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1.
EMBO J ; 42(12): e112675, 2023 06 15.
Artículo en Inglés | MEDLINE | ID: mdl-37092319

RESUMEN

Tumor cells surviving hypoxic stress acquire the ability to drive cancer progression. To explore the contribution of dehydrogenases to the low oxygen concentration response, we used siRNAs targeting 163 dehydrogenase-coding genes and discovered that glutamate dehydrogenase 1 (GDH1) plays a critical role in regulating colorectal cancer (CRC) cell survival under hypoxia. We observed that GDH1 deficiency had an inhibitory effect on CRC occurrence and impaired hypoxia-inducible factor 1-alpha (HIF-1α) stability even under hypoxia. Mechanistically, hypoxia triggered p300 recruitment to GDH1, promoting its acetylation at K503 and K527. GDH1 acetylation at K527 induced the formation of a GDH1 complex with EGLN1/HIF-1α; in contrast, GDH1 acetylation at K503 reinforced its affinity for α-ketoglutarate (αKG), and glutamate production. In line with this view, αKG is a product of GDH1 under normoxia, but hypoxia stimulation reversed GDH1 enzyme activity and αKG consumption by the EGLN1/HIF-1α complex, increasing HIF-1α stability and promoting CRC progression. Clinically, hypoxia-modulated GDH1 AcK503/527 can be used as a biomarker of CRC progression and is a potential target for CRC treatment.


Asunto(s)
Neoplasias Colorrectales , Ácido Glutámico , Humanos , Ácido Glutámico/metabolismo , Hipoxia , Hipoxia de la Célula/genética , Transformación Celular Neoplásica , Carcinogénesis , Neoplasias Colorrectales/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Línea Celular Tumoral
2.
BMC Bioinformatics ; 25(1): 91, 2024 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-38429654

RESUMEN

BACKGROUND: Uncovering functional genetic variants from an allele-specific perspective is of paramount importance in advancing our understanding of gene regulation and genetic diseases. Recently, various allele-specific events, such as allele-specific gene expression, allele-specific methylation, and allele-specific binding, have been explored on a genome-wide scale due to the development of high-throughput sequencing methods. RNA secondary structure, which plays a crucial role in multiple RNA-associated processes like RNA modification, translation and splicing, has emerged as an essential focus of relevant research. However, tools to identify genetic variants associated with allele-specific RNA secondary structures are still lacking. RESULTS: Here, we develop a computational tool called 'AStruct' that enables us to detect allele-specific RNA secondary structure (ASRS) from RT-stop based structuromic probing data. AStruct shows robust performance in both simulated datasets and public icSHAPE datasets. We reveal that single nucleotide polymorphisms (SNPs) with higher AStruct scores are enriched in coding regions and tend to be functional. These SNPs are highly conservative, have the potential to disrupt sites involved in m6A modification or protein binding, and are frequently associated with disease. CONCLUSIONS: AStruct is a tool dedicated to invoke allele-specific RNA secondary structure events at heterozygous SNPs in RT-stop based structuromic probing data. It utilizes allelic variants, base pairing and RT-stop information under different cell conditions to detect dynamic and functional ASRS. Compared to sequence-based tools, AStruct considers dynamic cell conditions and outperforms in detecting functional variants. AStruct is implemented in JAVA and is freely accessible at: https://github.com/canceromics/AStruct .


Asunto(s)
Regulación de la Expresión Génica , ARN , ARN/genética , ARN/química , Alelos , Empalme del ARN , Secuenciación de Nucleótidos de Alto Rendimiento/métodos
3.
Mol Ther ; 31(2): 517-534, 2023 02 01.
Artículo en Inglés | MEDLINE | ID: mdl-36307991

RESUMEN

N6-methyladenosine (m6A) is the most pervasive RNA modification and is recognized as a novel epigenetic regulation in RNA metabolism. Although the m6A modification involves various physiological processes, its roles in drug resistance in colorectal cancer (CRC) still remain unknown. We analyzed the RNA expression profile of m6A/A (%) with MRM mass spectrometry in human 5-fluorouracil (5-FU)-resistant CRC tissues, and used the m6A RNA immunoprecipitation assay to validate the m6A-regulated target. Our results have shown that the m6A demethylase FTO was up-regulated in human primary and 5-FU-resistant CRC. Depletion of FTO decreased cell growth, colony formation and metastasis in 5-FU-resistant CRC cells in vitro and in vivo. Mechanistically, we identified SIVA1, a critical apoptotic gene, as a key downstream target of the FTO-mediated m6A demethylation. The m6A demethylation of SIVA1 at the CDS region induced its mRNA degradation via a YTHDF2-dependent mechanism. The SIVA1 levels were negatively correlated with the FTO levels in clinical CRC tissues. Notably, inhibition of FTO significantly reduced the tolerance of 5-FU in 5-FU-resistant CRC cells via the FTO-SIVA1 axis, whereas SIVA1-depletion could restore the m6A-dependent 5-FU sensitivity in CRC cells. In summary, our findings demonstrate a critical role of FTO as an m6A demethylase enhancing chemo-resistance in CRC cells, and suggest that FTO inhibition may restore the sensitivity of chemo-resistant CRC cells to 5-FU.


Asunto(s)
Neoplasias Colorrectales , Epigénesis Genética , Humanos , ARN , Fluorouracilo/farmacología , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/metabolismo , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato/genética , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato/metabolismo
4.
Drug Resist Updat ; 66: 100909, 2023 01.
Artículo en Inglés | MEDLINE | ID: mdl-36525936

RESUMEN

Oxaliplatin chemoresistance is a major challenge in the clinical treatment of colorectal cancer (CRC), which is one of the most common malignancies worldwide. In this study, we identified the tryptophan-aspartate repeat domain 43 (WDR43) as a potentially critical oncogenic factor in CRC pathogenesis through bioinformatics analysis. It was found that WDR43 is highly expressed in CRC tissues, and WDR43 overexpression is associated with poor prognosis of CRC patients. WDR43 knockdown significantly inhibits cell growth by arresting cell cycle and enhancing the effect of oxaliplatin chemotherapy both in vitro and in vivo. Mechanistically, upon oxaliplatin stimulation, c-MYC promotes the transcriptional regulation and expression of WDR43. WDR43 enhances the ubiquitination of p53 by MDM2 through binding to RPL11, thereby reducing the stability of the p53 protein, which induces proliferation and chemoresistance of CRC cells. Thus, the overexpression of WDR43 promotes CRC progression, and could be a potential therapeutic target of chemoresistance in CRC.


Asunto(s)
Neoplasias Colorrectales , Proteína p53 Supresora de Tumor , Humanos , Línea Celular Tumoral , Proliferación Celular , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica , Oxaliplatino/farmacología , Oxaliplatino/uso terapéutico , Transducción de Señal , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo
5.
Mol Biol Rep ; 50(11): 9441-9452, 2023 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-37838622

RESUMEN

BACKGROUND: Glutamate exposure was fatal to HT-22 neuronal cells that derived from mouse hippocampus. This is often used as a model for hippocampus neurodegeneration in vitro. The targets relevant to glutamate-induced neuronal toxicity is not fully understood. In this study, we aimed to identify crucial factors associated with glutamate-induced cytotoxicity in HT-22 cells. METHODS: HT-22 cells were treated with 7.5 mM glutamate for 24 h and isobaric tags for relative and absolute quantitation (iTRAQ) proteomic analysis conducted to identify the differentially expressed proteins. Differential proteins were subjected to Gene Ontology analyses. Upregulation of barrier to autointegration factor (BANF1/BANF1) protein was confirmed by RT-qPCR and western blotting. Cell viability was measured by CKK-8 and MTT assays. Cell apoptosis rates and intracellular reactive oxygen species (ROS) levels were detected using flow cytometry. RESULTS: A total of 5811 proteins were quantified by iTRAQ, 50 of which were recognized as significantly differential proteins (fold change ≥ 1.5 and P ≤ 0.05); 26 proteins were up-regulated and 24 were down-regulated after exposure to glutamate. GO enrichment analysis showed that the apoptotic signaling pathway was involved in cell death induced by glutamate. BANF1 expression level was markedly increased in HT-22 cells after glutamate treatment. Further, knockdown of BANF1 alleviated glutamate-mediated cell death with lower ROS levels. CONCLUSIONS: In conclusion, we successfully filtered out differential proteins relevant to glutamate-mediated cytotoxicity. BANF1 upregulation promoted glutamate-induced apoptosis of HT-22 cells by enhancing ROS generation.


Asunto(s)
Ácido Glutámico , Proteómica , Ratones , Animales , Ácido Glutámico/toxicidad , Ácido Glutámico/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Línea Celular , Neuronas/metabolismo , Apoptosis , Hipocampo/metabolismo
6.
J Neurochem ; 152(2): 221-234, 2020 01.
Artículo en Inglés | MEDLINE | ID: mdl-31390677

RESUMEN

Histone deacetylase 6 (HDAC6) activity contributes to the malignant proliferation, invasion, and migration of glioma cells (GCs), but the molecular mechanisms underlying the processes remains elusive. Here, we reported that HDAC6 inhibition by Ricolinostat (ACY-1215) or CAY10603 led to a remarkable decrease in the phosphorylation of c-Jun N-terminal kinase (JNK) and c-Jun, which preceded its suppressive effects on glioma cell growth. Further investigation showed that these effects resulted from HDAC6 inhibitor-induced suppression of MAPK kinase 7 (MKK7), which was identified to be critical for JNK activation and exerts the oncogenic roles in GCs. Selectively silencing HDAC6 by siRNAs had the same responses, whereas transient transfections expressing HDAC6 promoted MKK7 expression. Interestingly, by performing Q-PCR, HDAC6 inhibition did not cause a down-regulation of MKK7 mRNA level, whereas the suppressive effects on MKK7 protein can be efficiently blocked by the proteasomal inhibitor MG132. As a further test, elevating MKK7-JNK activity was sufficient to rescue HDAC6 inhibitor-mediated-suppressive effects on c-Jun activation and the malignant features. The suppression of both MKK7 expression and JNK/c-Jun activities was involved in the tumor-growth inhibitory effects induced by CAY10603 in U87-xenograft mice. Collectively, our findings provide new insights into the molecular mechanism of glioma malignancy regarding HDAC6 in the selective regulation of MKK7 expression and JNK/c-Jun activity. MKK7 protein stability critically depends on HDAC6 activity, and inhibition of HDAC6 probably presents a potential strategy for suppressing the oncogenic roles of MKK7/JNK/c-Jun axis in GCs.


Asunto(s)
Procesos de Crecimiento Celular/fisiología , Glioblastoma/metabolismo , Histona Desacetilasa 6/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , MAP Quinasa Quinasa 4/metabolismo , MAP Quinasa Quinasa 7/metabolismo , Animales , Procesos de Crecimiento Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Movimiento Celular/fisiología , Relación Dosis-Respuesta a Droga , Femenino , Glioblastoma/patología , Histona Desacetilasa 6/antagonistas & inhibidores , Inhibidores de Histona Desacetilasas/farmacología , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Ensayos Antitumor por Modelo de Xenoinjerto/métodos
7.
Int J Cancer ; 145(9): 2496-2508, 2019 11 01.
Artículo en Inglés | MEDLINE | ID: mdl-30963560

RESUMEN

JNK activity has been implicated in the malignant proliferation, invasion and drug-resistance of glioma cells (GCs), but the molecular mechanisms underlying JNK activation are currently unknown. Here, we reported that MKK7, not MKK4, directly activates JNK in GCs and exerts oncogenic effects on tumor formation. Notably, MKK7 expression in glioma tissues was closely correlated with the grade of the glioma and JNK/c-Jun activation. Mechanistically, MKK7 transcription critically depends on the complexes formed by HDAC4 and the transcriptional factors SP1 and Krüppel-like factor-5 (KLF5), wherein HDAC4 directly deacetylates both SP1 and KLF5 and synergistically upregulates MKK7 transcription through two SP1 sites located on its promoter. In contrast, the increases in acetylated-SP1 and acetylated-KLF5 after HDAC4 inhibition switched to transcriptionally suppress MKK7. Selective inhibition of HDAC4 by LMK235, siRNAs or blockage of SP1 and KLF5 by the ectopic dominant-negative SP1 greatly reduced the malignant capacity of GCs. Furthermore, suppression of both MKK7 expression and JNK/c-Jun activities was involved in the tumor-growth inhibitory effects induced by LMK235 in U87-xenograft mice. Interestingly, HDAC4 is highly expressed in glioma tissues, and the rate of HDAC4 nuclear import is closely correlated with glioma grade, as well as with MKK7 expression. Collectively, these findings demonstrated that highly expressed MKK7 contributes to JNK/c-Jun signaling-mediated glioma formation. MKK7 transcription, regulated by SP1 and KLF5, critically depends on HDAC4 activity, and inhibition of HDAC4 presents a potential strategy for suppressing the oncogenic roles of MKK7/JNK/c-Jun signaling in GCs.


Asunto(s)
Glioma/genética , Histona Desacetilasas/genética , Factores de Transcripción de Tipo Kruppel/genética , MAP Quinasa Quinasa 7/genética , Proteínas Represoras/genética , Factor de Transcripción Sp1/genética , Animales , Línea Celular Tumoral , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/genética , Sistema de Señalización de MAP Quinasas/genética , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Regiones Promotoras Genéticas/genética , Transcripción Genética/genética , Activación Transcripcional/genética , Regulación hacia Arriba/genética
8.
Clin Proteomics ; 16: 33, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31384238

RESUMEN

BACKGROUND: Diabetic nephropathy is the most frequent cause of end-stage renal disease worldwide. Identification of biomarkers for diabetic nephropathy for early diagnosis may be the key to avoiding damage from this condition. METHODS: Proteomic iTRAQ technology was first used to identify differentially expressed plasma proteins in type 2 incipient diabetic nephropathy (IDN) using a Q-Exactive mass spectrometer. RESULTS: Compared with controls, 57 proteins (32 upregulated and 25 downregulated proteins) were identified. Furthermore, the gelsolin, collectin-11, PTPRJ, and AKAP-7 proteins were confirmed by Western blots as candidate biomarkers for type 2 IDN through ROC analysis. CONCLUSIONS: These findings offer a theoretical basis for the early treatment of diabetic nephropathy.

9.
BMC Ophthalmol ; 17(1): 253, 2017 Dec 19.
Artículo en Inglés | MEDLINE | ID: mdl-29258473

RESUMEN

BACKGROUND: The aim of the present study was to identify the proteomic differences among human lenses in different physiopathological states and to screen for susceptibility genes/proteins via proteogenomic characterization. METHODS: The total proteomes identified across the regenerative lens with secondary cataract (RLSC), congenital cataract (CC) and age-related cataract (ARC) groups were compared to those of normal lenses using isobaric tagging for relative and absolute protein quantification (iTRAQ). The up-regulated proteins between the groups were subjected to biological analysis. Whole exome sequencing (WES) was performed to detect genetic variations. RESULTS: The most complete human lens proteome to date, which consisted of 1251 proteins, including 55.2% previously unreported proteins, was identified across the experimental groups. Bioinformatics functional annotation revealed the common involvement of cellular metabolic processes, immune responses and protein folding disturbances among the groups. RLSC-over-expressed proteins were characteristically enriched in the intracellular immunological signal transduction pathways. The CC groups featured biological processes relating to gene expression and vascular endothelial growth factor (VEGF) signaling transduction, whereas the molecular functions corresponding to external stress were specific to the ARC groups. Combined with WES, the proteogenomic characterization narrowed the list to 16 candidate causal molecules. CONCLUSIONS: These findings revealed common final pathways with diverse upstream regulation of cataractogenesis in different physiopathological states. This proteogenomic characterization shows translational potential for detecting susceptibility genes/proteins in precision medicine.


Asunto(s)
Catarata/metabolismo , Proteínas del Ojo/metabolismo , Cristalino/metabolismo , Proteoma/análisis , Adulto , Preescolar , Cromatografía Liquida , Femenino , Humanos , Masculino , Persona de Mediana Edad , Proteogenómica , Proteoma/genética , Proteómica , Espectrometría de Masas en Tándem , Adulto Joven
10.
Comput Struct Biotechnol J ; 23: 2173-2189, 2024 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-38827229

RESUMEN

The vast neuronal diversity in the human neocortex is vital for high-order brain functions, necessitating elucidation of the regulatory mechanisms underlying such unparalleled diversity. However, recent studies have yet to comprehensively reveal the diversity of neurons and the molecular logic of neocortical origin in humans at single-cell resolution through profiling transcriptomic or epigenomic landscapes, owing to the application of unimodal data alone to depict exceedingly heterogeneous populations of neurons. In this study, we generated a comprehensive compendium of the developing human neocortex by simultaneously profiling gene expression and open chromatin from the same cell. We computationally reconstructed the differentiation trajectories of excitatory projection neurons of cortical origin and inferred the regulatory logic governing lineage bifurcation decisions for neuronal diversification. We demonstrated that neuronal diversity arises from progenitor cell lineage specificity and postmitotic differentiation at distinct stages. Our data paves the way for understanding the primarily coordinated regulatory logic for neuronal diversification in the neocortex.

11.
J Neurochem ; 125(5): 685-97, 2013 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-23470087

RESUMEN

Neuronal activity shapes the dendritic arbour; however, most of the molecular players in this process remain to be identified. We observed that depolarization-induced neuronal activity causes an increase in the phosphorylation of glycogen synthase kinase-3 (GSK-3)α/ß on Ser21/9 in cerebellar granule neurons. Using several approaches, including gene knockdown and GSK-3α/ß(S21A/S21A/S9A/S9A) double knockin mice, we demonstrated that both GSK-3ß and GSK-3α mediate activity-dependent dendritic growth and that Ser21/9 phosphorylation of GSK-3α/ß plays an important role in this process. Collapsin response mediator protein-2 (CRMP-2), which is crucial for axon development, is phosphorylated at Thr514 and inactivated by GSK-3. We found CRMP-2 was located mainly in the dendrites of cerebellar granule neurons, in contrast to the axonal distribution in hippocampal neurons. Over-expression of CRMP-2 promoted and knockdown of CRMP-2 impaired dendritic growth, suggesting that CRMP-2 is necessary and sufficient for activity-dependent dendritic development. Furthermore, silencing CRMP-2 completely blocked the dendritic growth-promoting effects of GSK-3 knockdown, and expression of Thr514 nonphosphorylated form of CRMP-2 counteracted the inhibitory effect of constitutively active GSK-3. This data indicate that CRMP-2 functions downstream of GSK-3. Together, these findings identify a novel GSK-3/CRMP-2 pathway that connects neuronal activity to dendritic growth.


Asunto(s)
Dendritas/fisiología , Glucógeno Sintasa Quinasa 3/fisiología , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Animales , Animales Recién Nacidos , Células Cultivadas , Técnicas de Sustitución del Gen , Glucógeno Sintasa Quinasa 3 beta , Péptidos y Proteínas de Señalización Intercelular/genética , Ratones , Ratones Endogámicos C57BL , Proteínas del Tejido Nervioso/genética , Fosforilación/fisiología , Ratas , Ratas Sprague-Dawley
12.
Parasitol Res ; 112(3): 1287-97, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-23274491

RESUMEN

The excretory secretory products (ESP) of Clonorchis sinensis are the causative agents of clonorchiasis and biliary diseases. The parasites' ESP play important roles in host-parasite interactions. The protein compositions of ESP at different secretory times are different and have not been systemically investigated so far. In this study, we collected ESP from six different periods (0-3 h, 3-6 h, 6-12 h, 12-24 h, 24-36 h, and 36-48 h) from C. sinensis adults. Using a shotgun LC-MS/MS analysis, we found 187, 80, 103, 58, 248, and 383 proteins, respectively. Among these proteins, we selected methionine aminopeptidase 2 (MAP-2, presented in 24-36 h and 36-48 h ESP) and acid phosphatase (AP, presented in 3-6 h, 12-24 h, 24-36 h, and 36-48 h ESP) for further study. Bioinformatics analysis showed that CsMAP-2 has metallopeptidase family M24, unique lysine residue-rich and acidic residue-rich domain, SGTS motif, and auto-cleavage point; and that CsAP has possible signal sequence cleavage site, acid phosphate domain, and two histidine acid phosphatases active regions. CsMAP-2 and CsAP's cDNA have 1,425 bp and1,410 bp ORF, encoding 475 and 470 amino acid proteins and weighing 55.3840 kDa and 55.2875 kDa, respectively. MAP-2 and AP were identified as antigens present in the ESP and circulating antigens by immunoblot analysis, which were also found expressing in the eggs, metacercaria, and adult stages of C. sinensis. Immunofluorescence analysis showed that they were located in tegument and intestinal cecum of adult. MTT assay showed that they could inhibit hepatic stellate cell line (LX-2) proliferation. These findings presented the compositions of different period excretory secretary products from C. sinensis adults.


Asunto(s)
Fosfatasa Ácida/análisis , Aminopeptidasas/análisis , Antígenos Helmínticos/análisis , Clonorchis sinensis/química , Metaloendopeptidasas/análisis , Proteoma/análisis , Fosfatasa Ácida/química , Aminopeptidasas/química , Estructuras Animales/química , Animales , Antígenos Helmínticos/química , Cromatografía Liquida , Metaloendopeptidasas/química , Microscopía Fluorescente , Peso Molecular , Ratas , Espectrometría de Masas en Tándem , Factores de Tiempo
13.
Int J Stem Cells ; 16(3): 326-341, 2023 Aug 30.
Artículo en Inglés | MEDLINE | ID: mdl-36310024

RESUMEN

Background and Objectives: Osteoarthritis (OA) is a degenerative disease that leads to the progressive destruction of articular cartilage. Current clinical therapeutic strategies are moderately effective at relieving OA-associated pain but cannot induce chondrocyte differentiation or achieve cartilage regeneration. We investigated the ability of wedelolactone, a biologically active natural product that occurs in Eclipta alba (false daisy), to promote chondrogenic differentiation. Methods and Results: Real-time reverse transcription-polymerase chain reaction, immunohistochemical staining, and immunofluorescence staining assays were used to evaluate the effects of wedelolactone on the chondrogenic differentiation of mesenchymal stem cells (MSCs). RNA sequencing, microRNA (miRNA) sequencing, and isobaric tags for relative and absolute quantitation analyses were performed to explore the mechanism by which wedelolactone promotes the chondrogenic differentiation of MSCs. We found that wedelolactone facilitates the chondrogenic differentiation of human induced pluripotent stem cell-derived MSCs and rat bone-marrow MSCs. Moreover, the forkhead box O (FOXO) signaling pathway was upregulated by wedelolactone during chondrogenic differentiation, and a FOXO1 inhibitor attenuated the effect of wedelolactone on chondrocyte differentiation. We determined that wedelolactone reduces enhancer of zeste homolog 2 (EZH2)-mediated histone H3 lysine 27 trimethylation of the promoter region of FOXO1 to upregulate its transcription. Additionally, we found that wedelolactone represses miR-1271-5p expression, and that miR-1271-5p post-transcriptionally suppresses the expression of FOXO1 that is dependent on the binding of miR-1271-5p to the FOXO1 3'-untranscribed region. Conclusions: These results indicate that wedelolactone suppresses the activity of EZH2 to facilitate the chondrogenic differentiation of MSCs by activating the FOXO1 signaling pathway. Wedelolactone may therefore improve cartilage regeneration in diseases characterized by inflammatory tissue destruction, such as OA.

14.
Comput Struct Biotechnol J ; 21: 4540-4551, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37810279

RESUMEN

Tumor mutation burden (TMB) has emerged as an essential biomarker for assessing the efficacy of cancer immunotherapy. However, due to the inherent complexity of tumors, TMB is not always correlated with the responsiveness of immune checkpoint inhibitors (ICIs). Thus, refining the interpretation and contextualization of TMB is a requisite for enhancing clinical outcomes. In this study, we conducted a comprehensive investigation of the relationship between TMB and multi-omics data across 33 human cancer types. Our analysis revealed distinct biological changes associated with varying TMB statuses in STAD, COAD, and UCEC. While multi-omics data offer an opportunity to dissect the intricacies of tumors, extracting meaningful biological insights from such massive information remains a formidable challenge. To address this, we developed and implemented the PGLCN, a biologically informed graph neural network based on pathway interaction information. This model facilitates the stratification of patients into subgroups with distinct TMB statuses and enables the evaluation of driver biological processes through enhanced interpretability. By integrating multi-omics data for TMB prediction, our PGLCN model outperformed previous traditional machine learning methodologies, demonstrating superior TMB status prediction accuracy (STAD AUC: 0.976 ± 0.007; COAD AUC: 0.994 ± 0.007; UCEC AUC: 0.947 ± 0.023) and enhanced interpretability (BA-House: 1.0; BA-Community: 0.999; BA-Grid: 0.994; Tree-Cycles: 0.917; Tree-Grids: 0.867). Furthermore, the biological interpretability inherent to PGLCN identified the Toll-like receptor family and DNA repair pathways as potential combined biomarkers in conjunction with TMB status in gastric cancer. This finding suggests a potential synergistic targeting strategy with immunotherapy for gastric cancer, thus advancing the field of precision oncology.

15.
Nat Commun ; 14(1): 2631, 2023 05 06.
Artículo en Inglés | MEDLINE | ID: mdl-37149708

RESUMEN

Although long-read single-cell RNA isoform sequencing (scISO-Seq) can reveal alternative RNA splicing in individual cells, it suffers from a low read throughput. Here, we introduce HIT-scISOseq, a method that removes most artifact cDNAs and concatenates multiple cDNAs for PacBio circular consensus sequencing (CCS) to achieve high-throughput and high-accuracy single-cell RNA isoform sequencing. HIT-scISOseq can yield >10 million high-accuracy long-reads in a single PacBio Sequel II SMRT Cell 8M. We also report the development of scISA-Tools that demultiplex HIT-scISOseq concatenated reads into single-cell cDNA reads with >99.99% accuracy and specificity. We apply HIT-scISOseq to characterize the transcriptomes of 3375 corneal limbus cells and reveal cell-type-specific isoform expression in them. HIT-scISOseq is a high-throughput, high-accuracy, technically accessible method and it can accelerate the burgeoning field of long-read single-cell transcriptomics.


Asunto(s)
Isoformas de ARN , ARN , Isoformas de ARN/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Consenso , Isoformas de Proteínas/genética , Análisis de Secuencia de ADN/métodos , Análisis de Secuencia de ARN
16.
Parasitol Res ; 109(3): 737-44, 2011 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-21424807

RESUMEN

Clonorchis sinensis is a food-borne zoonotic parasite that resides in bile ducts and causes clonorchiasis, which may result in cholelithiasis, cholecystitis, hepatic fibrosis, and liver tumors. Although total excretory secretory products (ESP) of C. sinensis adults induce hepatic fibrosis in vivo in rats, the causative mechanism is not well understood. To study components of the ESP, C. sinensis culture medium was collected and analyzed using shotgun LC-MS/MS. We identified a total of 110 proteins, including glycometabolic enzymes (such as fructose-1,6-bisphosphatase (FBPase) and enolase), detoxification enzymes (such as glutamate dehydrogenase, dihydrolipoamide dehydrogenase and cathepsin B endopeptidase), and a number of RAB family proteins. To identify a potential causative agent for hepatic fibrosis, we expressed and purified a recombinant FBPase, a 1,041-bp gene product that encodes a 41.7-kDa protein with prototypical FBPase domains and that can form a tetramer with a molecular mass of 166.8 kDa. In addition, we found that FBPase is an antigen present in the ESP and in circulation. Immunofluorescence showed that FBPase localizes to the intestinal cecum and vitellarium in C. sinensis adults. Our results describe the components of the excretory secretory products from C. sinensis adult worms and suggest that FBPase may be an important antigen present in the ESP of C. sinensis and may lay the foundation for additional studies on the development of clonorchiasis-associated hepatic fibrosis.


Asunto(s)
Antígenos Helmínticos/análisis , Clonorchis sinensis/química , Fructosa-Bifosfatasa/análisis , Proteoma/análisis , Estructuras Animales/química , Animales , Antígenos Helmínticos/química , Antígenos Helmínticos/inmunología , Cromatografía Liquida , Clonación Molecular , Medios de Cultivo/química , Fructosa-Bifosfatasa/química , Fructosa-Bifosfatasa/inmunología , Expresión Génica , Humanos , Peso Molecular , Espectrometría de Masas en Tándem
17.
Zhonghua Wai Ke Za Zhi ; 49(5): 445-9, 2011 May 01.
Artículo en Zh | MEDLINE | ID: mdl-21733404

RESUMEN

OBJECTIVE: To investigate and search correlative proteins of keloid by comparing the results of differential proteomic analysis between keloid and normal skin. METHODS: From January 2010 to June 2010 two-dimensional gel electrophoresis was used to define patterns of protein expression in keloid skin from 8 patients and matched normal skin from 3 patients. Differential expression protein spots were showed and analyzed by matrix-assisted laser desorption ionization-time of flying/time of flying (MALDI-TOF/TOF) mass spectrometry. RESULTS: This study succeeded to provide a two-dimensional protein profiling comparison between normal skin and keloid. Gel-analysis software identified an average of 2978 spots in keloid while 3053 spots in normal skin and statistical filtering yielded 40 spots of a 4-fold change, 32 of which were identified by using mass spectrometry, 20 were up-regulated and 12 were down-regulated. Functional analysis revealed that these proteins could be fractionated to carrier proteins (3 proteins), signal transduction proteins (4 proteins), proliferation and apoptosis related proteins (2 proteins), cytoskeleton proteins (6 proteins), extracellular matrix proteins (8 proteins), immunity related proteins (3 proteins), tumor related proteins (2 proteins), and function unknown protein (4 proteins). CONCLUSIONS: Proteomic analysis can identify the proteins with variance of keloid versus normal skin. The further research to these differential proteins may help reveal the pathogenesis of keloid and provide new treatments for keloid.


Asunto(s)
Queloide/metabolismo , Proteoma/análisis , Piel/metabolismo , Adolescente , Adulto , Femenino , Humanos , Masculino , Proteínas/metabolismo , Proteómica/métodos , Adulto Joven
18.
Front Mol Neurosci ; 13: 81, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32581704

RESUMEN

Glycogen synthase kinase-3 (GSK-3) dysregulation has been implicated in nigral dopaminergic neurodegeneration, one of the main pathological features of Parkinson's disease (PD). The two isoforms, GSK-3α and GSK-3ß, have both been suggested to play a detrimental role in neuronal death. To date, several studies have focused on the role of GSK-3ß on PD pathogenesis, while the role of GSK-3α has been largely overlooked. Here, we report in situ observations that both GSK-3α and GSK-3ß are dephosphorylated at a negatively acting regulatory serine, indicating kinase activation, selectively in nigral dopaminergic neurons following exposure of mice to 1-methyl-4-pheny-1,2,3,6-tetrahydropyridine (MPTP). To identify whether GSK-3α and GSK-3ß display functional redundancy in regulating parkinsonian dopaminergic cell death, we analysed dopaminergic neuron-specific Gsk3a null (Gsk3a ΔDat ) and Gsk3b null (Gsk3b ΔDat ) mice, respectively. We found that Gsk3b ΔDat , but not Gsk3a ΔDat , showed significant resistance to MPTP insult, revealing non-redundancy of GSK-3α and GSK-3ß in PD pathogenesis. In addition, we tested the neuroprotective effect of tideglusib, the most clinically advanced inhibitor of GSK-3, in the MPTP model of PD. Administration of higher doses (200 mg/kg and 500 mg/kg) of tideglusib exhibited significant neuroprotection, whereas 50 mg/kg tideglusib failed to prevent dopaminergic neurodegeneration from MPTP toxicity. Administration of 200 mg/kg tideglusib improved motor symptoms of MPTP-treated mice. Together, these data demonstrate GSK-3ß and not GSK-3α is critical for parkinsonian neurodegeneration. Our data support the view that GSK-3ß acts as a potential therapeutic target in PD and tideglusib would be a candidate drug for PD neuroprotective therapy.

19.
Int J Biol Sci ; 16(14): 2542-2558, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32792856

RESUMEN

Cancer-associated fibroblasts (CAFs) play crucial roles in enhancing cell survival, proliferation, invasion, and metastasis. We previously showed that hepatocellular carcinoma-derived CAFs (H-CAFs) promoted proliferation of hepatocellular carcinoma (HCC) cells. This study aimed to further explore the role of CAFs in HCC epithelial-mesenchymal transition (EMT) and the underlying mechanism. High CAF density was significantly associated with liver cirrhosis, inferior clinicopathologic characteristics, elevated EMT-associated markers, and poorer survival in human HCC. Within HCC cells, EMT was induced after co-culture with H-CAFs. Secretomic analysis showed that IL-6 and HGF were the key EMT-stimulating cytokines secreted by H-CAFs. Proteomic analysis revealed that TG2 was significantly upregulated in HCC cells with EMT phenotypes. Overexpression of TG2 promoted EMT of HCC cells, and knockdown of TG2 remarkably attenuated the H-CAF-induced EMT. Furthermore, during EMT, TG2 expression was enhanced after HCC cells were stimulated by IL-6, but not HGF. Inhibition of the IL-6/STAT3 signaling decreased TG2 expression. The principal TG2 transcription control element and a potential STAT3 binding site were identified using promoter analysis. Hence, H-CAFs facilitates HCC cells EMT mediated by IL-6, which in turn activates IL-6/IL6R/STAT3 axis to promote TG2 expression.


Asunto(s)
Fibroblastos Asociados al Cáncer/metabolismo , Carcinoma Hepatocelular/metabolismo , Transición Epitelial-Mesenquimal , Proteínas de Unión al GTP/metabolismo , Neoplasias Hepáticas/metabolismo , Transglutaminasas/metabolismo , Adulto , Anciano , Carcinoma Hepatocelular/mortalidad , Carcinoma Hepatocelular/patología , Línea Celular Tumoral , China/epidemiología , Femenino , Factor de Crecimiento de Hepatocito/metabolismo , Humanos , Interleucina-6/metabolismo , Hígado/patología , Neoplasias Hepáticas/mortalidad , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Proteína Glutamina Gamma Glutamiltransferasa 2 , Receptores de Interleucina-6/metabolismo , Factor de Transcripción STAT3/metabolismo , Adulto Joven
20.
Redox Biol ; 36: 101642, 2020 09.
Artículo en Inglés | MEDLINE | ID: mdl-32863238

RESUMEN

Normal high-density lipoprotein (nHDL) in normal, healthy subjects is able to promote angiogenesis, but the mechanism remains incompletely understood. HDL from patients with coronary artery disease may undergo a variety of oxidative modifications, rendering it dysfunctional; whether the angiogenic effect is mitigated by such dysfunctional HDL (dHDL) is unknown. We hypothesized that dHDL compromises angiogenesis. The angiogenic effects of nHDL and dHDL were assessed using endothelial cell culture, endothelial sprouts from cardiac tissue from C57BL/6 mice, zebrafish model for vascular growth and a model of impaired vascular growth in hypercholesterolemic low-density lipoprotein receptor null(LDLr-/-)mice. MiRNA microarray and proteomic analyses were used to determine the mechanisms. Lipid hydroperoxides were greater in dHDL than in nHDL. While nHDL stimulated angiogenesis, dHDL attenuated these responses. Protein and miRNA profiles in endothelial cells differed between nHDL and dHDL treatments. Moreover, nHDL suppressed miR-24-3p expression to increase vinculin expression resulting in nitric oxide (NO) production, whereas dHDL delivered miR-24-3p to inhibit vinculin expression leading to superoxide anion (O2•-) generation via scavenger receptor class B type 1. Vinculin was required for endothelial nitric oxide synthase (eNOS) expression and activation and modulated the PI3K/AKT/eNOS and ERK1/2 signaling pathways to regulate nHDL- and VEGF-induced angiogenesis. Vinculin overexpression or miR-24-3p inhibition reversed dHDL-impaired angiogenesis. The expressions of vinculin and eNOS and angiogenesis were decreased, but the expression of miR-24-3p and lipid hydroperoxides in HDL were increased in the ischemic lower limbs of hypercholesterolemic LDLr-/- mice. Overexpression of vinculin or miR-24-3p antagomir restored the impaired-angiogenesis in ischemic hypercholesterolemic LDLr-/- mice. Collectively, nHDL stimulated vinculin and eNOS expression to increase NO production by suppressing miR-24-3p to induce angiogenesis, whereas dHDL inhibited vinculin and eNOS expression to enhance O2•- generation by delivering miR-24-3p to impair angiogenesis, and that vinculin and miR-24-3p may be therapeutic targets for dHDL-impaired angiogenesis.


Asunto(s)
Enfermedad de la Arteria Coronaria , MicroARNs , Animales , Enfermedad de la Arteria Coronaria/tratamiento farmacológico , Enfermedad de la Arteria Coronaria/genética , Células Endoteliales , Voluntarios Sanos , Humanos , Lipoproteínas HDL , Ratones , Ratones Endogámicos C57BL , MicroARNs/genética , Neovascularización Fisiológica , Óxido Nítrico Sintasa de Tipo III/genética , Fosfatidilinositol 3-Quinasas , Proteómica , Pez Cebra
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