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1.
J Cell Sci ; 137(9)2024 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-38606629

RESUMO

The ADP-ribosylation factors (ARFs) and ARF-like (ARL) GTPases serve as essential molecular switches governing a wide array of cellular processes. In this study, we used proximity-dependent biotin identification (BioID) to comprehensively map the interactome of 28 out of 29 ARF and ARL proteins in two cellular models. Through this approach, we identified ∼3000 high-confidence proximal interactors, enabling us to assign subcellular localizations to the family members. Notably, we uncovered previously undefined localizations for ARL4D and ARL10. Clustering analyses further exposed the distinctiveness of the interactors identified with these two GTPases. We also reveal that the expression of the understudied member ARL14 is confined to the stomach and intestines. We identified phospholipase D1 (PLD1) and the ESCPE-1 complex, more precisely, SNX1, as proximity interactors. Functional assays demonstrated that ARL14 can activate PLD1 in cellulo and is involved in cargo trafficking via the ESCPE-1 complex. Overall, the BioID data generated in this study provide a valuable resource for dissecting the complexities of ARF and ARL spatial organization and signaling.


Assuntos
Fatores de Ribosilação do ADP , Fosfolipase D , Transdução de Sinais , Fatores de Ribosilação do ADP/metabolismo , Fatores de Ribosilação do ADP/genética , Humanos , Fosfolipase D/metabolismo , Fosfolipase D/genética , Células HEK293 , Animais , Nexinas de Classificação/metabolismo , Nexinas de Classificação/genética , Mapeamento de Interação de Proteínas
2.
Anticancer Drugs ; 35(3): 251-262, 2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38164802

RESUMO

The potential treatment option of targeting DNA methyltransferase 1 (DNMT1) has been explored, but further investigation is required to assess the efficacy of combination therapy in acute myeloid leukemia (AML). In this study, bioinformatics and online databases were utilized to select the combined therapeutic targets. The potential kinases associated with DNMT1-related genes in AML were analyzed using the Cancer Genome Atlas (TCGA) database and X2K Appyter (Expression2Kinases) database. In-vitro evaluations were conducted to assess the synergistic effects between DNMT1 and ATR/ATM in five AML cell lines (MOLM-16, NB-4, HEL 92.1.7, HEL, EOL-1). In our study, ATR and ATM are primarily the kinases associated with DNMT1-related genes in AML. We observed a significant upregulation of DNMT1, ATR, and ATM expression in AML tissues and cell lines. The five AML cell lines demonstrated sensitivity to monotherapy with GSK-368, AZD-1390, or AZD-6738 (EC50 value ranges from 5.461 to 7.349 nM, 5.821 to 10.120 nM, and 7.618 to 10.100 nM, respectively). A considerable synergistic effect was observed in AML cell lines when combining GSK-368 and AZD-1390, GSK-368 and AZD-6738, or AZD-1390 and AZD-6738, resulting in induced cell apoptosis and inhibited cell growth. DNMT1, ATM, and ATR possess potential as therapeutic targets for AML. Both individual targeting and combination targeting of these molecules have been confirmed as promising therapeutic approaches for AML.


Assuntos
Indóis , Leucemia Mieloide Aguda , Pirimidinas , Sulfonamidas , Humanos , Linhagem Celular Tumoral , Pirimidinas/farmacologia , Morfolinas/farmacologia , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Apoptose , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo
3.
Planta Med ; 90(2): 154-165, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37931776

RESUMO

Astragaloside IV (AS-IV) has been shown to provide renal protection in various kidney injury models. However, the metabolic profile variation of AS-IV in pathological models in vivo is not well established. This study aims to explore the metabolic pathway of AS-IV in vivo in the classical puromycin aminonucleoside (PAN)-induced kidney injury in a rat model. Twelve Wistar rats were randomly divided into the AS-IV (CA) and the PAN+AS-IV (PA) treatment groups. PAN was injected by a single tail intravenous (i. v.) injection at 5 mg/100 g body weight, and AS-IV was administered intragastrically (i. g.) at 40 mg/kg for 10 days. Fecal samples of these rats were collected, and metabolites of AS-IV were detected by ultra-performance liquid chromatography coupled with quadrupole/time-of-flight mass spectrometry (UPLC-Q-TOF-MS/MS) to explore the AS-IV metabolic pathway. The metabolic differences between the AS-IV and PAN+AS-IV groups were compared. A total of 25 metabolites were detected, and deglycosylation, deoxygenation, and methyl oxidation were found to be the main metabolic pathways of AS-IV in vivo. The abundance of most of these metabolites in the PAN+AS-IV group was lower than that in the AS-IV treatment group, and differences for seven of them were statistically significant. Our study indicates that AS-IV metabolism is affected in the PAN-induced kidney injury rat model.


Assuntos
Saponinas , Espectrometria de Massas em Tandem , Triterpenos , Ratos , Animais , Espectrometria de Massas em Tandem/métodos , Cromatografia Líquida de Alta Pressão/métodos , Ratos Wistar , Puromicina
4.
J Ultrasound Med ; 2024 Oct 04.
Artigo em Inglês | MEDLINE | ID: mdl-39365033

RESUMO

OBJECTIVE: Fetal growth restriction (FGR) is a condition where fetuses fail to reach their genetic potential for growth, posing a significant health challenge for newborns. The aim of this research was to explore the efficacy of texture-based analysis of neurosonographic images in identifying FGR in fetuses, which may provide a promising tool for early assessment of FGR. METHODS: A retrospective analysis collected 100 intrauterine neurosonographic images from 50 FGR and 50 gestational age-appropriate fetuses. Using MaZda software, approximately 300 texture features were extracted from occipital white matter (OWM) and cerebellum of intrauterine neurosonographic images, respectively. Then 10 optimal features were separately selected by 3 algorithms, including the Fisher coefficient method, the method of minimizing classification error probability and average correlation coefficients, and the mutual information coefficient method. Further, the 10 statistically most significant features were selected from these sets to form the mixed feature set. After nonlinear discriminant analysis was performed to reduce feature dimensionality, the artificial neural network (ANN) classifier was conducted, respectively. RESULTS: For OWM and cerebellum, a total of 11 and 14 statistically significant features were selected. When the mixed feature sets of OWM and cerebellum were applied to ANN classifier, classification accuracy were 90.00% (κ = 0.800; P < .001) and 93.00% (κ = 0.860; P < .001), and the receiver operating characteristic curve for identifying FGR showed an area under the curve of 0.82 and 0.87. CONCLUSIONS: Texture analysis of fetal intrauterine neurosonographic images is a feasible and noninvasive strategy for evaluating FGR fetuses.

5.
Mar Drugs ; 22(5)2024 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-38786610

RESUMO

Octocoral of the genus Clavularia is a kind of marine invertebrate possessing abundant cytotoxic secondary metabolites, such as prostanoids and dolabellanes. In our continuous natural product study of C. spp., two previously undescribed prostanoids [clavulone I-15-one (1) and 12-O-deacetylclavulone I (2)] and eleven known analogs (3-13) were identified. The structures of these new compounds were elucidated based on analysis of their 1D and 2D NMR, HRESIMS, and IR data. Additionally, all tested prostanoids (1 and 3-13) showed potent cytotoxic activities against the human oral cancer cell line (Ca9-22). The major compound 3 showed cytotoxic activity against the Ca9-22 cells with the IC50 value of 2.11 ± 0.03 µg/mL, which echoes the cytotoxic effect of the coral extract. In addition, in silico tools were used to predict the possible effects of isolated compounds on human tumor cell lines and nitric oxide production, as well as the pharmacological potentials.


Assuntos
Antozoários , Antineoplásicos , Prostaglandinas , Humanos , Antozoários/química , Animais , Linhagem Celular Tumoral , Prostaglandinas/farmacologia , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Óxido Nítrico/metabolismo , Concentração Inibidora 50 , Organismos Aquáticos , Estrutura Molecular
6.
Proc Natl Acad Sci U S A ; 118(48)2021 11 30.
Artigo em Inglês | MEDLINE | ID: mdl-34810254

RESUMO

Enzymes that produce second messengers are highly regulated. Revealing the mechanisms underlying such regulation is critical to understanding both how cells achieve specific signaling outcomes and return to homeostasis following a particular stimulus. Pooled genome-wide CRISPR screens are powerful unbiased approaches to elucidate regulatory networks, their principal limitation being the choice of phenotype selection. Here, we merge advances in bioorthogonal fluorescent labeling and CRISPR screening technologies to discover regulators of phospholipase D (PLD) signaling, which generates the potent lipid second messenger phosphatidic acid. Our results reveal glycogen synthase kinase 3 as a positive regulator of protein kinase C and PLD signaling. More generally, this work demonstrates how bioorthogonal, activity-based fluorescent tagging can expand the power of CRISPR screening to uncover mechanisms regulating specific enzyme-driven signaling pathways in mammalian cells.


Assuntos
Quinase 3 da Glicogênio Sintase/metabolismo , Fosfolipase D/metabolismo , Proteína Quinase C-alfa/metabolismo , Fenômenos Biológicos , Sistemas CRISPR-Cas/genética , Química Click/métodos , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Quinase 3 da Glicogênio Sintase/fisiologia , Células HEK293 , Humanos , Células K562 , Ácidos Fosfatídicos/metabolismo , Fosfolipase D/fisiologia , Proteína Quinase C-alfa/fisiologia , Sistemas do Segundo Mensageiro , Transdução de Sinais
7.
Endocr Res ; 49(4): 213-222, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38867680

RESUMO

BACKGROUND: Diabetes mellitus (DM) and complications such as chronic kidney disease and cardiovascular symptoms pose a substantial public health burden. Increasing studies have shown that circular RNAs (circRNAs) regulate many gene expressions that are essential in diverse pathological and biological procedures. However, the roles of particular circRNAs in DM are unclear. METHODS: In the current investigation, endothelial progenitor cells (EPCs) were used to search for abnormal expression of circRNAs by using high-throughput sequencing under high glucose (HG) conditions. The regulatory mechanisms and targets were then studied through bioinformatics analysis, luciferase reporter analysis, angiogenic differentiation experiments, flow cytometry detection of apoptosis and RT-qPCR analysis. RESULTS: The circ-Astn1 expression in EPCs decreased after HG treatment. Overexpression or circ-Astn1 suppressed HG induced endothelial cell damage. MicroRNA (miR)-138-5p and SIRT5 were found to be the downstream targets of circ-Astn1 through luciferase reporter analysis. SIRT5 downregulation or miR-138-5p overexpression reversed circ-Astn1's protective effect against HG induced endothelial cell dysfunction, including apoptosis and abnormal vascular differentiation. Furthermore, circ-Astn1 overexpression promoted autophagy activation by increasing SIRT5 expression under HG conditions. Our findings suggest that circ-Astn1 mediated promotion of SIRT5 facilitates autophagy by sponging miR-138-5p. CONLUSION: Together, our findings show that the overexpression of circ-Astn1 suppresses HG induced endothelial cell damage by targeting miR-138-5p/SIRT5 axis.


Assuntos
Autofagia , Células Progenitoras Endoteliais , Glucose , RNA Circular , Células Progenitoras Endoteliais/efeitos dos fármacos , Células Progenitoras Endoteliais/metabolismo , RNA Circular/metabolismo , RNA Circular/genética , Autofagia/efeitos dos fármacos , Autofagia/fisiologia , Glucose/farmacologia , Humanos , MicroRNAs/metabolismo , Células Cultivadas , Apoptose/efeitos dos fármacos , Apoptose/fisiologia
8.
Int J Mol Sci ; 25(15)2024 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-39125630

RESUMO

Melanosomes are specialized membrane-bound organelles where melanin is synthesized and stored. The levels of melanin can be effectively reduced by inhibiting melanin synthesis or promoting melanosome degradation via autophagy. Ceramide, a key component in the metabolism of sphingolipids, is crucial for preserving the skin barrier, keeping it hydrated, and warding off the signs of aging. Our preliminary study indicated that a long-chain C22-ceramide compound (Ehux-C22) isolated from the marine microalga Emiliania huxleyi, reduced melanin levels via melanosomal autophagy in B16 cells. Recently, microRNAs (miRNAs) were shown to act as melanogenesis-regulating molecules in melanocytes. However, whether the ceramide Ehux-C22 can induce melanosome autophagy at the post-transcriptional level, and which potential autophagy-dependent mechanisms are involved, remains unknown. Here, miR-199a-3p was screened and identified as a novel upregulated miRNA in Ehux-C22-treated B16 cells. An in vitro high melanin expression model in cultured mouse melanoma cells (B16 cells) was established by using 0.2 µM alpha-melanocyte-stimulating hormone(α-MSH) and used for subsequent analyses. miR-199a-3p overexpression significantly enhanced melanin degradation, as indicated by a reduction in the melanin level and an increase in melanosome autophagy. Further investigation demonstrated that in B16 cells, Ehux-C22 activated miR-199a-3p and inhibited mammalian target of rapamycin(mTOR) level, thus activating the mTOR-ULK1 signaling pathway by promoting the expression of unc-51-like autophagy activating kinase 1 (ULK1), B-cell lymphoma-2 (Bcl-2), Beclin-1, autophagy-related gene 5 (ATG5), and microtubule-associated protein light chain 3 (LC3-II) and degrading p62. Therefore, the roles of Ehux-C22-regulated miR-199a-3p and the mTOR pathway in melanosomal autophagy were elucidated. This research may provide novel perspectives on the post-translational regulation of melanin metabolism, which involves the coordinated control of melanosomes.


Assuntos
Autofagia , Ceramidas , Melaninas , Melanoma Experimental , Melanossomas , MicroRNAs , Transdução de Sinais , Serina-Treonina Quinases TOR , MicroRNAs/genética , MicroRNAs/metabolismo , Animais , Camundongos , Serina-Treonina Quinases TOR/metabolismo , Melanossomas/metabolismo , Ceramidas/metabolismo , Melaninas/metabolismo , Melaninas/biossíntese , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Melanoma Experimental/genética , Linhagem Celular Tumoral , alfa-MSH/metabolismo , Melanócitos/metabolismo , Melanócitos/efeitos dos fármacos
9.
Molecules ; 29(5)2024 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-38474566

RESUMO

In light of industrial developments, water pollution by heavy metals as hazardous chemicals has garnered attention. Addressing the urgent need for efficient heavy metal removal from aqueous environments, this study delves into using poly-γ-glutamic acid (γ-PGA) for the bioflocculation of heavy metals. Utilizing γ-PGA variants from Bacillus subtilis with different molecular weights and salt forms (Na-bonded and Ca-bonded), the research evaluates their adsorption capacities for copper (Cu), lead (Pb), and cadmium (Cd) ions. It was found that Na-bonded γ-PGA with a high molecular weight showed the highest heavy metal adsorption (92.2-98.3%), particularly at a 0.5% concentration which exhibited the highest adsorption efficiency. Additionally, the study investigated the interaction of γ-PGA in mixed heavy metal environments, and it was discovered that Na-γ-PGA-HM at a 0.5% concentration showed a superior adsorption efficiency for Pb ions (85.4%), highlighting its selectivity as a potential effective biosorbent for wastewater treatment. This research not only enlightens the understanding of γ-PGA's role in heavy metal remediation but also underscores its potential as a biodegradable and non-toxic alternative for environmental cleanup. The findings pave the way for further exploration into the mechanisms and kinetics of γ-PGA's adsorption properties.


Assuntos
Metais Pesados , Ácido Poliglutâmico/análogos & derivados , Poluentes Químicos da Água , Cádmio/química , Ácido Glutâmico , Chumbo , Peso Molecular , Metais Pesados/química , Água , Íons , Cloreto de Sódio , Adsorção , Concentração de Íons de Hidrogênio , Cinética
10.
Isr J Chem ; 63(1-2)2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37588264

RESUMO

In less than a decade, CRISPR screening has revolutionized forward genetics and cell and molecular biology. Advances in screening technologies, including sgRNA libraries, Cas9-expressing cell lines, and streamlined sequencing pipelines, have democratized pooled CRISPR screens at genome-wide scale. Initially, many such screens were survival-based, identifying essential genes in physiological or perturbed processes. With the application of new chemical biology tools to CRISPR screening, the phenotypic space is no longer limited to live/dead selection or screening for levels of conventional fluorescent protein reporters. Further, the resolution has been increased from cell populations to single cells or even the subcellular level. We highlight advances in pooled CRISPR screening, powered by chemical biology, that have expanded phenotypic space, resolution, scope, and scalability as well as strengthened the CRISPR/Cas enzyme toolkit to enable biological hypothesis generation and discovery.

11.
Curr Issues Mol Biol ; 44(5): 2243-2256, 2022 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-35678681

RESUMO

Oral squamous cell carcinoma (OSCC) is one of the most common cancers in the world, and the incidence and death rate of OSCC in men is twice that of women. CD47 is a ubiquitous cell surface transmembrane protein, also known as integrin-related protein (IAP). Previous studies have pointed out that CD47 can inhibit the growth of OSCC, but the detailed mechanism is not clear. This study aimed to explore the effect of CD47 gene expression profiles in OSCC. The OSCC cell lines, OECM-1 and OC-2, overexpressed CD47, and the expression profiles of mRNAs were analyzed through next-generation sequencing (NGS) with a bioinformatic approach. A total of 14 differentially expressed genes (DEGs) were listed. In addition, ingenuity pathway analysis (IPA) was used to analyze the molecular function (MF), biological process (BP), and cellular component (CC) network signaling. The human protein atlas (HPA) database was used to analyze gene expression and the survivability of human cancer. The results found that HSPA5, HYOU1, and PDIA4 were involved in the IPA network and when highly expressed, mediated the survivability of cancer. In addition, HSPA5 was positively and significantly correlated with CD47 expression (p < 0.0001) and induced by CD47-overexpression in the OECM-1 and OC-2 OSCC cancer cell lines. These findings provide important insights into possible new diagnostic strategies, including unfolded protein for OSCC-targeting CD47.

12.
Acta Pharmacol Sin ; 43(5): 1251-1263, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-34417577

RESUMO

Transcriptional factor EB (TFEB), a master regulator of autophagy and lysosomal biogenesis, is generally regarded as a pro-survival factor. Here, we identify that besides its effect on autophagy induction, TFEB exerts a pro-apoptotic effect in response to the cyclopentenone prostaglandin 15-deoxy-∆-12,14-prostaglandin J2 (15d-PGJ2). Specifically, 15d-PGJ2 promotes TFEB translocation from the cytoplasm into the nucleus to induce autophagy and lysosome biogenesis via reactive oxygen species (ROS) production rather than mTORC1 inactivation. Surprisingly, TFEB promotes rather than inhibits apoptosis in response to 15d-PGJ2. Mechanistically, ROS-mediated TFEB translocation into the nucleus transcriptionally upregulates the expression of ATF4, which is required for apoptosis elicited by 15d-PGJ2. Additionally, inhibition of TFEB activation by ROS scavenger N-acetyl cysteine or inhibition of protein synthesis by cycloheximide effectively compromises ATF4 upregulation and apoptosis in response to 15d-PGJ2. Collectively, these results indicate that ROS-induced TFEB activation exerts a novel role in promoting apoptosis besides its role in regulating autophagy in response to 15d-PGJ2. This work not only evidences how TFEB is activated by 15d-PGJ2, but also unveils a previously unexplored role of ROS-dependent activation of TFEB in modulating cell apoptosis in response to 15d-PGJ2.


Assuntos
Prostaglandina D2 , Prostaglandinas , Apoptose , Autofagia , Ciclopentanos , Prostaglandina D2/análogos & derivados , Prostaglandina D2/farmacologia , Prostaglandinas/farmacologia , Espécies Reativas de Oxigênio/metabolismo
13.
Biosci Biotechnol Biochem ; 86(5): 646-654, 2022 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-35218182

RESUMO

Atopic dermatitis (AD) is a chronic inflammatory and pruritic disease; it can be treated by inhibiting inflammation. Sarcodia suiae sp. is an edible, artificially cultivable red algae with multiple bioactivities. We assessed the anti-inflammatory activity of the ethyl acetate fraction of S. suiae sp. ethanol extract (PD1) on 1-chloro-2,4-dinitrochlorobenzene (DNCB)-induced AD-like lesions. Results show that PD1 alleviated symptoms and significantly decreased clinical dermatitis score. PD1 inhibited serum immunoglobulin E expression and alleviated swelling in the spleen and subiliac lymph nodes. In skin tissues, PD1 alleviated aberrant hyperplasia, decreased epidermal thickness, and decreased the accumulation of mast cells. PD1 mediated the recovery of skin barrier-related proteins, such as claudin-1 and filaggrin. Our study demonstrated that PD1 has anti-inflammatory effects, alleviates AD symptoms, inhibits inflammatory responses in skin tissues, and restores barrier function in DNCB-induced AD mice. These findings reveal that S. suiae sp. extract provides an alternative protective option against AD.


Assuntos
Dermatite Atópica , Rodófitas , Acetatos , Animais , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Citocinas/metabolismo , Dermatite Atópica/induzido quimicamente , Dermatite Atópica/tratamento farmacológico , Dermatite Atópica/metabolismo , Dinitroclorobenzeno/metabolismo , Dinitroclorobenzeno/farmacologia , Dinitroclorobenzeno/uso terapêutico , Etanol/metabolismo , Inflamação/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Extratos Vegetais/metabolismo , Rodófitas/metabolismo , Pele
14.
J Stroke Cerebrovasc Dis ; 31(2): 106221, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34837757

RESUMO

OBJECTIVE: To assess the potential effect of dl-3-N-butylphthalide (dl-NBP) for the proliferation and differentiation of neural stem cells (NSCs) against hypoxia and the underlying mechanism. MATERIALS AND METHODS: Hippocampal NSCs were obtained from fetal rats. NSCs combined with dl-NBP and single NSCs were cultured. The impact of siRNA-mediated hypoxia-inducible factor-1alpha (HIF-1α) knockdown on NSCs was detected with western blotting (WB) and quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR). Cell-counting kit-8 assay was used for evaluating the viability of NSCs. Levels of HIF-1α protein were measured using WB, and vascular endothelial growth factor (VEGF) expression was quantified using RT-qPCR and enzyme-linked immunosorbent assay. RESULTS: Compared with 7 different concentrations of dl-NBP, 0.25 g/L was determined as the optimal concentration to significantly increase the viability of NSCs (p < 0.001). Dl-NBP can significantly increase the viability of hypoxic NSCs (p < 0.001) and improve the differentiation of hypoxic NSCs into astrocytes (p = 0.001) and oligodendrocytes (p < 0.001). Meanwhile, Dl-NBP can significantly elevate levels of HIF-1α protein (p < 0.001) and VEGF mRNA (p = 0.001) / protein (p < 0.001) in NSCs in the hypoxic environment. However, after transfection with HIF-1α siRNA in NSCs, the viability and differentiation of NSCs was not recovered using dl-NBP under the hypoxic condition, as well as levels of HIF-1α and VEGF. CONCLUSION: Dl-NBP can reverse the weaker proliferation and differentiation power of NSCs in the hypoxic environment. The HIF-1α - VEGF pathway may be implicated in this protective effect of dl-NBP.


Assuntos
Benzofuranos , Subunidade alfa do Fator 1 Induzível por Hipóxia , Hipóxia , Células-Tronco Neurais , Animais , Benzofuranos/farmacologia , Hipóxia/prevenção & controle , Subunidade alfa do Fator 1 Induzível por Hipóxia/efeitos dos fármacos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Células-Tronco Neurais/patologia , Fármacos Neuroprotetores/farmacologia , Ratos
15.
Int J Mol Sci ; 23(3)2022 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-35163437

RESUMO

Insulin-like growth factor 1 (IGF-1) not only regulates neuronal function and development but also is neuroprotective in the setting of acute ischemic stroke. G-protein-coupled receptor 17 (GPR17) expression in brain tissue serves as an indicator of brain damage. As whether IGF-1 regulates GPR17 expression remains unknown, the aim of this study is to investigate how IGF-1 regulates GPR17 expression in vitro. Human neuroblastoma SK-N-SH cells were used. Lentivirus-mediated short hairpin RNA (shRNA) was constructed to mediate the silencing of FoxO1, while adenoviral vectors were used for its overexpression. Verification of the relevant signaling cascade was performed using a FoxO1 inhibitor (AS1842856), a phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002), and a GPR17 antagonist (cangrelor). Cell proliferation was analyzed using EdU staining; immunofluorescence staining was used to detect the expression and subcellular localization of FoxO1. Chromatin immunoprecipitation was used to analyze the binding of FoxO1 to the GPR17 promoter in SK-N-SH cells. The expression of FoxO1, GPR17, and protein kinase B (also known as Akt) mRNA and protein as well as the levels of FoxO1 and Akt phosphorylation were investigated in this study. IGF-1 was found to downregulate FoxO1 and GPR17 expression in SK-N-SH cells while promoting cell viability and proliferation. Inhibition of FoxO1 and antagonism of GPR17 were found to play a role similar to that of IGF-1. Silencing of FoxO1 by lentivirus-mediated shRNA resulted in the downregulation of FoxO1 and GPR17 expression. The overexpression of FoxO1 via adenoviral vectors resulted in the upregulation of FoxO1 and GPR17 expression. Blocking of PI3K signaling by LY294002 inhibited the effect of IGF-1 on GPR17 suppression. Results from chromatin immunoprecipitation revealed that IGF-1 promotes FoxO1 nuclear export and reduces FoxO1 binding to the GPR17 promoter in SK-N-SH cells. Here, we conclude that IGF-1 enhances cell viability and proliferation in SK-N-SH cells via the promotion of FoxO1 nuclear export and reduction of FoxO1 binding to the GPR17 promoter via PI3K/Akt signaling. Our findings suggest that the enhancement of IGF-1 signaling to antagonize GPR17 serves as a potential therapeutic strategy in the management of acute ischemic stroke.


Assuntos
Regulação para Baixo , Proteína Forkhead Box O1/genética , Fator de Crescimento Insulin-Like I/metabolismo , Neurônios/citologia , Receptores Acoplados a Proteínas G/genética , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/farmacologia , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Cromonas/farmacologia , Regulação para Baixo/efeitos dos fármacos , Proteína Forkhead Box O1/metabolismo , Técnicas de Inativação de Genes , Humanos , Lentivirus/fisiologia , Morfolinas/farmacologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação/efeitos dos fármacos , Regiões Promotoras Genéticas/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Quinolonas/farmacologia , RNA Interferente Pequeno/farmacologia , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais/efeitos dos fármacos
16.
BMC Plant Biol ; 21(1): 542, 2021 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-34800993

RESUMO

BACKGROUND: Rice is a crop that is very sensitive to low temperature, and its morphological development and production are greatly affected by low temperature. Therefore, understanding the genetic basis of cold tolerance in rice is of great significance for mining favorable genes and cultivating excellent rice varieties. However, there have been limited studies focusing on cold tolerance at the bud burst stage; therefore, considerable attention should be given to the genetic basis of cold tolerance at this stage. RESULTS: In this study, a natural population consisting of 211 rice landraces collected from 15 provinces in China and other countries was used for the first time to evaluate cold tolerance at the bud burst stage. Population structure analysis showed that this population was divided into two groups and was rich in genetic diversity. Our evaluation results confirmed that japonica rice was more tolerant to cold at the bud burst stage than indica rice. A genome-wide association study (GWAS) was performed with the phenotypic data of 211 rice landraces and a 36,727 SNP dataset under a mixed linear model. Twelve QTLs (P < 0.0001) were identified for the seedling survival rate (SR) after treatment at 4 °C, in which there were five QTLs (qSR2-2, qSR3-1, qSR3-2, qSR3-3 and qSR9) that were colocalized with those from previous studies and seven QTLs (qSR2-1, qSR3-4, qSR3-5, qSR3-6, qSR3-7, qSR4 and qSR7) that were reported for the first time. Among these QTLs, qSR9, harboring the most significant SNP, explained the most phenotypic variation. Through bioinformatics analysis, five genes (LOC_Os09g12440, LOC_Os09g12470, LOC_Os09g12520, LOC_Os09g12580 and LOC_Os09g12720) were identified as candidates for qSR9. CONCLUSION: This natural population consisting of 211 rice landraces combined with high-density SNPs will serve as a better choice for identifying rice QTLs/genes in the future, and the detected QTLs associated with cold tolerance at the bud burst stage in rice will be conducive to further mining favorable genes and breeding rice varieties under cold stress.


Assuntos
Temperatura Baixa , Resposta ao Choque Frio/genética , Flores/crescimento & desenvolvimento , Flores/genética , Oryza/crescimento & desenvolvimento , Oryza/genética , Locos de Características Quantitativas/genética , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Variação Genética , Estudo de Associação Genômica Ampla , Genótipo
17.
Arch Biochem Biophys ; 700: 108774, 2021 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-33548212

RESUMO

Homoharringtonine (HHT), an approved anti-leukemic alkaloid, has been reported effectively in many types of tumor cells. However, its effect on melanoma cells has not been investigated. And the anti-melanoma mechanism of HHT is still unknown. In this study, we detected the effects of HHT on two melanoma cell lines (A375 and B16F10) and on the A375 xenograft mouse model. HHT significantly inhibited the proliferation of melanoma cells as investigated by the CCK8 method, cell cloning assay, and EdU experiment. HHT induced A375 and B16F10 cells DNA damage, apoptosis, and G2/M cell cycle arrest as proved by TdT-mediated dUTP Nick-End Labeling (TUNEL) and flow cytometry assay. Additionally, the loss of mitochondrial membrane potential in HHT-treated cells were visualized by JC-1 fluorescent staining. For the molecule mechanism study, western blotting results indicated the protein expression levels of ATM, P53, p-P53, p-CHK2, γ-H2AX, PARP, cleaved-PARP, cleaved caspase-3, cleaved caspase-9, Bcl-2, Bax, Aurka, p-Aurka, Plk1, p-Plk1, Cdc25c, CDK1, cyclin B1, and Myt1 were regulated by HHT. And the relative mRNA expression level of Aurka, Plk1, Cdc25c, CDK1, cyclin B1, and Myt1 were ascertained by q-PCR assay. The results in vivo experiment showed that HHT can slow down the growth rate of tumors. At the same time, the protein expression levels in vivo were consistent with that in vitro. Collectively, our study provided evidence that HHT could be considered an effective anti-melanoma agent by inducing DNA damage, apoptosis, and cell cycle arrest.


Assuntos
Dano ao DNA/efeitos dos fármacos , DNA de Neoplasias/metabolismo , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Mepesuccinato de Omacetaxina/farmacologia , Pontos de Checagem da Fase M do Ciclo Celular/efeitos dos fármacos , Melanoma Experimental , Animais , Apoptose , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Proteínas de Neoplasias/biossíntese
18.
Anticancer Drugs ; 32(3): 314-322, 2021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33394687

RESUMO

Evodiamine (Evo), a quinazoline alkaloid and one of the most typical polycyclic heterocycles, is mainly isolated from Evodia rugulosa. Vasculogenic mimicry (VM) is a newly identified way of angiogenesis during tumor neovascularization, which is prevalent in a variety of highly invasive tumors. The purpose of this study was to investigate the effect and mechanism of Evo on VM in human colorectal cancer (CRC) cells. The number of VM structures was calculated by the three-dimensional culture of human CRC cells. Wound-healing was used to detect the migration of HCT116 cells. Gene expression was detected by reverse transcription-quantitative PCR assay. CD31/PAS staining was used to identify VM. Western blotting and immunofluorescence were used to detect protein levels. The results showed that Evo inhibited the migration of HCT116 cells, as well as the formation of VM. Furthermore, Evo reduced the expression of hypoxia-inducible factor 1-alpha (HIF-1α), VE-cadherin, VEGF, MMP2, and MMP9. In a model of subcutaneous xenotransplantation, Evo also inhibited tumor growth and VM formation. Our study demonstrates that Evo could inhibit VM in CRC cells HCT116 and reduce the expression of HIF-1α, VE-cadherin, VEGF, MMP2, and MMP9.


Assuntos
Neovascularização Patológica/tratamento farmacológico , Quinazolinas/farmacologia , Animais , Antígenos CD/efeitos dos fármacos , Caderinas/efeitos dos fármacos , Movimento Celular , Sobrevivência Celular , Transição Epitelial-Mesenquimal , Feminino , Células HCT116 , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/efeitos dos fármacos , Camundongos Endogâmicos BALB C , Neovascularização Patológica/patologia , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacos , Cicatrização/efeitos dos fármacos
19.
BMC Urol ; 21(1): 149, 2021 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-34736451

RESUMO

BACKGROUND: To explore whether opening the external urethral orifice in the coronal sulcus can reduce the incidence of epididymitis after operating on hypospadias with prostatic utricle cyst (PUC) connecting to the vas deferens. Group A consisted of 3 patients with severe hypospadias and PUC undergoing cystostomy, hypospadias correction and urethroplasty, along with the relocation of the external orifice of the urethra to the coronal sulcus. Group B consisted of 4 patients having initial hypospadias repaired with meatus in the orthotopic position in the glans, presenting with multiple epididymitis after hypospadias surgery and unsuccessful conservative treatment. MR confirmed that all the Group B patients had PUC connecting to the vas deferens. Group B patients underwent urethral dilatation along with urethral catheterization, cutting of the original corpus cavernosum that encapsulated the urethra, and extension of the position of the external urethral orifice to the coronal sulcus. RESULTS: In group A, 3 children underwent bladder fistula removal 2 weeks after the operation. The penis developed normally without any complications. Four children in group B underwent stent removal 12 weeks after operation, and one patient was still stenosed and dilated again. All patients in group B were followed without epididymitis recurrence. CONCLUSIONS: For patients with hypospadias complicating with a PUC, connecting to one side of the vas deferens, the positioning of the external urethral orifice in the coronary sulcus would be helpful to reduce the occurrence of epididymitis.


Assuntos
Cistos/cirurgia , Hipospadia/cirurgia , Procedimentos de Cirurgia Plástica/métodos , Doenças Prostáticas/cirurgia , Procedimentos Cirúrgicos Urológicos Masculinos/métodos , Cateterismo , Pré-Escolar , Cistostomia , Cistos/complicações , Cistos/diagnóstico por imagem , Dilatação , Epididimite/etiologia , Epididimite/prevenção & controle , Humanos , Hipospadia/complicações , Hipospadia/diagnóstico por imagem , Masculino , Complicações Pós-Operatórias , Doenças Prostáticas/complicações , Doenças Prostáticas/diagnóstico por imagem , Procedimentos de Cirurgia Plástica/efeitos adversos , Stents , Procedimentos Cirúrgicos Urológicos Masculinos/efeitos adversos
20.
Metab Brain Dis ; 36(7): 1889-1901, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34417941

RESUMO

Nowadays, similar strategies have been used for the treatment and prevention of acute stroke in both diabetes mellitus (DM) and non-DM populations. These strategies were analyzed to provide an experimental basis for the clinical prevention and treatment of stroke in patients both with and without DM. Tree shrews were randomly divided into control, DM, ischemic stroke (IS), and DMIS groups with 18 animals in each group. Serum biochemical indicators were used to assess metabolic status. Neural tissue damage was determined using triphenyl tetrazolium chloride staining, H-E staining, and electron microscopy. Differential gene expression of neural tissue between the DM and control groups and the IS and DMIS groups was measured using RNA-seq analysis. The serum glucose levels of the DM and DMIS groups were significantly higher than other groups. In the DMIS group, the infarct size was significantly larger than in the IS group (19.56 ± 1.25%), with a more obvious abnormal ultrastructure of neural cells. RNA-seq analysis showed that the expression of IL-8, C-C motif chemokine 2 (CCL2), and alpha-1-antichymotrypsin was significantly higher in the DM group than in the control group. The CCL7, ATP-binding cassette sub-family A member 12, and adhesion G protein-coupled receptor E2 levels were significantly higher in the DMIS group than in the IS group. For the prevention and treatment of stroke in patients with DM, reducing the inflammatory state of the nervous system may reduce the incidence of stroke and improve the prognosis of neurological function after IS.


Assuntos
Isquemia Encefálica , Diabetes Mellitus Tipo 2 , Diabetes Mellitus , AVC Isquêmico , Acidente Vascular Cerebral , Animais , Encéfalo/metabolismo , Isquemia Encefálica/genética , Isquemia Encefálica/metabolismo , Diabetes Mellitus Tipo 2/genética , Isquemia , AVC Isquêmico/genética , Análise de Sequência de RNA , Acidente Vascular Cerebral/genética , Acidente Vascular Cerebral/terapia , Tupaia/genética , Tupaiidae/genética
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