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1.
Heliyon ; 7(5): e07064, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-34136678

RESUMO

Cancer cells are dependent on glutamine for their metabolism and growth. Despite being the most abundant amino acid in the blood, glutamine deprivation occurs in the core of the tumor rendering less access to glutamine to the nearby tumor cells. Tumor cells mostly use the glutamine for mitochondrial oxidative phosphorylation (OXPHOS) to produce energy and the ingredients of the biomass required for the highly proliferating and metastatic ovarian cancer cells. But there is a lack of reports on the regulation of glutamine starvation on metastatic behavior and epithelial to mesenchymal transition (EMT) of ovarian cancer cells. We found that glutamine starvation reduced the migration and invasion properties of the ovarian cancer cells, PA1 and SKOV3. The expression of the invasion-inducing proteins, like matrix metalloproteinases (MMP2 and MMP9), were downregulated upon glutamine starvation. MMP genes are mostly regulated by the ETS1 oncogenic transcription factor in invasive tumor cells. Here we demonstrated the significant involvement of ETS1 on EMT and invasion in glutamine-deprived cells. We have further shown that the regulation of ETS1 expression and nuclear localization upon glutamine starvation is controlled in a cell type-specific manner. In PA1 cells, glutamine-induced ETS1 over-expression is HIF1α-dependent, while in SKOV3, its translocation to the nucleus is regulated through the mTOR pathway. Considering all, our study suggests that glutamine plays a very significant role in migration and invasion in ovarian cancer cells and ETS1 plays a key role in inducing such oncogenic parameters.

2.
Cell Mol Life Sci ; 78(10): 4821-4845, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33895866

RESUMO

Glutamine is essential for maintaining the TCA cycle in cancer cells yet they undergo glutamine starvation in the core of tumors. Cancer stem cells (CSCs), responsible for tumor recurrence are often found in the nutrient limiting cores. Our study uncovers the molecular basis and cellular links between glutamine deprivation and stemness in the cancer cells. We showed that glutamine is dispensable for the survival of ovarian and colon cancer cells while it is required for their proliferation. Glutamine starvation leads to the metabolic reprogramming in tumor cells with enhanced glycolysis and unaltered oxidative phosphorylation. Production of reactive oxygen species (ROS) in glutamine limiting condition induces MAPK-ERK1/2 signaling pathway to phosphorylate dynamin-related protein-1(DRP1) at Ser616. Moreover, p-DRP1 promotes mitochondrial fragmentation and enhances numbers of CD44 and CD117/CD45 positive CSCs. Besides the established features of cancer stem cells, glutamine deprivation induces perinuclear localization of fragmented mitochondria and reduction in proliferation rate which are usually observed in CSCs. Treatment with glutaminase inhibitor (L-DON) mimics the effects of glutamine starvation without altering cell survival in in vitro as well as in in vivo model. Interestingly, the combinatorial treatment of L-DON with DRP1 inhibitor (MDiVi-1) reduces the stem cell population in tumor tissue in mouse model. Collectively our data suggest that glutamine deficiency in the core of tumors can increase the cancer stem cell population and the combination therapy with MDiVi-1 and L-DON is a useful approach to reduce CSCs population in tumor.


Assuntos
Resistencia a Medicamentos Antineoplásicos/fisiologia , Dinaminas/metabolismo , Glutamina/metabolismo , Mitocôndrias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Animais , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Feminino , Células HCT116 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Dinâmica Mitocondrial/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/fisiologia
5.
Cell Physiol Biochem ; 51(4): 1658-1678, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30504730

RESUMO

BACKGROUND/AIMS: The conformation, folding and lipid binding properties of the intestinal fatty acid binding proteins (IFABP) have been extensively investigated. In contrast, the functional aspects of these proteins are not understood and matter of debates. In this study, we aim to address the deleterious effects of FA overload on cellular components, particularly mitochondria; and how IFABP helps in combating this stress by restoring the mitochondrial dynamics. METHODS: In the present study the functional aspect of IFABP under conditions of lipid stress was studied by a string of extensive in-cell studies; flow cytometry by fluorescence-activated cell sorting (FACS), confocal imaging, western blotting and quantitative real time PCR. We deployed ectopic expression of IFABP in rescuing cells under the condition of lipid stress. Again in order to unveil the mechanistic insights of functional traits, we arrayed extensive computational approaches by means of studying centrality calculations along with protein-protein association and ligand induced cluster dissociation. While addressing its functional importance, we used FCS and in-silico computational analyses, to show the structural distribution and the underlying mechanism of IFABP's action. RESULTS: Ectopic expression of IFABP in HeLa cells has been found to rescue mitochondrial morphological dynamics and restore membrane potential, partially preventing apoptotic damage induced by the increased FAs. These findings have been further validated in the functionally relevant intestinal Caco-2 cells, where the native expression of IFABP protects mitochondrial morphology from abrogation induced by FA overload. However, this native level expression is insufficient to protect against apoptotic cell death, which is rescued, at least partially in cells overexpressing IFABP. In addition, shRNA mediated IFABP knockdown in Caco-2 cells compromises mitochondrial dynamics and switches on intrinsic apoptotic pathways under FA-induced metabolic stress. CONCLUSION: To summarize, the present study implicates functional significance of IFABP in controlling ligand-induced damage in mitochondrial dynamics and apoptosis.


Assuntos
Apoptose , Proteínas de Ligação a Ácido Graxo/metabolismo , Ácidos Graxos/metabolismo , Dinâmica Mitocondrial , Células CACO-2 , Células HeLa , Humanos , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Modelos Moleculares , Estresse Fisiológico
6.
ACS Omega ; 3(7): 7703-7714, 2018 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-30221238

RESUMO

Biomimetic synthesis of multifunctional fluorescent gold nanoclusters (Au NCs) is of great demand because of their ever-increasing applications. In this study, we have used self-assembled bovine serum albumin (BSA) amyloid-like nanofibers as the bioinspired scaffold for the synthesis of Au NCs. The amyloid fibril stabilized gold nanocluster (Fib-Au NC) has been found to have appreciable enhancement of fluorescence emission and a large 25 nm red shift in its emission maxima when compared to its monomeric protein counterpart (BSA-Au NC). The underlying mechanism accountable for the fluorescence behavior and its spectral shift has been thoroughly investigated by a combined use of spectroscopic and microscopic techniques. We have subsequently demonstrated the use of Fib-Au NCs for cysteine (Cys) sensing both in vitro and inside live cells. Additionally, cellular uptake and postpermeation effect of Fib-Au NCs have also been ascertained by detailed flow cytometry analysis, viability assay, and real-time apoptotic gene expression profiling.

7.
FEBS J ; 285(3): 432-443, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-28971574

RESUMO

Metabolic activity of malignant cells is very different from that of their nontransformed equivalents, which establishes metabolic reprogramming as an important hallmark of every transformed cell. In particular, the current arena of research in this field aims to understand the regulatory effect of oncogenic signaling on metabolic rewiring in transformed cells in order to exploit this for therapeutic benefit. Alterations in lipid metabolism are one of the main aspects of metabolic rewiring of transformed cells. Up-regulation of several lipogenic enzymes has been reported to be a characteristic of various cancer types. Lysophosphatidic acid (LPA), a simple byproduct of the lipid biosynthesis pathway, has gained immense importance due to its elevated level in several cancers and associated growth-promoting activity. Importantly, a current study revealed its role in increased de novo lipid synthesis through up-regulation of sterol regulatory element-binding protein 1, a master regulator of lipid metabolism. This review summarizes the recent insights in the field of oncolipid LPA-mediated signaling in regard to lipid metabolism in cancers. Future work in this domain is required to understand the up-regulation of the de novo synthesis pathway and the role of its end products in malignant cells. This will open a new arena of research toward the development of specific metabolic inhibitors that can add to the pre-existing chemotherapeutics in order to increase the efficacy of clinical output in cancer patients.


Assuntos
Metabolismo dos Lipídeos , Lipídeos/antagonistas & inibidores , Lisofosfolipídeos/antagonistas & inibidores , Modelos Biológicos , Neoplasias/metabolismo , Transdução de Sinais , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Drogas em Investigação/farmacologia , Drogas em Investigação/uso terapêutico , Ácido Graxo Sintase Tipo I/antagonistas & inibidores , Ácido Graxo Sintase Tipo I/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipídeos/sangue , Lipogênese/efeitos dos fármacos , Lisofosfolipídeos/sangue , Lisofosfolipídeos/metabolismo , Terapia de Alvo Molecular , Proteínas de Neoplasias/agonistas , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/sangue , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Diester Fosfórico Hidrolases/química , Diester Fosfórico Hidrolases/genética , Diester Fosfórico Hidrolases/metabolismo , Receptores de Ácidos Lisofosfatídicos/antagonistas & inibidores , Receptores de Ácidos Lisofosfatídicos/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Estearoil-CoA Dessaturase/antagonistas & inibidores , Estearoil-CoA Dessaturase/genética , Estearoil-CoA Dessaturase/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/agonistas , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo
8.
Environ Pollut ; 233: 596-603, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29107899

RESUMO

Arsenic induced senescence (AIS) has been identified in the population of West Bengal, India very recently. Also there is a high incidence of arsenic induced peripheral neuropathy (PN) throughout India. However, the epigenetic regulation of AIS and its contribution in arsenic induced PN remains unexplored. We recruited seventy two arsenic exposed and forty unexposed individuals from West Bengal to evaluate the role of senescence associated miRNAs (SA-miRs) in AIS and their involvement if any, in PN. The downstream molecules of the miRNA associated with the disease outcome, was also checked by immuoblotting. In vitro studies were conducted with HEK 293 cells and sodium arsenite exposure. Our results show that all the SA-miRs were upregulated in comparison to unexposed controls. miR-29a was the most significantly altered, highest expression being in the arsenic exposed group with PN, suggesting its association with the occurrence of PN. We looked for the expression of peripheral myelin protein 22 (PMP22), a specific target of miR-29a associated with myelination and found that both in vitro and in vivo results showed over-expression of the protein. Since this was quite contrary to miRNA regulation, we checked for intermediate players ß-catenin and GSK-3ß upon arsenic exposure which affects PMP22 expression. We found that ß-catenin was upregulated in vitro and was also highest in the arsenic exposed group with PN while GSK-3ß followed the reverse pattern. Our findings suggest that arsenic exposure alters the expression of SA-miRs and the mir-29a/beta catenin/PMP22 axis might be responsible for arsenic induced PN.


Assuntos
Arsênio/toxicidade , Poluentes Ambientais/toxicidade , Doenças do Sistema Nervoso Periférico/genética , Arsênio/análise , Intoxicação por Arsênico/epidemiologia , Epigênese Genética , Quinase 3 da Glicogênio Sintase , Células HEK293 , Humanos , Índia/epidemiologia , MicroRNAs/metabolismo , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Doenças do Sistema Nervoso Periférico/epidemiologia , Regulação para Cima , beta Catenina
9.
Liver Int ; 38(6): 1084-1094, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29164820

RESUMO

BACKGROUND & AIMS: Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) are common clinico-pathological conditions that affect millions of patients worldwide. In this study, the efficacy of saroglitazar, a novel PPARα/γ agonist, was assessed in models of NAFLD/NASH. METHODS & RESULTS: HepG2 cells treated with palmitic acid (PA;0.75 mM) showed decreased expression of various antioxidant biomarkers (SOD1, SOD2, glutathione peroxidase and catalase) and increased expression of inflammatory markers (TNFα, IL1ß and IL6). These effects were blocked by saroglitazar, pioglitazone and fenofibrate (all tested at 10µM concentration). Furthermore, these agents reversed PA-mediated changes in mitochondrial dysfunction, ATP production, NFkB phosphorylation and stellate cell activation in HepG2 and HepG2-LX2 Coculture studies. In mice with choline-deficient high-fat diet-induced NASH, saroglitazar reduced hepatic steatosis, inflammation, ballooning and prevented development of fibrosis. It also reduced serum alanine aminotransferase, aspartate aminotransferase and expression of inflammatory and fibrosis biomarkers. In this model, the reduction in the overall NAFLD activity score by saroglitazar (3 mg/kg) was significantly more prominent than pioglitazone (25 mg/kg) and fenofibrate (100 mg/kg). Pioglitazone and fenofibrate did not show any improvement in steatosis, but partially improved inflammation and liver function. Antifibrotic effect of saroglitazar (4 mg/kg) was also observed in carbon tetrachloride-induced fibrosis model. CONCLUSIONS: Saroglitazar, a dual PPARα/γ agonist with predominant PPARα activity, shows an overall improvement in NASH. The effects of saroglitazar appear better than pure PPARα agonist, fenofibrate and PPARγ agonist pioglitazone.


Assuntos
Biomarcadores/sangue , Fígado/patologia , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , PPAR alfa/agonistas , Fenilpropionatos/farmacologia , Pirróis/farmacologia , Alanina Transaminase/sangue , Animais , Aspartato Aminotransferases/sangue , Dieta Hiperlipídica , Fenofibrato/farmacocinética , Células Hep G2 , Humanos , Macrófagos do Fígado/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/patologia , Pioglitazona/farmacologia , Fator de Necrose Tumoral alfa/sangue
10.
J Cell Biochem ; 119(4): 3373-3383, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29130517

RESUMO

Increased metastasis and a precipitous recurrence contribute to the lethality of ovarian cancer (OC). Several molecular mechanisms including aberrant-splicing have been closely associated with the extent of cancer progression. Numerous gene transcripts are differentially spliced in cancer cells, CD44 being one of them. CD44 splice isoforms contribute to the aggressiveness and gain of stem-like properties in different cancer types, but their role in ovarian cancer remains to be elucidated. We observed augmented CD44 levels in human ovarian cancer patient samples correlated with enhanced expression of the mesenchymal spliced variant CD44s (standard) and a concurrent decrease in the epithelial variants (CD44v). Moreover, CD44s was upregulated upon TGFß1-induced EMT, which was mediated through the downregulation of the splicing factor, ESRP1. Furthermore, overexpression of this mesenchymal isoform in the OC cells induced EMT and invasion, followed by the gain of stem-like characteristics and chemoresistance. Since all these phenomena render lethality to this disease type, CD44s can be attributed for playing a major role in deregulated-splicing mediated ovarian cancer progression.


Assuntos
Processamento Alternativo , Receptores de Hialuronatos/genética , Receptores de Hialuronatos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Neoplasias Ovarianas/metabolismo , Proteínas de Ligação a RNA/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Invasividade Neoplásica , Células-Tronco Neoplásicas/metabolismo , Neoplasias Ovarianas/genética
11.
Cell Physiol Biochem ; 41(4): 1336-1345, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28427047

RESUMO

BACKGROUND/AIMS: The aggressive property of ovarian cancer (OC) in terms of epithelial-mesenchymal transition (EMT), proliferation and metastasis are of major concern. Different growth factors including TGFß are associated with regulating these molecular events but the underlying mechanisms remain unclear. The aim of this report is to decipher the regulation of EMT by co-activation of TGFß and Wnt signalling cascades in gaining malignancy. METHODS: The expression of the different components of signalling events were analyzed by QPCR, Western blot, Immunofluorescence microscopy and flow cytometry. ß-catenin promoter activity was checked by luciferase assay. RESULTS: We observed reduced EMT in ovarian cancer cells upon co-activation with TGFß1 and LiCl as shown by the expressions of epithelial/mesenchymal markers and the EMT promoting factor, Snail1, accompanied by decrease in the invasion and migration of the cells compared to individual pathway activation. A detailed study of the mechanism suggested reduction in the ß-catenin and p-GSK3b (Ser 9) levels to be the driving cause of this phenomenon, which was reversed upon co-activation with higher concentrations of LiCl. CONCLUSIONS: Therefore, tumourigenesis might be affected by the concentration of ligand/ growth factors for the respective signalling pathways activated in the tumour microenvironment and interaction between them might alter tumourigenesis.


Assuntos
Transição Epitelial-Mesenquimal/efeitos dos fármacos , Cloreto de Lítio/farmacologia , Proteínas de Neoplasias/metabolismo , Neoplasias Ovarianas/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos , Linhagem Celular Tumoral , Feminino , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/patologia , Fatores de Transcrição da Família Snail/metabolismo
12.
J Pineal Res ; 62(4)2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28247434

RESUMO

Lipid generates reactive oxygen species (ROS) in consequence to mitochondrial fission followed by inflammation in propagating hepatic fibrosis. The interaction of SIRT1/Mitofusin2 is critical for maintaining mitochondrial integrity and functioning, which is disrupted upon excess lipid infiltration during the progression of steatohepatitis. The complex interplay between hepatic stellate cells and steatotic hepatocytes is critically regulated by extracellular factors including increased circulating free fatty acids during fibrogenesis. Melatonin, a potent antioxidant, protects against lipid-mediated mitochondrial ROS generation. Lipotoxicity induces disruption of SIRT1 and Mitofusin2 interaction leading to mitochondrial morphological disintegration in hepatocytes. Further, fragmented mitochondria leads to mitochondrial permeability transition pore opening, cell cycle arrest and apoptosis and melatonin protects against all these lipotoxicity-mediated dysfunctions. These impaired mitochondrial dynamics also enhances the cellular glycolytic flux and reduces mitochondrial oxygen consumption rate that potentiates ROS production. High glycolytic flux generates metabolically unfavorable milieu in hepatocytes leading to inflammation, which is abrogated by melatonin. The melatonin-mediated protection against mitochondrial dysfunction was also observed in high-fat diet (HFD)-fed mice through restoration of enzymatic activities associated with respiratory chain and TCA cycle. Subsequently, melatonin reduces hepatic fat deposition and inflammation in HFD-fed mice. Thus, melatonin disrupts the interaction between steatotic hepatocyte and stellate cells, leading to the activation of the latter to abrogate collagen deposition. Altogether, the results of the current study document that the pharmacological intervention with low dose of melatonin could abrogate lipotoxicity-mediated hepatic stellate cell activation and prevent the fibrosis progression.


Assuntos
Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Cirrose Hepática/tratamento farmacológico , Cirrose Hepática/metabolismo , Melatonina/uso terapêutico , Mitocôndrias Hepáticas/efeitos dos fármacos , Mitocôndrias Hepáticas/patologia , Animais , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Dieta Hiperlipídica/efeitos adversos , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Masculino , Melatonina/farmacologia , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio
13.
Food Funct ; 8(4): 1577-1586, 2017 Apr 19.
Artigo em Inglês | MEDLINE | ID: mdl-28282095

RESUMO

Consumption of food that surpasses the metabolic necessity of the body leads to an epidemic condition termed obesity, which causes several metabolic disorders including oxidative damage. Dietary intervention can enlighten the mechanisms and therapeutics associated with these metabolic disorders. The reported studies related to diet include fat of different kinds and from different sources, however they lack dose response aspects. Our study highlighted the importance of dietary fat modification in modulating oxidative stress-induced glucose intolerance. Animals were maintained on a diet with a varied content of fat (30%/45%/60%) for 12 weeks and the 'withdrawal' group was fed a standard diet for another 10 weeks. The diet containing 60 energy% of fat displayed glucose intolerance, high ALT, low GSH levels and tissue-specific modulation of the prooxidant/antioxidant enzymatic activities in the liver/muscles. Prolonged sustenance of the 60 energy% fat containing diet-fed rats on standard diet led to the alteration of antioxidant activities, reversing the oxidative damage. Notably, the 'withdrawal' group displayed an organ-specific response towards dietary modification where the recovery of the antioxidant activities was observed to be much more pronounced in the liver as compared to the muscle. Further, we identified the differential expression of liver/muscle-specific genes associated with oxidative stress and mitochondrial biogenesis in response to the differing fat content. These genes can serve as markers for HFD-induced metabolic complications involving the liver/muscle. Altogether, our study has highlighted the novel area where obesity-induced oxidative stress linked alterations expressed diet and organ specific responses that are recovered by altering the dietary regimen. Future investigation of dietary modulation will open nascent avenues for developing therapeutic modalities addressing obesity-related metabolic complications.


Assuntos
Gorduras na Dieta/efeitos adversos , Intolerância à Glucose/genética , Mitocôndrias/genética , Estresse Oxidativo , Animais , Antioxidantes/metabolismo , Gorduras na Dieta/metabolismo , Intolerância à Glucose/etiologia , Intolerância à Glucose/metabolismo , Humanos , Fígado/metabolismo , Masculino , Mitocôndrias/metabolismo , Músculos/metabolismo , Obesidade/genética , Obesidade/metabolismo , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1/genética , Superóxido Dismutase-1/metabolismo
14.
Mol Oncol ; 11(5): 491-516, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28236660

RESUMO

Extravasation and metastatic progression are two main reasons for the high mortality rate associated with cancer. The metastatic potential of cancer cells depends on a plethora of metabolic challenges prevailing within the tumor microenvironment. To achieve higher rates of proliferation, cancer cells reprogram their metabolism, increasing glycolysis and biosynthetic activities. Just why this metabolic reprogramming predisposes cells towards increased oncogenesis remains elusive. The accumulation of myriad oncolipids in the tumor microenvironment has been shown to promote the invasiveness of cancer cells, with lysophosphatidic acid (LPA) being one such critical factor enriched in ovarian cancer patients. Cellular bioenergetic studies confirm that oxidative phosphorylation is suppressed and glycolysis is increased with long exposure to LPA in ovarian cancer cells compared with non-transformed epithelial cells. We sought to uncover the regulatory complexity underlying this oncolipid-induced metabolic perturbation. Gene regulatory networking using RNA-Seq analysis identified the oncogene ETS-1 as a critical mediator of LPA-induced metabolic alterations for the maintenance of invasive phenotype. Moreover, LPA receptor-2 specific PtdIns3K-AKT signaling induces ETS-1 and its target matrix metalloproteases. Abrogation of ETS-1 restores cellular bioenergetics towards increased oxidative phosphorylation and reduced glycolysis, and this effect was reversed by the presence of LPA. Furthermore, the bioenergetic status of LPA-treated ovarian cancer cells mimics hypoxia through induction of hypoxia-inducible factor-1α, which was found to transactivate ets-1. Studies in primary tumors generated in syngeneic mice corroborated the in vitro findings. Thus, our study highlights the phenotypic changes induced by the pro-metastatic factor ETS-1 in ovarian cancer cells. The relationship between enhanced invasiveness and metabolic plasticity further illustrates the critical role of metabolic adaptation of cancer cells as a driver of tumor progression. These findings reveal oncolipid-induced metabolic predisposition as a new mechanism of tumorigenesis and propose metabolic inhibitors as a potential approach for future management of aggressive ovarian cancer.


Assuntos
Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Lisofosfolipídeos/farmacologia , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Adenocarcinoma/genética , Animais , Linhagem Celular Tumoral , Feminino , Perfilação da Expressão Gênica , Redes Reguladoras de Genes/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Inibidores de Metaloproteinases de Matriz/farmacologia , Inibidores de Metaloproteinases de Matriz/uso terapêutico , Camundongos , Camundongos Endogâmicos BALB C , Mitocôndrias/metabolismo , Invasividade Neoplásica , Neoplasias Ovarianas/genética , Fosforilação Oxidativa/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Proteína Proto-Oncogênica c-ets-1/antagonistas & inibidores , Proteína Proto-Oncogênica c-ets-1/genética , Proteína Proto-Oncogênica c-ets-1/metabolismo , Análise de Sequência de RNA
15.
Tumour Biol ; 39(2): 1010428317694314, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28240052

RESUMO

Paclitaxel (Tx) is one of the first-line chemotherapeutic drugs used against lung cancer, but acquired resistance to this drug is a major challenge against successful chemotherapy. In this work, we have focused on the chronological changes of various cellular parameters and associated effect on Tx (10 nM) resistance development in A549 cell line. It was observed, at initial stage, the cell death percentage due to drug treatment had increased up to 20 days, and thereafter, it started declining and became completely resistant by 40 days. Expressions of ßIII tubulin and drug efflux pumps also increased over the period of resistance development. Changes in cellular autophagy and reactive oxygen species generation showed a biphasic pattern and increased gradually over the course of upto 20 days, thereafter declined gradually; however, their levels remained higher than untreated cells when resistance was acquired. Increase in extracellular acidification rates and oxygen consumption rates was found to be directly correlated with acquisition of resistance. The depolarisation of mitochondrial membrane potential was also biphasic; first, it increased with increase of cell death up to 20 days, thereafter, it gradually decreased to normal level along with resistance development. Increase in activity of catalase, glutathione peroxidase and glutathione content over these periods may attribute in bringing down the reactive oxygen species levels and normalisation of mitochondrial membrane potential in spite of comparatively higher reactive oxygen species production by the Tx-resistant cells.


Assuntos
Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/metabolismo , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Potencial da Membrana Mitocondrial/fisiologia , Paclitaxel/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Células A549 , Adenocarcinoma/patologia , Adenocarcinoma de Pulmão , Antineoplásicos Fitogênicos/farmacologia , Autofagia , Caspase 3/metabolismo , Ciclo Celular/fisiologia , Resistencia a Medicamentos Antineoplásicos , Metabolismo Energético , Humanos , Neoplasias Pulmonares/patologia , Microscopia de Fluorescência
16.
Cell Physiol Biochem ; 41(2): 795-805, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28214851

RESUMO

BACKGROUND/AIMS: Epithelial-to-mesenchymal transition (EMT) plays an essential role in the transition from early to invasive phenotype, however the underlying mechanisms still remain elusive. Herein, we propose a mechanism through which the class-III deacetylase SIRT1 regulates EMT in ovarian cancer (OC) cells. METHODS: Expression analysis was performed using Q-PCR, western blot, immunofluorescence and fluorescence-IHC study. Matrigel invasion assay was used for the invasion study. Morphological alterations were observed by phalloidin-staining. Co-immunoprecipitation study was performed to analyze protein-protein interaction. RESULTS: Overexpression of SIRT1-WT as well as Resveratrol-mediated SIRT1 activation antagonized the invasion of OC cells by suppressing EMT. SIRT1 deacetylates HIF1α, to inactivate its transcriptional activity. To further validate HIF1α inactivation, its target gene, i.e. ZEB1, an EMT-inducing factor was found to attenuate upon SIRT1 activation. To uncover the regulatory factor governing SIRT1 expression, lysophosphatidic acid (LPA), a highly enriched oncolipid in ascites/serum of OC patients, was found to down-regulate SIRT1 expression. Importantly, LPA was found to induce the mesenchymal switch in OC cells through suppression of SIRT1. Decreased level of SIRT1 was further validated in ovarian tissue samples of OC patients. CONCLUSION: We have identified a mechanism that relates SIRT1 down-regulation to LPA-induced EMT in OC cells and may open new arenas on developing novel anti-cancer therapeutics.


Assuntos
Transição Epitelial-Mesenquimal/efeitos dos fármacos , Lisofosfolipídeos/farmacologia , Sirtuína 1/metabolismo , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Feminino , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Microscopia de Fluorescência , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Sirtuína 1/antagonistas & inibidores , Sirtuína 1/genética
17.
Biochem Biophys Res Commun ; 479(4): 933-939, 2016 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-27702549

RESUMO

Insulin resistance (IR) is an important determinant of type-2 diabetes mellitus (T2DM). Free fatty acids (FFAs) induce IR by various mechanisms. A surfeit of circulating FFA leads to intra-myocellular lipid accumulation that induces mitochondrial ROS generation and worsens IR. However, the molecular mechanisms behind are unclear. We identified thioredoxin interacting protein (TxNIP), which is overexpressed in T2DM, to be a promoter of ROS-induced IR. We observed upregulation of TxNIP upon palmitate treatment in skeletal muscle cells that led to ROS generation and Glut-4 downregulation resulting in impaired glucose-uptake. FFA-induced overexpression of TxNIP gene was mediated through the activation of its bona-fide trans activator, ChREBP. Further, Palmitate-induced impairment in AMPK-SIRT-1 pathway resulted in overexpression of ChREBP. While Fenofibrate, abrogated PA-induced TxNIP expression and ROS generation in skeletal muscle cells, Saroglitazar, a dual PPARα/γ-agonist, not only inhibited PA-induced TXNIP expression but also led to greater improvement in glucose uptake. Taken together, TxNIP appears to be an important factor in FFA-induced ROS generation and IR in skeletal muscle cells, which can be modulated for the management of this complex disorder.


Assuntos
Proteínas de Transporte/metabolismo , Glucose/metabolismo , Fibras Musculares Esqueléticas/efeitos dos fármacos , Fibras Musculares Esqueléticas/metabolismo , Ácido Palmítico/farmacologia , Tiorredoxinas/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos , Transporte Biológico Ativo/efeitos dos fármacos , Proteínas de Transporte/genética , Linhagem Celular , Diabetes Mellitus Tipo 2/metabolismo , Ácidos Graxos não Esterificados/metabolismo , Fenofibrato/farmacologia , Transportador de Glucose Tipo 4/metabolismo , Resistência à Insulina , Camundongos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fenilpropionatos/farmacologia , Pirróis/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Sirtuína 1/metabolismo , Tiorredoxinas/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
18.
Cell Physiol Biochem ; 37(4): 1315-28, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26488284

RESUMO

BACKGROUNDS/AIMS: The lipid induced insulin resistance is a major pathophysiologic mechanism underlying glucose intolerance of varying severity. PPARα-agonists are proven as effective hypolipidemic agents. The aim of this study was to see if impaired glucose uptake in palmitate treated myotubes is reversed by fenofibrate. METHODS: Palmitate-treated myotubes were used as a model for insulin resistance, impaired glucose uptake, fatty acid oxidation and ceramide synthesis. mRNA levels of CPT1 and CPT2 were determined by PCR array and Q-PCR. RESULTS: The incubation of myotubes with 750 uM palmitate not only reduced glucose uptake but also impaired fatty acid oxidation and cytosolic ceramide accumulation. Palmitate upregulated CPT1b expression in L6 myotubes, while CPT2 expression level remained unchanged. The altered stoichiometric ratio between the two CPT isoforms led to reduced fatty acid oxidation (FAO), ceramide accumulation and impaired glucose uptake, whereas administration of 200 µM fenofibrate significantly reversed the above abnormalities by increasing CPT2 mRNA levels and restoring CPT1b to CPT2 ratio. CONCLUSION: Palmitate-induced alteration in the stoichiometric ratio of mitochondrial CPT isoforms leads to incomplete FAO and enhanced cytosolic ceramide accumulation that lead to insulin resistance. Fenofibrate ameliorated insulin resistance by restoring the altered stoichiometry by upregulating CPT2 and preventing, cytoplasmic ceramide accumulation.


Assuntos
Ceramidas/metabolismo , Ácidos Graxos/metabolismo , Fenofibrato/farmacologia , Glucose/metabolismo , Hipolipemiantes/farmacologia , Palmitatos/farmacologia , Animais , Carnitina O-Palmitoiltransferase/genética , Carnitina O-Palmitoiltransferase/metabolismo , Linhagem Celular , Citosol/metabolismo , Dieta Hiperlipídica , Transportador de Glucose Tipo 4/metabolismo , Resistência à Insulina , Peroxidação de Lipídeos , Camundongos , Mitocôndrias/metabolismo , Músculo Esquelético/citologia , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , RNA Mensageiro/metabolismo , Ratos
19.
Mol Cancer ; 14: 162, 2015 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-26298390

RESUMO

BACKGROUND: Most ovarian cancers are highly invasive in nature and the high burden of metastatic disease make them a leading cause of mortality among all gynaecological malignancies. The homeodomain transcription factor, PITX2 is associated with cancer in different tissues. Our previous studies demonstrated increased PITX2 expression in human ovarian tumours. Growing evidence linking activation of TGF-ß pathway by homeodomain proteins prompted us to look for the possible involvement of this signalling pathway in PITX2-mediated progression of ovarian cancer. METHODS: The status of TGF-ß signalling in human ovarian tissues was assessed by immunohistochemistry. The expression level of TGFB/INHBA and other invasion-associated genes was measured by quantitative-PCR (Q-PCR) and Western Blot after transfection/treatments with clones/reagents in normal/cancer cells. The physiological effect of PITX2 on invasion/motility was checked by matrigel invasion and wound healing assay. The PITX2- and activin-induced epithelial-mesenchymal transition (EMT) was evaluated by Q-PCR of respective markers and confocal/phase-contrast imaging of cells. RESULTS: Human ovarian tumours showed enhanced TGF-ß signalling. Our study uncovers the PITX2-induced expression of TGFB1/2/3 as well as INHBA genes (p < 0.01) followed by SMAD2/3-dependent TGF-ß signalling pathway. PITX2-induced TGF-ß pathway regulated the expression of invasion-associated genes, SNAI1, CDH1 and MMP9 (p < 0.01) that accounted for enhanced motility/invasion of ovarian cancers. Snail and MMP9 acted as important mediators of PITX2-induced invasiveness of ovarian cancer cells. PITX2 over-expression resulted in loss of epithelial markers (p < 0.01) and gain of mesenchymal markers (p < 0.01) that contributed significantly to ovarian oncogenesis. PITX2-induced INHBA expression (p < 0.01) contributed to EMT in both normal and ovarian cancer cells. CONCLUSIONS: Overall, our findings suggest a significant contributory role of PITX2 in promoting invasive behaviour of ovarian cancer cells through up-regulation of TGFB/INHBA. We have also identified the previously unknown involvement of activin-A in promoting EMT. Our work provides novel mechanistic insights into the invasive behavior of ovarian cancer cells. The extension of this study have the potential for therapeutic applications in future.


Assuntos
Proteínas de Homeodomínio/genética , Subunidades beta de Inibinas/genética , Neoplasias Ovarianas/genética , Fatores de Transcrição/genética , Fator de Crescimento Transformador beta/genética , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/metabolismo , Humanos , Subunidades beta de Inibinas/biossíntese , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Neoplasias Ovarianas/patologia , Transdução de Sinais , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/biossíntese
20.
Mol Carcinog ; 54(9): 800-9, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24665044

RESUMO

Arsenic (As) induces pre-malignant and malignant dermatological lesions, non-dermatological health effects and cancers in humans. Senescence involves telomere length changes and acquisition of senescence-associated secretory phenotype (SASP), which promotes carcinogenesis. Though in vitro studies have shown that As induces senescence, population based studies are lacking. We investigated the arsenic-induced senescence, telomere length alteration and its contribution towards development of As-induced skin cancer. The study participants included 60 each of As-exposed individuals with skin lesion (WSL), without skin lesions (WOSL) and 60 unexposed controls. Exposure assessment of drinking water and urine was done. SA ß-gal activity, ELISA, and quantification of senescence proteins, alternative lengthening of telomere (ALT) associated proteins and telomerase activity were performed. Relative telomere length (RTL) was determined by qPCR. A significantly higher number of senescent cells, over-expression of p53 and p21 were observed in the As-exposed individuals when compared to unexposed. SASP markers, MMP-1/MMP-3 were significantly higher in the WSL but not IL-6/IL-8. A significant increase of RTL was observed in the WSL group, which was telomerase-independent but exhibited an over-expression of ALT associated proteins TRF-1 and TRF-2 with higher increase in TRF-2. An increased risk for developing As-induced skin lesions was found for individuals having RTL greater than 0.827 (odds ratio, 13.75; 95% CI: 5.66-33.41; P < 0.0001). Arsenic induces senescence in vivo, but the SASP markers are not strictly over-expressed in the As-induced skin lesion group, whereas telomerase-independent elongation of telomere length might be useful for predicting the risk of development of As-induced skin lesions.


Assuntos
Envelhecimento/efeitos dos fármacos , Arsênio/toxicidade , Água Potável/efeitos adversos , Telômero/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Adulto , Carcinogênese/induzido quimicamente , Carcinogênese/metabolismo , Carcinogênese/patologia , Estudos de Casos e Controles , Água Potável/análise , Feminino , Humanos , Índia/epidemiologia , Masculino , Pessoa de Meia-Idade , Transdução de Sinais/efeitos dos fármacos , Pele/efeitos dos fármacos , Pele/metabolismo , Pele/patologia , Dermatopatias/induzido quimicamente , Dermatopatias/epidemiologia , Dermatopatias/metabolismo , Dermatopatias/patologia , Telomerase/metabolismo , Telômero/patologia , Proteína Supressora de Tumor p53/metabolismo
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