intramyocardial injection. The beneficial effects of hADSCs on MI-induced histological and cognitive changes were abolished after adding SIN-1. MI-induced ROS attacked the hippocampus to induce neurodegeneration, resulting in cognitive deficit. The remotely intramyocardial administration of hADSCs has the capacity of improved synaptic neuroplasticity in the hippocampus mediated by ROS, not the cell engraftment, after MI. KEY MESSAGES: Human adipose-derived stem cells (hADSCs) ameliorated injury after myocardial infarction by attenuating reactive oxygen species (ROS) levels. Intramyocardial administration of hADSCs remotely exerted therapeutic benefits in improving cognitive impairment after myocardial infarction. The improved synaptic neuroplasticity in the hippocampus was mediated by hADSC-inhibiting ROS, not by the stem cell engraftment.
Asunto(s)
Tejido Adiposo/citología , Disfunción Cognitiva/terapia , Infarto del Miocardio/terapia , Trasplante de Células Madre , Animales , Encéfalo/metabolismo , Encéfalo/patología , Disfunción Cognitiva/metabolismo , Disfunción Cognitiva/patología , Disfunción Cognitiva/fisiopatología , Humanos , Masculino , Infarto del Miocardio/metabolismo , Infarto del Miocardio/patología , Infarto del Miocardio/fisiopatología , Miocardio/metabolismo , Estrés Oxidativo , Ratas Wistar , Células Madre , Superóxidos/sangre , Superóxidos/metabolismo , Microglobulina beta-2/metabolismoRESUMEN
Amyotrophic lateral sclerosis (ALS) is a progressive nervous system disease that causes motor neuron (MN) degeneration and results in patient death within a few years. To recapitulate the cytopathies of ALS patients' MNs, SOD1G85R mutant and corrected SOD1G85G isogenic-induced pluripotent stem cell (iPSC) lines were established. Two SOD1 mutant ALS (SOD1G85R and SOD1D90A), two SOD1 mutant corrected (SOD1G85G and SOD1D90D), and one sporadic ALS iPSC lines were directed toward MNs. After receiving ~90% purity for MNs, we first demonstrated that SOD1G85R mutant ALS MNs recapitulated ALS-specific nerve fiber aggregates, similar to SOD1D90A ALS MNs in a previous study. Moreover, we found that both SOD1 mutant MNs showed ALS-specific neurite degenerations and neurotransmitter-induced calcium hyperresponsiveness. In a small compound test using these MNs, we demonstrated that gastrodin, a major ingredient of Gastrodia elata, showed therapeutic effects that decreased nerve fiber cytopathies and reverse neurotransmitter-induced hyperresponsiveness. The therapeutic effects of gastrodin applied not only to SOD1 ALS MNs but also to sporadic ALS MNs and SOD1G93A ALS mice. Moreover, we found that coactivation of the GSK3ß and IGF-1 pathways was a mechanism involved in the therapeutic effects of gastrodin. Thus, the coordination of compounds that activate these two mechanisms could reduce nerve fiber cytopathies in SOD1 ALS MNs. Interestingly, the therapeutic role of GSK3ß activation on SOD1 ALS MNs in the present study was in contrast to the role previously reported in research using cell line- or transgenic animal-based models. In conclusion, we identified in vitro ALS-specific nerve fiber and neurofunctional markers in MNs, which will be useful for drug screening, and we used an iPSC-based model to reveal novel therapeutic mechanisms (including GSK3ß and IGF-1 activation) that may serve as potential targets for ALS therapy.
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Esclerosis Amiotrófica Lateral/metabolismo , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Factor I del Crecimiento Similar a la Insulina/metabolismo , Neuronas Motoras/patología , Mutación/genética , Fibras Nerviosas/patología , Superóxido Dismutasa-1/genética , Animales , Axones/efectos de los fármacos , Axones/patología , Alcoholes Bencílicos/farmacología , Calcio/metabolismo , Diferenciación Celular , Glucósidos/farmacología , Ácido Glutámico/farmacología , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Ratones Transgénicos , Degeneración Nerviosa , Neuritas/patología , Análisis de SupervivenciaRESUMEN
Human pluripotent stem cells (hPSCs) are sources of several somatic cell types for human developmental studies, in vitro disease modeling, and cell transplantation therapy. Improving strategies of derivation of high-purity specific neural and glial lineages from hPSCs is critical for application to the study and therapy of the nervous system. Here, we will focus on the principles behind establishment of neuron and glia differentiation methods according to developmental studies. We will also highlight the limitations and challenges associated with the differentiation of several "difficult" neural lineages and delay in neuronal maturation and functional integration. To overcome these challenges, we will introduce strategies and novel technologies aimed at improving the differentiation of various neural lineages to expand the application potential of hPSCs to the study of the nervous system.
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Neurogénesis/genética , Neuronas/metabolismo , Células Madre Pluripotentes/metabolismo , Diferenciación Celular , HumanosRESUMEN
Despite the improved overall survival rates in most cancers, pancreatic cancer remains one of the deadliest cancers in this decade. The rigid microenvironment, which majorly comprises cancer-associated fibroblasts (CAFs), plays an important role in the obstruction of pancreatic cancer therapy. To overcome this predicament, the signaling of receptor tyrosine kinases (RTKs) and TGF beta receptor (TGFßR) in both pancreatic cancer cell and supporting CAF should be considered as the therapeutic target. The activation of receptors has been reported to be aberrant to cell cycle regulation, and signal transduction pathways, such as growth-factor induced proliferation, and can also influence the apoptotic sensitivity of tumor cells. In this article, the regulation of RTKs/TGFßR between pancreatic ductal adenocarcinoma (PDAC) and CAFs, as well as the RTKs/TGFßR inhibitor-based clinical trials on pancreatic cancer are reviewed.
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Antineoplásicos/uso terapéutico , Carcinoma Ductal Pancreático/tratamiento farmacológico , Neoplasias Pancreáticas/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Receptores de Factores de Crecimiento Transformadores beta/antagonistas & inhibidores , Transducción de Señal/efectos de los fármacos , Fibroblastos Asociados al Cáncer/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Humanos , Neoplasias Pancreáticas/metabolismo , Proteínas Tirosina Quinasas Receptoras/metabolismo , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Resultado del Tratamiento , Microambiente Tumoral/efectos de los fármacosRESUMEN
Spinocerebellar ataxia type 3 (SCA3), a hereditary and lethal neurodegenerative disease, is attributed to the abnormal accumulation of undegradable polyglutamine (polyQ), which is encoded by mutated ataxin-3 gene (ATXN3). The toxic fragments processed from mutant ATXN3 can induce neuronal death, leading to the muscular incoordination of the human body. Some treatment strategies of SCA3 are preferentially focused on depleting the abnormal aggregates, which led to the discovery of small molecule n-butylidenephthalide (n-BP). n-BP-promoted autophagy protected the loss of Purkinje cell in the cerebellum that regulates the network associated with motor functions. We report that the n-BP treatment may be effective in treating SCA3 disease. n-BP treatment led to the depletion of mutant ATXN3 with the expanded polyQ chain and the toxic fragments resulting in increased metabolic activity and alleviated atrophy of SCA3 murine cerebellum. Furthermore, n-BP treated animal and HEK-293GFP-ATXN3-84Q cell models could consistently show the depletion of aggregates through mTOR inhibition. With its unique mechanism, the two autophagic inhibitors Bafilomycin A1 and wortmannin could halt the n-BP-induced elimination of aggregates. Collectively, n-BP shows promising results for the treatment of SCA3.
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Autofagia , Enfermedad de Machado-Joseph/tratamiento farmacológico , Enfermedad de Machado-Joseph/patología , Anhídridos Ftálicos/uso terapéutico , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismo , Adenilato Quinasa/metabolismo , Animales , Ataxina-3/genética , Autofagia/efectos de los fármacos , Cerebelo/patología , Femenino , Células HEK293 , Humanos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Enfermedad de Machado-Joseph/fisiopatología , Ratones Endogámicos C57BL , Actividad Motora/efectos de los fármacos , Mutación/genética , Anhídridos Ftálicos/farmacología , Agregado de Proteínas/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Células de Purkinje/efectos de los fármacos , Células de Purkinje/patología , Transducción de Señal/efectos de los fármacosRESUMEN
Alzheimer's disease (AD) is a neurodegenerative disease with a variety of causes. Traditional Chinese medicine (TCM), which includes the two main approaches of acupuncture and herbal medication, views the human body as a self-controlled system network. Fundamental theories, including "qi," the five elements, and the theory of viscera, form the basis for classification. Diseases in humans are considered to be caused by an imbalance of "yang qi" and "yin qi" that lead to the nonhomeostasis of organs. Acupuncture is derived from 12 main meridians and 365 acupuncture points characterized by "blood and qi." Needling of different positions corresponds to specific disease treatments to increase qi. Treatment with Chinese herbal medicines is based on syndrome differentiation characterized as "Zheng" which differs from the cause orientation approach of Western medicine. In this article, we review basic and clinical research studies that describe TCM herbs and acupuncture for the treatment of AD. Moreover, we propose that these two approaches be integrated to improve the outcomes for AD patients.
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The phenotypic switch of vascular smooth muscle cells (VSMCs) plays a pivotal role in the development of vascular disorders, such as atherosclerosis, stenosis and restenosis, after vascular intervention. In our previous study, n-butylidenephthalide (BP) was reported to have anti-proliferating and apoptotic effects on VSMCs. The purpose of the current study is to further investigate its role in platelet-derived growth factor (PDGF)-induced VSMC phenotypic modulation in an arteriovenous fistula model. In vitro, we observed that BP inhibited the PDGF-induced cytoskeleton reorganization of the VSMCs. The enhanced expression of vimentin and collagen, as well as the migration ability induced by PDGF, were also inhibited by BP. By cell cycle analysis, we found that BP inhibited the PDGF-induced VSMCs proliferation and arrested the VSMCs in the G0/G1 phase. In an arteriovenous fistula rat model, the formation of stenosis, which was coupled with a thrombus, and the expression of vimentin and collagen in VSMCs, were also inhibited by administration of BP, indicating that BP inhibited the PDGF-induced phenotypic switch and the migration of VSMCs. Besides, the inhibitory effects of BP on the phenotypic switch were found to accompany the activated 5' AMP-activated protein kinase (AMPK) as well as the inhibited phosphorylation of mTOR. Knockdown of AMPK by gene silencing conflicted the effects of BP and further exacerbated the PDGF-induced VSMCs phenotypic switch, confirming the modulating effect that BP exerted on the VSMCs by this pathway. These findings suggest that BP may contribute to the vasculoprotective potential in vasculature.
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Proteínas Quinasas Activadas por AMP/metabolismo , Plasticidad de la Célula/efectos de los fármacos , Músculo Liso Vascular/citología , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Fenotipo , Anhídridos Ftálicos/farmacología , Animales , Fístula Arteriovenosa/etiología , Fístula Arteriovenosa/metabolismo , Fístula Arteriovenosa/patología , Biomarcadores , Movimiento Celular/efectos de los fármacos , Constricción Patológica/etiología , Constricción Patológica/metabolismo , Constricción Patológica/prevención & control , Técnica del Anticuerpo Fluorescente , Hiperplasia , Inmunofenotipificación , Neointima/metabolismo , Ratas , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
Patients with multiple system atrophy (MSA), a progressive neurodegenerative disorder of adult onset, were found less than 9 years of life expectancy after onset. The disorders include bradykinesia and rigidity commonly seen in Parkinsonism disease and additional signs such as autonomic dysfunction, ataxia, or dementia. In clinical treatments, MSA poorly responds to levodopa, the drug used to remedy Parkinsonism disease. The exact cause of MSA is still unknown, and exploring a therapeutic solution to MSA remains critical. A transgenic mouse model was established to study the feasibility of human adipose-derived stem cell (ADSC) therapy in vivo. The human ADSCs were transplanted into the striatum of transgenic mice via intracerebral injection. As compared with sham control, we reported significantly enhanced rotarod performance of transgenic mice treated with ADSC at an effective dose, 2 × 105 ADSCs/mouse. Our ex vivo feasibility study supported that intracerebral transplantation of ADSC might alleviate striatal degeneration in MSA transgenic mouse model by improving the nigrostriatal pathway for dopamine, activating autophagy for α-synuclein clearance, decreasing inflammatory signal, and further cell apoptosis, improving myelination and cell survival at caudate-putamen.
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Tejido Adiposo/citología , Cuerpo Estriado/patología , Atrofia de Múltiples Sistemas/terapia , Degeneración Nerviosa/patología , Células Madre/citología , Animales , Apoptosis , Rastreo Celular , Modelos Animales de Enfermedad , Factor Neurotrófico Derivado de la Línea Celular Glial/metabolismo , Humanos , Ratones Transgénicos , Modelos Biológicos , Atrofia de Múltiples Sistemas/complicaciones , Proteína Básica de Mielina/genética , Proteína Básica de Mielina/metabolismo , Degeneración Nerviosa/complicaciones , Regiones Promotoras Genéticas/genética , Agregado de Proteínas , Prueba de Desempeño de Rotación con Aceleración Constante , alfa-Sinucleína/metabolismoRESUMEN
Functional decline of stem cell transplantation in ageing hosts is well documented. The mechanism for this is poorly understood, although it is known that advancing age does not provide an optimal milieu for exogenous stem cells to survive, engraft and differentiate. We showed that n-butylidenephthalide improved human adipose-derived stem cell (hADSC) engraftment via attenuating the production of reactive oxygen species (ROS). It remained unclear whether pre-treated hosts with n-butylidenephthalide can rejuvenate the ageing heart and improve hADSC engraftment by regulating the ROS/NLRP3 inflammasome-mediated cardiac fibrosis after myocardial infarction. One hour after coronary ligation, hADSCs were transplanted into the hearts of young and ageing Wistar rats that were pre-treated with or without n-butylidenephthalide for 3 days. At day 3 after infarction, myocardial infarction was associated with an increase in ROS levels and NLRP3 inflammasome activity with age. hADSC transplant effectively provided a significant decrease in ROS levels, NLRP3 inflammasome activity, IL-1ß levels and cardiac fibrosis in either young or old infarcted rats. However, the beneficial effects of hADSCs were greater in young compared with old rats in terms of NLRP3 inflammasome activity. The infarcted ageing rats pre-conditioned by n-butylidenephthalide improved engraftment and differentiation of hADSCs and additionally attenuated cardiac fibrosis compared with hADSCs alone. The anti-inflammation effects of n-butylidenephthalide were reversed by SIN-1. In conclusions, the increased NLRP3 inflammasome activity plays the pathogenesis of ageing-related functional hADSC decline in the ageing hosts. n-butylidenephthalide-pre-treated ageing hosts reversibly ameliorate the harsh microenvironments, improve stem cell engraftment and attenuate cardiac fibrosis after myocardial infarction.
Asunto(s)
Tejido Adiposo/citología , Envejecimiento , Inflamasomas/metabolismo , Precondicionamiento Isquémico Miocárdico , Infarto del Miocardio/fisiopatología , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Trasplante de Células Madre , Animales , Diferenciación Celular , Fibrosis , Hemodinámica , Humanos , Interleucina-1beta/metabolismo , Masculino , Miocardio/patología , Proteínas de Neoplasias/metabolismo , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Fenotipo , Anhídridos Ftálicos/farmacología , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismo , Células Madre/citología , Superóxidos/metabolismoRESUMEN
Although butylidenephthalide (BP) is an efficient anticancer drug, its poor bioavailability renders it ineffective for treating drug-resistant brain tumors. However, this problem is overcome through the use of noninvasive delivery systems, including intranasal administration. Herein, the bioavailability, drug stability, and encapsulation efficiency (EE, up to 95%) of BP were improved by using cyclodextrin-encapsulated BP in liposomal formulations (CDD1). The physical properties and EE of the CDD1 system were investigated via dynamic light scattering, transmission electron microscopy, UV-Vis spectroscopy, and nuclear magnetic resonance spectroscopy. The cytotoxicity was examined via MTT assay, and the cellular uptake was observed using fluorescence microscopy. The CDD1 system persisted for over 8 h in tumor cells, which was a considerable improvement in the retention of the BP-containing cyclodextrin or the BP-containing liposomes, thereby indicating a higher BP content in CDD1. Nanoscale CDD1 formulations were administered intranasally to nude mice that had been intracranially implanted with temozolomide-resistant glioblastoma multiforme cells, resulting in increased median survival time. Liquid chromatography-mass spectrometry revealed that drug biodistribution via intranasal delivery increased the accumulation of BP 10-fold compared to oral delivery methods. Therefore, BP/cyclodextrin/liposomal formulations have potential clinical applications for treating drug-resistant brain tumors.
Asunto(s)
Antineoplásicos/farmacocinética , Encéfalo/metabolismo , Sistemas de Liberación de Medicamentos , Anhídridos Ftálicos/farmacocinética , Animales , Antineoplásicos/administración & dosificación , Disponibilidad Biológica , Encéfalo/efectos de los fármacos , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/metabolismo , Ciclodextrinas/química , Glioblastoma/tratamiento farmacológico , Glioblastoma/metabolismo , Liposomas/química , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , Anhídridos Ftálicos/administración & dosificación , Distribución TisularRESUMEN
Neurodegenerative diseases represent a significant unmet medical need in our aging society. There are no effective treatments for most of these diseases, and we know comparatively little regarding pathogenic mechanisms. Among the challenges faced by those involved in developing therapeutic drugs for neurodegenerative diseases, the syndromes are often complex, and small animal models do not fully recapitulate the unique features of the human nervous system. Human induced pluripotent stem cells (iPSCs) are a novel technology that ideally would permit us to generate neuronal cells from individual patients, thereby eliminating the problem of species-specificity inherent when using animal models. Specific phenotypes of iPSC-derived cells may permit researchers to identify sub-types and to distinguish among unique clusters and groups. Recently, iPSCs were used for drug screening and testing for neurologic disorders including Alzheimer's disease (AD), amyotrophic lateral sclerosis (ALS), spinocerebellar atrophy (SCA), and Zika virus infection. However, there remain many challenges still ahead, including how one might effectively recapitulate sporadic disease phenotypes and the selection of ideal phenotypes and for large-scale drug screening. Fortunately, quite a few novel strategies have been developed that might be combined with an iPSC-based model to solve these challenges, including organoid technology, single-cell RNA sequencing, genome editing, and deep learning artificial intelligence. Here, we will review current applications and potential future directions for iPSC-based neurodegenerative disease models for critical drug screening.
Asunto(s)
Descubrimiento de Drogas , Evaluación Preclínica de Medicamentos , Células Madre Pluripotentes Inducidas/efectos de los fármacos , Células Madre Pluripotentes Inducidas/metabolismo , Animales , Diferenciación Celular , Susceptibilidad a Enfermedades , Descubrimiento de Drogas/métodos , Evaluación Preclínica de Medicamentos/métodos , Humanos , Células Madre Pluripotentes Inducidas/citología , Enfermedades Neurodegenerativas/tratamiento farmacológico , Enfermedades Neurodegenerativas/etiología , Enfermedades Neurodegenerativas/metabolismo , Neuroglía/efectos de los fármacos , Neuroglía/metabolismo , Neuronas/efectos de los fármacos , Neuronas/metabolismo , FenotipoRESUMEN
Exosomes, 60-200-nm extracellular vesicles secreted from cells, have been used as an active pharmaceutical ingredient or drug carrier in disease treatment. Human- and plant-derived exosomes are registered in clinical trials, but more complete reports are available for human-derived exosomes. Because exosomes act as vesicles and carry cell secreting components, they have been used as drug or peptide vehicles to treat diseases. The dendritic cells (DCs) and mesenchymal stem cells (MSCs) are two popular cell sources for exosome preparation. Exosomes from DCs can initiate inflammation in patients, particularly in patients with cancer, as they contain the tumor antigen to induce specific inflammation response. A well-established cell bank of MSCs is available, and these cells can be used as an alternative source for exosome preparation. The major application of MSC-derived exosomes is in inflammation treatment. Exosomes in clinical trials need to comply with good manufacturing practice (GMP). Three important issues are prevalent in GMP for exosomes, i.e., upstream of cell cultivation process, downstream of the purification process, and exosome quality control. This paper concisely reviews exosome development, including exosome generation and clinical trial application.