RESUMO
The vegetation phenology of the Qinghai-Xizang Plateau is changing significantly in the context of climate change. However, there are many hydrothermal factors affecting the phenology, and few studies have focused on the effects of multiple factors on the phenology of the Qinghai-Xizang Plateau, resulting in a lack of understanding of the mechanisms underlying phenological changes on the Qinghai-Xizang Plateau. In this study, we used remote sensing data interpretation to analyze the spatial and temporal variability of grassland phenology on the Qinghai-Xizang Plateau from 2002 to 2021, focusing on precipitation, temperature, altitude, soil, and other aspects to reveal the dominant factors of phenological variability using an interpretable machine learning method (SHAP) and to quantify the interactive effects of multiple factors on phenology. The results showed that:â The growing season start (SOS) of grasslands on the Qinghai-Xizang Plateau mostly ranged from 110 to 150 d, with 56.32 % of grasslands showing an early SOS trend; the growing season end (EOS) mostly ranged from 290-320 d, with 67.65 % of grasslands showing a delayed EOS trend; and the growing season length (LOS) mostly ranged from 120 to 210 d, with 65.50 % of the grasslands showing a trend towards longer growing season lengths. â¡ SOS in grasslands on the Qinghai-Xizang Plateau was mainly influenced by moisture conditions, in which soil moisture between 10 and 25 kg·m-2 in the 0-10 cm soil layer in March promoted the advancement of SOS and peaked at approximately 20 kg·m-2. EOS was mainly influenced by temperature, with higher temperatures in September and October having a stronger effect on EOS latency promotion and peaking at over 8 â and -0.5 â, respectively. The main influencing factors of LOS were more consistent with SOS, in which soil moisture between 15 and 25 kg·m-2 in the 0-10 cm soil layer in March promoted the prolongation of LOS and peaked at approximately 18 kg·m-2. ⢠There was an obvious interactive effect of water and heat and other factors on phenology; after soil moisture reached 20 kg·m-2 in the 0-10 cm soil layer in March, SOS was more advanced in low-precipitation and low-altitude areas. Better moisture conditions were more conducive to EOS delay at temperatures above 0 â in October, and soil moisture in high precipitation areas promoted LOS prolongation more when soil moisture was between 12 and 22 kg·m-2 in 0-10 cm in March. The results also demonstrated that interpretable machine learning methods could provide a new approach to the analysis of the multifactorial effects of phenological change.
Assuntos
Mudança Climática , Pradaria , Aprendizado de Máquina , Estações do Ano , China , Altitude , Tecnologia de Sensoriamento Remoto , Monitoramento Ambiental/métodos , Solo/química , Temperatura , Chuva , Poaceae/crescimento & desenvolvimentoRESUMO
Microplastics (MPs) as emerging contaminants have accumulated extensively in agricultural ecosystems and are known to exert important effects on biogeochemical processes. However, how MPs in paddy soils influence the conversion of mercury (Hg) to neurotoxic methylmercury (MeHg) remains poorly understood. Here, we evaluated the effects of MPs on Hg methylation and associated microbial communities in microcosms using two typical paddy soils in China (i.e., yellow and red soils). Results showed that the addition of MPs significantly increased MeHg production in both soils, which could be related to higher Hg methylation potential in the plastisphere than in the bulk soil. We found significant divergences in the community composition of Hg methylators between the plastisphere and the bulk soil. In addition, the plastisphere had higher proportions of Geobacterales in the yellow soil and Methanomicrobia in the red soil compared with the bulk soil, respectively; and plastisphere also had more densely connected microbial groups between non-Hg methylators and Hg methylators. These microbiota in the plastisphere are different from those in the bulk soil, which could partially account for their distinct MeHg production ability. Our findings suggest plastisphere as a unique biotope for MeHg production and provide new insights into the environment risks of MP accumulation in agricultural soils.
Assuntos
Mercúrio , Compostos de Metilmercúrio , Microbiota , Oryza , Poluentes do Solo , Compostos de Metilmercúrio/química , Solo/química , Plásticos , Poluentes do Solo/análise , Mercúrio/análise , Oryza/químicaRESUMO
Angiogenesis, the formation of new blood vessels from existing vessels, has been associated with more than 70 diseases. Although numerous studies have established angiogenesis models, only a few indicators can be used to analyze angiogenic structures. In the present study, we developed an image-processing pipeline based on deep learning to analyze and quantify angiogenesis. We utilized several image-processing algorithms to quantify angiogenesis, including a deep learning-based cell nuclear segmentation algorithm and image skeletonization. This method could quantify and measure changes in blood vessels in response to biochemical gradients using 16 indicators, including length, width, number, and nuclear distribution. Moreover, this procedure is highly efficient for the three-dimensional quantitative analysis of angiogenesis and can be applied to diverse angiogenesis investigations.
Assuntos
Aprendizado Profundo , Processamento de Imagem Assistida por Computador/métodos , Algoritmos , Dispositivos Lab-On-A-ChipRESUMO
A traditional Chinese medicinal fungus, Antrodia salmonea (AS), with antioxidant properties is familiar in Taiwan but anti-cancer activity of AS in human colon cancer is ambiguous. Hence, we explored the anti-cancer activity of AS in colon cancer cells. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay revealed that AS showed a remarkable effect on cell viability in colon cancer cells; SW620, HCT116, and HT29. Annexin V/propidium iodide (PI) stained cells indicated that AS induced both early/late apoptosis in SW620 cells. Additionally, cells treated with AS induced caspase-3 activation, poly (ADP-ribose) polymerase (PARP) cleavage, mitochondrial dysfunction, and Bcl-2 associated X (Bax)/B-cell lymphoma (Bcl-2) dysregulation. Microtubule- associated protein 1A/1B-light chain 3B (LC3-II) accumulation, sequestosome 1 (p62/SQSTM1) activation, autophagy related 4B cysteine peptidase (ATG4B) inactivation, acidic vesicular organelles (AVOs) formation, and Beclin-1/Bcl-2 dysregulation revealed that AS-induced autophagy. Interestingly, cells pretreated with 3-methyladenine (3-MA) strengthened AS-induced caspase-3/apoptosis. Suppression of apoptosis by z-Val-Ala-Asp fluoromethyl ketone (Z-VAD-FMK) did not however block AS-induced autophagy, suggesting that autophagy was not attenuated by the AS-induced apoptosis. Application of N-acetylcysteine (NAC) prevented AS-induced cell death, caspase-3 activation, LC3-II accumulation, and AVOs formation, indicating that AS-induced apoptosis and autophagy was mediated by reactive oxygen species (ROS). Furthermore, AS-induced cytoprotective autophagy and apoptosis through extracellular signal-regulated kinase (ERK) signaling cascades. Moreover, in vivo data disclosed that AS inhibited colitis-associated tumorigenesis in azoxymethane (AOM)-dextran sodium sulphate (DSS)-treated mice. For the first time, we report the anti-cancer properties of this potentially advantageous mushroom for the treatment of human colon cancer.
Assuntos
Apoptose , Autofagia , Neoplasias do Colo/patologia , Citoproteção , Polyporales/química , Adenina/análogos & derivados , Adenina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Azoximetano , Proteína Beclina-1/metabolismo , Carcinogênese/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cloroquina/farmacologia , Colite/induzido quimicamente , Colite/complicações , Neoplasias do Colo/etiologia , Citoproteção/efeitos dos fármacos , Sulfato de Dextrana , Progressão da Doença , Humanos , Inflamação/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos Endogâmicos ICR , Proteínas Associadas aos Microtúbulos/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , NF-kappa B/metabolismo , Tamanho do Órgão/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Proteína X Associada a bcl-2/metabolismoRESUMO
Phase-sensitive surface plasmon resonance biosensors are known for their high sensitivity. One of the technology bottle-necks of such sensors is that the phase sensorgram, when measured at fixed angle set-up, can lead to low reproducibility as the signal conveys multiple data. Leveraging the sensitivity, while securing satisfying reproducibility, is therefore is an underdiscussed key issue. One potential solution is to map the phase sensorgram into refractive index unit by the use of sensor calibration data, via a simple non-linear fit. However, basic fitting functions poorly portray the asymmetric phase curve. On the other hand, multi-layer reflectivity calculation based on the Fresnel coefficient can be employed for a precise mapping function. This numerical approach however lacks the explicit mathematical formulation to be used in an optimization process. To this end, we aim to provide a first methodology for the issue, where mapping functions are constructed from Bayesian optimized multi-layer model of the experimental data. The challenge of using multi-layer model as optimization trial function is addressed by meta-modeling via segmented polynomial approximation. A visualization approach is proposed for assessment of the goodness-of-the-fit on the optimized model. Using metastatic cancer exosome sensing, we demonstrate how the present work paves the way toward better plasmonic sensors.
Assuntos
Técnicas Biossensoriais , Ressonância de Plasmônio de Superfície , Algoritmos , Teorema de Bayes , Desenho de Equipamento , Refratometria , Reprodutibilidade dos TestesRESUMO
This study investigated the epidemiological and clinical peculiarities of BCL2 and BCL6 rearrangement in patients with high grade B-cell lymphoma (HGBL) from Taiwan, compared with data from Western countries. Two hundred and eighty-two DLBCL cases from Taipei Medical University-affiliated hospitals (n = 179) and Tri-Service General Hospital (n = 103) were enrolled for this study. From the 282, 47 (16.7%) had MYC translocation; 24 of these harbored concurrent BCL2 and/or BCL6 translocation (double-hit, DH or triple-hit, TH). Twelve DH-HGBL cases had simultaneous MYC and BCL6 translocations, 8 harbored MYC and BCL2 rearrangement, while the remaining 4 patients exhibited TH. Together, 66.7% of DH/TH-HGBL patients were BCL6 rearrangement positive. Among these BCL6-rearranged DH/TH-HGBL patients, only 6 (37.5%) overexpressed MYC and BCL6 proteins simultaneously, indicating that MYC-BCL6 co-overexpression may not be plausible surrogate biomarker for screening BCL6-rearranged DH-HGBL. By the end of year 5, all patients with TH-HGBL, BCL2 DH-HGBL and all but one BCL6 DH-HGBL cases had expired or were lost to follow-up. Progression-free survival (PFS) was longer for the non-DH/TH-HGBL group compared with the DH/TH-HGBL group. While the patients with BCL2 DH-HGBL were lost to follow-up by day 800, their remaining TH-HGBL and BCL6 DH-HGBL peers exhibited very poor PFS, regardless of age strata. More so, patients with BCL6 rearrangement were 5.5-fold more likely associated with extranodal involvement compared with their BCL2-rearranged peers. Moreover, ~60.0% of the BCL6-rearranged DH-HGBL cases were non-GCB, suggesting that including screening for BCL6 rearrangement in patients with the non-GCB phenotype may aid medical decision-making and therapeutic strategy. Contrary to contemporary data from western countries, 2 in every 3 patients with DH/TH-HGBL in Taiwan harbor BCL6 rearrangement. Consistent with present findings, we recommend mandatory screening for BCL6 rearrangement in patients with aggressive HGBL in Taiwan.
RESUMO
Surface Plasmon Resonance (SPR) is widely used in biological and chemical sensing with fascinating properties. However, the application of SPR to detect trace targets is hampered by non-specific binding and poor signal. A variety of approaches for amplification have been explored to overcome this deficiency including DNA aptamers as versatile target detection tools. Hybridization chain reaction (HCR) is a high-efficiency enzyme-free DNA amplification method operated at room temperature, in which two stable species of DNA hairpins coexist in solution until the introduction of the initiator strand triggers a cascade of hybridization events. At an optimal salt condition, as the concentrations of H1 and H2 increased, the HCR signals were enhanced, leading to signal amplification reaching up to 6.5-fold of the detection measure at 30 min. This feature enables DNA to act as an amplifying transducer for biosensing applications to provide an enzyme-free alternative that can easily detect complex DNA sequences. Improvement of more diverse recognition events can be achieved by integrating HCR with a phase-sensitive SPR (pSPR)-tested aptamer stimulus. This work seeks to establish pSPR aptamer system for highly informative sensing by means of an amplification HCR. Thus, combining pSPR and HCR technologies provide an expandable platform for sensitive biosensing.
Assuntos
Técnicas Biossensoriais , Hibridização de Ácido Nucleico , Ressonância de Plasmônio de Superfície , Aptâmeros de Nucleotídeos/química , DNA/química , Ouro/química , Limite de Detecção , Técnicas de Amplificação de Ácido NucleicoRESUMO
BACKGROUND: Triple-negative breast cancer (TNBC) has a more aggressive phenotype and poorer prognosis than hormone receptor (HR+) and human epidermal growth factor receptor (HER2 -) subtypes. Inhibition of cyclin-dependent kinase (CDK)4 and CDK6 was successful in patients with advanced metastatic HR+/HER2- breast cancer, but those with TNBC exhibited low or no response to this therapeutic approach. This study investigated the dual therapeutic targeting of CDK2 and CDK4 by using 4-acetyl-antroquinonol B (4-AAQB) against TNBC cells. METHODS: We examined the effects of CDK2, CDK4, and CDK6 inhibition through 4-AAQB treatment on TNBC cell lines and established an orthotropic xenograft mouse model to confirm the in vitro results of inhibiting CDK2, CDK4, and CDK6 by 4-AAQB treatment. RESULTS: High expression and alteration of CDK2 and CDK4 but not CDK6 significantly correlated with poor overall survival of patients with breast cancer. CDK2 and CDK4 were positively correlated with damage in DNA replication and repair pathways. Docking results indicated that 4-AAQB was bound to CDK2 and CDK4 with high affinity. Treatment of TNBC cells with 4-AAQB suppressed the expression of CDK2 and CDK4 in vitro. Additionally, 4-AAQB induced cell cycle arrest, DNA damage, and apoptosis in TNBC cells. In vivo study results confirmed that the anticancer activity of 4-AAQB suppressed tumor growth through the inhibition of CDK2 and CDK4. CONCLUSION: The expression level of CDK2 and CDK4 and DNA damage response (DDR) signaling are prominent in TNBC cell cycle regulation. Thus, 4-AAQB is a potential agent for targeting CDK2/4 and DDR in TNBC cells.
Assuntos
4-Butirolactona/análogos & derivados , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Quinase 2 Dependente de Ciclina/metabolismo , Quinase 4 Dependente de Ciclina/metabolismo , Cicloexanonas/farmacologia , Dano ao DNA , Reparo do DNA/efeitos dos fármacos , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , 4-Butirolactona/farmacologia , Animais , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Quinase 2 Dependente de Ciclina/genética , Quinase 4 Dependente de Ciclina/genética , Regulação para Baixo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos Endogâmicos NOD , Camundongos SCID , Transdução de Sinais , Neoplasias de Mama Triplo Negativas/enzimologia , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The regulation of mTOR and the dimethylation of histone H3 on lysine 9 (H3K9me2) H3K9me2 by Uhrf1 and the mechanism of autophagy regulation in myocardial ischemia-reperfusion injury (MIRI) were studied in vivo and in vitro. An in vitro I/R injury model was established using the primary mouse cardiomyocytes treated with H2O2. Subsequent analysis by qRT-PCR, western blot, and immunofluorescence indicated that overexpression of Uhrf1 significantly inhibited apoptosis of the H2O2-treated cardiomyocytes, reduced expression of apoptosis factors caspase-3 and Bax, and increased expression of apoptosis inhibitory factor Bcl-2. Furthermore, Uhrf1 was found to increase cardiomyocyte proliferation and promote the expression of mTOR, while the four expression peaks of H3K9me2 on the mTOR gene were inhibited by overexpression of Uhrf1. The expression of autophagy factors LC3, Beclin-1, and p-mTOR in Uhrf1-overexpressed cardiomyocytes was dramatically increased, and P62 expression was dramatically decreased. When an H3K9me2 inhibitor was added to the Uhrf1-knockdown cardiomyocytes, the expression of mTOR was increased, the expression of LC3, Beclin-1, and p-mTOR was decreased, and P62 expression was significantly increased. In the present study, Uhrf1 exhibits a protective function in MIRI, reducing the apoptosis of cardiomyocytes while increasing their proliferation and viability.
Assuntos
Autofagia/fisiologia , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Histonas/metabolismo , Traumatismo por Reperfusão Miocárdica/metabolismo , Miócitos Cardíacos/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Autofagia/efeitos dos fármacos , Caspase 3/metabolismo , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Peróxido de Hidrogênio/farmacologia , Camundongos , Miócitos Cardíacos/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Proteína X Associada a bcl-2/metabolismoRESUMO
Developing materials with remote controllability of macroscale ligand presentation can mimic extracellular matrix (ECM) remodeling to regulate cellular adhesion in vivo. Herein, we designed charged mobile nanoligands with superparamagnetic nanomaterials amine-functionalized and conjugated with polyethylene glycol linker and negatively charged RGD ligand. We coupled negatively a charged nanoligand to a positively charged substrate by optimizing electrostatic interactions to allow reversible planar movement. We demonstrate the imaging of both macroscale and in situ nanoscale nanoligand movement by magnetically attracting charged nanoligand to manipulate macroscale ligand density. We show that in situ magnetic control of attracting charged nanoligand facilitates stem cell adhesion, both in vitro and in vivo, with reversible control. Furthermore, we unravel that in situ magnetic attraction of charged nanoligand stimulates mechanosensing-mediated differentiation of stem cells. This remote controllability of ECM-mimicking reversible ligand variations is promising for regulating diverse reparative cellular processes in vivo.
Assuntos
Adesão Celular , Fenômenos Magnéticos , Oligopeptídeos , Células-Tronco , Diferenciação Celular , Matriz ExtracelularRESUMO
In drug delivery to the human brain, blood vessels are a significant hurdle because they restrict the entry of most solutes to protect brain. To overcome this hurdle, an in vitro 3D model for brain endothelial barrier is developed using a microfluidic device with hydrogel providing a 3D extracellular matrix scaffold. Using the model, peptides known to utilize receptor-mediated transcytosis are verified, which has been one of the most promising mechanisms for brain-specific penetration. The cytotoxicity and cellular damage to the peptide are investigated and the receptor-mediated transcytosis and brain endothelial specific penetrating abilities of the peptides in a quantitative manner are demonstrated. As a preclinical test, applying the quantification assays conducted in this study are suggested, including the penetrating ability, cytotoxicity, endothelial damage, and receptor specificity. Using this microfluidic device as an in vitro platform for evaluating various brain targeting drugs and drug carrier candidates is also proposed.
Assuntos
Barreira Hematoencefálica/metabolismo , Peptídeos Penetradores de Células , Células Endoteliais/metabolismo , Dispositivos Lab-On-A-Chip , Técnicas Analíticas Microfluídicas , Modelos Cardiovasculares , Barreira Hematoencefálica/citologia , Linhagem Celular , Peptídeos Penetradores de Células/farmacocinética , Peptídeos Penetradores de Células/farmacologia , Células Endoteliais/citologia , Humanos , TranscitoseRESUMO
Particulate matter (PM) is able to induce airway epithelial injury, while the detailed mechanisms remain unclear. Here we demonstrated that PM exposure inactivated MTOR (mechanistic target of rapamycin kinase), enhanced macroautophagy/autophagy, and impaired lysosomal activity in HBE (human bronchial epithelial) cells and in mouse airway epithelium. Genetic or pharmaceutical inhibition of MTOR significantly enhanced, while inhibition of autophagy attenuated, PM-induced IL6 expression in HBE cells. Consistently, club-cell-specific deletion of Mtor aggravated, whereas loss of Atg5 in bronchial epithelium reduced, PM-induced airway inflammation. Interestingly, the augmented inflammatory responses caused by MTOR deficiency were markedly attenuated by blockage of downstream autophagy both in vitro and in vivo. Mechanistically, the dysregulation of MTOR-autophagy signaling was partially dependent on activation of upstream TSC2, and interacted with the TLR4-MYD88 to orchestrate the downstream NFKB activity and to regulate the production of inflammatory cytokines in airway epithelium. Moreover, inhibition of autophagy reduced the expression of EPS15 and the subsequent endocytosis of PM. Taken together, the present study provides a mechanistic explanation for how airway epithelium localized MTOR-autophagy axis regulates PM-induced airway injury, suggesting that activation of MTOR and/or suppression of autophagy in local airway might be effective therapeutic strategies for PM-related airway disorders.Abbreviations: ACTB: actin beta; AKT: AKT serine/threonine kinase; ALI: air liquid interface; AP2: adaptor related protein complex 2; ATG: autophagy related; BALF: bronchoalveolar lavage fluid; COPD: chronic obstructive pulmonary disease; CXCL: C-X-C motif chemokine ligand; DOX: doxycycline; EGF: epidermal growth factor; EGFR: epidermal growth factor receptor; EPS15: epidermal growth factor receptor pathway substrate 15; HBE: human bronchial epithelial; H&E: hematoxylin & eosin; IKK: IKB kinase; IL: interleukin; LAMP2: lysosomal-associated membrane protein 2; LPS: lipopolysaccharide; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3 beta; MTEC: mouse tracheal epithelial cells; MTOR: mechanistic target of rapamycin kinase; MYD88: MYD88 innate immune signal transduction adaptor; NFKB: nuclear factor of kappa B; NFKBIA: NFKB inhibitor alpha; PM: particulate matter; PtdIns3K: phosphatidylinositol 3-kinase; Rapa: rapamycin; RELA: RELA proto-oncogene, NFKB subunit; SCGB1A1: secretoglobin family 1A member 1; siRNA: small interfering RNAs; SQSTM1: sequestosome 1; TEM: transmission electronic microscopy; TLR4: toll like receptor 4; TSC2: TSC complex subunit 2.
Assuntos
Autofagia , Células Epiteliais/patologia , Material Particulado/toxicidade , Pneumonia/induzido quimicamente , Pneumonia/patologia , Serina-Treonina Quinases TOR/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteína 5 Relacionada à Autofagia/metabolismo , Brônquios/patologia , Linhagem Celular , Citocinas/metabolismo , Endocitose/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Deleção de Genes , Humanos , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Camundongos , Proteínas Associadas aos Microtúbulos/deficiência , Proteínas Associadas aos Microtúbulos/metabolismo , Modelos Biológicos , Fator 88 de Diferenciação Mieloide/metabolismo , NF-kappa B/metabolismo , Ligação Proteica/efeitos dos fármacos , Proto-Oncogene Mas , Transdução de Sinais , Receptor 4 Toll-Like/metabolismo , Proteína 2 do Complexo Esclerose Tuberosa/metabolismoRESUMO
Engraftment of oligodendrocyte progenitor cells (OPCs), which form myelinating oligodendrocytes, has the potential to treat demyelinating diseases such as multiple sclerosis. However, conventional strategies for generating oligodendrocytes have mainly focused on direct differentiation into forebrain- or spinal cord-restricted oligodendrocytes without establishing or amplifying stem/progenitor cells. Taking advantage of a recently established culture system, we generated expandable EN1- and GBX2-positive glial-restricted progenitor-like cells (GPLCs) near the anterior hindbrain. These cells expressed PDGFRα, CD9, S100ß, and SOX10 and mostly differentiated into GFAP-positive astrocytes and MBP-positive oligodendrocytes. RNA-seq analysis revealed that the transcriptome of GPLCs was similar to that of O4-positive OPCs, but distinct from that of rosette-type neural stem cells. Notably, engrafted GPLCs not only differentiated into GFAP-positive astrocytes but also myelinated the brains of adult shiverer mice 8 weeks after transplantation. Our strategy for establishing anterior hindbrain-specific GPLCs with gliogenic potency will facilitate their use in the treatment of demyelinating diseases and studies of the molecular mechanisms underlying glial development in the hindbrain.
Assuntos
Astrócitos/citologia , Doenças Desmielinizantes/terapia , Bainha de Mielina/metabolismo , Células Precursoras de Oligodendrócitos/citologia , Células Precursoras de Oligodendrócitos/transplante , Oligodendroglia/citologia , Células-Tronco Pluripotentes/citologia , Animais , Astrócitos/metabolismo , Perfilação da Expressão Gênica , Proteína Glial Fibrilar Ácida/metabolismo , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Camundongos Transgênicos , Oligodendroglia/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Rombencéfalo/citologia , Subunidade beta da Proteína Ligante de Cálcio S100/metabolismo , Fatores de Transcrição SOXE/metabolismo , Tetraspanina 29/metabolismoRESUMO
Axon degeneration, a hallmark of chemotherapy-induced peripheral neuropathy (CIPN), is thought to be caused by a loss of the essential metabolite nicotinamide adenine dinucleotide (NAD+) via the prodegenerative protein SARM1. Some studies challenge this notion, however, and suggest that an aberrant increase in a direct precursor of NAD+, nicotinamide mononucleotide (NMN), rather than loss of NAD+, is responsible. In support of this idea, blocking NMN accumulation in neurons by expressing a bacterial NMN deamidase protected axons from degeneration. We hypothesized that protection could similarly be achieved by reducing NMN production pharmacologically. To achieve this, we took advantage of an alternative pathway for NAD+ generation that goes through the intermediate nicotinic acid mononucleotide (NAMN), rather than NMN. We discovered that nicotinic acid riboside (NAR), a precursor of NAMN, administered in combination with FK866, an inhibitor of the enzyme nicotinamide phosphoribosyltransferase that produces NMN, protected dorsal root ganglion (DRG) axons against vincristine-induced degeneration as well as NMN deamidase. Introducing a different bacterial enzyme that converts NAMN to NMN reversed this protection. Collectively, our data indicate that maintaining NAD+ is not sufficient to protect DRG neurons from vincristine-induced axon degeneration, and elevating NMN, by itself, is not sufficient to cause degeneration. Nonetheless, the combination of FK866 and NAR, which bypasses NMN formation, may provide a therapeutic strategy for neuroprotection.
Assuntos
Acrilamidas/farmacologia , NAD/metabolismo , Degeneração Neural/prevenção & controle , Neurônios/efeitos dos fármacos , Niacinamida/análogos & derivados , Mononucleotídeo de Nicotinamida/análogos & derivados , Piperidinas/farmacologia , Vincristina/toxicidade , Animais , Antineoplásicos Fitogênicos/toxicidade , Combinação de Medicamentos , Francisella tularensis/enzimologia , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/metabolismo , Gânglios Espinais/patologia , Degeneração Neural/induzido quimicamente , Degeneração Neural/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Niacinamida/farmacologia , Mononucleotídeo de Nicotinamida/metabolismo , Nicotinamida Fosforribosiltransferase/antagonistas & inibidores , Nicotinamida Fosforribosiltransferase/metabolismo , Compostos de PiridínioRESUMO
Four wild Carex speices (C. rigescens, C. lancifolia, C. leucochlora and C. humilis) collected from Mount Taishan were used as experimental materials. The adaption across the winter and physiological characteristics resistant to cold stress were investigated, semi-lethal temperature (LT50) was calculated and fuzzy subordination method was used to generally evaluate the Carex resistant to cold stress. The results showed that 4 Carex species all survived in the winter safely and restored well to grow in the following spring. The green period of the species was between 260 d and 310 d, and percentage of the withered leaves was between 12% and 95%, the range of LT50 was from -18.65 â to -11.74 â. With intensifying cold stress, the contents of MDA, proline (Pro) and soluble protein increased at first and then decreased, while the soluble sugar content increased with the treatment time. C. rigescens with poor cold tolerance showed the early accumulation of MDA, Pro and soluble sugar. The value of soluble protein peaked at the late stage of low tempe-rature stress, and the Carex with stronger cold-resistance showed the smaller value. The SOD activity in the leaves of C. lancifolia was higher than that of the other three species in the beginning of treatments. The cold resistance of four Carex species was in order of C. lancifolia>C. leucochlora>C. humilis>C. rigescens.
Assuntos
Carex (Planta) , Temperatura Baixa , Aclimatação , Folhas de Planta , TemperaturaRESUMO
Translation regulation is a fundamental component of gene expression, allowing cells to respond rapidly to a variety of stimuli in the absence of new transcription. The lack of methods for profiling nascent proteomes in distinct cell populations in heterogeneous tissues has precluded an understanding of translational regulation in physiologically relevant contexts. Here, we describe a chemical genetic method that involves orthogonal enzyme-mediated incorporation of a clickable puromycin analogue into nascent polypeptides. Using this method, we show that we can label newly synthesized proteins in a cell-specific manner in cells grown in culture and in heterogeneous tissues. We also show that we can identify the nascent proteome in genetically targeted cell populations using affinity enrichment and tandem mass spectrometry. Our method has the potential to provide unprecedented insights into cell-specific translational regulation in heterogeneous tissues.
Assuntos
Adenosina/análogos & derivados , Proteoma/química , Puromicina/análogos & derivados , Tirosina/análogos & derivados , Adenosina/química , Adenosina/metabolismo , Animais , Biotinilação , Química Click , Corantes Fluorescentes/química , Células Secretoras de Glucagon/metabolismo , Células HEK293 , Humanos , Células Secretoras de Insulina/metabolismo , Camundongos , Penicilina Amidase/química , Peptídeos/química , Peptídeos/genética , Peptídeos/metabolismo , Proteoma/genética , Proteoma/metabolismo , Puromicina/química , Tirosina/química , Tirosina/metabolismoRESUMO
Early steps of gene expression are a composite of promoter recognition, promoter activation, RNA synthesis and RNA processing, and it is known that SUMOylation, a post-translational modification, is involved in transcription regulation. We previously found that SUMO-1 marks chromatin at the proximal promoter regions of some of the most active housekeeping genes during interphase in human cells, but the SUMOylated targets on the chromatin remained unclear. In this study, we found that SUMO-1 marks the promoters of ribosomal protein genes via modification of the Scaffold Associated Factor B (SAFB) protein, and the SUMOylated SAFB stimulated both the binding of RNA polymerase to promoters and pre-mRNA splicing. Depletion of SAFB decreased RNA polymerase II binding to promoters and nuclear processing of the mRNA, though mRNA stability was not affected. This study reveals an unexpected role of SUMO-1 and SAFB in the stimulatory coupling of promoter binding, transcription initiation and RNA processing.
Assuntos
Cromatina/metabolismo , Proteínas de Ligação à Região de Interação com a Matriz/fisiologia , Proteínas Associadas à Matriz Nuclear/fisiologia , Regiões Promotoras Genéticas , Splicing de RNA , Receptores de Estrogênio/fisiologia , Proteínas Ribossômicas/genética , Proteína SUMO-1/metabolismo , Transcrição Gênica/fisiologia , Regulação para Baixo , Células HeLa , Humanos , Processamento Pós-Transcricional do RNA , RNA Mensageiro/metabolismoRESUMO
This study was purposed to explore the correlation of regenerating Islet-derived 3-alpha(Reg3α) protein level in plasma with the diagnosis and prognosis of the gastrointestinal acute graft-versus-host disease (GI-aGVHD) after all-HSCT, 103 patients who underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT) were observed in our hospital from December 2011 to December 2012. Peripheral blood samples were routinely collected at 9 d before allo-HSCT, 0 d, 14 d, 28 d after allo-HSCT as well as in aGVHD and at the 1 and 4 weeks after aGVHD therapy. The plasma concentrations of Reg3α were measured by using ELISA kit. The results indicated that among the 103 patients, 17 cases never developed aGVHD symptoms (no-aGVHD), 27 cases presented with non-aGVHD associated diarrhea, 10 cases presented with isolated skin aGVHD, 17 cases developed grades I-II GI-aGVHD, 32 cases with grades III-IV GI-aGVHD. The plasma concentrations of Reg3α in group of patients with GI-aGVHD and group of non-aGVHD diarrhea were 111.5 (54.7-180.2) and 23.9 (14.5-89.5) ng/ml respectively with significant difference (P < 0.001). The plasma concentrations of Reg3α in 17 patients of grades III-IV GI-aGVHD who experienced a complete or partial response and 7 patients who had no response to therapy at 4 weeks were 137.2(51.7-205.4) and 679.4(122.3-896.8) ng/ml respectively with the significant difference (P = 0.028). All of the patients who had no response to therapy died of aGVHD associated multiple organ failure. The area under the ROC curve was 0.902 when plasma concentration of Reg3α was set at 87.73 ng/ml. The sensitivity was 81.48% and the specificity was 82.86% when the critical value was used in diagnosis of grades III-IV GI-aGVHD. The probability of grades III-IV GI-aGVHD had statistical difference above and below 87.73 ng/ml after allo-HSCT (P < 0.001). It is concluded that the increase of plasma Reg3α level after transplantation suggests the incidence of grades III-IV GI-aGVHD. The high level of plasma Reg3α protein in patients with grades III-IV GI-aGVHD after the immunosuppressive treatment for four weeks indicates a poor prognosis. The plasma concentrations of Reg3α can be used as a specific biomarker of GI-aGVHD.
Assuntos
Antígenos de Neoplasias/sangue , Biomarcadores Tumorais/sangue , Doença Enxerto-Hospedeiro/diagnóstico , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Enteropatias/diagnóstico , Lectinas Tipo C/sangue , Adolescente , Adulto , Feminino , Doença Enxerto-Hospedeiro/etiologia , Humanos , Enteropatias/etiologia , Masculino , Pessoa de Meia-Idade , Proteínas Associadas a Pancreatite , Plasma , Prognóstico , Transplante Homólogo , Adulto JovemRESUMO
This study was aimed to explore the method for induction and expansion of EB virus specific cytotoxic T lymphocytes (EBV-CTL) in vitro, and to detect their killing effect. Peripheral blood mononuclear cells (PBMNC) were collected from 6 EBV seropositive healthy donors, and EBV-transformed B lymphoblastoid cells (BLCL)were used as the antigen-presenting cells and antigen stimulant which was irradiated by 40 Gy (60)Co irradiator. The autologous PBMNC and irradiated BLCL were cultured to induce and expand the EBV-CTL, and the immunophenotype was identified by the flow cytometry. The killing effect of the EBV-CTL against the autologous BLCL (autoBLCL), the autologous PHA cultured B lymphoblastoid cells( PHA-BLCL), the allogeneic BLCL (alloBLCL) and the K562 cells were measured with LDH release assay under different effector-to-target ratio. The results showed that the 6 cell lines of EBV-CTL were induced and expanded from the EBV seropositive healthy donors, the overall increase in cell numbers varied from 18.6 to 55.0 times. After 10 stimulations, the specific killing efficiency of the EBV-CTL for the autoBLCL were 59.4%, 43.2% and 29.0% under the effector-to-target ratio of 20: 1, 10: 1 and 5: 1. The nonspecific killing efficiency for the PHA-blast, alloBLCL and K562 cells were 7.1%, 9.4% and 10.3% (P < 0.05) under the 20: 1 ratio; 6.6%, 8.3% and 8.1% (P < 0.05) under 10: 1; 5.4%, 7.3% and 6.3% (P < 0.05) under 5: 1, respectively. It is concluded that the EBV-CTL can be successfully induced and expanded ex vivo for specific killing of HLA matched BLCL and may become a potential treatment for EBV related post-transplant lymphoproliferative disorders.
Assuntos
Linfócitos B/imunologia , Herpesvirus Humano 4/imunologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/virologia , Linhagem Celular Transformada , Humanos , Células K562 , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/virologia , Linfócitos T Citotóxicos/citologiaRESUMO
Despite the chemotherapy is successful in inducing remission of hematologic malignancy, this disease also has a high probability of relapse; besides, the toxicity of chemotherapy for these patients can not be avoided. Researchers have been attempting to eliminate tumor cells by immunotherapy. Recently, various leukemia-associated antigens (LAA) that are recognized by cytotoxic T cell (CTL) in the context of HLA class I molecules have been identified. These LAA include WT1, PR-3, RHAMM, BCR-ABL and Aur-A. On the basis of these findings, various clinical trials of immunotherapy for hematologic malignancy including tumor peptide vaccination, adoptive T cell therapy, NK cell therapy and dendritic cells-cytokine induced killer (DC-CIK) cell therapy are on going. In this review, the current status and future feasibility of cellular immunotherapy for leukemia are discussed.