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2.
Life Sci ; 271: 119173, 2021 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-33556375

RESUMO

AIMS: Cell cycle arrest plays critical roles in preventing renal tubular epithelial cell (RTEC) injury and maladaptation after the onset of chronic kidney disease (CKD), but the underlying mechanism governing this arrest has not been fully elucidated. This study was designed to determine the underlying role of YB-1 in promoting cell cycle progression and nuclear translocation in HK-2 cells induced by trimethylamine N-oxide (TMAO). MAIN METHODS: YB-1 primarily accumulated in the cytoplasm in HK-2 cells after they were treated with TMAO for 30 min and 6 h. Gene expression was analysed using RNA sequencing in HK-2 cells treated with TMAO. Cell cycle progression was analysed via flow cytometry. Luciferase assay and ChIP-PCR were performed to determine the relationship between transcription factor YB-1 and Gadd45a promoter region. Additionally, mice were fed with TMAO to test renal dysfunction and measure the expression of YB-1, GADD45a and CCNA2 in the kidney sections through immunohistochemistry. KEY FINDINGS: YB-1 primarily accumulated in the cytoplasm in HK-2 cells after they were treated with TMAO for 30 min and 6 h. RNA sequencing analysis showed that the cell cycle checkpoint genes growth arrest and DNA damage (Gadd)45a, Gadd45g, cyclin (Ccn)a2, Ccnb1, Ccne1 and Ccnf were differentially expressed in HK-2 cells after treated with 400 µM TMAO for 30 min. Flow cytometry results demonstrated that cell cycle progression was blocked at the G2/M checkpoint. In animal models, elevated dietary TMAO directly led to progressive renal tubulointerstitial dysfunction and inhibited the expression of YB-1 in kidney. Moreover, YB-1 was determined to regulate Gadd45a expression by directly binding to its promoter region. YB-1 expression was negatively correlated with the expression of Gadd45a and Gadd45g but positively correlated with Ccna2, Ccnb1, Ccne1 and Ccnf in CKD. SIGNIFICANCE: YB-1 may be a reliable molecular target and an effective prognostic biomarker for CKD.


Assuntos
Proteínas de Ciclo Celular/biossíntese , Ciclo Celular/fisiologia , Regulação para Baixo/fisiologia , Metilaminas/toxicidade , Insuficiência Renal Crônica/metabolismo , Proteína 1 de Ligação a Y-Box/metabolismo , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Transporte Ativo do Núcleo Celular/fisiologia , Animais , Ciclo Celular/efeitos dos fármacos , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Expressão Gênica , Humanos , Túbulos Renais/citologia , Túbulos Renais/efeitos dos fármacos , Túbulos Renais/metabolismo , Camundongos Endogâmicos C57BL , Insuficiência Renal Crônica/induzido quimicamente , Insuficiência Renal Crônica/genética
3.
Mol Immunol ; 132: 165-171, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33592572

RESUMO

The therapeutic options of non-small cell lung cancer (NSCLC) are limited, although a combination of targeted therapy and immunotherapy is promising. To explore novel targets for immunotherapy, we explored the role of Ccr4-Not transcription complex subunit 4 (CNOT4) in NSCLC. The expression of CNOT4 in tumor tissues was determined by immunohistochemistry staining and western blotting. The cell lines that stably express CNOT4 were established in H1299 and A549 cells. Direct cell counting, MTT assay, and colony formation were used to determine the ability of cell proliferation. Cell apoptosis and cell cycle were next analyzed by PI/Annexin V staining. Cell invasion and migration were examined by transwell assays. To further explore the function of CNOT4 in cytotoxic T lymphocytes (CTLs) mediated cytotoxicity, an in vitro co-culture system of CNOT4 overexpressing and control H1299 cells with CTLs was developed. CNOT4 was down-regulated in tumor tissues compared with paired normal tissues from patients with lung cancers. CNOT4 overexpression significantly inhibited tumor cell proliferation, colony formation, cell migration, and invasion, but promoted cell apoptosis. Furthermore, overexpression of CNOT4 enhanced cytotoxicity of CTLs to H1299. CNOT4 functions as a potential tumor suppressor of NSCLC via inhibiting tumor cell function and increasing the sensitivity to CTLs.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Neoplasias Pulmonares/metabolismo , Linfócitos T Citotóxicos/metabolismo , Fatores de Transcrição/metabolismo , Células A549 , Apoptose/fisiologia , Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Regulação para Baixo/fisiologia , Humanos , MicroRNAs/metabolismo
4.
Cancer Sci ; 112(4): 1624-1632, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33540491

RESUMO

Lysophosphatidic acid receptor 5 (LPAR5) is involved in mediating thyroid cancer progression, but the underlying mechanism needs to be further revealed. In this study, we confirmed that LPAR5 is upregulated in papillary thyroid carcinoma (PTC), especially in BRAF-like PTC, by analyzing The Cancer Genome Atlas (TCGA) database and performing immunohistochemistry assay in human thyroid cancer tissues. LPAR5-specific antagonist TC LPA5 4 treatment inhibited CGTH-W3, TPC-1, B-CPAP, and BHT-101 cell proliferation, CGTH-W3 and TPC-1 cell migration significantly. In vivo, TC LPA5 4 treatment could delay CGTH-W3 xenograft growth in nude mice. We also found that LPAR5-specific antagonist TC LPA5 4, PI3K inhibitor wortmannin, or mTOR inhibitor rapamycin pretreatment abrogated phosphorylation of Akt and p70S6K1 stimulated by LPA in CGTH-W3 and TPC-1 cells. Stimulating CGTH-W3 cells transfected with pEGFPC1-Grp1-PH fusion protein with LPA resulted in the generation of phosphatidylinositol (3,4,5)-triphosphate, which indicates that PI3K was activated by LPA directly. The p110ß-siRNA instead of p110α-siRNA transfection abrogated the increase of levels of phosphorylated Akt and S6K1 stimulated by LPA. Furthermore, immunoprecipitation assay confirmed an interaction between LPAR5 and p110ß. Overall, we provide new insights that the downregulation of LPAR5 decreased the proliferation and migration phenotype via the PI3K/Akt pathway. Inhibition of LPAR5 or the PI3K/Akt signal may be a novel therapeutic strategy for treating thyroid cancer.


Assuntos
Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Classe Ia de Fosfatidilinositol 3-Quinase/metabolismo , Receptores de Ácidos Lisofosfatídicos/metabolismo , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/patologia , Animais , Domínio Catalítico/fisiologia , Linhagem Celular Tumoral , Regulação para Baixo/fisiologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Transdução de Sinais/fisiologia , Câncer Papilífero da Tireoide/metabolismo , Câncer Papilífero da Tireoide/patologia
5.
J Neurosci ; 41(6): 1331-1339, 2021 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-33443069

RESUMO

The dorsolateral prefrontal cortex (DLPFC) and ventrolateral PFC (VLPFC) are both crucial structures involved in voluntary emotional regulation. However, it remains unclear whether the functions of these two cortical regions that are involved in emotional regulation, which are usually active in non-social situations, could be generalized to the regulation of social pain as well. This study employed transcranial magnetic stimulation (TMS) to examine the causal relationship between the DLPFC/VLPFC and the emotional regulation of social pain via distraction and reappraisal. Ninety human participants (45 males and 45 females) initially underwent either active (DLPFC/VLPFC, n = 30/30) or sham (vertex, n = 30) TMS sessions. Participants were then instructed to use both distraction and reappraisal strategies to downregulate any negative emotions evoked by social exclusion pictures. Convergent results of the subjective emotional rating and electrophysiological indices demonstrated that: (1) both the DLPFC and VLPFC highly facilitate the downregulation of affective responses caused by social exclusion, revealing a causal role of these lateral PFCs in voluntary emotional regulation of both non-social and social pain; and (2) these two cortical regions showed relative functional specificity for distraction (DLPFC) and reappraisal (VLPFC) strategies, which helps to refine the cortical targeting of therapeutic protocols. In addition, the TMS effect was sustainable for at least 1 h, showcasing the potential feasibility of using this method in clinical practice. Together, these findings provide cognitive and neural evidence for the targeting of the VLPFC and/or the DLPFC to improve emotional regulation abilities, especially in social contexts.SIGNIFICANCE STATEMENT This study aimed to examine the role of the dorsolateral prefrontal cortex (DLPFC) and ventrolateral PFC (VLPFC) in emotional regulation, particularly in response to social pain through the use of distraction and reappraisal strategies, as this is a relatively underexplored area of inquiry. This study makes a significant contribution to the literature because our results provide novel empirical information on the role of these cortical structures in the processing of negative emotions elicited within certain social contexts. As such, our findings have potential clinical implications, paving the way for future clinicians to be able to accurately target specific brain regions among patients struggling with impaired social cognition abilities, including those diagnosed with posttraumatic stress disorder, autism spectrum disorder, social anxiety disorder, and depression.


Assuntos
Regulação para Baixo/fisiologia , Regulação Emocional/fisiologia , Estimulação Luminosa/métodos , Córtex Pré-Frontal/fisiologia , Isolamento Social/psicologia , Estimulação Magnética Transcraniana/métodos , Adolescente , Eletroencefalografia/métodos , Emoções/fisiologia , Feminino , Humanos , Masculino , Adulto Jovem
6.
Neurosci Lett ; 743: 135552, 2021 01 19.
Artigo em Inglês | MEDLINE | ID: mdl-33352285

RESUMO

Central sensitization is one of the important pathological mechanisms of chronic migraine (CM). Metabolic glutamate receptor 5 (mGluR5) mediates pain by activating various intracellular pathways. However, whether mGluR5 contributes to central sensitization in CM and the exact mechanism remains unclear. Male rats were used to establish a CM model by repeated infusions of inflammatory soup (IS) for 7 days to stimulate the activation of the dural nociceptor. The mechanical and thermal thresholds were used to evaluate allodynia, and central sensitization was assessed by measuring calcitonin gene-related peptide (CGRP) and substance P (SP). Microtubule associated protein 1 light chain 3 (LC3) and p62/SQSTM1 were used to assess autophagy. We found that the expression of mGluR5 in the trigeminal nucleus caudalis (TNC) of CM rats was significantly increased. In addition, the downregulation of mGluR5 activated autophagy by inhibiting the mTOR pathway. Moreover, the activation of autophagy alleviated allodynia and central sensitization in CM rats. This study identified a novel strategy for the treatment of CM; the downregulation of mGluR5 in a rat model of CM decreased the expression of the inflammatory factor interleukin-1 beta (IL-1ß) and the central sensitization-associated proteins CGRP and SP by activating autophagy via inhibiting the mTOR pathway.


Assuntos
Autofagia/fisiologia , Sensibilização do Sistema Nervoso Central/fisiologia , Regulação para Baixo/fisiologia , Transtornos de Enxaqueca/metabolismo , Receptor de Glutamato Metabotrópico 5/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Animais , Autofagia/efeitos dos fármacos , Sensibilização do Sistema Nervoso Central/efeitos dos fármacos , Doença Crônica , Regulação para Baixo/efeitos dos fármacos , Mediadores da Inflamação/metabolismo , Mediadores da Inflamação/toxicidade , Masculino , Transtornos de Enxaqueca/induzido quimicamente , Transtornos de Enxaqueca/patologia , Ratos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Serina-Treonina Quinases TOR/antagonistas & inibidores
7.
Chem Biol Interact ; 334: 109354, 2021 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-33309620

RESUMO

Lactosyl-Sepharose binding proteins (LSBPs) were recently described in human pancreatic ductal adenocarcinoma (PDAC) Suit2-007 cells regarding their lectin-like properties and role in metastasis. This study further investigated how calcium and galactose influence the binding of LSBPs to the lactosyl resin as well as their anti-proliferative effect in Suit2-007 cells. Altered binding of LSBPs to the lactosyl resin was evaluated by affinity chromatography and mass spectrometry. Calcium binding EF-hand proteins were aligned and identified with a motif derived from the Uniprot protein database. The antiproliferative effects of LSBPs and monosaccharides were determined by MTT assay. In addition, LSBPs and galactose effects were investigated by chip array and tumor take in nude rats. LSBPs reduced Suit2-007 cells' proliferation with an IC50 of 125 µg/mL. Coincubation of LSBPs with EGTA decreased the number of LSBPs binding to the lactosyl resin by ~50%. Ca2+ -sensitive LSBPs included subgroups of galactose-sensitive (10%) and EF-hand calcium binding motifs containing (2.5%) proteins. In vitro, the combination of LSBPs with monosaccharides including galactose synergistically decreased cell proliferation compared to single agents (p < 0.05). In addition, LSBPs in combination with galactose prevented the tumor growth of Suit2-007 cells in nude rats, as opposed to single treatments. At mRNA level, the combination treatment modulated 5% of Ca2+ -sensitive LSBPs and downregulated 216 genes, 18% of which were up-regulated during PDAC progression. This study highlights the importance of calcium and galactose in modulating the affinity and anti-proliferative activity of LSBPs and their potential application as therapeutic agents for metastatic PDAC.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Cálcio/metabolismo , Proliferação de Células/fisiologia , Galactose/metabolismo , Ligação Proteica/fisiologia , Sefarose/metabolismo , Animais , Linhagem Celular Tumoral , Regulação para Baixo/fisiologia , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Lectinas/metabolismo , Masculino , RNA Mensageiro/metabolismo , Ratos , Ratos Nus , Regulação para Cima/fisiologia
8.
Life Sci ; 258: 118222, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32768577

RESUMO

AIMS: We previously reported that fenugreek-derived 4-hydroxyisoleucine ameliorates insulin resistance via regulation of TNF-α converting enzyme (TACE) expression. In the present study, we further investigate the effects and mechanisms of fenugreek on obesity-induced inflammation and insulin signaling in the high-fat diet (HFD)-challenged obese mice. MAIN METHODS: After 12 weeks of HFD intervention, mice were treated with the low or high dosages of fenugreek. Serum levels of glucose, insulin, lipid profile, inflammation cytokines, and adipokines were detected. Macrophage infiltration and adipose tissue morphology were observed. Western blot was conducted to investigate the expressions of inactive rhomboid 2 (iRhom2) and TACE as well as other signaling pathways in subcutaneous adipose tissue. KEY FINDINGS: We showed that fenugreek significantly suppressed body weight gain and fat accumulation in HFD-challenged obese mice. Meanwhile, fasting glucose, insulin, and HOMA-IR in fenugreek-treated mice were remarkably decreased, which were properly explained by fenugreek-induced activation of the insulin receptor signaling pathway. Moreover, the anti-inflammatory properties of fenugreek were shown by the decrease of systemic and local expressions of pro-inflammatory cytokines as well as reduced macrophage infiltration into adipose tissue. Additionally, fenugreek markedly deactivated NF-κB and JNK pathways. Finally, we demonstrated that fenugreek strikingly repressed the transcriptions and expressions of iRhom2 and TACE. SIGNIFICANCE: Fenugreek shows an encouraging and promising property in ameliorating insulin resistance and suppressing inflammation in obesity, which might be realized by fenugreek-mediated inhibition of iRhom2/TACE axis-facilitated TNF-α release from adipocytes.


Assuntos
Proteína ADAM17/antagonistas & inibidores , Proteínas de Transporte/antagonistas & inibidores , Mediadores da Inflamação/antagonistas & inibidores , Resistência à Insulina/fisiologia , Obesidade/tratamento farmacológico , Trigonella , Proteína ADAM17/sangue , Animais , Proteínas de Transporte/sangue , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Inflamação/sangue , Inflamação/tratamento farmacológico , Mediadores da Inflamação/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/sangue , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Sementes
9.
Life Sci ; 258: 118223, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32768584

RESUMO

Kidney fibrosis is a common final pathway of chronic kidney diseases, which are characterized by renal architecture damage, inflammation, fibroblast expansion and myofibroblast formation. Endothelin converting enzyme-1 (ECE-1) contributes to activation of Endothelin-1 (ET-1), a potent vasoconstrictor and pro-fibrotic substance. This study elucidated the effect of ECE-1 knockout in kidney fibrosis model in mice in association of ET-1 downregulation. Kidney fibrosis was performed in ECE-1 knockout (ECE-1 KO) and vascular endothelial derived ET-1 KO (VEETKO) mice (2 months, 20-30 g, n = 30) and their wild type (WT) littermates using unilateral ureteral obstruction (UUO) procedure. Mice were euthanized on day-7 and day-14 after UUO. Histopathological analysis was conducted for fibrosis and tubular injury. Immunostainings were done to quantify macrophages (F4/80), fibroblasts (FSP-1) and myofibroblasts (α-SMA). Monocyte Chemoattractant Protein-1 (MCP-1), ECE-1 and preproET-1 (ppET-1) mRNA expression were quantified with qRT-PCR, while Transforming Growth Factor-ß1 (TGF-ß1) and α-SMA protein level were quantified with Western blot. ECE-1 KO mice demonstrated reduction of ECE-1 and ppET-1 mRNA expression, attenuation of kidney fibrosis, tubular injury, MCP-1 mRNA expression and macrophage number compared to WT. Double immunostaining revealed fibroblast to myofibroblast formation after UUO, while ECE-1 KO mice had significantly lower fibroblast number and myofibroblast formation compared to WT, which were associated with significantly lower TGF-ß1 and α-SMA protein levels in day-14 of UUO. VEETKO mice also demonstrated attenuation of ET-1 protein level, fibrosis and myofibroblast formation. In conclusion, ECE-1 knockout and ET-1 downregulation attenuated kidney fibrosis.


Assuntos
Regulação para Baixo/fisiologia , Endotelina-1/metabolismo , Enzimas Conversoras de Endotelina/deficiência , Rim/metabolismo , Animais , Enzimas Conversoras de Endotelina/genética , Fibrose , Rim/patologia , Masculino , Camundongos , Camundongos Knockout
10.
Nat Cell Biol ; 22(9): 1103-1115, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32839548

RESUMO

Plasticity of cancer invasion and metastasis depends on the ability of cancer cells to switch between collective and single-cell dissemination, controlled by cadherin-mediated cell-cell junctions. In clinical samples, E-cadherin-expressing and -deficient tumours both invade collectively and metastasize equally, implicating additional mechanisms controlling cell-cell cooperation and individualization. Here, using spatially defined organotypic culture, intravital microscopy of mammary tumours in mice and in silico modelling, we identify cell density regulation by three-dimensional tissue boundaries to physically control collective movement irrespective of the composition and stability of cell-cell junctions. Deregulation of adherens junctions by downregulation of E-cadherin and p120-catenin resulted in a transition from coordinated to uncoordinated collective movement along extracellular boundaries, whereas single-cell escape depended on locally free tissue space. These results indicate that cadherins and extracellular matrix confinement cooperate to determine unjamming transitions and stepwise epithelial fluidization towards, ultimately, cell individualization.


Assuntos
Neoplasias da Mama/patologia , Adesão Celular/fisiologia , Invasividade Neoplásica/patologia , Junções Aderentes/patologia , Animais , Linhagem Celular , Linhagem Celular Tumoral , Regulação para Baixo/fisiologia , Feminino , Regulação Neoplásica da Expressão Gênica/fisiologia , Células HEK293 , Humanos , Junções Intercelulares/patologia , Células MCF-7 , Camundongos Endogâmicos BALB C
11.
Anticancer Res ; 40(7): 3697-3705, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32620608

RESUMO

BACKGROUND/AIM: Time restricted feeding (TRF) mitigates the high-fat diet-enhanced mammary tumorigenesis in a MMTV-PyMT breast cancer model. MATERIALS AND METHODS: We performed untargeted metabolomic and targeted transcriptomic analyses on mammary tumors from MMTV-PyMT mice fed a standard AIN93G diet, a high-fat diet (HFD), or HFD with TRF (12 h, dark phase) and mammary glands from wild-type mice fed the AIN93G diet. RESULTS: The metabolic profile of mammary tumors differed from that of mammary glands; there was no impact of TRF upon tumor metabolome. TRF did reduce elevated expression of Hmgcr, Srebp1, Fads2, and Ppard in mammary tumors, indicating a down-regulation of lipid metabolism. CONCLUSION: The null effect of TRF on the metabolomic profile does not rule out changes in more refined intracellular signaling pathways. It suggests that the protection of TRF against mammary tumorigenesis may rely upon its action on the host rather than a direct effect on tumor metabolism.


Assuntos
Glândulas Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Experimentais/metabolismo , Metaboloma/fisiologia , Obesidade/metabolismo , Animais , Carcinogênese/metabolismo , Dieta Hiperlipídica/métodos , Modelos Animais de Doenças , Regulação para Baixo/fisiologia , Feminino , Metabolismo dos Lipídeos/fisiologia , Masculino , Camundongos
12.
Anticancer Res ; 40(7): 3751-3757, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32620614

RESUMO

BACKGROUND/AIM: Colorectal cancer is frequently associated with metabolic diseases. Adiponectin (APN) is an insulin-sensitizing adipokine circulating as low molecular weight (LMW), medium molecular weight (MMW) and high molecular weight (HMW) oligomers; the latter are the most bio-active oligomers. APN, through AdipoR1, AdipoR2 and T-cadherin receptors, regulates inflammation, and proliferation. Considering the anti-proliferative and anti-inflammatory properties of APN, we investigated the involvement of the "APN system" in colorectal cancer. MATERIALS AND METHODS: A total of 44 colorectal cancer patients and 51 healthy controls were recruited. We analysed APN and HMW oligomers in sera, AdipoR1, AdipoR2 and T-cadherin expression in non-cancerous and cancerous colon tissues. RESULTS: we found statistically lower levels of APN in patients compared to controls, with a specific decrease of HMW oligomers. Importantly, APN correlated to cancer grade. AdipoR1 was found overexpressed in cancerous compared to non-cancerous tissues while AdipoR2 and T-cadherin were down-regulated. CONCLUSION: The deregulated expression of the "APN system" in colorectal cancer with a specific correlation to tumor grade suggests APN as a promising biomarker in colorectal cancer.


Assuntos
Adiponectina/metabolismo , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/metabolismo , Caderinas/metabolismo , Estudos Transversais , Regulação para Baixo/fisiologia , Feminino , Humanos , Inflamação/metabolismo , Inflamação/patologia , Masculino , Pessoa de Meia-Idade , Gradação de Tumores/métodos
13.
Toxicol Appl Pharmacol ; 401: 115080, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32497533

RESUMO

Upregulation of ABCB1/MDR1 (P-gp) and BIRC5/Survivin promotes multidrug resistance in a variety of human cancers. LCL161 is an anti-cancer DIABLO/SMAC mimetic currently being tested in patients with solid tumors, but the molecular mechanism of action of LCL161 in cancer cells is still incompletely understood. It is still unclear whether LCL161 is therapeutically applicable for patients with ABCB1-overexpressing multidrug resistant tumors. In this study, we found that the potency of LCL161 is not affected by the expression of ABCB1 in KB-TAX50, KB-VIN10, and NTU0.017 cancer cells. Besides, LCL161 is equally potent towards the parental MCF7 breast cancer cells and its BIRC5 overexpressing, hormone therapy resistance subline MCF7-TamC3 in vitro. Mechanistically, we found that LCL161 directly modulates the ABCB1-ATPase activity and inhibits ABCB1 multi-drug efflux activity at low cytotoxic concentrations (i.e. 0.5xIC50 or less). Further analysis revealed that LCL161 also decreases intracellular ATP levels in part through BIRC5 downregulation. Therapeutically, co-treatment with LCL161 at low cytotoxic concentrations restored the sensitivity to the known ABCB1 substrate, paclitaxel, in ABCB1-expressing cancer cells and increased the sensitivity to tamoxifen in MCF7-TamC3 cells. In conclusion, LCL161 has the potential for use in the management of cancer patients with ABCB1 and BIRC5-related drug resistance. The findings of our study provide important information to physicians for designing a more "patient-specific" LCL161 clinical trial program in the future.


Assuntos
Adenosina Trifosfatases/antagonistas & inibidores , Antineoplásicos/farmacologia , Proteínas Reguladoras de Apoptose/farmacologia , Proteínas Mitocondriais/farmacologia , Survivina/antagonistas & inibidores , Tiazóis/farmacologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/antagonistas & inibidores , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Adenosina Trifosfatases/metabolismo , Antineoplásicos/química , Proteínas Reguladoras de Apoptose/química , Materiais Biomiméticos/química , Materiais Biomiméticos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Proteínas Mitocondriais/química , Estrutura Secundária de Proteína , Survivina/biossíntese , Survivina/genética , Tiazóis/química
14.
Metabolism ; 108: 154258, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32376130

RESUMO

RATIONALE: Tubulointerstitial fibrosis, which is closely related to functional injury of the kidney, can be observed in advanced stages of diabetic nephropathy (DN). Mammalian serine/threonine-protein kinase 4 (MST1), a core component of the Hippo pathway that is involved in cellular proliferation and differentiation, plays a crucial role in the pathogenesis of multiple metabolic diseases, kidney diseases and cancer. METHODS: In type 1 and type 2 diabetic animals, as well as in human proximal tubular epithelial cells (HK-2), activation of MST1 was analyzed by immunohistochemistry and western blotting. In db/db mice, MST1 protein was knocked down or overexpressed by shRNA, and renal function, fibrosis, and downstream signaling were then investigated. RNA silencing and overexpression were performed by using an MST1 or YAP knockdown/expression lentivirus to investigate the regulation of MST1-mediated YAP/TEAD signaling pathways in the fibrosis process in HK-2 cells. Luciferase and coimmunoprecipitation (co-IP) assays were used to identify whether YAP directly regulated TEAD activation by forming a YAP-TEAD heterodimer, which ultimately leads to tubulointerstitial fibrosis. RESULTS: MST1 activation was significantly decreased in type 1 and type 2 diabetic nephropathy. Notably, the downregulation of MST1 activation was also observed in HK-2 cells in a glucose- and time-dependent manner. In vivo, downregulation of MST1 was sufficient to promote renal dysfunction and fibrosis in db/m mice, whereas overexpression of MST1 ameliorated diabetic nephropathy-induced renal fibrosis. Further mechanistic study demonstrated that activated YAP induced by MST1 inhibition directly upregulated TEAD activation by binding to TEAD and forming a YAP-TEAD heterodimer, resulting in the promotion of epithelial-mesenchymal transition (EMT) and fibrosis in renal tubular epithelial. CONCLUSIONS: MST1 activation represents a potential therapeutic strategy to treat or prevent the progression of diabetic nephropathy-induced renal fibrosis.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/metabolismo , Diabetes Mellitus/metabolismo , Nefropatias Diabéticas/metabolismo , Transição Epitelial-Mesenquimal/fisiologia , Fibrose/metabolismo , Nefropatias/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Regulação para Baixo/fisiologia , Regulação da Expressão Gênica/fisiologia , Glucose/metabolismo , Túbulos Renais/metabolismo , Camundongos , Ratos , Transdução de Sinais/fisiologia , Fatores de Transcrição/metabolismo , Regulação para Cima/fisiologia
15.
Medicine (Baltimore) ; 99(19): e20183, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32384511

RESUMO

BACKGROUNDS: Lung adenocarcinoma (LUAD) is one of the most common malignancies, and is a serious threat to human health. The aim of the present study was to assess potential biomarkers for the prognosis of LUAD through the analysis of gene expression microarrays. METHODS: The gene expression data for GSE118370 was downloaded from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) between normal lung and LUAD samples were screened using the R language. The DAVID database was used to analyze the functions and pathways of DEGs. The STRING database was used to the map protein-protein interaction (PPI) networks, and these were visualized with the Cytoscape software. Finally, the prognostic analysis of the hub gene in the PPI network was performed using the Kaplan-Meier tool. RESULTS: A total of 406 downregulated and 203 upregulated DEGs were identified. The GO analysis results revealed that downregulated DEGs were significantly enriched in angiogenesis, calcium ion binding and cell adhesion. The upregulated DEGs were significantly enriched in the extracellular matrix disassembly, collagen catabolic process, chemokine-mediated signaling pathway and endopeptidase inhibitor activity. The KEGG pathway analysis revealed that downregulated DEGs were enriched in neuroactive ligand-receptor interaction, hematopoietic cell lineage and vascular smooth muscle contraction, while upregulated DEGs were enriched in phototransduction. In addition, the top 10 hub genes and the most closely interacting modules of the top 3 proteins in the PPI network were screened. Finally, the independent prognostic value of each hub gene in LUAD patients was analyzed through the Kaplan-Meier plotter. Seven hub genes (ADCY4, S1PR1, FPR2, PPBP, NMU, PF4, and GCG) were closely correlated to overall survival time. CONCLUSION: The discovery of these candidate genes and pathways reveals the etiology and molecular mechanisms of LUAD, providing ideas and guidance for the development of new therapeutic approaches to LUAD.


Assuntos
Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/mortalidade , Bases de Dados Genéticas , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidade , Biomarcadores Tumorais , Quimiocinas/metabolismo , Colágeno/metabolismo , Regulação para Baixo/fisiologia , Matriz Extracelular/metabolismo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Ontologia Genética , Humanos , Estimativa de Kaplan-Meier , Análise em Microsséries , Prognóstico , Inibidores de Proteases/metabolismo , Mapas de Interação de Proteínas , Regulação para Cima/fisiologia
16.
Cancer Immunol Immunother ; 69(9): 1891-1903, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32377817

RESUMO

The objective response rate of immune checkpoint blockade (ICB) in hepatocellular carcinoma (HCC) with anti PD-L1/PD-1 therapy is low. Discovering the signaling pathways regulating PD-L1 might help to improve ICB response rates. Here, we investigate transcription factors IRF-1 and IRF-2 signaling pathways regulating PD-L1 in HCC cells. In vivo studies show that IRF-1 and PD-L1 mRNA expression in human HCC tumors are significantly repressed compared with noncancerous background liver. IRF-1, IRF-2, and PD-L1 mRNA expression correlated positively in HCC tumors. Increased IRF-1 mRNA expression was observed in patients with well-differentiated or early stage HCC tumors. In vitro studies show that IFN-γ induces PD-L1 mRNA and protein expression through upregulation of IRF-1 in mouse and human HCC cells. IRF-1, IRF-2, and PD-L1 mRNA expression is upregulated in murine HCC by co-culture with effector T cells from spleen cells incubated with anti-CD3/CD28 antibodies. IRF-2 over-expression down-regulates IFN-γ induced PD-L1 promoter activity and protein levels in a dose-dependent manner. We identify two IRF-1 response elements (IRE1/IRE2) in the upstream 5'-flanking region of the CD274 (PD-L1) gene promoter. Site-directed mutagenesis shows both IRE1 and IRE2 are functional in transfection promoter assays. IRF-1 traditionally functions as tumor suppressor gene. However, these novel findings show a complex role for IRF-1 which upregulates PD-L1 in the inflammatory tumor microenvironment. IRF-1 antagonizes IRF-2 for binding to the IRE promoter element in PD-L1 which gives new insight to the regulation of PD-L1/PD-1 pathways in HCC ICB therapy.


Assuntos
Antígeno B7-H1/metabolismo , Carcinoma Hepatocelular/metabolismo , Fator Regulador 1 de Interferon/metabolismo , Fator Regulador 2 de Interferon/metabolismo , Neoplasias Hepáticas/metabolismo , Idoso , Animais , Linhagem Celular Tumoral , Regulação para Baixo/fisiologia , Feminino , Células Hep G2 , Humanos , Interferon gama/metabolismo , Masculino , Camundongos , Transdução de Sinais/fisiologia , Microambiente Tumoral/fisiologia , Regulação para Cima/fisiologia
17.
Adv Clin Exp Med ; 29(5): 565-572, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32421262

RESUMO

BACKGROUND: Thoracic aortic aneurysm (TAA) formation is accompanied by degradation of extracellular matrix components (EMC). Numerous matrix metalloproteinases (MMPs) have been implicated in the process, but the involvement of MMP-3 remains unclear. Additionally, the changes in proteoglycan (PG) structure can alter the signal transduction pathways in TAA, though the enzymatic systems which originate them are not fully understood. OBJECTIVES: To measure MMP-3 and sulfatase levels in aneurysmal tissue, comparing them with non-aneurysmal vessels, and to investigate possible correlations with patients' serum levels in order to evaluate their potential usefulness in aiding aneurysm detection and monitoring. MATERIAL AND METHODS: The study included 74 patients (TAA: n = 42; control group: n = 32). Sulfatase activity was measured colometrically and MMP-3 levels were measured immunoenzymatically. RESULTS: Sulfatase activities were higher (p = 0.03) and MMP-3 concentrations lower (p = 0.014) in aneurysmal tissue than in normal aortic tissue. Medium-sized dilatations were associated with lower tissue MMP-3 concentrations than small dilatations (p = 0.033). No differences in sulfatase activity or MMP-3 concentration in the serum of TAA patients were observed in comparison with the controls. The serum and tissue levels of MMP-3 were correlated (r = 0.41; p < 0.001). The serum levels of MMP-3 were significantly lower in the female patients than in the male patients (p = 0.006). CONCLUSIONS: Our studies confirmed the lower MMP-3 levels in aneurysmal tissue, but the lack of a statistically confirmed reduction of MMP-3 in the blood serum seems to preclude its usefulness for diagnostic purposes. Our study points to the differences in MMP-3 behavior between TAA and abdominal aortic aneurysms. Significantly higher sulfatase activity in TAA tissue suggests a possible impact of sulfatase on signal transduction pathways involved in aneurysm formation.


Assuntos
Aneurisma da Aorta Torácica/diagnóstico , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Sulfatases/metabolismo , Aorta , Aorta Torácica , Aneurisma da Aorta Torácica/sangue , Estudos de Casos e Controles , Regulação para Baixo/fisiologia , Feminino , Humanos , Masculino , Sulfatases/genética , Regulação para Cima/fisiologia
18.
Sci Rep ; 10(1): 5819, 2020 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-32242034

RESUMO

Climatic change is pointed as one of the major challenges for global food security. Based on current models of climate change, reduction in precipitations and in turn, increase in the soil salinity will be a sharp constraint for crops productivity worldwide. In this context, root fungi appear as a new strategy to improve plant ecophysiological performance and crop yield under abiotic stress. In this study, we evaluated the impact of the two fungal endophytes Penicillium brevicompactum and P. chrysogenum isolated from Antarctic plants on nutrients and Na+ contents, net photosynthesis, water use efficiency, yield and survival in tomato and lettuce, facing salinity stress conditions. Inoculation of plant roots with fungal endophytes resulted in greater fresh and dry biomass production, and an enhanced survival rate under salt conditions. Inoculation of plants with the fungal endophytes was related with a higher up/down-regulation of ion homeostasis by enhanced expression of the NHX1 gene. The two endophytes diminished the effects of salt stress in tomato and lettuce, provoked a higher efficiency in photosynthetic energy production and an improved sequestration of Na+ in vacuoles is suggested by the upregulating of the expression of vacuolar NHX1 Na+/H+ antiporters. Promoting plant-beneficial interactions with root symbionts appears to be an environmentally friendly strategy to mitigate the impact of climate change variables on crop production.


Assuntos
Produtos Agrícolas/metabolismo , Produtos Agrícolas/fisiologia , Endófitos/fisiologia , Raízes de Plantas/metabolismo , Raízes de Plantas/fisiologia , Estresse Salino/fisiologia , Sódio/metabolismo , Regiões Antárticas , Biomassa , Mudança Climática , Produtos Agrícolas/microbiologia , Regulação para Baixo/fisiologia , Homeostase/fisiologia , Íons/metabolismo , Alface/metabolismo , Alface/microbiologia , Alface/fisiologia , Lycopersicon esculentum/metabolismo , Lycopersicon esculentum/microbiologia , Lycopersicon esculentum/fisiologia , Penicillium chrysogenum/fisiologia , Fotossíntese/fisiologia , Raízes de Plantas/microbiologia , Salinidade , Trocadores de Sódio-Hidrogênio/metabolismo , Solo , Estresse Fisiológico/fisiologia , Taxa de Sobrevida , Regulação para Cima/fisiologia , Água/metabolismo
19.
Arch Biochem Biophys ; 687: 108385, 2020 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-32335050

RESUMO

MicroRNA-342-3p (miR-342) has been shown to act as a tumor-suppressor in different cancer types. However, the role and therapeutic implications of miR-342 via modulation of Cofilin 1 (CFL1) has not been studied in any type of cancer. Given the importance of Cofilin signalling in breast, this study was undertaken to explore the therapeutic implications of miR-342 and its target CFL1 in breast cancer. Herein, we found that miR-342 was significantly (P < 0.05) downregulated in breast cancer tissues and cell lines. Functional assays revealed that overexpression of miR-342 caused a significant (P < 0.05) inhibition of the proliferation, colony formation, invasion and migration of the MDA-MB-436 and CAMA-1 breast cancer cells via induction of apoptosis. Bioinformatic approaches and the dual luciferase reporter assay confirmed the interaction between miR-342 and its target CFL1. Moreover, we found that CFL1 was aberrantly overexpressed in breast cancer tissues and cell lines. Overexpression of miR-342 caused remarkable depletion in the expression of CFL1 in MDA-MB-436 breast cancer cells. Silencing of CFL1 in CAMA-1 and MDA-MB-436 cells caused remarkable decrease in the proliferation, colony formation and migration of these cells, similar to that of miR-342 ovexpression. However, overexpression of CFL1 in MDA-MB-346 cells could avoid the tumor suppressive effects of miR-342. Our data provide novel information about the implications of miR-342 and its target CFL1 in breast cancer treatment.


Assuntos
Neoplasias da Mama/fisiopatologia , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Cofilina 1/metabolismo , MicroRNAs/metabolismo , Adulto , Idoso , Apoptose/fisiologia , Linhagem Celular Tumoral , Regulação para Baixo/fisiologia , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Pessoa de Meia-Idade , Regulação para Cima/fisiologia
20.
Biochem Pharmacol ; 175: 113897, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32135158

RESUMO

Deoxynivalenol (DON) is a mycotoxin produced by multipleFusariumspecies that often contaminates cereals and threatens human and animal health. A wide range of cytotoxic effects, such as the induction of DNA damage, an increase in mitochondrial permeability and the inhibition of macromolecule synthesis, have been reported. However, the effects of DON on cell migration-a fundamental process in living cells critical for normal development, immune responses, and disease processes-and the mechanism underlying these effects are still unclear. Here, we showed that DONsignificantly inhibited the migration of MRC-5, CCD-18Co, HCT116 and WM793 cells at 50 ng/ml, 50 ng/ml, 400 ng/ml and 250 ng/ml, respectively, which maintained cell viability at 90%. Further analysis showed that DON inhibited the expression of tumour endothelial marker 8 (TEM8), a key gene in cell migration. Furthermore, we showed that DON inhibited the expression of TEM8 through increasing the level of H3K27me3 in the TEM8 promoter. Finally, overexpression of TEM8 or treating by H3K27me3-specific inhibitor GSK126 attenuated the inhibitory effect of DON on cell migration. In summary, low doses of DON at approximately dietary exposure significantly inhibited cell migration by downregulating the expression of TEM8 in a manner mediated by H3K27me3, which may generate increasing concerns for the risk of DON exposure.


Assuntos
Movimento Celular/fisiologia , Regulação para Baixo/fisiologia , Regulação Neoplásica da Expressão Gênica , Histonas/biossíntese , Proteínas dos Microfilamentos/biossíntese , Receptores de Superfície Celular/biossíntese , Tricotecenos/farmacologia , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Células HCT116 , Histonas/genética , Humanos , Proteínas dos Microfilamentos/antagonistas & inibidores , Proteínas dos Microfilamentos/genética , Receptores de Superfície Celular/antagonistas & inibidores , Receptores de Superfície Celular/genética
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