RESUMO
Hyponatremia is the most frequent electrolyte alteration among hospitalized patients and it has been reported in 20-40% of patients with SARS-CoV-2 (COVID-19) infection. Multiple causes of hyponatremia have been hypothesized in these patients. The syndrome of inappropriate antidiuresis (SIAD) has been considered one of the main reasons leading to hyponatremia in this condition. SIAD can be secondary to cytokines release, in particular IL-6. Positive pressure ventilation can be another cause of hyponatremia due to SIAD. Other possible etiologies of hyponatremia in COVID-19 patients can be related to secondary hypocortisolism, nausea, vomiting, heart and kidney damage. Similar to many other clinical conditions, there is strong evidence that hyponatremia is associated with a worse prognosis also in patients with COVID-19 infection. In particular, hyponatremia has been identified as an independent risk of ICU transfer, need of non-invasive ventilation and death. Hyponatremia in COVID-19 patients is in principle acute and symptomatic and should be treated as such, according to the published guidelines. Therefore, patients should be initially treated with i.v. hypertonic saline (3% NaCl) infusion and serum [Na+] should be frequently monitored, in order to remain within a safe rate of correction. There is evidence showing that serum [Na+] correction is associated with a better outcome in different pathologies, including COVID-19 infection.
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We have previously demonstrated that the vasopressin type 2 receptor (AVPR2) antagonist tolvaptan reduces cell proliferation and invasion and triggers apoptosis in different human cancer cell lines. To study this effect in vivo, a xenograft model of small cell lung cancer was developed in Fox1nu/nu nude mice through the subcutaneous inoculation of H69 cells, which express AVPR2. One group of mice (n = 5) was treated with tolvaptan for 60 days, whereas one group (n = 5) served as the control. A reduced growth was observed in the tolvaptan group in which the mean tumor volume was significantly smaller on day 60 compared to the control group. In the latter group, a significantly lower survival was observed. The analysis of excised tumors revealed that tolvaptan effectively inhibited the cAMP/PKA and PI3K/AKT signaling pathways. The expression of the proliferative marker proliferating cell nuclear antigen (PCNA) was significantly lower in tumors excised from tolvaptan-treated mice, whereas the expression levels of the apoptotic marker caspase-3 were higher than those in control animals. Furthermore, tumor vascularization was significantly lower in the tolvaptan group. Overall, these findings suggest that tolvaptan counteracts tumor progression in vivo and, if confirmed, might indicate a possible role of this molecule as an adjuvant in anticancer strategies.
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Antagonistas dos Receptores de Hormônios Antidiuréticos , Proliferação de Células , Neoplasias Pulmonares , Camundongos Nus , Receptores de Vasopressinas , Carcinoma de Pequenas Células do Pulmão , Tolvaptan , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Tolvaptan/farmacologia , Tolvaptan/uso terapêutico , Antagonistas dos Receptores de Hormônios Antidiuréticos/farmacologia , Camundongos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Humanos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Carcinoma de Pequenas Células do Pulmão/metabolismo , Carcinoma de Pequenas Células do Pulmão/patologia , Receptores de Vasopressinas/metabolismo , Apoptose/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacosRESUMO
Hyponatremia is the prevalent electrolyte imbalance in cancer patients, and it is associated with a worse outcome. Notably, emerging clinical evidence suggests that hyponatremia adversely influences the response to anticancer treatments. Therefore, this study aims to investigate how reduced extracellular [Na+] affects the responsiveness of different cancer cell lines (from human colon adenocarcinoma, neuroblastoma, and small cell lung cancer) to cisplatin and the underlying potential mechanisms. Cisplatin dose-response curves revealed higher IC50 in low [Na+] than normal [Na+]. Accordingly, cisplatin treatment was less effective in counteracting the proliferation and migration of tumor cells when cultured in low [Na+], as demonstrated by colony formation and invasion assays. In addition, the expression analysis of proteins involved in autophagosome-lysosome formation and the visualization of lysosomal areas by electron microscopy revealed that one of the main mechanisms involved in chemoresistance to cisplatin is the promotion of autophagy. In conclusion, our data first demonstrate that the antitumoral effect of cisplatin is markedly reduced in low [Na+] and that autophagy is an important mechanism of drug escape. This study indicates the role of hyponatremia in cisplatin chemoresistance and reinforces the recommendation to correct this electrolyte alteration in cancer patients.
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Antineoplásicos , Autofagia , Proliferação de Células , Cisplatino , Sódio , Humanos , Cisplatino/farmacologia , Autofagia/efeitos dos fármacos , Sódio/metabolismo , Linhagem Celular Tumoral , Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Hiponatremia/metabolismo , Movimento Celular/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/patologia , Lisossomos/metabolismo , Lisossomos/efeitos dos fármacosRESUMO
In cancer patients, hyponatremia is detected in about 40% of cases at hospital admission and has been associated to a worse outcome. We have previously observed that cancer cells from different tissues show a significantly increased proliferation rate and invasion potential, when cultured in low extracellular [Na+]. We have recently developed an animal model of hyponatremia using Foxn1nu/nu mice. The aim of the present study was to compare tumor growth and invasivity of the neuroblastoma cell line SK-N-AS in hyponatremic vs. normonatremic mice. Animals were subcutaneously implanted with luciferase-expressing SK-N-AS cells. When masses reached about 100 mm3, hyponatremia was induced in a subgroup of animals via desmopressin infusion. Tumor masses were significantly greater in hyponatremic mice, starting from day 14 and until the day of sacrifice (day 28). Immunohistochemical analysis showed a more intense vascularization and higher levels of expression of the proliferating cell nuclear antigen, chromogranin A and heme oxigenase-1 gene in hyponatremic mice. Finally, metalloproteases were also more abundantly expressed in hyponatremic animals compared to control ones. To our knowledge, this is the first demonstration in an experimental animal model that hyponatremia is associated to increased cancer growth by activating molecular mechanisms that promote proliferation, angiogenesis and invasivity.
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Hiponatremia , Neuroblastoma , Humanos , Camundongos , Animais , Hiponatremia/etiologia , Xenoenxertos , Sódio/metabolismo , HospitalizaçãoRESUMO
Apelin is an endogenous ligand for the G protein-coupled receptor APJ and has multiple biological activities in human tissues and organs, including the heart, blood vessels, adipose tissue, central nervous system, lungs, kidneys, and liver. This article reviews the crucial role of apelin in regulating oxidative stress-related processes by promoting prooxidant or antioxidant mechanisms. Following the binding of APJ to different active apelin isoforms and the interaction with several G proteins according to cell types, the apelin/APJ system is able to modulate different intracellular signaling pathways and biological functions, such as vascular tone, platelet aggregation and leukocytes adhesion, myocardial activity, ischemia/reperfusion injury, insulin resistance, inflammation, and cell proliferation and invasion. As a consequence of these multifaceted properties, the role of the apelinergic axis in the pathogenesis of degenerative and proliferative conditions (e.g., Alzheimer's and Parkinson's diseases, osteoporosis, and cancer) is currently investigated. In this view, the dual effect of the apelin/APJ system in the regulation of oxidative stress needs to be more extensively clarified, in order to identify new potential strategies and tools able to selectively modulate this axis according to the tissue-specific profile.
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Receptores de Apelina , Apelina , Estresse Oxidativo , Humanos , Apelina/metabolismo , Receptores de Apelina/metabolismo , Receptores Acoplados a Proteínas G/metabolismoRESUMO
Hyponatremia is the most common electrolyte disorder encountered in hospitalized patients. This applies also to cancer patients. Multiple causes can lead to hyponatremia, but most frequently this electrolyte disorder is due to the syndrome of inappropriate antidiuresis. In cancer patients, this syndrome is mostly secondary to ectopic secretion of arginine vasopressin by tumoral cells. In addition, several chemotherapeutic drugs induce the release of arginine vasopressin by the hypothalamus. There is evidence that hyponatremia is associated to a more negative outcome in several pathologies, including cancer. Many studies have demonstrated that in different cancer types, both progression-free survival and overall survival are negatively affected by hyponatremia, whereas the correction of serum [Na+] has a positive effect on patient outcome. In vitro studies have shown that cells grown in low [Na+] have a greater proliferation rate and motility, due to a dysregulation in intracellular signalling pathways. Noteworthy, vasopressin receptors antagonists, which were approved more than a decade ago for the treatment of euvolemic and hypervolemic hyponatremia, have shown unexpected antiproliferative effects. Because of this property, vaptans were also approved for the treatment of polycystic kidney disease. In vitro evidence indicated that this family of drugs effectively counteracts proliferation and invasivity of cancer cells, thus possibly opening a new scenario among the pharmacological strategies to treat cancer.
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Hyponatremia, i.e., the presence of a serum sodium concentration ([Na+]) < 136 mEq/L, is the most frequent electrolyte imbalance in the elderly and in hospitalized patients. Symptoms of acute hyponatremia, whose main target is the central nervous system, are explained by the "osmotic theory" and the neuronal swelling secondary to decreased extracellular osmolality, which determines cerebral oedema. Following the description of neurological and systemic manifestations even in mild and chronic hyponatremia, in the last decade reduced extracellular [Na+] was associated with detrimental effects on cellular homeostasis independently of hypoosmolality. Most of these alterations appeared to be elicited by oxidative stress. In this review, we focus on the role of oxidative stress on both osmolality-dependent and -independent impairment of cell and tissue functions observed in hyponatremic conditions. Furthermore, basic and clinical research suggested that oxidative stress appears to be a common denominator of the degenerative processes related to aging, cancer progression, and hyponatremia. Of note, low [Na+] is able to exacerbate multiple manifestations of senescence and to decrease progression-free and overall survival in oncologic patients.
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PURPOSE: Hyponatraemia is frequently observed in cancer patients and can be due to the syndrome of inappropriate anti-diuresis (SIAD), related to ectopic vasopressin secretion, particularly in small cell lung cancer (SCLC). Hyponatraemia is associated with a worse outcome in cancer patients. The vasopressin receptor antagonist tolvaptan effectively corrects hyponatraemia secondary to SIAD and there is in vitro evidence that it has also an antiproliferative effect in cancer cells. The purpose of this study was i) to analyse the effect of low serum sodium concentrations ([Na+]) in SCLC cells and ii) to determine whether tolvaptan counteracts tumor progression. METHODS: We evaluated cell proliferation, cell cycle, apoptosis, oxidative stress, invasivity in low [Na+] as well as after exposure to tolvaptan. We also analysed the intracellular signalling pathways involved. RESULTS: In reduced [Na+] cell proliferation was significantly increased compared to normal [Na+] and cells were mostly distributed in the G2/M phase. Apoptosis appeared reduced. In addition, the ability to cross matrigel-coated membranes markedly increased. As observed in other cancer cell models, the expression of the heme-oxigenase-1 gene was increased. Finally, we found that in cells cultured in low [Na+] the RhoA/ROCK1/2 pathway, which is involved in the regulation of actin cytoskeleton, was activated. On the other hand, we found that tolvaptan effectively inhibited cell proliferation, anchorage-independent growth, invasivity and promoted apoptosis. Accordingly, the RhoA/ROCK-1/2 pathway was inhibited. CONCLUSIONS: These findings demonstrate for the first time that low [Na+] favours tumor progression in SCLC cells, whereas tolvaptan effectively inhibits cell proliferation, survival and invasivity.
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Neoplasias Pulmonares/patologia , Carcinoma de Pequenas Células do Pulmão/patologia , Sódio/farmacologia , Tolvaptan/farmacologia , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Heme Oxigenase-1/metabolismo , Humanos , Invasividade Neoplásica , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Quinases Associadas a rho/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
OBJECTIVE: Hyponatremia is the most common electrolyte disorder in hospitalized patients and occurs in about 30% of patients with pneumonia. Hyponatremia has been associated with a worse outcome in several pathologic conditions The main objective of this study was to determine whether serum sodium alterations may be independent predictors of the outcome of hospitalized COVID-19 patients. DESIGN AND METHODS: In this observational study, data from 441 laboratory-confirmed COVID-19 patients admitted to a University Hospital were collected. After excluding 61 patients (no serum sodium at admission available, saline solution infusion before sodium assessment, transfer from another hospital), data from 380 patients were analyzed. RESULTS: 274 (72.1%) patients had normonatremia at admission, 87 (22.9%) patients had hyponatremia and 19 (5%) patients had hypernatremia. We found an inverse correlation between serum sodium and IL-6, whereas a direct correlation between serum sodium and PaO2/FiO2 ratio was observed. Patients with hyponatremia had a higher prevalence of non-invasive ventilation and ICU transfer than those with normonatremia or hypernatremia. Hyponatremia was an independent predictor of in-hospital mortality (2.7-fold increase vs normonatremia) and each mEq/L of serum sodium reduction was associated with a 14.4% increased risk of death. CONCLUSIONS: These results suggest that serum sodium at admission may be considered as an early prognostic marker of disease severity in hospitalized COVID-19 patients.
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COVID-19/sangue , SARS-CoV-2 , Índice de Gravidade de Doença , Sódio/sangue , Idoso , Idoso de 80 Anos ou mais , COVID-19/epidemiologia , COVID-19/mortalidade , Comorbidade , Cuidados Críticos/estatística & dados numéricos , Feminino , Fluorocarbonos/sangue , Mortalidade Hospitalar , Hospitalização/estatística & dados numéricos , Humanos , Hidrocarbonetos Bromados/sangue , Hipernatremia/epidemiologia , Hiponatremia/epidemiologia , Interleucina-6/sangue , Masculino , Pessoa de Meia-Idade , Respiração Artificial/estatística & dados numéricos , Estudos Retrospectivos , Coronavírus Relacionado à Síndrome Respiratória Aguda GraveRESUMO
PURPOSE: Hyponatremia is the most common electrolyte disorder in hospitalized patients, and its etiopathogenesis is related to an underlying tumor in 14% of cases. Hyponatremia has been associated with a worse outcome in several pathologies, including cancer, in which the leading cause of this electrolyte alteration is the syndrome of inappropriate antidiuresis. The aim of this study was to analyze in vitro the effects of low extracellular [Na+] in cancer progression. MATERIALS AND METHODS: We used a previously validated experimental model of chronic hyponatremia to characterize the effects of low extracellular [Na+] in different human cancer cell lines: pancreatic adenocarcinoma (PANC-1), neuroblastoma (SK-N-AS, SH-SY5Y), colorectal adenocarcinoma (HCT-8), chronic myeloid leukemia (K562). RESULTS: Our results demonstrate a direct relationship between low [Na+], reduced cell adhesion and increased invasion and proliferation in all cell lines tested. Accordingly, the number of tumor colonies grown in soft agar and the expression of collagenases type IV (metalloproteinases 2 and 9) were markedly higher in cancer cells exposed to reduced extracellular [Na+]. Gene analysis showed an upregulation of molecular pathways involved in oxidative stress (heme oxygenase 1) and in proliferation and invasion (RhoA, ROCK-1, ROCK-2). The activation of RhoA/ROCK pathway was paralleled by a deregulation of the cytoskeleton-associated proteins, resulting in the promotion of actin cytoskeletal remodeling and cell invasion. CONCLUSIONS: Overall, our data demonstrate for the first time that low [Na+] promotes cancer progression in vitro, thus suggesting that hyponatremia is not a simple bystander of disease severity in cancer.
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Adenocarcinoma , Neoplasias Pancreáticas , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Citoesqueleto , Humanos , Invasividade Neoplásica , SódioRESUMO
PURPOSE: GLP-1 receptor agonists are antidiabetic drugs currently used in the therapy of type 2 diabetes. Despite several in vitro and in vivo animal studies suggesting a beneficial effect of GLP-1 analogues on bone, in humans their skeletal effects are not clear and clinical studies report conflicting results. METHODS: We differentiated human mesenchymal stromal cells (hMSC) toward the adipogenic and the osteoblastic lineages, analysing the effect of Exendin-4 (EXE) before, during and after specific differentiations. RESULTS: We showed EXE ability to act selectively on a sub-population of hMSC characterised by a more stem potential, shifting them from G1 to S/M phase of cell cycle. We observed that EXE pre-treatment promotes both adipogenic and osteoblastic differentiations, possibly determined by an increased number of uncommitted progenitors. In fully differentiated cells, EXE affects mature adipocytes by increasing lipolysis, otherwise not altering osteoblasts metabolic activity. Moreover, the increased expression of osteoprotegerin, a modulator of the RANK/RANKL system, observed during osteogenic induction in presence of EXE, could negatively modulate osteoclastogenesis. CONCLUSIONS: Our data suggest a complex action of EXE on bone, targeting the proliferation of mesenchymal progenitors, the metabolism of mature adipocytes and the modulation of osteoclastogenesis. Thus, an overall positive effect of this molecule on bone quality might be hypothesised.
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Células da Medula Óssea/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Exenatida/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Adipogenia/efeitos dos fármacos , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Humanos , Lipólise/efeitos dos fármacos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Osteogênese/efeitos dos fármacos , Osteoprotegerina/metabolismoRESUMO
What is the central question of this study? Hyponatraemia, an electrolyte disorder encountered in hospitalized patients, can cause neurological symptoms usually attributed to a reduction in plasma osmolarity. Here, we investigated whether low [Na(+) ] per se can cause neuronal changes independent of osmolarity, focusing on involvement of the Na(+) -Ca(2+) exchanger. What is the main finding and its importance? We show that hyponatraemia per se causes alterations of neuronal properties. The novel finding of Na(+) -Ca(2+) exchanger involvement helps us to elucidate the volume regulation following hyponatraemia. This might have relevance in a translational perspective because Na(+) -Ca(2+) exchanger could be a target for novel therapies. Hyponatraemia is the most frequent electrolyte disorder encountered in hospitalized patients, and it can cause a wide variety of neurological symptoms. Most of the negative effects of this condition on neuronal cells are attributed to cell swelling because of the reduction of plasma osmolarity, although in hyponatraemia different membrane proteins are supposed to be involved in the conservation of neuronal volume. We have recently reported detrimental effects of hyponatraemia on two different neuronal cell lines, SK-N-AS and SH-SY5Y, independent of osmotic alterations. In this study we investigated, in the same cell lines, whether hyponatraemic conditions per se can cause electrophysiological alterations and whether these effects vary over time. Accordingly, we carried out experiments in low-sodium medium in either hyposmotic [Osm(-)] or isosmotic [Osm(+)] conditions, for a short (24 h) or long time (7 days). Using a patch pipette in voltage-clamp conditions, we recorded possible modifications of cell capacitance (Cm ) and membrane conductance (Gm ). Our results indicate that in both Osm(-) and Osm(+) medium, Cm and Gm show a similar increase, but such effects are dependent on the time in culture in different ways. Notably, regarding the possible mechanisms involved in the maintenance of Cm , Gm and Gm /Cm in Osm(+) conditions, we observed a greater contribution of the Na(+) -Ca(2+) exchanger compared with Osm(-) and control conditions. Overall, these novel electrophysiological results help us to understand the mechanisms of volume regulation after ionic perturbation. Our results might also have relevance in a translational perspective because the Na(+) -Ca(2+) exchanger can be considered a target for planning novel therapies.
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Membrana Celular/fisiologia , Hiponatremia/fisiopatologia , Neurônios/fisiologia , Cálcio/metabolismo , Contagem de Células/métodos , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Humanos , Hiponatremia/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Níquel/farmacologia , Concentração Osmolar , Técnicas de Patch-Clamp/métodos , Sódio/metabolismo , Trocador de Sódio e Cálcio/metabolismoRESUMO
The purpose of this study is to describe a case of concurrent medullary and papillary thyroid carcinoma (MTC and PTC) and cutaneous melanoma and to analyze BRAF(V600E) mutation in plasma and tissues. We report the clinical history and the laboratory, imaging, and histopathological findings of a 47-year-old man affected by multinodular goiter. BRAF(V600E)-mutated DNA was quantified in plasma samples and in cancer sections by quantitative real-time polymerase chain reaction (qPCR). At ultrasound examination, the dominant right nodule of the thyroid was weakly hyperechoic and hypervascularized, while the left one was hypoechoic without internal vascularization. Regional lymphadenomegalia was not detected. Basal plasma calcitonin was elevated, and the patient underwent total thyroidectomy and resection of central cervical lymph nodes. Histopathological examination identified two distinct foci of MTC and PTC and micrometastasis of well-differentiated carcinoma in one of the six resected lymph nodes. RET proto-oncogene germline mutations were not detected. Cutaneous melanoma of the thorax was subsequently diagnosed. BRAF(V600E) tissue DNA was detected in PTC and melanoma but not in MTC. The cell-free plasma percentage of BRAF(V600E) DNA was detected in pre-thyroidectomy peripheral blood and was drastically reduced after cancer treatments. This study confirms the occurrence of synchronous MTC and PTC and is the first evidence of the co-existence of melanoma and distinct thyroid cancers of different origin. BRAF(V600E) allele was detected in PTC and melanoma but not in MTC tissues. BRAF(V600E) molecular quantification in pre- and post-treatment blood supports our previous data, suggesting its possible role in diagnosis and follow-up of BRAF-positive tumors.
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Carcinoma Medular/patologia , Carcinoma Papilar/patologia , Neoplasias Primárias Múltiplas/patologia , Proteínas Proto-Oncogênicas B-raf/genética , Glândula Tireoide/patologia , Neoplasias da Glândula Tireoide/patologia , Alelos , Carcinoma Medular/genética , Carcinoma Papilar/genética , Análise Mutacional de DNA , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Neoplasias Primárias Múltiplas/genética , Proto-Oncogene Mas , Neoplasias da Glândula Tireoide/genéticaRESUMO
Exendin-4 is a molecule currently used, in its synthetic form exenatide, for the treatment of type 2 diabetes mellitus. Exendin-4 binds and activates the Glucagon-Like Peptide-1 Receptor (GLP-1R), thus inducing insulin release. More recently, additional biological properties have been associated to molecules that belong to the GLP-1 family. For instance, Peptide YY and Vasoactive Intestinal Peptide have been found to affect cell adhesion and migration and our previous data have shown a considerable actin cytoskeleton rearrangement after exendin-4 treatment. However, no data are currently available on the effects of exendin-4 on tumor cell motility. The aim of this study was to investigate the effects of this molecule on cell adhesion, differentiation and migration in two neuroblastoma cell lines, SH-SY5Y and SK-N-AS. We first demonstrated, by Extra Cellular Matrix cell adhesion arrays, that exendin-4 increased cell adhesion, in particular on a vitronectin substrate. Subsequently, we found that this molecule induced a more differentiated phenotype, as assessed by i) the evaluation of neurite-like protrusions in 3D cell cultures, ii) the analysis of the expression of neuronal markers and iii) electrophysiological studies. Furthermore, we demonstrated that exendin-4 reduced cell migration and counteracted anchorage-independent growth in neuroblastoma cells. Overall, these data indicate for the first time that exendin-4 may have anti-tumoral properties.
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Diferenciação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Peptídeos/farmacologia , Peçonhas/farmacologia , Canais de Cálcio/metabolismo , Adesão Celular/efeitos dos fármacos , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Exenatida , Expressão Gênica/efeitos dos fármacos , Receptor do Peptídeo Semelhante ao Glucagon 1 , Humanos , Hipoglicemiantes/farmacologia , Potenciais da Membrana/efeitos dos fármacos , Proteínas Associadas aos Microtúbulos/genética , Invasividade Neoplásica , Neuroblastoma/genética , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Ligação Proteica/efeitos dos fármacos , Receptores de Glucagon/genética , Receptores de Ativador de Plasminogênio Tipo Uroquinase/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sinaptofisina/genética , Vitronectina/metabolismo , Proteínas tau/genéticaRESUMO
There is evidence that chronic hyponatremia, even when mild, may cause neurological signs and symptoms. These have been traditionally associated with water movement into nervous cells, as a result of the hypotonic state. The aim of the present study was to determine whether low extracellular sodium directly exerts negative effects on human neuronal cells, independently of reduced osmolality. We exposed neuroblastoma SK-N-AS and SH-SY5Y cells to sustained low extracellular sodium, thus mimicking a condition of chronic hyponatremia, both in the presence of reduced and in the presence of unaltered osmolality. We found that very low sodium (i.e., 115 mmol/L in SK-N-AS and 90 mmol/L in SH-SY5Y) significantly reduced cell viability. However, intermediate low sodium was able to cause cell distress, as assessed by the altered expression of anti-apoptotic genes and the reduced ability to differentiate into a mature neuronal phenotype. Noteworthy, these effects were observed also in the presence of unaltered osmolality. Moreover, we performed a comprehensive microarray analysis in cells maintained in normal sodium or in low sodium and unaltered osmolality, and we found that the most altered pathway included genes involved in "cell death and survival." Among the more than 40 differentially expressed genes, the Heme oxygenase gene, which represents a transcriptional response to oxidative stress, showed the highest increase in the expression level. This study demonstrates that low extracellular sodium causes detrimental effects in neuronal cells that are at least in part independent of reduced osmolality. These findings further support the recommendation to effectively correct hyponatremia, even when mild and chronic.
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Líquido Extracelular/metabolismo , Hiponatremia/patologia , Neurônios/efeitos dos fármacos , Sódio/farmacologia , Apoptose/genética , Adesão Celular , Linhagem Celular Tumoral , Movimento Celular , Sobrevivência Celular , Doença Crônica , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Heme Oxigenase-1/biossíntese , Heme Oxigenase-1/genética , Humanos , Hiponatremia/metabolismo , Neuroblastoma/patologia , Neurogênese , Neurônios/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Concentração Osmolar , Pressão Osmótica , Reação em Cadeia da Polimerase em Tempo RealRESUMO
The bioactive form of vitamin D, 1,25-dihydroxyvitamin D(3), is a secosteroid hormone that binds to the vitamin D receptor (VDR), a member of the nuclear receptor superfamily, and modulates a variety of biological functions. The VDR is expressed by most cell types, including cells of the urogenital system, such as prostate and bladder cells. In particular, the prostate is a target organ of VDR agonists and represents an extrarenal synthesis site of 1,25-dihydroxyvitamin D(3). We have analyzed the capacity of VDR agonists to treat benign prostatic hyperplasia (BPH), a complex syndrome characterized by a static component related to prostate overgrowth, a dynamic component responsible for urinary irritative symptoms, and an inflammatory component. Data reviewed here demonstrate that VDR agonists, and notably elocalcitol, reduce the static component of BPH by inhibiting the activity of intraprostatic growth factors downstream of the androgen receptor, the dynamic component by targeting the RhoA/ROCK pathway in prostate and bladder cells, and the inflammatory component by targeting the NF-kappaB pathway.
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Inflamação/metabolismo , Próstata/metabolismo , Hiperplasia Prostática/metabolismo , Receptores de Calcitriol/agonistas , Substâncias de Crescimento/metabolismo , Humanos , Masculino , Modelos Biológicos , Próstata/efeitos dos fármacos , Vitamina D/análogos & derivados , Vitamina D/farmacologia , Quinases Associadas a rho/metabolismoRESUMO
INTRODUCTION: In male, lower urinary tract symptoms (LUTS) have been associated, beside benign prostatic hyperplasia, to some unexpected comorbidities (hypogonadism, obesity, metabolic syndrome), which are essentially characterized by an unbalance between circulating androgens/estrogens. Within the bladder, LUTS are linked to RhoA/Rho-kinase (ROCK) pathway overactivity. AIM: To investigate the effects of changing sex steroids on bladder smooth muscle. METHODS: ER α, ER ß, GPR30/GPER1 and aromatase mRNA expression was analyzed in male genitourinary tract tissues, and cells isolated from bladder, prostate, and urethra. Estrogen and G1 effect on RhoA/ROCK signaling output like cell migration, gene expression, and cytoskeletal remodeling, and [Ca(2+) ](i) was also studied in hB cells. Contractile studies on bladder strips from castrated male rats supplemented with estradiol and testosterone was also performed. MAIN OUTCOME MEASURES: The effects of classical (ER α, ER ß) and nonclassical (GPR30/GPER1) estrogen receptor ligands (17 ß-estradiol and G1, respectively) and androgens on RhoA/ROCK-.mediated cell functions were studied in hB cells. Contractility studies were also performed in bladder strips from castrated male rats supplemented with testosterone or estradiol. RESULTS: Aromatase and sex steroid receptors, including GPR30, were expressed in human bladder and mediates several biological functions. Both 17 ß-estradiol and G1 activated calcium transients and induced RhoA/ROCK signaling (cell migration, cytoskeleton remodeling and smooth muscle gene expression). RhoA/ROCK inhibitors blunted these effects. Estrogen-, but not androgen-supplementation to castrated rats increased sensitivity to the ROCK inhibitor, Y-27632 in isolated bladder strips. In hB cells, testosterone elicited effects similar to estrogen, which were abrogated by blocking its aromatization through letrozole. CONCLUSION: Our data indicate for the first time that estrogen-more than androgen-receptors up-regulate RhoA/ROCK signaling. Since an altered estrogen/androgen ratio characterizes conditions, such as aging, obesity and metabolic syndrome, often associated to LUTS, we speculate that a relative hyperestrogenism may induce bladder overactivity through the up-regulation of RhoA/ROCK pathway.
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Músculo Liso/fisiopatologia , Hiperplasia Prostática/genética , Hiperplasia Prostática/fisiopatologia , RNA Mensageiro/genética , Obstrução do Colo da Bexiga Urinária/genética , Obstrução do Colo da Bexiga Urinária/fisiopatologia , Bexiga Urinária Hiperativa/genética , Bexiga Urinária Hiperativa/fisiopatologia , Bexiga Urinária/fisiopatologia , Quinases Associadas a rho/genética , Quinases Associadas a rho/fisiologia , Proteína rhoA de Ligação ao GTP/genética , Proteína rhoA de Ligação ao GTP/fisiologia , Androgênios/sangue , Animais , Aromatase/genética , Aromatase/fisiologia , Movimento Celular/genética , Movimento Celular/fisiologia , Células Cultivadas , Citoesqueleto/genética , Citoesqueleto/fisiologia , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/fisiologia , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/fisiologia , Estrogênios/sangue , Genitália Masculina/fisiopatologia , Humanos , Hipogonadismo/genética , Hipogonadismo/fisiopatologia , Masculino , Síndrome Metabólica/genética , Síndrome Metabólica/fisiopatologia , Microscopia Confocal , Obesidade/genética , Obesidade/fisiopatologia , Ratos , Ratos Sprague-Dawley , Receptores de Estrogênio , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/fisiologia , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Testosterona/sangue , Regulação para Cima/genética , Regulação para Cima/fisiologiaRESUMO
INTRODUCTION: Phosphodiesterase type 5 (PDE5) inhibitors ameliorate low urinary tract (LUT) symptoms in men with ED and symptomatic benign prostatic hyperplasia (BPH). PDE5 is highly expressed in rat and human bladder, where it regulates cyclic guanosine monophosphate (cGMP) degradation, muscle tone, and proliferation. AIM: To investigate PDE5 tissue distribution and activity in human LUT tissues (urethra, prostate, and bladder). MAIN OUTCOME MEASURES: PDE5 expression and activity were analyzed and compared within the same BPH patient in LUT tissues and in smooth muscle cells (SMCs) cultured from urethra, prostate, and bladder. METHODS: In LUT tissues, PDE5 was localized by immunohistochemistry and mRNA expression by quantitative real-time polymerase chain reaction. Proliferation assay was used as readout of PDE5 activity, evaluated as ability of vardenafil to increase the antiproliferative effect of different nitric oxide (NO)/cGMP pathway activators [the PDE5-resistant cGMP analog Sp-8-Br-PET-cGMPS, the NO donor sodium nitroprusside (SNP), and the soluble guanylate cyclase (sGC) stimulator BAY 41-8543]. RESULTS: In all the LUT tissues, PDE5 was immunolocalized in blood vessels and in muscular fibres, but not in epithelium. PDE5 mRNA expression was higher in urethra and bladder than in prostate SMC. The antiproliferative effect of Sp-8-Br-PET-cGMPS was similar in all LUT SMC. In prostatic SMC, SNP and BAY 41-8543 show a dose-dependent antiproliferative effect that resulted marginally enhanced by vardenafil. Conversely, in urethra and bladder SMC the antiproliferative effect of SNP and BAY 41-8543 was lower than in prostatic SMC, but it was significantly enhanced by vardenafil. In urethral and bladder cells vardenafil half-maximal response inhibiting concentration was in the subnanomolar range, whereas in prostate cells it resulted significantly higher. CONCLUSIONS: The highest expression and biological activity of PDE5 was found in bladder. However, a consistent PDE5 expression and activity was also found in prostatic urethra. In contrast, the prostate gland showed the lowest PDE5 abundance and cultures derived from this tissue were less sensitive to vardenafil.
Assuntos
Nucleotídeo Cíclico Fosfodiesterase do Tipo 5/genética , Próstata/metabolismo , RNA Mensageiro/genética , Uretra/metabolismo , Bexiga Urinária/metabolismo , Idoso , Divisão Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Expressão Gênica/efeitos dos fármacos , Humanos , Imidazóis/farmacologia , Masculino , Morfolinas/farmacologia , Músculo Liso/efeitos dos fármacos , Músculo Liso/metabolismo , Doadores de Óxido Nítrico/farmacologia , Nitroprussiato/farmacologia , Piperazinas/farmacologia , Próstata/efeitos dos fármacos , Hiperplasia Prostática/genética , Hiperplasia Prostática/patologia , Pirimidinas/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sulfonas/farmacologia , Triazinas/farmacologia , Uretra/efeitos dos fármacos , Bexiga Urinária/efeitos dos fármacos , Dicloridrato de Vardenafila , Vasodilatadores/farmacologiaRESUMO
INTRODUCTION: In human bladder, phosphodiesterase type 5 (PDE5) is present not only in the muscular wall but also in the vascular beds, suggesting a role for PDE5 inhibitors in favoring bladder blood flow and tissue oxygenation. AIM: To investigate whether acute administration of vardenafil could affect bladder oxygenation in spontaneously hypertensive rats (SHR), an animal model of naturally occurring overactive bladder. MAIN OUTCOME MEASURES: The effect of vardenafil on hypoxia-induced alterations was studied in vivo in SHR by acute dosing (10 mg/kg, 90 minutes before sacrifice) and in vitro in human bladder smooth muscle cells (hBCs). METHODS: Bladder oxygenation was detected using the hypoxyprobe immunostaining. The expression of some hypoxia markers (vascular endothelial growth factor [VEGF] and endothelin-1 type B [ETB] receptor) was also evaluated by immunohistochemistry and Western blot. Gene expression in hBC was quantified by real-time reverse transcription-polymerase chain reaction. RESULTS: Rat bladder PDE5 immunopositivity was detected in the muscular wall and in the endothelial and smooth muscle cells of blood vessels. In SHR bladder, a significant increase of hypoxic cells, VEGF, and ETB expression was observed when compared with their normotensive counterpart Wistar Kyoto rats (WKY). Vardenafil treatment dramatically decreased hypoxyprobe staining, as well as VEGF and ETB expression in SHR bladder up to WKY level. Accordingly, in SHR bladder, vardenafil administration significantly blunted relaxation induced by the selective ETB agonist IRL-1620. In hBCs, experimental hypoxia significantly induced gene expression of hypoxia markers (carbonic anhydrase IX and VEGF), which was not changed by simultaneous treatment with vardenafil. Conversely, the hypoxia-related induction of smooth muscle-specific genes (alphaSMA, SM22alpha, and desmin) was significantly reduced by vardenafil. CONCLUSIONS: SHR showed bladder hypoxia which was significantly reduced by acute vardenafil treatment. Thus, besides relaxing muscular wall, PDE5 inhibition may positively affect urinary vesicle blood perfusion.
Assuntos
Imidazóis/farmacologia , Músculo Liso/irrigação sanguínea , Consumo de Oxigênio/efeitos dos fármacos , Consumo de Oxigênio/fisiologia , Piperazinas/farmacologia , Bexiga Urinária Hiperativa/fisiopatologia , Bexiga Urinária/irrigação sanguínea , Vasodilatadores/farmacologia , Animais , Células Cultivadas , Nucleotídeo Cíclico Fosfodiesterase do Tipo 5/genética , Nucleotídeo Cíclico Fosfodiesterase do Tipo 5/metabolismo , Esquema de Medicação , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Hipóxia/patologia , Hipóxia/fisiopatologia , Técnicas Imunoenzimáticas , Masculino , Músculo Liso/patologia , Hiperplasia Prostática/patologia , Hiperplasia Prostática/fisiopatologia , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptor de Endotelina B/efeitos dos fármacos , Receptor de Endotelina B/genética , Sulfonas/farmacologia , Triazinas/farmacologia , Bexiga Urinária/patologia , Bexiga Urinária Hiperativa/patologia , Dicloridrato de Vardenafila , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
INTRODUCTION: Metabolic syndrome (MetS) is a clustering of cardio-metabolic risk factors (hyperglycemia, hypertension, dyslipidemia, visceral fat accumulation) that is also associated with hypogonadism and erectile dysfunction (ED). AIM: To clarify the relationships among MetS, hypogonadism, and ED, we developed an animal model of MetS. METHODS: Male rabbits fed a high-fat diet (HFD), with or without testosterone (T) supplementation, were compared with control rabbits (fed a standard chow) and with rabbits made hypogonadal by a single injection of a long-acting GnRH-analog, triptorelin. MAIN OUTCOME MEASURES: Evaluation of metabolic disturbances (plasma glucose, cholesterol, triglycerides, testosterone, LH, FSH level, glucose tolerance, mean arterial pressure, visceral fat accumulation), and corpora cavernosa (CC) relaxant capacity (in vitro contractility study) in HFD animals as compared with control, GnRH analog-treated rabbits, and T-supplemented HFD rabbits. RESULTS: HFD rabbits showed all the features of MetS. HFD induced hypogonadotropic hypogonadism is characterized by a reduction of plasma T, FSH, LH levels, testis and seminal vesicles weight, and testicular steroidogenic enzymes. Such a phenotype is similar to that induced by triptorelin administration. A reduced GnRH immunopositivity in hypothalamus suggests a central origin of HFD-related hypogonadism. HFD also induced penile alterations, as demonstrated by a reduction of acetylcholine-and electrical field stimulation-induced CC relaxation, hyper-responsiveness to the NO donor, SNP, and unresponsiveness to PDE5 inhibitors. Similar penile alterations were observed in triptorelin treated rabbit. In HFD, as well as in triptorelin treated rabbits, PDE5 and eNOS mRNA expression quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) were significantly decreased. T administration prevented almost all penile alterations observed in HFD rabbits. T treatment dramatically reduced HFD-induced visceral obesity, partially ameliorating also the metabolic profile. CONCLUSION: We have developed an animal model of MetS associated with hypogonadotropic hypogonadism and penile alterations including unresponsiveness to PDE5 inhibitors. T supplementation was able to partially revert HFD-induced phenotype.