Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 443.978
Filtrar
Mais filtros








Intervalo de ano de publicação
1.
BMB Rep ; 57(2): 116-121, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38303564

RESUMO

We investigated the therapeutic potential of bone marrow-derived mesenchymal stem cell-conditioned medium (BMSC-CM) on immortalized renal proximal tubule epithelial cells (RPTEC/ TERT1) in a fibrotic environment. To replicate the increased stiffness characteristic of kidneys in chronic kidney disease, we utilized polyacrylamide gel platforms. A stiff matrix was shown to increase α-smooth muscle actin (α-SMA) levels, indicating fibrogenic activation in RPTEC/TERT1 cells. Interestingly, treatment with BMSC-CM resulted in significant reductions in the levels of fibrotic markers (α-SMA and vimentin) and increases in the levels of the epithelial marker E-cadherin and aquaporin 7, particularly under stiff conditions. Furthermore, BMSC-CM modified microRNA (miRNA) expression and reduced oxidative stress levels in these cells. Our findings suggest that BMSC-CM can modulate cellular morphology, miRNA expression, and oxidative stress in RPTEC/TERT1 cells, highlighting its therapeutic potential in fibrotic kidney disease. [BMB Reports 2024; 57(2): 116-121].


Assuntos
Nefropatias , MicroRNAs , Humanos , Meios de Cultivo Condicionados/farmacologia , Linhagem Celular , Nefropatias/tratamento farmacológico , Fibrose , MicroRNAs/genética
2.
Biofabrication ; 16(2)2024 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-38306674

RESUMO

Glandular cancers are amongst the most prevalent types of cancer, which can develop in many different organs, presenting challenges in their detection as well as high treatment variability and failure rates. For that purpose, anticancer drugs are commonly tested in cancer cell lines grown in 2D tissue culture on plastic dishesin vitro, or in animal modelsin vivo. However, 2D culture models diverge significantly from the 3D characteristics of living tissues and animal models require extensive animal use and time. Glandular cancers, such as prostate cancer-the second leading cause of male cancer death-typically exist in co-centrical architectures where a cell layer surrounds an acellular lumen. Herein, this spatial cellular position and 3D architecture, containing dual compartments with different hydrogel materials, is engineered using a simple co-axial nozzle setup, in a single step utilizing prostate as a model of glandular cancer. The resulting hydrogel soft structures support viable prostate cancer cells of different cell lines and enable over-time maturation into cancer-mimicking aggregates surrounding the acellular core. The biofabricated cancer mimicking structures are then used as a model to predict the inhibitory efficacy of the poly ADP ribose polymerase inhibitor, Talazoparib, and the antiandrogen drug, Enzalutamide, in the growth of the cancer cell layer. Our results show that the obtained hydrogel constructs can be adapted to quickly obtain 3D cancer models which combine 3D physiological architectures with high-throughput screening to detect and optimize anti-cancer drugs in prostate and potentially other glandular cancer types.


Assuntos
Antineoplásicos , Neoplasias da Próstata , Humanos , Animais , Masculino , Hidrogéis/química , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/metabolismo , Linhagem Celular
3.
Biol Direct ; 19(1): 13, 2024 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-38308285

RESUMO

BACKGROUND: Hepatocellular carcinoma (HCC) is a prevalent malignant tumor that poses a major threat to people's lives and health. Previous studies have found that multiple deubiquitinating enzymes are involved in the pathogenesis of HCC. The purpose of this work was to elucidate the function and mechanism of the deubiquitinating enzyme USP40 in HCC progression. METHODS: The expression of USP40 in human HCC tissues and HCC cell lines was investigated using RT-qPCR, western blotting and immunohistochemistry (IHC). Both in vitro and in vivo experiments were conducted to determine the crucial role of USP40 in HCC progression. The interaction between USP40 and Claudin1 was identified by immunofluorescence, co-immunoprecipitation and ubiquitination assays. RESULTS: We discovered that USP40 is elevated in HCC tissues and predicts poor prognosis in HCC patients. USP40 knockdown inhibits HCC cell proliferation, migration and stemness, whereas USP40 overexpression shows the opposite impact. Furthermore, we confirmed that Claudin1 is a downstream gene of USP40. Mechanistically, USP40 interacts with Claudin1 and inhibits its polyubiquitination to stabilize Claudin1 protein. CONCLUSIONS: Our study reveals that USP40 enhances HCC malignant development by deubiquitinating and stabilizing Claudin1, suggesting that targeting USP40 may be a novel approach for HCC therapy.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/genética , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Ubiquitinação
4.
Soft Matter ; 20(9): 1996-2007, 2024 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-38323652

RESUMO

In cell clusters, the prominent factors at play encompass contractility-based enhanced tissue surface tension and cell unjamming transition. The former effect pertains to the boundary effect, while the latter constitutes a bulk effect. Both effects share outcomes of inducing significant elongation in cells. This elongation is so substantial that it surpasses the limits of linear elasticity, thereby giving rise to additional effects. To investigate these effects, we employ atomic force microscopy (AFM) to analyze how the mechanical properties of individual cells change under such considerable elongation. Our selection of cell lines includes MCF-10A, chosen for its pronounced demonstration of the extended differential adhesion hypothesis (eDAH), and MDA-MB-436, selected due to its manifestation of cell unjamming behavior. In the AFM analyses, we observe a common trend in both cases: as elongation increases, both cell lines exhibit strain stiffening. Notably, this effect is more prominent in MCF-10A compared to MDA-MB-436. Subsequently, we employ AFM on a dynamic range of 1-200 Hz to probe the mechanical characteristics of cell spheroids, focusing on both surface and bulk mechanics. Our findings align with the results from single cell investigations. Specifically, MCF-10A cells, characterized by strong contractile tissue tension, exhibit the greatest stiffness on their surface. Conversely, MDA-MB-436 cells, which experience significant elongation, showcase their highest stiffness within the bulk region. Consequently, the concept of single cell strain stiffening emerges as a crucial element in understanding the mechanics of multicellular spheroids (MCSs), even in the case of MDA-MB-436 cells, which are comparatively softer in nature.


Assuntos
Esferoides Celulares , Linhagem Celular , Elasticidade , Células Cultivadas , Microscopia de Força Atômica/métodos
5.
Cell Rep Med ; 5(2): 101411, 2024 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-38325381

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) is characterized by extremely poor prognosis. PDAC presents with molecularly distinct subtypes, with the basal-like one being associated with enhanced chemoresistance. Splicing dysregulation contributes to PDAC; however, its involvement in subtype specification remains elusive. Herein, we uncover a subtype-specific splicing signature associated with prognosis in PDAC and the splicing factor Quaking (QKI) as a determinant of the basal-like signature. Single-cell sequencing analyses highlight QKI as a marker of the basal-like phenotype. QKI represses splicing events associated with the classical subtype while promoting basal-like events associated with shorter survival. QKI favors a plastic, quasi-mesenchymal phenotype that supports migration and chemoresistance in PDAC organoids and cell lines, and its expression is elevated in high-grade primary tumors and metastatic lesions. These studies identify a splicing signature that defines PDAC subtypes and indicate that QKI promotes an undifferentiated, plastic phenotype, which renders PDAC cells chemoresistant and adaptable to environmental changes.


Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Processamento Alternativo/genética , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , Linhagem Celular , Fenótipo
6.
Cell Mol Life Sci ; 81(1): 82, 2024 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-38340178

RESUMO

Interaction between programmed death-1 (PD-1) ligand 1 (PD-L1) on tumor cells and PD-1 on T cells allows tumor cells to evade T cell-mediated immune surveillance. Strategies targeting PD-1/PD-L1 have shown clinical benefits in a variety of cancers. However, limited response rates in hepatocellular carcinoma (HCC) have prompted us to investigate the molecular regulation of PD-L1. Here, we identify B cell lymphoma-2-associated transcription factor 1 (BCLAF1) as a key PD-L1 regulator in HCC. Specifically, BCLAF1 interacts with SPOP, an E3 ligase that mediates the ubiquitination and degradation of PD-L1, thereby competitively inhibiting SPOP-PD-L1 interaction and subsequent ubiquitination and degradation of PD-L1. Furthermore, we determined an SPOP-binding consensus (SBC) motif mediating the BCLAF1-SPOP interaction on BCLAF1 protein and mutation of BCLAF1-SBC motif disrupts the regulation of the SPOP-PD-L1 axis. In addition, BCLAF1 expression was positively correlated with PD-L1 expression and negatively correlated with biomarkers of T cell activation, including CD3 and CD8, as well as with the level of immune cell infiltration in HCC tissues. Besides, BCLAF1 depletion leads to a significant reduction of PD-L1 expression in vitro, and this reduction of PD-L1 promoted T cell-mediated cytotoxicity. Notably, overexpression of BCLAF1 sensitized tumor cells to checkpoint therapy in an in vitro HCC cells-Jurkat cells co-culture model, whereas BCLAF1-SBC mutant decreased tumor cell sensitivity to checkpoint therapy, suggesting that BCLAF1 and its SBC motif serve as a novel therapeutic target for enhancing anti-tumor immunity in HCC.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Carcinoma Hepatocelular/metabolismo , Linhagem Celular , Neoplasias Hepáticas/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Receptor de Morte Celular Programada 1 , Proteínas Repressoras/genética , Proteínas Supressoras de Tumor , Evasão da Resposta Imune/genética
7.
PLoS Pathog ; 20(2): e1012022, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38359079

RESUMO

Enterovirus A71 (EV-A71) infection involves a variety of receptors. Among them, two transmembrane protein receptors have been investigated in detail and shown to be critical for infection: P-selectin glycoprotein ligand-1 (PSGL-1) in lymphocytes (Jurkat cells), and scavenger receptor class B member 2 (SCARB2) in rhabdomyosarcoma (RD) cells. PSGL-1 and SCARB2 have been reported to be expressed on the surface of Jurkat and RD cells, respectively. In the work reported here, we investigated the roles of PSGL-1 and SCARB2 in the process of EV-A71 entry. We first examined the expression of SCARB2 in Jurkat cells, and detected it within the cytoplasm, but not on the cell surface. Further, using PSGL-1 and SCARB2 knockout cells, we found that although both PSGL-1 and SCARB2 are essential for virus infection of Jurkat cells, virus attachment to these cells requires only PSGL-1. These results led us to evaluate the cell surface expression and the roles of SCARB2 in other EV-A71-susceptible cell lines. Surprisingly, in contrast to the results of previous studies, we found that SCARB2 is absent from the surface of RD cells and other susceptible cell lines we examined, and that although SCARB2 is essential for infection of these cells, it is dispensable for virus attachment. These results indicate that a receptor other than SCARB2 is responsible for virus attachment to the cell and probably for internalization of virions, not only in Jurkat cells but also in RD cells and other EV-A71-susceptible cells. SCARB2 is highly concentrated in lysosomes and late endosomes, where it is likely to trigger acid-dependent uncoating of virions, the critical final step of the entry process. Our results suggest that the essential interactions between EV-A71 and SCARB2 occur, not at the cell surface, but within the cell.


Assuntos
Enterovirus Humano A , Infecções por Enterovirus , Enterovirus , Humanos , Enterovirus/metabolismo , Enterovirus Humano A/genética , Enterovirus Humano A/metabolismo , Membrana Celular/metabolismo , Linhagem Celular , Receptores Depuradores/genética , Receptores Depuradores/metabolismo , /genética
8.
Toxins (Basel) ; 16(2)2024 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-38393158

RESUMO

Glioblastoma multiforme (GBM) is the most aggressive type of brain cancer, with a poor prognosis. GBM cells, which develop in the environment of neural tissue, often exploit neurotransmitters and their receptors to promote their own growth and invasion. Nicotinic acetylcholine receptors (nAChRs), which play a crucial role in central nervous system signal transmission, are widely represented in the brain, and GBM cells express several subtypes of nAChRs that are suggested to transmit signals from neurons, promoting tumor invasion and growth. Analysis of published GBM transcriptomes revealed spatial heterogeneity in nAChR subtype expression, and functional nAChRs of α1*, α7, and α9 subtypes are demonstrated in our work on several patient-derived GBM microsphere cultures and on the U87MG GBM cell line using subtype-selective neurotoxins and fluorescent calcium mobilization assay. The U87MG cell line shows reactions to nicotinic agonists similar to those of GBM patient-derived culture. Selective α1*, α7, and α9 nAChR neurotoxins stimulated cell growth in the presence of nicotinic agonists. Several cultivating conditions with varying growth factor content have been proposed and tested. The use of selective neurotoxins confirmed that cell cultures obtained from patients are representative GBM models, but the use of media containing fetal bovine serum can lead to alterations in nAChR expression and functioning.


Assuntos
Glioblastoma , Receptores Nicotínicos , Humanos , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Receptor Nicotínico de Acetilcolina alfa7/genética , Receptor Nicotínico de Acetilcolina alfa7/metabolismo , Agonistas Nicotínicos/farmacologia , Proteínas/metabolismo , Peptídeos/farmacologia , Linhagem Celular , Proliferação de Células , Antagonistas Nicotínicos/farmacologia
9.
Comput Biol Med ; 170: 108052, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38308868

RESUMO

The imbalance of epigenetic regulatory mechanisms such as DNA methylation, which can promote aberrant gene expression profiles without affecting the DNA sequence, may cause the deregulation of signaling, regulatory, and metabolic processes, contributing to a cancerous phenotype. Since some metabolites are substrates and cofactors of epigenetic regulators, their availability can be affected by characteristic cancer cell metabolic shifts, feeding cancer onset and progression through epigenetic deregulation. Hence, there is a need to study the influence of cancer metabolic reprogramming in DNA methylation to design new effective treatments. In this study, a generic Genome-Scale Metabolic Model (GSMM) of a human cell, integrating DNA methylation or demethylation reactions, was obtained and used for the reconstruction of Genome-Scale Metabolic Models enhanced with Enzymatic Constraints using Kinetic and Omics data (GECKOs) of 31 cancer cell lines. Furthermore, cell-line-specific DNA methylation levels were included in the models, as coefficients of a DNA composition pseudo-reaction, to depict the influence of metabolism over global DNA methylation in each of the cancer cell lines. Flux simulations demonstrated the ability of these models to provide simulated fluxes of exchange reactions similar to the equivalent experimentally measured uptake/secretion rates and to make good functional predictions. In addition, simulations found metabolic pathways, reactions and enzymes directly or inversely associated with the gene promoter methylation. Two potential candidates for targeted cancer epigenetic therapy were identified.


Assuntos
Metilação de DNA , Neoplasias , Humanos , Metilação de DNA/genética , Epigênese Genética , Linhagem Celular , Neoplasias/genética , Genoma
10.
Hum Cell ; 37(2): 523-530, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38329694

RESUMO

Atypical teratoid/rhabdoid (AT/RT) is a rare and highly malignant tumor of the central nervous system (CNS). It is most commonly found in children less than 5 years of age and is associated with inactivation of loss of function of SMARCB1/INI1. An experimental model for AT/RT is necessary to develop new and effective therapies. We established a patient-derived new cell line (MZ611ATRT), which showed loss of BAF-47. MZ611ATRT genetically features somatic heterozygous deletion of SMARCB1 and single nucleotide deletion of the residual allele, exon 5 ([c.541delC]), resulting in a stop codon at codon 954 by frameshift. We assessed the RNA-sequencing data of the other two AT/RT cell lines with forced expression of SMARCB1 available from public databases. We found SMARCB1 overexpression significantly down-regulates the expression of a group of enzymes related to cholesterol biosynthesis. Simvastatin was highly sensitive against MZ611ATRT cells and induced apoptosis (IC50 was 3.098 µM for MZ611ATRT, 41.88uM for U-87 MG, 23.34uM for IOMM-Lee, and 18.12uM for U-251 MG.). Pathways involved in cholesterol biosynthesis may be new targets for adjuvant therapy of AT/RT.


Assuntos
Colesterol , Criança , Humanos , Linhagem Celular , Éxons
11.
J Ethnopharmacol ; 325: 117870, 2024 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-38331121

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Masson Pine pollen (Pinus massoniana; MP) are used in Traditional Chinese Medicine to treat gut conditions. Early in vivo work supports this claim and suggests interaction of the material with the gastrointestinal immune system. AIM OF THE STUDY: The present study tested if and how MP material activates HD11 chicken macrophages in vitro using material from different production sites and harvest years. MATERIAL & METHODS: We applied twelve batches of MP from different Chinese production sites and harvest years. Materials were subjected to LAL tests (endotoxic activity), GC-MS (fatty acid analysis), and plate techniques (microbiological background, antimicrobial activity). Furthermore, HD11 chicken macrophages were challenged (6 h, 37 °C) with MP or LPS (E. coli O111:B4), respectively, to quantify nitric oxide (NO) production and immune gene expression (RT-qPCR). RESULTS: MP material promoted strong signals in LAL tests and contained significant amounts of 3-hydroxydodecanoic acid and 3-hydroxymyristic acid, irrespective of processing, harvest year, or origin. The pollen material activated HD11 chicken macrophages, which was confirmed by spikes of NO release and k-means cluster analysis of TLR-signaling pathway gene expression data. Response of NO production to Log2-titration of MP and LPS-treated media was in any case linear and significant. The response was reduced by polymyxin-B (PMB) and the inhibition was twice as strong for LPS than MP. No or minor microbiological background was detected on the majority of MP samples. Three samples showed presence of spoilage microorganisms and Gram-negative bacteria, but this did not correlate to LAL data or bacterial DNA counts. No antimicrobial activity of MP was evident. CONCLUSION: Pollen of the Masson Pine activated HD11 chicken macrophages in vitro, which is likely partially due to a background of bacterial LPS associated with the pollen material. However, as most of the effect (appr. 80%) could not be blocked by PMB this is certainly due to other stimuli. We hypothesize that polysaccharides and oligosaccharides of the pollen matrix have the potential to interact with certain immune receptors presented on the plasma membrane of chicken macrophages.


Assuntos
Galinhas , Pinus , Animais , Lipopolissacarídeos/farmacologia , Lipopolissacarídeos/metabolismo , Escherichia coli , Linhagem Celular , Macrófagos , Pólen
12.
Front Immunol ; 15: 1337557, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38390321

RESUMO

Introduction: The clinical efficacy of CAR-NK cells against CD19-expressing blood cancers has been demonstrated, and they have shown potential for treating solid tumors as well. However, the efficacy of CAR-NK cells for treating human oral tongue squamous cell carcinoma (OTSCC) has not been examined. Methods: We assessed MUC1 expression in human OTSCC tissue and a cell line using immunohistochemistry and immunofluorescence. We constructed NK cells that express CAR targeted to MUC1 from pluripotent stem cells (iPSC-derived MUC1-targeted CAR-NK cells) and evaluated their effectiveness against OTSCC in vitro using the xCELLigence Real-Time Cell Analysis system and CCK8 assay, and in vivo by measuring xenograft growth daily in BNDG mice treated with MUC1-targeted CAR-NK cells. As controls, we used iPSC-derived NK cells and NK-free media, which were CAR-free and blank, respectively. Results: MUC1 expression was detected in 79.5% (66/83) of all OTSCC patients and 72.7% (24/33) of stage III and IV. In stage III and IV MUC1 positive OTSCC, 63.6% (21/33) and 48.5% (16/33) patients had a MUC1-positive cancer cell rate of more than 50% and 80%, respectively. The iPSC-derived MUC1-targeted CAR-NK cells exhibited significant cytotoxicity against MUC1-expressing OTSCC cells in vitro, in a time- and dose-dependent manner, and showed a significant inhibitory effect on xenograft growth compared to both the iPSC-derived NK cells and the blank controls. We observed no weight loss, severe hematological toxicity or NK cell-mediated death in the BNDG mice. Conclusion: The MUC1-targeted CAR-NK cells had significant efficacy against human OTSCC, and their promising therapeutic response warrants further clinical trials.


Assuntos
Carcinoma de Células Escamosas , Neoplasias da Língua , Humanos , Animais , Camundongos , Carcinoma de Células Escamosas/terapia , Neoplasias da Língua/terapia , Células Matadoras Naturais , Linhagem Celular , Língua/metabolismo , Mucina-1/genética , Mucina-1/metabolismo
13.
Cells ; 13(4)2024 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-38391916

RESUMO

Base excision repair (BER) is the predominant pathway for the removal of most forms of hydrolytic, oxidative, and alkylative DNA lesions. The precise functioning of BER is achieved via the regulation of each step by regulatory/accessory proteins, with the most important of them being poly(ADP-ribose) polymerase 1 (PARP1). PARP1's regulatory functions extend to many cellular processes including the regulation of mRNA stability and decay. PARP1 can therefore affect BER both at the level of BER proteins and at the level of their mRNAs. Systematic data on how the PARP1 content affects the activities of key BER proteins and the levels of their mRNAs in human cells are extremely limited. In this study, a CRISPR/Cas9-based technique was used to knock out the PARP1 gene in the human HEK 293FT line. The obtained cell clones with the putative PARP1 deletion were characterized by several approaches including PCR analysis of deletions in genomic DNA, Sanger sequencing of genomic DNA, quantitative PCR analysis of PARP1 mRNA, Western blot analysis of whole-cell-extract (WCE) proteins with anti-PARP1 antibodies, and PAR synthesis in WCEs. A quantitative PCR analysis of mRNAs coding for BER-related proteins-PARP2, uracil DNA glycosylase 2, apurinic/apyrimidinic endonuclease 1, DNA polymerase ß, DNA ligase III, and XRCC1-did not reveal a notable influence of the PARP1 knockout. The corresponding WCE catalytic activities evaluated in parallel did not differ significantly between the mutant and parental cell lines. No noticeable effect of poly(ADP-ribose) synthesis on the activity of the above WCE enzymes was revealed either.


Assuntos
Reparo do DNA , Humanos , Extratos Celulares , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , DNA/metabolismo , Linhagem Celular , Proteína 1 Complementadora Cruzada de Reparo de Raio-X/genética
14.
Cells ; 13(4)2024 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-38391968

RESUMO

In this study, we investigated the beneficial effects of grapefruit IntegroPectin, derived from industrial waste grapefruit peels via hydrodynamic cavitation, on microglia cells exposed to oxidative stress conditions. Grapefruit IntegroPectin fully counteracted cell death and the apoptotic process induced by cell exposure to tert-butyl hydroperoxide (TBH), a powerful hydroperoxide. The protective effects of the grapefruit IntegroPectin were accompanied with a decrease in the amount of ROS, and were strictly dependent on the activation of the phosphoinositide 3-kinase (PI3K)/Akt cascade. Finally, IntegroPectin treatment inhibited the neuroinflammatory response and the basal microglia activation by down-regulating the PI3K- nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB)- inducible nitric oxide synthase (iNOS) cascade. These data strongly support further investigations aimed at exploring IntegroPectin's therapeutic role in in vivo models of neurodegenerative disorders, characterized by a combination of chronic neurodegeneration, oxidative stress and neuroinflammation.


Assuntos
Citrus paradisi , Microglia , Humanos , Microglia/metabolismo , Citrus paradisi/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/metabolismo , Linhagem Celular
15.
Signal Transduct Target Ther ; 9(1): 33, 2024 Feb 19.
Artigo em Inglês | MEDLINE | ID: mdl-38369543

RESUMO

Pyrogen, often as a contaminant, is a key indicator affecting the safety of almost all parenteral drugs (including biologicals, chemicals, traditional Chinese medicines and medical devices). It has become a goal to completely replace the in vivo rabbit pyrogen test by using the in vitro pyrogen test based on the promoted 'reduction, replacement and refinement' principle, which has been highly considered by regulatory agencies from different countries. We used NF-κB, a central signalling molecule mediating inflammatory responses, as a pyrogenic marker and the monocyte line THP-1 transfected with a luciferase reporter gene regulated by NF-κB as an in vitro model to detect pyrogens by measuring the intensity of a fluorescence signal. Here, we show that this test can quantitatively and sensitively detect endotoxin (lipopolysaccharide from different strains) and nonendotoxin (lipoteichoic acid, zymosan, peptidoglycan, lectin and glucan), has good stability in terms of NF-κB activity and cell phenotypes at 39 cell passages and can be applied to detect pyrogens in biologicals (group A & C meningococcal polysaccharide vaccine; basiliximab; rabies vaccine (Vero cells) for human use, freeze-dried; Japanese encephalitis vaccine (Vero cells), inactivated; insulin aspart injection; human albumin; recombinant human erythropoietin injection (CHO Cell)). The within-laboratory reproducibility of the test in three independent laboratories was 85%, 80% and 80% and the interlaboratory reproducibility among laboratories was 83.3%, 95.6% and 86.7%. The sensitivity (true positive rate) and specificity (true negative rate) of the test were 89.9% and 90.9%, respectively. In summary, the test provides a novel alternative for pyrogen detection.


Assuntos
NF-kappa B , Pirogênios , Animais , Chlorocebus aethiops , Coelhos , Humanos , Pirogênios/farmacologia , Pirogênios/química , Células Vero , Reprodutibilidade dos Testes , Linhagem Celular
16.
Sci Rep ; 14(1): 4000, 2024 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-38369625

RESUMO

Autophagy is activated in response to a variety of stress conditions including anti-cancer therapies, and tumors cells often depend on autophagy for survival. In this study, we have evaluated inhibition of autophagy as therapeutic strategy in acute lymphoblastic leukemia (ALL) in children, both as a single treatment and in combination with glucocorticoid (GC) Dexamethasone (Dexa). Analysis of proteomics and RNA-seq of ALL cell lines and primary samples identified an upregulation of Vps34 and ATG14 proteins and autophagy and lysosomal pathway enrichment in a genetic subgroup with a recurrent t(12;21) translocation. Cells from this sugbroup were also significantly more sensitive to the selective autophagy or lysosomal inhibitors than cells with other genetic rearrangements. Further, combination of Dexa with either lysosomal or autophagy inhibitors was either synergistic or additive in killing leukemic cells across various genetic and lineage backgrounds, for both cell lines and primary samples, as assessed using viability assays and SynergyFinder as well as apoptotic caspase 3/7-based live-cell assays. Our data demonstrate that targeting autophagy represents a promising strategy for the treatment of pediatric ALL, both as a selective modality for the t(12;21) pre-B-ALL subgroup, and in combination treatments to sensitize to GC-induced cytotoxicity.


Assuntos
Leucemia-Linfoma Linfoblástico de Células Precursoras B , Leucemia-Linfoma Linfoblástico de Células Precursoras , Criança , Humanos , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Autofagia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Linhagem Celular , Glucocorticoides/uso terapêutico , Linhagem Celular Tumoral , Apoptose
17.
Vet Med Sci ; 10(2): e1350, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38373050

RESUMO

BACKGROUND: Feline large granular lymphocyte (LGL) lymphoma is an aggressive neoplasia characterised by short survival and poor response to chemotherapy. OBJECTIVES: In this study, the effect of different chemotherapeutic agents on the growth kinetics of the feline cell line S87, a non-MHC-restricted feline LGL cell line, was investigated. Where possible, IC50 (inhibitory concentration 50) values were determined. The IC50 values of the cell line as lymphoma models can provide clues to the situation in vivo and serve as a basis for studying resistance mechanisms. METHODS: Cells were incubated with various concentrations of vincristine, doxorubicin, 4-hydroperoxycyclophosphamide, prednisolone, methotrexate and L-asparaginase for 24 and 48 h, respectively. RESULTS: The IC50 values could be determined as 14.57 (7.49-28.32) µg/mL at 24 h incubation and 5.72 (4.05-8.07) µg/mL at 48 h incubation for doxorubicin and 9.12 (7.72-10.76) µg/mL at 24 h incubation and 4.53 (3.74-5.47) µg/mL at 48 h incubation for 4-hydroperpoxycyclophosphamide. Treatment with vincristine and methotrexate resulted in relatively high cell resistance whereas L-asparaginase and prednisolone treatment led to a reduction in cell number compared to control while cell viability was not affected (cytostatic effect). CONCLUSION: Overall, the feline LGL cell line S87 proves to be relatively sensitive to doxorubicin and 4-hydroperoxycyclophosphamide and relatively resistant to treatment with vincristine, prednisolone, methotrexate and L-asparaginase. The results of this study can be used for further investigations on resistance mechanisms in feline LGL lymphoma. Doxorubicin and cyclophosphamide can be interpreted as promising candidates for the therapy of feline LGL lymphomas.


Assuntos
Doenças do Gato , Ciclofosfamida/análogos & derivados , Linfoma , Gatos , Animais , Vincristina , Asparaginase/uso terapêutico , Metotrexato/uso terapêutico , Linfoma/tratamento farmacológico , Linfoma/veterinária , Linfoma/patologia , Doxorrubicina/farmacologia , Linhagem Celular , Prednisolona/farmacologia , Prednisolona/uso terapêutico , Linfócitos/patologia
18.
Pediatr Surg Int ; 40(1): 55, 2024 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-38347163

RESUMO

PURPOSE: In this research, we analyzed the expression of serpinB9 in hepatoblastoma and investigated the factors which enhance its expression. METHOD: SerpinB9 expression in hepatoblastoma cell lines and macrophages co-cultured with each other or stimulated by anticancer agents was examined using RT-qPCR and western blotting. Immunohistochemistry for SerpinB9 in hepatoblastoma specimens was performed. Single-cell RNA-sequence data for hepatoblastoma from an online database were analyzed to investigate which types of cells express SerpinB9. RESULT: HepG2, a hepatoblastoma cell line, exhibited increased expression of SerpinB9 when indirectly co-cultured with macrophages. Immunohistochemistry for the specimens demonstrated that serpinB9 is positive not in hepatoblastoma cells but in macrophages. Single-cell RNA sequence analysis in tissues from hepatoblastoma patients showed that macrophages expressed SerpinB9 more than tumor cells did. Co-culture of macrophages with hepatoblastoma cell lines led to the enhanced expression of SerpinB9 in both macrophages and cell lines. Anticancer agents induced an elevation of SerpinB9 in hepatoblastomas cell lines. CONCLUSION: In hepatoblastoma, SerpinB9 is thought to be more highly expressed in macrophages and enhanced by interaction with hepatoblastoma cell.


Assuntos
Antineoplásicos , Hepatoblastoma , Neoplasias Hepáticas , Humanos , Linhagem Celular , Hepatoblastoma/patologia , Imuno-Histoquímica , Neoplasias Hepáticas/patologia , Microambiente Tumoral/genética
19.
Invest Ophthalmol Vis Sci ; 65(2): 27, 2024 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-38349785

RESUMO

Purpose: Epigenetic alterations in uveal melanoma (UM) are still neither well characterized, nor understood. In this pilot study, we sought to provide a deeper insight into the possible role of epigenetic alterations in the pathogenesis of UM and their potential prognostic relevance. To this aim, we comprehensively profiled histone post-translational modifications (PTMs), which represent epigenetic features regulating chromatin accessibility and gene transcription, in UM formalin-fixed paraffin-embedded (FFPE) tissues, control tissues, UM cell lines, and healthy melanocytes. Methods: FFPE tissues of UM (n = 24), normal choroid (n = 4), human UM cell lines (n = 7), skin melanocytes (n = 6), and uveal melanocytes (n = 2) were analyzed through a quantitative liquid chromatography-mass spectrometry (LC-MS) approach. Results: Hierarchical clustering showed a clear separation with several histone PTMs that changed significantly in a tumor compared to normal samples, in both tissues and cell lines. In addition, several acetylations and H4K20me1 showed lower levels in BAP1 mutant tumors. Some of these changes were also observed when we compared GNA11 mutant tumors with GNAQ tumors. The epigenetic profiling of cell lines revealed that the UM cell lines MP65 and UPMM1 have a histone PTM pattern closer to the primary tissues than the other cell lines analyzed. Conclusions: Our results suggest the existence of different histone PTM patterns that may be important for diagnosis and prognosis in UM. However, further analyses are needed to confirm these findings in a larger cohort. The epigenetic characterization of a panel of UM cell lines suggested which cellular models are more suitable for epigenetic investigations.


Assuntos
Melanoma , Neoplasias Uveais , Humanos , Histonas , Projetos Piloto , Melanoma/metabolismo , Melanócitos/metabolismo , Neoplasias Uveais/patologia , Linhagem Celular , Espectrometria de Massas
20.
J Biomed Mater Res B Appl Biomater ; 112(2): e35381, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38348489

RESUMO

Recent studies show good osteoinductive properties of polyurethanes modified with polyhedral oligomeric silsesquioxanes (POSS). In this work, three types of POSS; propanediolisobutyl-POSS (PHI-POSS), disilanolisobutyl-POSS (DSI-POSS), and octahydroxybutyl-POSS (OCTA-POSS) were chemically incorporated into linear polyurethane based on an aliphatic isocyanate, hexamethylene diisocyanate (HDI), to obtain new nanohybrid PU-POSS materials. The full conversion of POSS was confirmed by Fourier transform infrared spectroscopy (FTIR-ATR) spectra of the model reactions with pure HDI. The materials obtained were investigated by FTIR, SEM-EDS, and DSC. The DSC studies showed the thermoplasticity of the obtained materials and apparently good recovery. 30-day immersion in SBF (simulated body fluid) revealed an increase in the rate of deposition of hydroxyapatite (HAp) for the highest POSS loadings, resulting in thick layers of hydroxyapatite (~60-40 µm), and the Ca/P ratio 1.67 (even 1.785). The structure and properties of the inorganic layer depend on the type of POSS, the number of hard segments, and those containing POSS, which can be tailored by changing the HDI/poly(tetramethylene glycol) (PTMG) ratio. Furthermore, the obtained composites revealed good biocompatibility, as confirmed by cytotoxicity tests conducted on two cell lines; normal human dermal fibroblasts (NHDF) and primary human osteoblasts (HOB). Adherent cells seeded on the tested materials showed viability even after a 48-h incubation. After this time, the population of viable, and proliferating cells exceeded 90%. Bioimaging studies have shown the fibroblast and osteoblast cells were well attached to the surface of the tested materials.


Assuntos
Durapatita , Isocianatos , Poliuretanos , Humanos , Poliuretanos/farmacologia , Poliuretanos/química , Espectroscopia de Infravermelho com Transformada de Fourier , Osteoblastos , Linhagem Celular
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA