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1.
Fitoterapia ; 175: 105861, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38354824

RESUMO

Humulus lupulus extracts have in their composition different molecules, such as polyphenols, α-acids, ß-acids, and hydrocarbons, which contribute to the plant's medicinal properties. These molecules are associated with antimicrobial, antioxidant and anti-inflammatory activities. OBJECTIVE: This work focuses on the evaluation of H. lupulus biological activities, with the aim of evaluating its potential for inclusion in cosmetic formulations. METHODS: Two distinct aqueous extracts and two hydrolates obtained via hydrodistillation were evaluated. These include the flower parts (FE, FH) and the mix of aboveground parts (ME, MH). The chemical profiles for both aqueous extracts and hydrolates were identified by high performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS). Antimicrobial, antioxidant, cytotoxicity, and anti-inflammatory activity were tested in vitro using standard methods. RESULTS: Rutin was the major compound found in FE (40.041 µg mg-1 of extract) and ME (2.909 µg mg-1 of extract), while humulenol II was the most abundant compound in hydrolates (FH: 20.83%; MH: 46.80%). Furthermore, FE was able to inhibit the growth of Staphylococcus aureus and Staphylococcus epidermis with MIC values of 50% and 25% (v/v), respectively. FH showed the same effect in Staphylococcus aureus (50% v/v). FH evidenced poor antioxidant potential in DPPH scavenging test and demonstrated significant antioxidant and anti-inflammatory effects by reducing (***p < 0.001) intracellular reactive oxygen species (ROS), NO (nitric oxide) levels (***p < 0.001) and cyclooxygenase-2 (COX-2) protein expression (***p < 0.001) in lipopolysaccharide (LPS)-stimulated macrophages. Nevertheless, it is important to note that FH exhibited cytotoxicity at high concentrations in 3T3 fibroblasts and RAW 264.7 macrophages. CONCLUSION: The studied H. lupulus aqueous extracts and hydrolates revealed that FH stands out as the most promising bioactive source for cosmetic formulations. However, future research addressing antimicrobial activity is necessary to confirm its potential incorporation into dermatological and cosmetic formulations.


Assuntos
Anti-Inflamatórios , Antioxidantes , Cosméticos , Humulus , Extratos Vegetais , Humulus/química , Extratos Vegetais/farmacologia , Extratos Vegetais/química , Antioxidantes/farmacologia , Anti-Inflamatórios/farmacologia , Camundongos , Animais , Células RAW 264.7 , Flores/química , Anti-Infecciosos/farmacologia , Anti-Infecciosos/química , Compostos Fitoquímicos/farmacologia , Compostos Fitoquímicos/isolamento & purificação , Macrófagos/efeitos dos fármacos , Testes de Sensibilidade Microbiana
3.
Pathol Res Pract ; 240: 154181, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36327818

RESUMO

BACKGROUND: Androgens, the known drivers of prostate cancer (PCa), have been indicated as important metabolic regulators with a relevant role in stimulating lipid metabolism. Also, the relationship between obesity and the aggressiveness of PCa has been established. However, it is unknown if the androgenic hormonal environment may alter the response of PCa cells to lipid availability. PURPOSE: The present study evaluated the effect of 5α-dihydrotestosterone (DHT) in regulating lipid metabolism, and the interplay between this hormone and low-density lipoprotein (LDL)-cholesterol in modulating PCa cells fate. METHODS: Non-neoplastic and neoplastic PCa cells were treated with 10 nM DHT, and the expression of fatty acids transporter, fatty acid synthase (FASN), and carnitine palmitoyltransferase 1A (CPT1A) evaluated. PCa cells were also exposed to LDL (100 µg/ml) in the presence or absence of DHT. RESULTS: Treatment with DHT upregulated the expression of FASN and CPT1A in androgen-sensitive PCa cells. In contrast, LDL supplementation suppressed FASN expression regardless of the presence of DHT, whereas augmenting CPT1A levels. Our results also showed that LDL-cholesterol increased PCa cells viability, proliferation, and migration dependently on the presence of DHT. Moreover, LDL and DHT synergistically enhanced the accumulation of lipid droplets in PCa cells. CONCLUSIONS: The obtained results show that androgens deregulate lipid metabolism and enhance the effects of LDL increasing PCa cells viability, proliferation and migration. The present findings support clinical data linking obesity with PCa and first implicate androgens in this relationship. Also, they sustain the application of pharmacological approaches targeting cholesterol availability and androgens signaling simultaneously.


Assuntos
Androgênios , Neoplasias da Próstata , Masculino , Humanos , Androgênios/farmacologia , LDL-Colesterol/uso terapêutico , Neoplasias da Próstata/metabolismo , Di-Hidrotestosterona/farmacologia , Obesidade , Receptores Androgênicos/metabolismo
4.
Molecules ; 26(10)2021 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-34063349

RESUMO

Sweet cherries (Prunus avium L.) are among the most appreciated fruits worldwide because of their organoleptic properties and nutritional value. The accurate phytochemical composition and nutritional value of sweet cherries depends on the climatic region, cultivar, and bioaccessibility and bioavailability of specific compounds. Nevertheless, sweet cherry extracts are highly enriched in several phenolic compounds with relevant bioactivity. Over the years, technological advances in chemical analysis and fields as varied as proteomics, genomics and bioinformatics, have allowed the detailed characterization of the sweet cherry bioactive phytonutrients and their biological function. In this context, the effect of sweet cherries on suppressing important events in the carcinogenic process, such as oxidative stress and inflammation, was widely documented. Interestingly, results from our research group and others have widened the action of sweet cherries to many hallmarks of cancer, namely metabolic reprogramming. The present review discusses the anticarcinogenic potential of sweet cherries by addressing their phytochemical composition, the bioaccessibility and bioavailability of specific bioactive compounds, and the existing knowledge concerning the effects against oxidative stress, chronic inflammation, deregulated cell proliferation and apoptosis, invasion and metastization, and metabolic alterations. Globally, this review highlights the prospective use of sweet cherries as a dietary supplement or in cancer treatment.


Assuntos
Antineoplásicos Fitogênicos/química , Compostos Fitoquímicos/química , Prunus avium/química , Antineoplásicos Fitogênicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Suplementos Nutricionais , Humanos , Estresse Oxidativo/efeitos dos fármacos , Compostos Fitoquímicos/farmacologia
5.
Cell Oncol (Dordr) ; 44(2): 385-403, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33464483

RESUMO

PURPOSE: Resistance to androgen-deprivation therapies and progression to so-called castrate-resistant prostate cancer (CRPC) remain challenges in prostate cancer (PCa) management and treatment. Among other alterations, CRPC has been associated with metabolic reprogramming driven by androgens. Here, we investigated the role of androgens in regulating glutaminolysis in PCa cells and determined the relevance of this metabolic route in controlling the survival and growth of androgen-sensitive (LNCaP) and CRPC (DU145 and PC3) cells. METHODS: PCa cells (LNCaP, DU145 and PC3) and 3-month old rats were treated with 5α-dihydrotestosterone (DHT). Alternatively, LNCaP cells were exposed to the glutaminase inhibitor BPTES, alone or in combination with the anti-androgen bicalutamide. Biochemical, Western blot and extracellular flux assays were used to evaluate the viability, proliferation, migration and metabolism of PCa cells in response to DHT treatment or glutaminase inhibition. RESULTS: We found that DHT up-regulated the expression of the glutamine transporter ASCT2 and glutaminase, both in vitro in LNCaP cells and in vivo in rat prostate cells. BPTES diminished the viability and migration of PCa cells, while increasing caspase-3 activity. CRPC cells were found to be more dependent on glutamine and more sensitive to glutaminase inhibition. BPTES and bicalutamide co-treatment had an additive effect on suppressing LNCaP cell viability. Finally, we found that inhibition of glutaminolysis differentially affected glycolysis and lipid metabolism in both androgen-sensitive and CRPC cells. CONCLUSION: Our data reveal glutaminolysis as a central metabolic route controlling PCa cell fate and highlight the relevance of targeting glutaminase for CRPC treatment.


Assuntos
Di-Hidrotestosterona/farmacologia , Glutamina/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Sistema ASC de Transporte de Aminoácidos/genética , Sistema ASC de Transporte de Aminoácidos/metabolismo , Androgênios/farmacologia , Anilidas/farmacologia , Animais , Caspase 3/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glucose/metabolismo , Glutaminase/metabolismo , Glicólise/efeitos dos fármacos , Humanos , Ácido Láctico/biossíntese , Metabolismo dos Lipídeos/efeitos dos fármacos , Masculino , Antígenos de Histocompatibilidade Menor/genética , Antígenos de Histocompatibilidade Menor/metabolismo , Modelos Biológicos , Proteínas de Neoplasias/metabolismo , Nitrilas/farmacologia , Neoplasias de Próstata Resistentes à Castração/patologia , Ratos , Sulfetos/farmacologia , Tiadiazóis/farmacologia , Compostos de Tosil/farmacologia
6.
Med Res Rev ; 41(3): 1499-1538, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33274768

RESUMO

Prostate cancer (PCa), one of the most commonly diagnosed cancers worldwide, still presents important unmet clinical needs concerning treatment. In the last years, the metabolic reprogramming and the specificities of tumor cells emerged as an exciting field for cancer therapy. The unique features of PCa cells metabolism, and the activation of specific metabolic pathways, propelled the use of metabolic inhibitors for treatment. The present work revises the knowledge of PCa metabolism and the metabolic alterations that underlie the development and progression of the disease. A focus is given to the role of bioenergetic sources, namely, glucose, lipids, and glutamine sustaining PCa cell survival and growth. Moreover, it is described as the action of oncogenes/tumor suppressors and sex steroid hormones in the metabolic reprogramming of PCa. Finally, the status of PCa treatment based on the inhibition of metabolic pathways is presented. Globally, this review updates the landscape of PCa metabolism, highlighting the critical metabolic alterations that could have a clinical and therapeutic interest.


Assuntos
Neoplasias da Próstata , Humanos , Masculino , Redes e Vias Metabólicas , Oncogenes , Neoplasias da Próstata/tratamento farmacológico
7.
Theriogenology ; 157: 472-482, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32898822

RESUMO

Age-related changes, namely the increase in oxidative stress (OS) with the consequent sperm damage, result in decreased male fertility. Regucalcin (RGN) is a Ca2+-binding protein that has been shown to have beneficial effects on spermatogenesis by suppressing OS and chemical/radiation-induced damage. This work aims to evaluate whether RGN overexpression reduces the ageing-associated decline of male reproductive function. Sperm and testicular function analysis were performed in young-adult and senescent transgenic rats overexpressing RGN (Tg-RGN) comparatively with their wild-type (Wt) littermates. The gonadosomatic index (GI), tubular differentiation index and the expression levels of RGN and other proliferation regulators were evaluated. Moreover, the sperm parameters, OS analysis and immunolocalization of RGN were assessed, as well as morphometric evaluation of epididymal tubules. Both GI and sperm counts were reduced in the senescent Wt rats, but maintained in the Tg-RGN. Also, the levels of stem cell factor (SCF), c-Kit, and Akt were maintained in the testis of aged Tg-RGN rats, suggesting that the normal spermatogenic output was preserved over time in these animals, an effect not observed in Wt. Senescent Tg-RGN rats also presented lower sperm lipid peroxidation and total oxidant status relative to the Wt. Furthermore, aged Tg-RGN rats displayed higher sperm viability, higher frequency of sperm with normal morphology, and reduced incidence of head and neck/midpiece defects when compared with Wt, which may be a consequence of the lower OS levels found in the sperm of these animals. Interestingly, RGN expression increased with ageing in sperm, being mainly localized in the acrosome. Altogether, these findings indicate that the modulation of RGN levels may alleviate the age-related decline in sperm quality and testicular function.


Assuntos
Proteínas de Ligação ao Cálcio , Peptídeos e Proteínas de Sinalização Intracelular , Envelhecimento , Animais , Proteínas de Ligação ao Cálcio/genética , Hidrolases de Éster Carboxílico , Masculino , Estresse Oxidativo , Ratos , Ratos Sprague-Dawley , Contagem de Espermatozoides/veterinária , Espermatozoides/metabolismo , Testículo/metabolismo
8.
Nutr Cancer ; 72(6): 917-931, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31507215

RESUMO

The present work evaluated the anticancer properties of sweet cherry (Prunus avium) extract on human prostate cells. Several sweet cherry cultivars from Fundão (Portugal) were methanol-extracted and their phytochemical composition characterized. The Saco "late harvest" extract was highly-enriched in anthocyanins and selected for use in biological assays. Non-neoplastic (PNT1A) and neoplastic (LNCaP and PC3) human prostate cells were treated with 0-2,000 µg/ml of extract for 48-96 h. Cell viability was evaluated by the MTT assay. Apoptosis, oxidative stress, and glycolytic metabolism were assessed by Western blotting and enzymatic assays. Glucose consumption and lactate production were measured spectrophotometrically. Saco cherry extract diminished the viability of neoplastic and non-neoplastic cells, whereas enhancing apoptosis in LNCaP. Cherry extract-treatment also diminished oxidative damage and suppressed glycolytic metabolism in LNCaP cells. These findings widened the knowledge on the mechanisms by which cherry extract modulate cell physiology, demonstrating their broad action over the hallmarks of cancer.


Assuntos
Neoplasias , Prunus avium , Antocianinas/farmacologia , Apoptose , Humanos , Masculino , Próstata
9.
Eur J Med Chem ; 171: 343-363, 2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-30928707

RESUMO

The last decade has witnessed the peculiarities of metabolic reprogramming in tumour onset and progression, and their relevance in cancer therapy. Also, it has been indicated that the metastatic process may depend on the metabolic rewiring and adaptation of cancer cells to the pressure of tumour microenvironment and limiting nutrient availability. The present review gatherers the existent knowledge on the influence of tumour microenvironment and metabolic routes driving metastasis. A focus will be given to glycolysis, fatty acid metabolism, glutaminolysis, and amino acid handling. In addition, the role of metabolic waste driving metastasization will be explored. Finally, we discuss the status of cancer treatment approaches targeting metabolism. This knowledge revision will highlight the critical metabolic targets in metastasis and the chemicals already used in preclinical studies and clinical trials, providing clues that would be further exploited in medicinal chemistry research.


Assuntos
Antineoplásicos/farmacologia , Neoplasias/tratamento farmacológico , Antineoplásicos/química , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Neoplasias/metabolismo , Relação Estrutura-Atividade , Microambiente Tumoral/efeitos dos fármacos
10.
Life Sci ; 218: 274-283, 2019 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-30605651

RESUMO

AIMS: The tyrosine kinase inhibitor imatinib has been used in prostate cancer treatment with outcomes that did not follow the in vitro findings. The glycolytic environment has been shown to influence the efficacy of anti-cancer drugs. This study aimed to evaluate the effect of imatinib on cell viability, apoptosis, and metabolism in cell line models of castrate-resistant prostate cancer (CRPC) under hyperglycemic and hypoglycemic conditions. MAIN METHODS: DU145 and PC3 CRPC cell lines were exposed to 20 µM imatinib under 5 mM (hypoglycemia) or 30 mM glucose (hyperglycemia) for 48-72 h. Cell viability was assessed by the MTS assay. The expression of apoptosis regulators and glycolytic metabolism-related proteins was analysed by Western blot, and the activity of caspase-3 and lactate dehydrogenase (LDH) was determined spectrophotometrically. Glucose consumption and lactate production were determined using biochemical assays. KEY FINDINGS: Imatinib decreased CRPC cells viability, whereas increasing apoptosis; effects only observed in hyperglycemic conditions. Glucose consumption and lactate production were significantly increased in imatinib-treated DU145 and PC3 cells, and independently of glucose availability. Accordingly, LDH expression and activity were significantly increased in response to imatinib. SIGNIFICANCE: Higher glucose availability improved the effectiveness of imatinib suppressing survival and growth of CRPC cells. It was also shown that imatinib treatment stimulated the glycolytic metabolism of CRPC cells. This study first demonstrated that a glucose-enriched environment intensifies the effect of imatinib, which stimulates the interest for testing this compound into the clinical setting, namely in hyperglycemia conditions (diabetic patients) or in co-administration with inhibitors of glycolytic metabolism.


Assuntos
Glucose/metabolismo , Hiperglicemia/patologia , Mesilato de Imatinib/farmacologia , Ácido Láctico/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Inibidores de Proteínas Quinases/farmacologia , Apoptose , Proliferação de Células , Glicólise , Humanos , Hiperglicemia/tratamento farmacológico , Hiperglicemia/metabolismo , Masculino , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/metabolismo , Células Tumorais Cultivadas
11.
Sci Rep ; 8(1): 10321, 2018 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-29985416

RESUMO

Sertoli cells (SCs) possess the unparalleled ability to provide the germ line with growth factors and nutrients. Although SCs can oxidize amino acids, e.g., glutamine, they mostly metabolize glucose, producing high amounts of lactate, the germ cells preferential substrate. Regucalcin (RGN) is a calcium-binding protein that has been indicated as a regulator of cell metabolism. In this study, we investigated glucose and glutamine handling in the SCs of transgenic rats overexpressing RGN (Tg-RGN) comparatively with wild-type (Wt) littermates. Primary SCs isolated from adult Tg-RGN animals and maintained in culture for 24 hours, produced and exported more lactate, despite consuming less glucose. These observations were underpinned by increased expression of alanine transaminase, and augmented glutamine consumption, suggesting that alternative routes are contributing to the enhanced lactate production in the SCs of Tg-RGN rats. Moreover, lactate seems to be used by germ cells, with diminished apoptosis being detected in the seminiferous tubules of Tg-RGN animals cultured ex vivo. The obtained results showed a distinct metabolism in the SCs of Wt and Tg-RGN rats widening the roles assigned to RGN in spermatogenesis. These findings also highlighted the plasticity of SCs metabolism, a feature that would be exploited in the context of male infertility.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Glucose/metabolismo , Glutamina/metabolismo , Ácido Láctico/metabolismo , Animais , Apoptose , Proteínas de Ligação ao Cálcio/genética , Células Cultivadas , Transportador de Glucose Tipo 2/genética , Transportador de Glucose Tipo 2/metabolismo , Infertilidade Masculina/metabolismo , Infertilidade Masculina/patologia , Masculino , Fosfofrutoquinase-1/genética , Fosfofrutoquinase-1/metabolismo , Ratos , Ratos Sprague-Dawley , Ratos Transgênicos , Túbulos Seminíferos/metabolismo , Células de Sertoli/citologia , Células de Sertoli/metabolismo , Espermatogênese
12.
J Appl Toxicol ; 37(2): 159-166, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27109168

RESUMO

Regucalcin (RGN) is a calcium (Ca2+ )-binding protein with multiple physiological roles and has also been linked to the suppression of oxidative stress. It is widely known that oxidative stress adversely affects spermatogenesis, disrupting the development of germ cells, and interfering with sperm function. The present study aims to analyze the role of RGN modulating testicular oxidative stress. To address this issue, seminiferous tubules (SeT) from transgenic rats overexpressing RGN (Tg-RGN) and wild-type (WT) were cultured ex vivo for 24 h in the presence/absence of pro-oxidant stimuli, tert-butyl hydroperoxide (TBHP, 250 and 500 µM) and cadmium chloride (Cd, 10 and 20 µM). Noteworthy, SeT from Tg-RGN animals displayed a significantly higher antioxidant capacity and diminished levels of thiobarbituric acid reactive substances relatively to their WT counterparts, both in control and experimental conditions. Regarding the antioxidant defense systems, a significant increase in the activity of glutathione-S-transferase was found in the SeT of Tg-RGN whereas no differences were observed in superoxide dismutase activity throughout experimental conditions. The activity of apoptosis executioner caspase-3 was significantly increased in the SeT of WT rats treated with 250 µM of TBHP or 10 µM of Cd, an effect not seen in Tg-RGN animals. These results showed that the SeT of Tg-RGN animals displayed lower levels of oxidative stress and increased antioxidant defenses, exhibiting protection against oxidative damage and apoptosis. Moreover, the present findings support the antioxidant role of RGN in spermatogenesis, which may be an important issue of further research in the context of male infertility. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Cádmio/toxicidade , Proteínas de Ligação ao Cálcio/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Estresse Oxidativo/efeitos dos fármacos , Testículo/efeitos dos fármacos , terc-Butil Hidroperóxido/toxicidade , Animais , Antioxidantes/metabolismo , Proteínas de Ligação ao Cálcio/genética , Hidrolases de Éster Carboxílico , Técnicas In Vitro , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Ratos Sprague-Dawley , Ratos Transgênicos , Túbulos Seminíferos/efeitos dos fármacos , Túbulos Seminíferos/metabolismo , Testículo/metabolismo
13.
Transgenic Res ; 25(2): 139-48, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26553531

RESUMO

Regucalcin (RGN) is a calcium-binding protein underexpressed in human prostate cancer cases, and it has been associated with the suppression of cell proliferation and the regulation of several metabolic pathways. On the other hand, it is known that the metabolic reprogramming with augmented glycolytic metabolism and enhanced proliferative capability is a characteristic of prostate cancer cells. The present study investigated the influence of RGN on the glycolytic metabolism of rat prostate by comparing transgenic adult animals overexpressing RGN (Tg-RGN) with their wild-type counterparts. Glucose consumption was significantly decreased in the prostate of Tg-RGN animals relatively to wild-type, and accompanied by the diminished expression of glucose transporter 3 and glycolytic enzyme phosphofructokinase. Also, prostates of Tg-RGN animals displayed lower lactate levels, which resulted from the diminished expression/activity of lactate dehydrogenase. The expression of the monocarboxylate transporter 4 responsible for the export of lactate to the extracellular space was also diminished with RGN overexpression. These results showed the effect of RGN in inhibiting the glycolytic metabolism in rat prostate, which was underpinned by a reduced cell proliferation index. The present findings also suggest that the loss of RGN may predispose to a hyper glycolytic profile and fostered proliferation of prostate cells.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Proliferação de Células/genética , Glucose/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Próstata/metabolismo , Animais , Apoptose/genética , Proteínas de Ligação ao Cálcio/biossíntese , Hidrolases de Éster Carboxílico , Regulação da Expressão Gênica , Glucose/genética , Transportador de Glucose Tipo 3/biossíntese , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/biossíntese , Masculino , Transportadores de Ácidos Monocarboxílicos/biossíntese , Proteínas Musculares/biossíntese , Fosfofrutoquinase-1/biossíntese , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Ratos , Ratos Transgênicos
14.
Biochem Pharmacol ; 99: 73-87, 2016 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-26592659

RESUMO

The development of prostate cancer (PCa) is intimately associated with the hormonal environment, and the sex steroids estrogens have been implicated in prostate malignancy. However, if some studies identified estrogens as causative agents of PCa, others indicated that these steroids have a protective role counteracting prostate overgrowth. The tyrosine kinase receptor c-KIT and its ligand, the stem cell factor (SCF), have been associated with the control of cell proliferation/apoptosis and prostate carcinogenesis, and studies show that estrogens regulate their expression in different tissues, though, in the case of prostate this remains unknown. The present study aims to evaluate the role of 17ß-estradiol (E2) in regulating the expression of SCF/c-KIT in human prostate cell lines and rat prostate, and to investigate the consequent effects on prostate cell proliferation and apoptosis. qPCR, Western Blot, and immuno(cito)histochemistry analysis showed that E2-treatment decreased the expression of SCF and c-KIT both in human prostate cells and rat prostate. Furthermore, the diminished expression of SCF/c-KIT was underpinned by the diminished prostate weight and reduced proliferation index. On the other hand, the results of TUNEL labelling, the increased activity of caspase-3, and the augmented expression of caspase-8 and Fas system in the prostate of E2-treated animals indicated augmented apoptosis in response to E2. The obtained results demonstrated that E2 down-regulated the expression of SCF/c-KIT system in prostate cells, which was associated with antiproliferative and proapoptotic effects. Moreover, these findings support the protective role of estrogens in PCa and open new perspectives on the application of estrogen-based therapies.


Assuntos
Apoptose/fisiologia , Proliferação de Células/fisiologia , Estrogênios/farmacologia , Próstata/metabolismo , Proteínas Proto-Oncogênicas c-kit/metabolismo , Fator de Células-Tronco/metabolismo , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Humanos , Masculino , Próstata/citologia , Próstata/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-kit/antagonistas & inibidores , Ratos , Ratos Wistar , Fator de Células-Tronco/antagonistas & inibidores
15.
J Cancer Res Clin Oncol ; 142(1): 5-16, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26048031

RESUMO

PURPOSE: The present study aims to investigate the role of androgens in controlling the glycolytic metabolism and lactate efflux in prostate cancer (PCa) cells. METHODS: Androgen-responsive LNCaP cells were treated with 5α-dihydrotestosterone (DHT, 10 nM) for 12-48 h, and their glycolytic metabolism, lactate production and viability were analyzed. Intracellular and extracellular levels of glucose and lactate were determined spectrophotometrically, and the expression of glucose transporters (GLUT1/GLUT3), phosphofructokinase 1, lactate dehydrogenase (LDH) and monocarboxylate transporter (MCT4) was analyzed by real-time PCR and Western blot. The enzymatic activity of LDH was determined by means of a colorimetric assay. Experiments were reproduced in androgen-non-responsive DU145 and PC3 cells. RESULTS: Androgens stimulated glucose consumption in LNCaP cells by increasing the expression of GLUT3, GLUT1 and PFK, which was underpinned by increased cell viability. Accordingly, lactate production by LNCaP cells was enhanced upon DHT stimulation as evidenced by the increased levels of lactate found in cell culture medium. Although LDH enzymatic activity decreased in LNCaP cells treated with DHT, the expression of MCT4 was significantly increased with androgenic treatment, which sustains the increase on lactate export. Glucose consumption and the expression of GLUTs and PFK remained unchanged in DHT-treated DU145 and PC3 cells. CONCLUSIONS: The results obtained establish androgens as modulators of glycolytic metabolism in PCa cells by stimulating glucose consumption, as well as the production and export of lactate, which may represent a crucial issue-driven prostate tumor development. These findings also highlight the importance of PCa therapies targeting AR and metabolism-related proteins.


Assuntos
Androgênios/farmacologia , Transportador de Glucose Tipo 1/metabolismo , Transportador de Glucose Tipo 3/metabolismo , Glucose/metabolismo , L-Lactato Desidrogenase/metabolismo , Ácido Láctico/metabolismo , Transportadores de Ácidos Monocarboxílicos/metabolismo , Proteínas Musculares/metabolismo , Fosfofrutoquinase-1/metabolismo , Neoplasias da Próstata/metabolismo , Apoptose/efeitos dos fármacos , Western Blotting , Proliferação de Células/efeitos dos fármacos , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 3/genética , Glicólise/efeitos dos fármacos , Humanos , Técnicas Imunoenzimáticas , L-Lactato Desidrogenase/genética , Masculino , Transportadores de Ácidos Monocarboxílicos/genética , Proteínas Musculares/genética , Fosfofrutoquinase-1/genética , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais Cultivadas
16.
Transl Res ; 166(6): 693-705, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26397424

RESUMO

Regucalcin (RGN) is a calcium (Ca(2+))-binding protein that displays a characteristic downregulated expression with aging in several tissues. Besides its role in regulating intracellular Ca(2+) homeostasis, RGN has been associated with the control of oxidative stress, cell proliferation, and apoptosis. Thus, the diminished expression of RGN with aging may contribute to the age-associated deterioration of cell function. In the present study, we hypothesized that the maintenance of high expression levels of RGN may prevent age-related alterations in the processes mentioned previously. First, we confirmed that RGN expression is significantly diminished in the prostate of 8-, 9-, 12-, and 24-months wild-type rats. Then, the effect of aging on lipid peroxidation, antioxidant defenses, cell proliferation, and apoptosis in the prostate of wild-type controls and transgenic rats overexpressing RGN (Tg-RGN) was investigated. The activity of glutathione and the antioxidant capacity were increased in Tg-RGN rats in response to the age-associated increase in thiobarbituric acid reactive substances levels, an effect not seen in wild type. Overexpression of RGN also counteracted the effect of aging increasing prostate cell proliferation. In contrast to wild-type animals, the prostate weight of Tg-RGN did not change with aging and was underpinned by the diminished expression of stem cell factor and c-kit, and increased expression of p53. In addition, aged Tg-RGN animals displayed increased expression (activity) of apoptosis regulators, therefore not showing the age-induced resistance to apoptosis observed in wild type. Altogether, these findings indicate the protective role of RGN against the development of age-related pathologies, such as, for example, prostate cancer.


Assuntos
Envelhecimento/metabolismo , Apoptose , Proteínas de Ligação ao Cálcio/metabolismo , Proliferação de Células , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Estresse Oxidativo , Próstata/patologia , Animais , Hidrolases de Éster Carboxílico , Masculino , Tamanho do Órgão , Próstata/metabolismo , Ratos , Ratos Sprague-Dawley , Ratos Transgênicos
17.
Med Oncol ; 32(9): 228, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26255063

RESUMO

Androgens have been associated with the development of normal breast, and their role in mammary gland carcinogenesis has also been described. Several studies reported that androgens inhibit breast cancer cell growth, whereas others linked their action with the modulation of calcium (Ca(2+)) pumps, Ca(2+) channels and Ca(2+)-binding proteins. Also, it is known that deregulated Ca(2+) homeostasis has been implicated in the pathophysiology of breast. The L-type Ca(2+) channels (LTCCs) were found to be up-regulated in colon, colorectal and prostate cancer, but their presence in breast tissues remains uncharacterized. On the other hand, regucalcin (RGN) is a Ca(2+)-binding protein involved in the control of mammary gland cell proliferation, which has been identified as an androgen target gene in distinct tissues except breast. This study aimed to confirm the expression and activity of LTCCs in human breast cancer cells and investigate the effect of androgens in regulating the expression of α1C subunit (Cav1.2) of LTCCs and Ca(2+)-binding protein RGN. PCR, Western blot, immunofluorescence and electrophysiological experiments demonstrated the expression and activity of Cav1.2 subunit in MCF-7 cells. The MCF-7 cells were treated with 1, 10 or 100 nM of 5α-dihydrotestosterone (DHT) for 24-72 h. The obtained results showed that 1 nM DHT up-regulated the expression of Cav1.2 subunit while diminishing RGN protein levels, which was underpinned by reduced cell viability. These findings first confirmed the presence of LTCCs in breast cancer cells and opened new perspectives for the development of therapeutic approaches targeting Ca(2+) signaling.


Assuntos
Canais de Cálcio Tipo L/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Di-Hidrotestosterona/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Canais de Cálcio Tipo L/análise , Canais de Cálcio Tipo L/genética , Proteínas de Ligação ao Cálcio/análise , Proteínas de Ligação ao Cálcio/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Di-Hidrotestosterona/farmacologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/análise , Peptídeos e Proteínas de Sinalização Intracelular/genética
18.
Biochim Biophys Acta ; 1853(10 Pt A): 2621-8, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26171977

RESUMO

Extracellular calcium (Ca2+o) and its receptor, the Ca2+-sensing receptor (CaSR), play an important role in prostate physiology, and it has been shown that the deregulation of Ca2+ homeostasis and the overexpression of CaSR are involved in prostate cancer (PCa). Regucalcin (RGN), a Ca2+-binding protein that plays a relevant role in intracellular Ca2+ homeostasis, was identified as an under-expressed protein in human PCa. Moreover, RGN was associated with suppression of cell proliferation, suggesting that the loss of RGN may favor development and progression of PCa. This work aims to unveil the role of Ca2+o on RGN expression and viability of non-neoplastic (PNT1A) and neoplastic (LNCaP) prostate cell lines. It was demonstrated that Ca2+o up-regulates RGN expression in both cell lines, but important differences were found between cells for dose- and time-responses to Ca2+o treatment. It was also shown that high [Ca2+]o triggers different effects on cell proliferation of neoplastic and non-neoplastic PCa cells, which seems to be related with RGN expression levels. This suggests the involvement of RGN in the regulation of cell proliferation in response to Ca2+o treatment. Also, the effect of Ca2+o on CaSR expression seems to be dependent of RGN expression, which is strengthened by the fact that RGN-knockdown in PNT1A cells increases the CaSR expression, whereas transgenic rats overexpressing RGN exhibit low levels of CaSR. Overall, our results highlighted the importance of RGN as a regulatory protein in Ca2+-dependent signaling pathways and its deregulation of RGN expression by Ca2+o may contribute for onset and progression of PCa.


Assuntos
Sinalização do Cálcio/efeitos dos fármacos , Proteínas de Ligação ao Cálcio/biossíntese , Cálcio , Proliferação de Células/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intracelular/biossíntese , Próstata/metabolismo , Neoplasias da Próstata/metabolismo , Animais , Cálcio/metabolismo , Cálcio/farmacologia , Sinalização do Cálcio/genética , Proteínas de Ligação ao Cálcio/genética , Hidrolases de Éster Carboxílico , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Próstata/patologia , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Ratos , Ratos Sprague-Dawley , Receptores de Detecção de Cálcio/biossíntese , Receptores de Detecção de Cálcio/genética
19.
Prostate ; 75(9): 923-35, 2015 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-25786656

RESUMO

BACKGROUND: Imatinib mesylate is a chemotherapeutic drug that inhibits the tyrosine kinase activity of c-KIT and has been successfully used to treat leukemias and some solid tumors. However, its application for treatment of hormone-refractory prostate cancer (HRPC) has shown modest effectiveness and did not follow the outcomes in cultured cells or animal models. Moreover, the molecular pathways by which imatinib induces cytotoxicity in prostate cancer cells are poorly characterized. METHODS: Two cell line models of HRPC (DU145 and PC3) were exposed to 20 µM of imatinib for 6-72 hr. MTS assay was used to assess cell viability during the course of experiment. Gene expression analysis of c-KIT, cell-cycle and apoptosis regulators, and angiogenic factors was determined by means of real-time PCR, western blot, and/or immunocytochemistry. The enzymatic activity of the apoptosis effector, caspase-3, was determined by a colorimetric assay. RESULTS: Imatinib significantly decreased the viability of DU145 cells but paradoxically augmented the viability of PC3 cells. DU145 cells displayed diminished expression of anti-apoptotic Bcl-2 protein and augmented levels of caspase-8 and -9, as well as, increased enzymatic activity of caspase-3 in response to imatinib. No differences existed on the expression levels of apoptosis-related proteins in PC3 cells treated with imatinib, though the activity of caspase-3 was decreased. The mRNA levels of angiogenic factor VEGF were decreased in DU145-treated cells, whereas an opposite effect was seen in PC3. In addition, it was shown that DU145 and PC3 cells present a differential expression of c-KIT protein variants. CONCLUSION: DU145 and PC3 cells displayed a contradictory behavior in response to imatinib, which was underpinned by a distinct expression pattern (or activity) of target regulators of cell-cycle, apoptosis, and angiogenesis. The paradoxical effect of imatinib in PC3 cells may be related with the differential expression of c-KIT protein variants. Moreover, the present findings helped to understand the discrepancies in the efficacy of imatinib as therapeutic option in HRPC.


Assuntos
Antineoplásicos/farmacologia , Benzamidas/farmacologia , Piperazinas/farmacologia , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Proteínas Proto-Oncogênicas c-kit/biossíntese , Pirimidinas/farmacologia , Western Blotting , Caspases/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Mesilato de Imatinib , Imuno-Histoquímica , Masculino , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas c-kit/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-kit/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Fator A de Crescimento do Endotélio Vascular/biossíntese , Fator A de Crescimento do Endotélio Vascular/genética , Quinases Ativadas por p21/biossíntese , Quinases Ativadas por p21/genética
20.
Exp Cell Res ; 330(2): 325-335, 2015 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-25128811

RESUMO

Regucalcin (RGN) is a calcium-binding protein, which has been shown to be underexpressed in cancer cases. This study aimed to determine the association of RGN expression with clinicopathological parameters of human breast cancer. In addition, the role of RGN in malignancy of mammary gland using transgenic rats overexpressing the protein (Tg-RGN) was investigated. Wild-type (Wt) and Tg-RGN rats were treated with 7,12-dimethylbenz[α]anthracene (DMBA). Carcinogen-induced tumors were histologically classified and the Ki67 proliferation index was estimated. Immunohistochemistry analysis showed that RGN immunoreactivity was negatively correlated with the histological grade of breast infiltrating ductal carcinoma suggesting that progression of breast cancer is associated with loss of RGN. Tg-RGN rats displayed lower incidence of carcinogen-induced mammary gland tumors, as well as lower incidence of invasive forms. Moreover, higher proliferation was observed in non-invasive tumors of Wt animals comparatively with Tg-RGN. Overexpression of RGN was associated with diminished expression of cell-cycle inhibitors and increased expression of apoptosis inducers. Augmented activity of apoptosis effector caspase-3 was found in the mammary gland of Tg-RGN. RGN overexpression protected from carcinogen-induced mammary gland tumor development and was linked with reduced proliferation and increased apoptosis. These findings indicated the protective role of RGN in the carcinogenesis of mammary gland.


Assuntos
Neoplasias da Mama/patologia , Proteínas de Ligação ao Cálcio/biossíntese , Carcinogênese/genética , Peptídeos e Proteínas de Sinalização Intracelular/biossíntese , Glândulas Mamárias Animais/patologia , 9,10-Dimetil-1,2-benzantraceno/farmacologia , Animais , Apoptose/genética , Proteínas de Ligação ao Cálcio/genética , Carcinógenos/farmacologia , Caspase 3/biossíntese , Pontos de Checagem do Ciclo Celular/genética , Proliferação de Células , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Antígeno Ki-67/metabolismo , Pessoa de Meia-Idade , Gradação de Tumores , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Ratos , Ratos Sprague-Dawley , Ratos Transgênicos
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