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1.
Gene ; 807: 145964, 2022 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-34530087

RESUMO

AIMS: We aimed to investigate the role of G protein subunit alpha Z(GNAZ) in the progression and prognosis of patients with hepatocellular carcinoma (HCC). METHODS: Oncomine, GEO, TCGA, GEPIA2, Kaplan-Meier Plotter, TIMER2, Metascape, CCLE, LinkedOmics, and UALCAN databases were used to analyze the differential expression of GNAZ in HCC and normal liver tissues, relationship between GNAZ expression and prognosis of patients with HCC, and expression of GNAZ in common human HCC cell lines. Western blotting was performed to analyze GNAZ expression, while the Cell Counting Kit 8 assay was used to determine cell proliferation, and flow cytometry was used to evaluate the cell cycle and apoptosis. Wound healing and transwell invasion assays were used to investigate cell metastasis and invasion. RESULTS: Using Oncomine, Gene Expression Omnibus (GEO), and GEPIA2 databases, GNAZ was found to be overexpressed in HCC tissues compared with that in adjacent normal liver tissues, and western blotting analysis showed GNAZ overexpression in seven patients with HCC who underwent surgical resection of HCC and para-cancerous tissues (p < 0.01). Survival analysis revealed that high GNAZ expression was negatively associated with overall survival (OS), recurrence-free survival, progression-free survival, and disease-specific survival in patients with HCC (p < 0.05). GNAZ overexpression was associated with worse 4- month, 6- month, 12- month, 24- month, 36- month, 48- month, and 60-month OS, as well as with different clinicopathological characteristics of patients with HCC, including hepatitis virus infection state; alcohol consumption state; male; female; Asian; microvascular invasion, Stage I-II, Stage II-III, and Stage III-IV; and grade II (Cox regression, p < 0.05). KEGG/GO biological process enrichment indicated that the genes similar to GNAZ in HCC were mainly enriched in the cell cycle, cell cycle phase transition, DNA replication checkpoint, and regulation of G0 to G1 transition. siRNA-GNAZ significantly reduced the viability of JHH-2 and SNU-761 cells from 12 to 96 h; increased the percentage of cells in the G0/G1 phase and decreased that of cells in the S and G2/M phases (p < 0.05); and markedly downregulated the expression of cyclin D, cyclin E, and CDK2 protein. siRNA-GNAZ also significantly increased the percentage of JHH-2 and SNU-761 cell apoptosis at late stages, while the number of surviving cells decreased (p < 0.05), and upregulated the expression of apoptosis-related proteins Bax and caspase 3 protein. Furthermore, siRNA-GNAZ remarkably reduced the healing of scratch wounds in JHH-2 and SNU-761 cells and the number of invasive cells compared with that in the control group (p < 0.001). CONCLUSION: Our study demonstrated that GNAZ plays a pivotal role as a potential oncogene and predicts poor prognosis in patients with HCC. It promotes tumor proliferation via cell cycle arrest, apoptosis, migration, and invasion. Thus, GNAZ may be a potential candidate biomarker providing useful insight into hepatocarcinogenesis and aggressiveness.


Assuntos
Carcinoma Hepatocelular/genética , Pontos de Checagem da Fase G1 do Ciclo Celular/genética , Subunidades alfa de Proteínas de Ligação ao GTP/genética , Idoso , Apoptose/genética , Grupo com Ancestrais do Continente Asiático/genética , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/mortalidade , Ciclo Celular/genética , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , China , Feminino , Subunidades alfa de Proteínas de Ligação ao GTP/metabolismo , Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/mortalidade , Masculino , Pessoa de Meia-Idade , Prognóstico , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Transcriptoma/genética
2.
DNA Cell Biol ; 40(11): 1396-1406, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34767734

RESUMO

Sepsis has become a major public health problem worldwide. Methylprednisolone sodium succinate (MP) is a commonly used drug to prevent inflammation. However, the role and underlying mechanism of MP in sepsis remain vague. MP inhibited the lipopolysaccharide (LPS)-induced production of tumor necrosis factor-alpha (TNF-α) and interleukin (IL)-17 and suppressed cell growth in alveolar type II epithelial cells (ATII cells). Small nucleolar RNA host gene 5 (SNHG5) expression was inhibited by LPS and restored by MP. Upregulation of SNHG5 inhibited the cellular role of LPS in ATII cells, and further, downregulation of SNHG5 inhibited the cellular role of MP in ATII cells under LPS conditions. SNHG5 elevated the expression of Copine 1 (CPNE1) by enhancing the mRNA stability of CPNE1. Increasing CPNE1 expression restored the silenced SNHG5-induced inhibitor role of MP in ATII cells under LPS conditions. Finally, MP attenuated lung injury and TNF-α and IL-17 secretion in an LPS-induced sepsis mouse model. Overall, this study investigated the mechanism underlying the effect of MP treatment in sepsis and, for the first time, revealed the important role of the SNHG5/CPNE1 pathway in the development and treatment of sepsis and the potential to serve as a diagnostic and therapeutic target for sepsis.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Metilprednisolona/farmacologia , Sepse/tratamento farmacológico , Células Epiteliais Alveolares/efeitos dos fármacos , Células Epiteliais Alveolares/metabolismo , Animais , Apoptose/efeitos dos fármacos , Proteínas de Ligação ao Cálcio/efeitos dos fármacos , Proteínas de Ligação ao Cálcio/genética , Proteínas de Transporte , Ciclo Celular/efeitos dos fármacos , Proliferação de Células/genética , Feminino , Inflamação , Lipopolissacarídeos/farmacologia , Metilprednisolona/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , RNA Longo não Codificante/genética , RNA Nucleolar Pequeno/genética , Sepse/metabolismo , Transdução de Sinais/efeitos dos fármacos
3.
Int J Mol Sci ; 22(19)2021 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-34639098

RESUMO

Synchronous cell populations are commonly used for the analysis of various aspects of cellular metabolism at specific stages of the cell cycle. Cell synchronization at a chosen cell cycle stage is most frequently achieved by inhibition of specific metabolic pathway(s). In this respect, various protocols have been developed to synchronize cells in particular cell cycle stages. In this review, we provide an overview of the protocols for cell synchronization of mammalian cells based on the inhibition of synthesis of DNA building blocks-deoxynucleotides and/or inhibition of DNA synthesis. The mechanism of action, examples of their use, and advantages and disadvantages are described with the aim of providing a guide for the selection of suitable protocol for different studied situations.


Assuntos
Ciclo Celular , Divisão Celular , Replicação do DNA , DNA/biossíntese , Animais , DNA/antagonistas & inibidores , Humanos
4.
Int J Mol Sci ; 22(19)2021 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-34639115

RESUMO

Psoriasis is a chronic inflammatory skin disease. Recently, lysophosphatidic acid (LPA)/LPAR5 signaling has been reported to be involved in both NLRP3 inflammasome activation in macrophages and keratinocyte activation to produce inflammatory cytokines, contributing to psoriasis pathogenesis. However, the effect and molecular mechanisms of LPA/LPAR signaling in keratinocyte proliferation in psoriasis remain unclear. In this study, we investigated the effects of LPAR1/3 inhibition on imiquimod (IMQ)-induced psoriasis-like mice. Treatment with the LPAR1/3 antagonist, ki16425, alleviated skin symptoms in IMQ-induced psoriasis-like mouse models and decreased keratinocyte proliferation in the lesion. It also decreased LPA-induced cell proliferation and cell cycle progression via increased cyclin A2, cyclin D1, cyclin-dependent kinase (CDK)2, and CDK4 expression and decreased p27Kip1 expression in HaCaT cells. LPAR1 knockdown in HaCaT cells reduced LPA-induced proliferation, suppressed cyclin A2 and CDK2 expression, and restored p27Kip1 expression. LPA increased Rho-associated protein kinase 2 (ROCK2) expression and PI3K/AKT activation; moreover, the pharmacological inhibition of ROCK2 and PI3K/AKT signaling suppressed LPA-induced cell cycle progression. In conclusion, we demonstrated that LPAR1/3 antagonist alleviates IMQ-induced psoriasis-like symptoms in mice, and in particular, LPAR1 signaling is involved in cell cycle progression via ROCK2/PI3K/AKT pathways in keratinocytes.


Assuntos
Proliferação de Células , Regulação da Expressão Gênica/efeitos dos fármacos , Imiquimode/toxicidade , Queratinócitos/citologia , Lisofosfolipídeos/farmacologia , Psoríase/tratamento farmacológico , Animais , Apoptose , Biomarcadores/metabolismo , Ciclo Celular , Células Cultivadas , Humanos , Indutores de Interferon/toxicidade , Queratinócitos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Psoríase/induzido quimicamente , Psoríase/metabolismo , Psoríase/patologia , Receptores de Ácidos Lisofosfatídicos/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Quinases Associadas a rho/genética , Quinases Associadas a rho/metabolismo
5.
Nat Commun ; 12(1): 5977, 2021 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-34645816

RESUMO

Muscle diseases and aging are associated with impaired myogenic stem cell self-renewal and fewer proliferating progenitors (MPs). Importantly, distinct metabolic states induced by glycolysis or oxidative phosphorylation have been connected to MP proliferation and differentiation. However, how these energy-provisioning mechanisms cooperate remain obscure. Herein, we describe a mechanism by which mitochondrial-localized transcriptional co-repressor p107 regulates MP proliferation. We show p107 directly interacts with the mitochondrial DNA, repressing mitochondrial-encoded gene transcription. This reduces ATP production by limiting electron transport chain complex formation. ATP output, controlled by the mitochondrial function of p107, is directly associated with the cell cycle rate. Sirt1 activity, dependent on the cytoplasmic glycolysis product NAD+, directly interacts with p107, impeding its mitochondrial localization. The metabolic control of MP proliferation, driven by p107 mitochondrial function, establishes a cell cycle paradigm that might extend to other dividing cell types.


Assuntos
Lactato Desidrogenase 5/genética , Mitocôndrias/genética , Músculo Esquelético/metabolismo , Mioblastos/metabolismo , Proteína p107 Retinoblastoma-Like/genética , Células-Tronco/metabolismo , Trifosfato de Adenosina/biossíntese , Animais , Ciclo Celular/genética , Linhagem Celular , Proliferação de Células , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Complexo de Proteínas da Cadeia de Transporte de Elétrons/genética , Complexo de Proteínas da Cadeia de Transporte de Elétrons/metabolismo , Regulação da Expressão Gênica , Glicólise , Humanos , Lactato Desidrogenase 5/antagonistas & inibidores , Lactato Desidrogenase 5/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/metabolismo , Músculo Esquelético/citologia , Mioblastos/citologia , Fosforilação Oxidativa , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteína p107 Retinoblastoma-Like/metabolismo , Sirtuína 1/genética , Sirtuína 1/metabolismo , Células-Tronco/citologia , Transcrição Genética
6.
Int J Mol Sci ; 22(19)2021 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-34638889

RESUMO

The response to DNA damage is the mechanism that allows the interaction between stress signals, inflammatory secretions, DNA repair, and maintenance of cell and tissue homeostasis. Adipocyte dysfunction is the cellular trigger for various disease states such as insulin resistance, diabetes, and obesity, among many others. Previously, our group demonstrated that adipogenesis per se, from mesenchymal/stromal stem cells derived from human adipose tissue (hASCs), involves an accumulation of DNA damage and a gradual loss of the repair capacity of oxidative DNA damage. Therefore, our objective was to identify whether healthy adipocytes differentiated for the first time from hASCs, when receiving inflammatory signals induced with TNFα, were able to persistently activate the DNA Damage Response and thus trigger adipocyte dysfunction. We found that TNFα at similar levels circulating in obese humans induce a sustained response to DNA damage response as part of the Senescence-Associated Secretory Phenotype. This mechanism shows the impact of inflammatory environment early affect adipocyte function, independently of aging.


Assuntos
Adipócitos/metabolismo , Diferenciação Celular , Dano ao DNA , Fator de Necrose Tumoral alfa/metabolismo , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Tecido Adiposo/citologia , Tecido Adiposo/metabolismo , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Ensaio Cometa/métodos , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Fator de Necrose Tumoral alfa/farmacologia
7.
Int J Mol Sci ; 22(19)2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34639030

RESUMO

During the cell cycle, DNA suffers several lesions that need to be repaired prior to entry into mitosis to preserve genome integrity in daughter cells. Toward this aim, cells have developed complex enzymatic machinery, the so-called DNA damage response (DDR), which is able to repair DNA, temporarily stopping the cell cycle to provide more time to repair, or if the damage is too severe, inducing apoptosis. This DDR mechanism is considered the main source of resistance to DNA-damaging therapeutic treatments in oncology. Recently, cancer stem cells (CSCs), which are a small subset of tumor cells, were identified as tumor-initiating cells. CSCs possess self-renewal potential and persistent tumorigenic capacity, allowing for tumor re-growth and relapse. Compared with cancer cells, CSCs are more resistant to therapeutic treatments. Wee1 is the principal gatekeeper for both G2/M and S-phase checkpoints, where it plays a key role in cell cycle regulation and DNA damage repair. From this perspective, Wee1 inhibition might increase the effectiveness of DNA-damaging treatments, such as radiotherapy, forcing tumor cells and CSCs to enter into mitosis, even with damaged DNA, leading to mitotic catastrophe and subsequent cell death.


Assuntos
Biomarcadores Tumorais , Proteínas de Ciclo Celular/genética , Resistencia a Medicamentos Antineoplásicos/genética , Proteínas Tirosina Quinases/genética , Tolerância a Radiação/genética , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/metabolismo , Terapia Combinada , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/efeitos da radiação , Progressão da Doença , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Terapia de Alvo Molecular , Família Multigênica , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Especificidade de Órgãos/genética , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/metabolismo
8.
Sheng Li Xue Bao ; 73(5): 761-771, 2021 Oct 25.
Artigo em Chinês | MEDLINE | ID: mdl-34708233

RESUMO

Nutrient overload-caused deregulation of glucose and lipid metabolism leads to insulin resistance and metabolic disorders, which increases the risk of several types of cancers. CREB/ATF bZIP transcription factor (CREBZF), a novel transcription factor of the ATF/CREB family, has emerged as a critical mechanism bridging the gap between metabolism and cell growth. CREBZF forms a heterodimer with other proteins and functions as a coregulator for gene expression. CREBZF deficiency in the liver attenuates hepatic steatosis in high fat diet-induced insulin-resistant mice, while the expression levels of CREBZF are increased in the livers of obese mice and humans with hepatic steatosis. Intriguingly, CREBZF also regulates cell proliferation and apoptosis via interaction with several transcription factors including STAT3, p53 and HCF-1. Knockout of CREBZF in hepatocytes results in enhanced cell cycle progression and proliferation capacity in mice. Here we highlight how the CREBZF signaling network contributes to the deregulation of metabolism and cell growth, and discuss the potential of targeting these molecules for the treatment of insulin resistance, diabetes, fatty liver disease and cancer.


Assuntos
Fatores de Transcrição de Zíper de Leucina Básica , Resistência à Insulina , Fígado , Animais , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Ciclo Celular , Proliferação de Células , Dieta Hiperlipídica , Hepatócitos , Metabolismo dos Lipídeos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais
9.
Nat Commun ; 12(1): 5784, 2021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34599161

RESUMO

Cardiac regeneration involves the generation of new cardiomyocytes from cycling cardiomyocytes. Understanding cell-cycle activity of pre-existing cardiomyocytes provides valuable information to heart repair and regeneration. However, the anatomical locations and in situ dynamics of cycling cardiomyocytes remain unclear. Here we develop a genetic approach for a temporally seamless recording of cardiomyocyte-specific cell-cycle activity in vivo. We find that the majority of cycling cardiomyocytes are positioned in the subendocardial muscle of the left ventricle, especially in the papillary muscles. Clonal analysis revealed that a subset of cycling cardiomyocytes have undergone cell division. Myocardial infarction and cardiac pressure overload induce regional patterns of cycling cardiomyocytes. Mechanistically, cardiomyocyte cell cycle activity requires the Hippo pathway effector YAP. These genetic fate-mapping studies advance our basic understanding of cardiomyocyte cell cycle activity and generation in cardiac homeostasis, repair, and regeneration.


Assuntos
Miócitos Cardíacos/citologia , Animais , Southern Blotting , Ciclo Celular/fisiologia , Proliferação de Células/fisiologia , Feminino , Citometria de Fluxo , Coração/fisiologia , Masculino , Camundongos , Microscopia de Fluorescência , Miocárdio/metabolismo , Miocárdio/patologia , Miócitos Cardíacos/metabolismo
10.
Science ; 374(6565): 263-264, 2021 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-34648332

RESUMO

[Figure: see text].


Assuntos
Ciclo Celular
11.
Int J Mol Sci ; 22(19)2021 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-34638565

RESUMO

Malignant pleural mesothelioma (MPM) is an aggressive malignancy with limited effective treatment options. Focal adhesion kinase (FAK) inhibitors have been shown to efficiently suppress MPM cell growth initially, with limited utility in the current clinical setting. In this study, we utilised a large collection of MPM cell lines and MPM tissue samples to study the role of E-cadherin (CDH1) and microRNA on the efficacy of FAK inhibitors in MPM. The immunohistochemistry (IHC) results showed that the majority of MPM FFPE samples exhibited either the absence of, or very low, E-cadherin protein expression in MPM tissue. We showed that MPM cells with high CDH1 mRNA levels exhibited resistance to the FAK inhibitor PND-1186. In summary, MPM cells that did not express CDH1 mRNA were sensitive to PND-1186, and MPM cells that retained CDH1 mRNA were resistant. A cell cycle analysis showed that PND-1186 induced cell cycle disruption by inducing the G2/M arrest of MPM cells. A protein-protein interaction study showed that EGFR is linked to the FAK pathway, and a target scan of the microRNAs revealed that microRNAs (miR-17, miR221, miR-222, miR137, and miR148) interact with EGFR 3'UTR. Transfection of MPM cells with these microRNAs sensitised the CHD1-expressing FAK-inhibitor-resistant MPM cells to the FAK inhibitor.


Assuntos
Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Quinase 1 de Adesão Focal/antagonistas & inibidores , Mesotelioma Maligno/tratamento farmacológico , Mesotelioma Maligno/genética , MicroRNAs/fisiologia , Inibidores de Proteínas Quinases/farmacologia , Aminopiridinas/farmacologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Receptores ErbB/metabolismo , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Mapas de Interação de Proteínas
12.
Anticancer Res ; 41(10): 4807-4820, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34593430

RESUMO

BACKGROUND/AIM: LY2835219 (LY), a novel CDK4/6 inhibitor, prevents cell proliferation through G1 arrest. Docetaxel (DTX) and paclitaxel (PTX) are cytotoxic drugs targeting tubulin-mediated apoptotic cell death via G2/M arrest. We evaluated the antitumor effects of DTX/PTX and LY individually and in combination in lung adenocarcinoma cells with or without KRAS mutations and xenograft mice harboring KRAS mutations. MATERIALS AND METHODS: We investigated in vitro/in vivo changes in signaling molecules and analyzed cell proliferation, cycle, and apoptosis via flow cytometry and western blotting. RESULTS: LY cytotoxicity was dose-dependent and varied with KRAS mutation status. DTX→LY showed synergistic cytotoxicity regardless of KRAS mutation. Furthermore, the synergistic effect of PTX→LY was significantly greater than that of PTX+LY. DTX→LY remarkably reduced the number of G0/G1 cells and increased the number of G2/M arrested cells, resulting in an increase in apoptosis and subG1 cells. CONCLUSION: DTX→LY has synergistic antitumor effect in lung cancer cells and xenograft mice regardless of KRAS mutation.


Assuntos
Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Neoplasias Pulmonares/tratamento farmacológico , Proteínas Proto-Oncogênicas p21(ras)/genética , Taxoides/farmacologia , Aminopiridinas/farmacologia , Aminopiridinas/uso terapêutico , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Benzimidazóis/farmacologia , Benzimidazóis/uso terapêutico , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sinergismo Farmacológico , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Camundongos , Mutação , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Transdução de Sinais/efeitos dos fármacos , Taxoides/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Int J Mol Sci ; 22(19)2021 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-34638879

RESUMO

Colorectal cancer (CRC) is one of the most frequently diagnosed cancers in humans. At early stages CRC is treated by surgery and at advanced stages combined with chemotherapy. We examined here the potential effect of glucosylceramide synthase (GCS)-inhibition on CRC biology. GCS is the rate-limiting enzyme in the glycosphingolipid (GSL)-biosynthesis pathway and overexpressed in many human tumors. We suppressed GSL-biosynthesis using the GCS inhibitor Genz-123346 (Genz), NB-DNJ (Miglustat) or by genetic targeting of the GCS-encoding gene UDP-glucose-ceramide-glucosyltransferase- (UGCG). GCS-inhibition or GSL-depletion led to a marked arrest of the cell cycle in Lovo cells. UGCG silencing strongly also inhibited tumor spheroid growth in Lovo cells and moderately in HCT116 cells. MS/MS analysis demonstrated markedly elevated levels of sphingomyelin (SM) and phosphatidylcholine (PC) that occurred in a Genz-concentration dependent manner. Ultrastructural analysis of Genz-treated cells indicated multi-lamellar lipid storage in vesicular compartments. In mice, Genz lowered the incidence of experimentally induced colorectal tumors and in particular the growth of colorectal adenomas. These results highlight the potential for GCS-based inhibition in the treatment of CRC.


Assuntos
Ciclo Celular/efeitos dos fármacos , Neoplasias do Colo , Dioxanos/farmacologia , Glicoesfingolipídeos , Pirrolidinas/farmacologia , Esferoides Celulares , Animais , Neoplasias do Colo/induzido quimicamente , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Glucosiltransferases/antagonistas & inibidores , Glucosiltransferases/metabolismo , Glicoesfingolipídeos/biossíntese , Glicoesfingolipídeos/genética , Células HCT116 , Humanos , Camundongos , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/metabolismo , Neoplasias Experimentais/induzido quimicamente , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/genética , Neoplasias Experimentais/metabolismo , Esferoides Celulares/metabolismo , Esferoides Celulares/patologia
14.
Int J Mol Sci ; 22(19)2021 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-34638937

RESUMO

Acute myeloid leukemia is characterized by uncontrolled clonal proliferation of abnormal myeloid progenitor cells. Despite recent advances in the treatment of this disease, the prognosis and overall long-term survival for patients remain poor, which drives the search for new chemotherapeutics and treatment strategies. Piceatannol, a polyphenolic compound present in grapes and wine, appears to be a promising chemotherapeutic agent in the treatment of leukemia. The aim of the present study was to examine whether piceatannol induces autophagy and/or apoptosis in HL-60 human acute myeloid leukemia cells and whether HL-60 cells are able to acquire resistance to piceatannol toxicity. We found that piceatannol at the IC90 concentration of 14 µM did not induce autophagy in HL-60 cells. However, it induced caspase-dependent apoptosis characterized by phosphatidylserine externalization, disruption of the mitochondrial membrane potential, caspase-3 activation, internucleosomal DNA fragmentation, PARP1 cleavage, chromatin condensation, and fragmentation of cell nuclei. Our findings also imply that HL-60 cells are able to acquire resistance to piceatannol toxicity via mechanisms related to MRP1 activity. Our results suggest that the use of piceatannol as a potential chemotherapeutic agent may be associated with the risk of multidrug resistance, warranting its use in combination with other chemotherapeutic agents.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Leucemia Mieloide Aguda/metabolismo , Resveratrol/análogos & derivados , Estilbenos/farmacologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Antineoplásicos/química , Autofagia/efeitos dos fármacos , Caspase 3/metabolismo , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Fragmentação do DNA/efeitos dos fármacos , Células HL-60 , Humanos , Concentração Inibidora 50 , Leucemia Mieloide Aguda/patologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Estrutura Molecular , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Estilbenos/química
15.
Molecules ; 26(20)2021 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-34684829

RESUMO

Wound healing is a great challenge in many health conditions, especially in non-healing conditions. The search for new wound healing agents continues unabated, as the use of growth factors is accompanied by several limitations. Medicinal plants have been used for a long time in would healing, despite the lack of scientific evidence veryfying their efficacy. Up to now, the number of reports about medicinal plants with wound healing properties is limited. Urtica dioica L. is a well-known plant, widely used in many applications. Reports regarding its wound healing potential are scant and sparse. In this study, the effect of an Urtica dioica L. extract (containing fewer antioxidant compounds compared to methanolic or hydroalcoholic extracts) on cell proliferation, the cell cycle, and migration were examined. Additionally, antioxidant and anti-inflammatory properties were examined. Finally, in vivo experiments were carried out on full-thickness wounds on Wistar rats. It was found that the extract increases the proliferation rate of HEK-293 and HaCaT cells up to 39% and 30% after 24 h, respectively, compared to control cells. The extract was found to increase the population of cells in the G2/M phase by almost 10%. Additionally, the extract caused a two-fold increase in the cell migration rate of both cell lines compared to control cells. Moreover, the extract was found to have anti-inflammatory properties and moderate antioxidant properties that augment its overall wound healing potential. Results from the in vivo experiments showed that wounds treated with an ointment of the extract healed in 9 days, while wounds not treated with the extract healed in 13 days. Histopathological examination of the wound tissue revealed, among other findings, that inflammation was significantly reduced compared to the control. Urtica dioica L. extract application results in faster wound healing, making the extract ideal for wound healing applications and a novel drug candidate for wound healing.


Assuntos
Extratos Vegetais/farmacologia , Plantas Medicinais/química , Urtica dioica/química , Cicatrização/efeitos dos fármacos , Animais , Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células HEK293 , Humanos , Masculino , Ratos , Ratos Wistar , Pele/efeitos dos fármacos , Pele/lesões , Pele/patologia
16.
Nat Cell Biol ; 23(10): 1095-1104, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34616022

RESUMO

BRCA2-mutant cells are defective in homologous recombination, making them vulnerable to the inactivation of other pathways for the repair of DNA double-strand breaks (DSBs). This concept can be clinically exploited but is currently limited due to insufficient knowledge about how DSBs are repaired in the absence of BRCA2. We show that DNA polymerase θ (POLθ)-mediated end joining (TMEJ) repairs DSBs arising during the S phase in BRCA2-deficient cells only after the onset of the ensuing mitosis. This process is regulated by RAD52, whose loss causes the premature usage of TMEJ and the formation of chromosomal fusions. Purified RAD52 and BRCA2 proteins both block the DNA polymerase function of POLθ, suggesting a mechanism explaining their synthetic lethal relationships. We propose that the delay of TMEJ until mitosis ensures the conversion of originally one-ended DSBs into two-ended DSBs. Mitotic chromatin condensation might further serve to juxtapose correct break ends and limit chromosomal fusions.


Assuntos
Proteína BRCA2/metabolismo , Quebras de DNA de Cadeia Dupla , Reparo do DNA por Junção de Extremidades , DNA Polimerase Dirigida por DNA/metabolismo , Recombinação Homóloga , Mitose , Proteína Rad52 de Recombinação e Reparo de DNA/metabolismo , Proteína BRCA2/genética , Ciclo Celular , DNA Polimerase Dirigida por DNA/genética , Células HeLa , Humanos , Proteína Rad52 de Recombinação e Reparo de DNA/genética
17.
Nat Commun ; 12(1): 5864, 2021 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-34620876

RESUMO

Pausing of RNA polymerase II (Pol II) close to promoters is a common regulatory step in RNA synthesis, and is coordinated by a ribonucleoprotein complex scaffolded by the noncoding RNA RN7SK. The function of RN7SK-regulated gene transcription in adult tissue homoeostasis is currently unknown. Here, we deplete RN7SK during mouse and human epidermal stem cell differentiation. Unexpectedly, loss of this small nuclear RNA specifically reduces transcription of numerous cell cycle regulators leading to cell cycle exit and differentiation. Mechanistically, we show that RN7SK is required for efficient transcription of highly expressed gene pairs with bidirectional promoters, which in the epidermis co-regulated cell cycle and chromosome organization. The reduction in transcription involves impaired splicing and RNA decay, but occurs in the absence of chromatin remodelling at promoters and putative enhancers. Thus, RN7SK is directly required for efficient Pol II transcription of highly transcribed bidirectional gene pairs, and thereby exerts tissue-specific functions, such as maintaining a cycling cell population in the epidermis.


Assuntos
Regulação da Expressão Gênica , RNA Nuclear Pequeno/genética , RNA Nuclear Pequeno/metabolismo , Pele/metabolismo , Transcrição Genética , Animais , Ciclo Celular , Diferenciação Celular , Proliferação de Células , Cromatina , Montagem e Desmontagem da Cromatina , Epiderme , Feminino , Humanos , Queratinócitos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Regiões Promotoras Genéticas , RNA Polimerase II/metabolismo , Splicing de RNA , Pele/patologia , Células-Tronco
18.
Molecules ; 26(20)2021 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-34684858

RESUMO

A series of new Knoevenagel adducts, bearing two indolinone systems, has been synthesized and evaluated on 60 human cancer cell lines according to protocols available at the National Cancer Institute (Bethesda, MD, USA). Some derivatives proved to be potent antiproliferative agents, showing GI50 values in the submicromolar range. Compound 5b emerged as the most active and was further studied in Jurkat cells in order to determine the effects on cell-cycle phases and the kind of cell death induced. Finally, oxidative stress and DNA damage induced by compound 5b were also analyzed.


Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Oxindóis/farmacologia , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Dano ao DNA/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Humanos , Células Jurkat , Estrutura Molecular , Estresse Oxidativo/efeitos dos fármacos
20.
Biomed Res Int ; 2021: 5340449, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34545328

RESUMO

Objective: To identify significant pathways and genes in intervertebral disc degeneration (IDD) based on bioinformatics analysis. Design: The GEO database was used to download the GSE124272 dataset. Differentially expressed genes (DEGs) were analyzed using Limma package in R language. Then, gene ontologies (GO), Kyoto encyclopedia of genes and genomes (KEGG), and protein-protein interaction (PPI) networks were used to further identify hub genes. The mRNA expression levels of top six hub genes were verified. Results: We found 563 DEGs, of which 214 were upregulated and 349 were downregulated. The top 5 GO terms and pathways were shown including immune response, cell cycle, and p53 pathway. Based on the PPI analysis, we verified the mRNA expression levels of 6 hub genes. The mRNA levels of CHEK1, CDCA2, SKA3, and KIF20A were upregulated in degenerative NP tissue than in healthy NP tissue. However, the mRNA level of BUB1 and SPC25 was downregulated. Conclusions: This study may provide new biomarkers for the IDD and treatments to repair IDD related to CHEK1, CDCA2, SKA3, BUB1, KIF20A, and SPC25.


Assuntos
Redes Reguladoras de Genes/genética , Degeneração do Disco Intervertebral/genética , Mapas de Interação de Proteínas/genética , Biomarcadores/metabolismo , Proteínas de Transporte/genética , Ciclo Celular , Proteínas de Ciclo Celular/genética , Quinase 1 do Ponto de Checagem/genética , China , Proteínas Cromossômicas não Histona/genética , Biologia Computacional/métodos , Bases de Dados Genéticas , Expressão Gênica/genética , Perfilação da Expressão Gênica/métodos , Ontologia Genética , Humanos , Degeneração do Disco Intervertebral/metabolismo , Cinesina/genética , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas Nucleares/genética , Mapeamento de Interação de Proteínas/métodos , Proteínas Serina-Treonina Quinases/genética , Transdução de Sinais/genética , Transcriptoma/genética
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