RESUMO
Osteosarcoma (OS) is the most common primary malignant bone tumor and its etiology has recently been associated with osteogenic differentiation dysfunctions. OS cells keep a capacity for uncontrolled proliferation showing a phenotype similar to undifferentiated osteoprogenitors with abnormal biomineralization. Within this context, both conventional and X-ray synchrotron-based techniques have been exploited to deeply characterize the genesis and evolution of mineral depositions in a human OS cell line (SaOS-2) exposed to an osteogenic cocktail for 4 and 10 days. A partial restoration of the physiological biomineralization, culminating with the formation of hydroxyapatite, was observed at 10 days after treatment together with a mitochondria-driven mechanism for calcium transportation within the cell. Interestingly, during differentiation, mitochondria showed a change in morphology from elongated to rounded, indicating a metabolic reprogramming of OS cells possibly linked to an increase in glycolysis contribution to energy metabolism. These findings add a dowel to the genesis of OS giving new insights on the development of therapeutic strategies able to restore the physiological mineralization in OS cells.
Assuntos
Neoplasias Ósseas , Osteossarcoma , Humanos , Osteogênese , Biomineralização , Linhagem Celular Tumoral , Osteossarcoma/metabolismo , Diferenciação Celular/fisiologia , Mitocôndrias/metabolismo , Neoplasias Ósseas/metabolismo , Proliferação de Células/fisiologiaRESUMO
γ-terpinene, α-terpinene, p-cymene, and myrcene are monoterpenes found in many essential oils extracted from a variety of plants and spices. Myrcene also occurs naturally in plants such as hops, cannabis, lemongrass, and verbena and is used as a flavoring agent in food and beverage manufacturing. In this research, the biological efficacy of γ-terpinene, α-terpinene, p-cymene, and myrcene was studied in human cell lines (HeLa, SH-SY5Y, and HDFa). Cytotoxicity, cell proliferation, cell migration, and morphology assays were performed to obtain detailed information on the anticancer properties. Our results show that myrcene has potential biological activity, especially in HeLa cells. In this cell line, it leads to an arrest of proliferation, a decrease in motility and morphological changes with loss of sphericity and thickness, and DNA damage. In addition, the interaction of γ-terpinene, α-terpinene, p-terpinene, and myrcene with calf thymus DNA (ct-DNA) was studied by UV-visible spectrophotometry. DNA binding experiments show that only myrcene can interact with DNA with an apparent dissociation constant (Kd) of 29 × 10-6 M.
RESUMO
Metallodrugs represent a combination of multifunctionalities that are present concomitantly and can act differently on diverse biotargets. Their efficacy is often related to the lipophilic features exhibited both by long carbo-chains and the phosphine ligands. Three Ru(II) complexes containing hydroxy stearic acids (HSAs) were successfully synthesized in order to evaluate possible synergistic effects between the known antitumor activity of HSA bio-ligands and the metal center. HSAs were reacted with [Ru(H)2CO(PPh3)3] selectively affording O,O-carboxy bidentate complexes. The organometallic species were fully characterized spectroscopically using ESI-MS, IR, UV-Vis, and NMR techniques. The structure of the compound Ru-12-HSA was also determined using single crystal X-ray diffraction. The biological potency of ruthenium complexes (Ru-7-HSA, Ru-9-HSA, and Ru-12-HSA) was studied on human primary cell lines (HT29, HeLa, and IGROV1). To obtain detailed information about anticancer properties, tests for cytotoxicity, cell proliferation, and DNA damage were performed. The results demonstrate that the new ruthenium complexes, Ru-7-HSA and Ru-9-HSA, possess biological activity. Furthermore, we observed that the Ru-9-HSA complex shows increased antitumor activity on colon cancer cells, HT29.
Assuntos
Antineoplásicos , Complexos de Coordenação , Rutênio , Humanos , Antineoplásicos/farmacologia , Antineoplásicos/química , Rutênio/química , Ligantes , Células HeLa , Proliferação de Células , Complexos de Coordenação/farmacologia , Complexos de Coordenação/química , Linhagem Celular TumoralRESUMO
Indoles constitute a large family of heterocyclic compounds widely occurring in nature which are present in a number of bioactive natural and synthetic compounds, including anticancer agents or atypical opioid agonists. As a result, exponential increases in the development of novel methods for the synthesis of indole-containing compounds have been reported in the literature. A series of indole-aryl amide derivatives 1-7 containing tryptamine or an indolylacetic acid nucleus were designed, synthesized, and evaluated as opioid ligands. These new indole derivatives showed negligible to very low affinity for µ- and δ-opioid receptor (OR). On the other hand, compounds 2, 5 and 7 showed Ki values in the low µM range for κ-OR. Since indoles are well known for their anticancer potential, their effect against a panel of tumor cell lines was tested. The target compounds were evaluated for their in vitro cytotoxicity in HT29, HeLa, IGROV-1, MCF7, PC-3, and Jurkat J6 cells. Some of the synthesized compounds showed good activity against the selected tumor cell lines, with the exception of IGROV1. In particular, compound 5 showed a noteworthy selectivity towards HT29 cells, a malignant colonic cell line, without affecting healthy human intestinal cells. Further studies revealed that 5 caused the cell cycle arrest in the G1 phase and promoted apoptosis in HT29 cells.
Assuntos
Antineoplásicos , Neoplasias , Humanos , Amidas/farmacologia , Analgésicos Opioides/farmacologia , Linhagem Celular Tumoral , Pontos de Checagem do Ciclo Celular , Indóis/farmacologia , Proliferação de Células , Ensaios de Seleção de Medicamentos Antitumorais , Relação Estrutura-Atividade , Estrutura Molecular , ApoptoseRESUMO
A series of naphthoquinones, namely, 1,4-naphthoquinone, menadione, plumbagin, juglone, naphthazarin, and lawsone, were reacted with N-acetyl-L-cysteine, and except for lawsone, which did not react, the related adducts were obtained. After the tuning of the solvent and reaction conditions, the reaction products were isolated as almost pure from the complex reaction mixture via simple filtration and were fully characterized. Therefore, the aim of this work was to evaluate whether the antitumor activity of new compounds of 1,4-naphthoquinone derivatives leads to an increase in ROS in tumor cell lines of cervical carcinoma (HeLa), neuroblastoma (SH-SY5Y), and osteosarcoma (SaOS2, U2OS) and in normal dermal fibroblast (HDFa). The MTT assay was used to assay cell viability, the DCF-DA fluorescent probe to evaluate ROS induction, and cell-cycle analysis to measure the antiproliferative effect. Compounds 8, 9, and 12 showed a certain degree of cytotoxicity towards all the malignant cell lines tested, while compound 11 showed biological activity at higher IC50 values. Compounds 8 and 11 induced increases in ROS generation after 1 h of exposure, while after 48 h of treatment, only 8 induced an increase in ROS formation in HeLa cells. Cell-cycle analysis showed that compound 8 caused an increase in the number of G0/G1-phase cells in the HeLa experiment, while for the U2OS and SH-SY5Y cell lines, it led to an accumulation of S-phase cells. Therefore, these novel 1,4-naphthoquinone derivatives may be useful as antitumoral agents in the treatment of different cancers.
Assuntos
Naftoquinonas , Neuroblastoma , Acetilcisteína/farmacologia , Linhagem Celular Tumoral , Células HeLa , Humanos , Naftoquinonas/metabolismo , Naftoquinonas/farmacologia , Espécies Reativas de Oxigênio/metabolismoRESUMO
A series of regioisomers of the hydroxystearic acid (HSA) was prepared, and the effect of the position of the hydroxyl group along the chain on a panel of human cancer cell lines was investigated. Among the various regioisomers, those carrying the hydroxyl at positions 5, 7, and 9 had growth inhibitor activity against various human tumor cell lines, including CaCo-2, HT29, HeLa, MCF7, PC3, and NLF cells. 10-HSA and 11-HSA showed a very weak effect. 8-HSA did not show inhibitory activity in all cell lines. The biological role of 7-HSA and 9-HSA is widely recognized, while little is known about the effects of 5-HSA. Therefore, the biological effects of 5-HSA in HeLa, HT29, MCF7, and NLF cell lines were investigated using the Livecyte's ptychography technology, which allows correlating changes in proliferation, motility, and morphology as a function of treatment at the same time. 5-HSA not only reduces cell proliferation but also induces changes in cell displacement, directionality, and speed. It is important to characterize the biological effects of 5-HSA, this molecule being an important component of fatty acyl esters of hydroxy fatty acids (FAHFA), a class of endogenous mammalian lipids with noticeable anti-diabetic and anti-inflammatory effects.
Assuntos
Ácidos Graxos , Neoplasias , Animais , Células CACO-2 , Linhagem Celular Tumoral , Proliferação de Células , Ésteres/farmacologia , Ácidos Graxos/farmacologia , Humanos , MamíferosRESUMO
Osteosarcoma (OS) is a malignant disease characterized by poor prognosis due to a high incidence of metastasis and chemoresistance. Recently, Licochalcone A (Lic-A) has been reported as a promising agent against OS. Starting from chalcones selected from a wide in-house library, a new series was designed and synthetized. The antitumor activity of the compounds was tested on the MG63 OS cell line through the innovative Quantitative Phase Imaging technique and MTT assay. To further investigate the biological profile of active derivatives, cell cycle progression and apoptosis induction were evaluated. An earlier and more consistent arrest in the G2-M phase with respect to Lic-A was observed. Moreover, apoptosis was assessed by Annexin V staining as well as by the detection of typical morphological features of apoptotic cells. Among the selected compounds, 1e, 1q, and 1r proved to be the most promising antitumor molecules. This study pointed out that an integrated methodological approach may constitute a valuable platform for the rapid screening of large series of compounds.
Assuntos
Antineoplásicos , Neoplasias Ósseas , Chalcona , Chalconas , Osteossarcoma , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Apoptose , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Proliferação de Células , Chalcona/farmacologia , Chalconas/farmacologia , Chalconas/uso terapêutico , Humanos , Osteossarcoma/patologiaRESUMO
Valorization of wild plants to obtain botanical ingredients could be a strategy for sustainable production of cosmetics. This study aimed to select the rosehip extract containing the greatest amounts of bioactive compounds and to encapsulate it in vesicular systems capable of protecting their own antioxidant activity. Chemical analysis of Rosa canina L. extracts was performed by LC-DAD-MS/MS and 1H-NMR and vitamins, phenolic compounds, sugars, and organic acids were detected as the main compounds of the extracts. Liposomes, prepared by the film hydration method, together with hyalurosomes and ethosomes, obtained by the ethanol injection method, were characterized in terms of vesicle size, polydispersity index, entrapment efficiency, zeta potential, in vitro release and biocompatibility on WS1 fibroblasts. Among all types of vesicular systems, ethosomes proved to be the most promising nanocarriers showing nanometric size (196 ± 1 nm), narrow polydispersity (0.20 ± 0.02), good entrapment efficiency (92.30 ± 0.02%), and negative zeta potential (-37.36 ± 0.55 mV). Moreover, ethosomes showed good stability over time, a slow release of polyphenols compared with free extract, and they were not cytotoxic. In conclusion, ethosomes could be innovative carriers for the encapsulation of rosehip extract.
Assuntos
Rosa , Antioxidantes/química , Lipídeos , Lipossomos/química , Polifenóis/farmacologia , Rosa/química , Espectrometria de Massas em TandemRESUMO
A small library of derivatives carrying a polycyclic scaffold recently identified by us as a new privileged structure in medicinal chemistry was designed and synthesized, aiming at obtaining potent MDR reverting agents also endowed with antitumor properties. In particular, as a follow-up of our previous studies, attention was focused on the role of the spacer connecting the polycyclic core with a properly selected nitrogen-containing group. A relevant increase in reverting potency was observed, going from the previously employed but-2-ynyl- to a pent-3-ynylamino moiety, as in compounds 3d and 3e, while the introduction of a triazole ring proved to differently impact on the activity of the compounds. The docking results supported the data obtained by biological tests, showing, for the most active compounds, the ability to establish specific bonds with P-glycoprotein. Moreover, a multifaceted anticancer profile and dual in vitro activity was observed for all compounds, showing both revertant and antitumor effects on leukemic cells. In this respect, 3c emerged as a "triple-target" agent, endowed with a relevant reverting potency, a considerable antiproliferative activity and a collateral sensitivity profile.
Assuntos
Antracenos/farmacologia , Antineoplásicos/farmacologia , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Succinimidas/farmacologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Antracenos/síntese química , Antracenos/metabolismo , Antineoplásicos/síntese química , Antineoplásicos/metabolismo , Apoptose/efeitos dos fármacos , Hidrocarbonetos Aromáticos com Pontes/síntese química , Hidrocarbonetos Aromáticos com Pontes/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Ligação Proteica , Bibliotecas de Moléculas Pequenas/síntese química , Bibliotecas de Moléculas Pequenas/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Relação Estrutura-Atividade , Succinimidas/síntese química , Succinimidas/metabolismoRESUMO
Biomineralization is the process by which living organisms generate organized mineral crystals. In human cells, this phenomenon culminates with the formation of hydroxyapatite, which is a naturally occurring mineral form of calcium apatite. The mechanism that explains the genesis within the cell and the propagation of the mineral in the extracellular matrix still remains largely unexplained, and its characterization is highly controversial, especially in humans. In fact, up to now, biomineralization core knowledge has been provided by investigations on the advanced phases of this process. In this study, we characterize the contents of calcium depositions in human bone mesenchymal stem cells exposed to an osteogenic cocktail for 4 and 10 days using synchrotron-based cryo-soft-X-ray tomography and cryo-XANES microscopy. The reported results suggest crystalline calcite as a precursor of hydroxyapatite depositions within the cells in the biomineralization process. In particular, both calcite and hydroxyapatite were detected within the cell during the early phase of osteogenic differentiation. This striking finding may redefine most of the biomineralization models published so far, taking into account that they have been formulated using murine samples while studies in human cell lines are still scarce.
Assuntos
Biomineralização/efeitos dos fármacos , Carbonato de Cálcio/farmacologia , Diferenciação Celular/efeitos dos fármacos , Durapatita/farmacologia , Células-Tronco Mesenquimais/citologia , Humanos , Masculino , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/ultraestrutura , Distribuição NormalRESUMO
The roots of two cultivars of Paeonia, namely Paeonia officinalis "Rubra Plena" and Paeonia "Pink Hawaiian Coral", have been extracted with chloroform. The composition of the lipid fraction, analyzed by GC-MS technique, revealed the absence of paeonol and the presence of phenol, benzoic acid, fatty acid-and some sterol-derivatives. The chloroformic extracts have been tested on normal and several cancer cell lines but showed antiproliferative activity only on the ovarian carcinoma and the osteosarcoma. The biological activity of extracts was investigated mainly by confocal microscopy, flow cytometry and quantum phase imaging. The results indicated that the root extracts induced a hyperpolarization of mitochondria and an increase in reactive oxygen species levels, without inducing cell death. These effects are associated to an increased doubling time and a retarded confluence.
Assuntos
Lipídeos/química , Lipídeos/farmacologia , Paeonia/química , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Raízes de Plantas/química , Ácido Benzoico/química , Ácido Benzoico/farmacologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ácidos Graxos/química , Ácidos Graxos/farmacologia , Feminino , Havaí , Células HeLa , Humanos , Células MCF-7 , Mitocôndrias/efeitos dos fármacos , Osteossarcoma/tratamento farmacológico , Neoplasias Ovarianas/tratamento farmacológico , Fenóis/química , Fenóis/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Esteróis/química , Esteróis/farmacologiaRESUMO
At present, there is no vaccine or effective standard treatment for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection (or coronavirus disease-19 (COVID-19)), which frequently leads to lethal pulmonary inflammatory responses. COVID-19 pathology is characterized by extreme inflammation and amplified immune response with activation of a cytokine storm. A subsequent progression to acute lung injury (ALI) or acute respiratory distress syndrome (ARDS) can take place, which is often followed by death. The causes of these strong inflammatory responses in SARS-CoV-2 infection are still unknown. As uncontrolled pulmonary inflammation is likely the main cause of death in SARS-CoV-2 infection, anti-inflammatory therapeutic interventions are particularly important. Fenretinide N-(4-hydroxyphenyl) retinamide is a bioactive molecule characterized by poly-pharmacological properties and a low toxicity profile. Fenretinide is endowed with antitumor, anti-inflammatory, antiviral, and immunomodulating properties other than efficacy in obesity/diabetic pathologies. Its anti-inflammatory and antiviral activities, in particular, could likely have utility in multimodal therapies for the treatment of ALI/ARDS in COVID-19 patients. Moreover, fenretinide administration by pulmonary delivery systems could further increase its therapeutic value by carrying high drug concentrations to the lungs and triggering a rapid onset of activity. This is particularly important in SARS-CoV-2 infection, where only a narrow time window exists for therapeutic intervention.
Assuntos
Anti-Inflamatórios/uso terapêutico , Infecções por Coronavirus/tratamento farmacológico , Fenretinida/uso terapêutico , Pneumonia Viral/tratamento farmacológico , Animais , Anti-Inflamatórios/farmacologia , Betacoronavirus/isolamento & purificação , COVID-19 , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Citocinas , Fenretinida/farmacologia , Humanos , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Pandemias , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Sistema Respiratório/efeitos dos fármacos , Sistema Respiratório/metabolismo , SARS-CoV-2 , Transdução de Sinais/efeitos dos fármacosRESUMO
In this study, we explore the behaviour of intracellular magnesium during bone phenotype modulation in a 3D cell model built to mimic osteogenesis. In addition, we measured the amount of magnesium in the mineral depositions generated during osteogenic induction. A two-fold increase of intracellular magnesium content was found, both at three and seven days from the induction of differentiation. By X-ray microscopy, we characterized the morphology and chemical composition of the mineral depositions secreted by 3D cultured differentiated cells finding a marked co-localization of Mg with P at seven days of differentiation. This is the first experimental evidence on the presence of Mg in the mineral depositions generated during biomineralization, suggesting that Mg incorporation occurs during the bone forming process. In conclusion, this study on the one hand attests to an evident involvement of Mg in the process of cell differentiation, and, on the other hand, indicates that its multifaceted role needs further investigation.
Assuntos
Magnésio/análise , Osteogênese , Fósforo/análise , Técnicas de Cultura de Células/métodos , Diferenciação Celular , Linhagem Celular Tumoral , Humanos , Magnésio/metabolismo , Fósforo/metabolismoRESUMO
The focus of this work was to prepare Spanish Broom, flax, and hemp dressings impregnated with glycyrrhetinic acid (GA) liposomes or hyalurosomes to promote the healing process and protect the skin wounds. Vesicles were prepared by the film hydration method and characterized in terms of size, particle size distribution, ζ potential, encapsulation efficiency, in vitro release, and biocompatibility on 3T3 fibroblasts. Loaded liposomes and hyalurosomes showed nanometric size (355 ± 19 nm and 424 ± 32 nm, respectively), good size distribution (lower than 0.3), and appropriate encapsulation efficiency (58.62 ± 3.25% and 59.22 ± 8.18%, respectively). Hyalurosomes showed good stability during the storage period, which can be correlated to the negative ζ potential, and allowed a fast and complete release of GA. Preliminary biological studies revealed that both kinds of loaded vesicles were not cytotoxic and that hyalurosomes could exert a slight stimulating effect on fibroblast proliferation. Finally, in vitro release studies from the different dressings impregnated with the loaded vesicles demonstrated that a high amount of GA could be reached at the wound site after 60 min from application. In conclusion, the results suggested that the developed dressings, especially those impregnated with hyalurosomes, can be efficiently used to promote the healing process.
Assuntos
Cannabis/química , Linho/química , Ácido Glicirretínico/química , Ácido Hialurônico/química , Lipossomos/química , Spartium/química , Cicatrização/efeitos dos fármacos , Células 3T3 , Animais , Materiais Biocompatíveis , Ciclo Celular , Proliferação de Células , Sobrevivência Celular , Portadores de Fármacos , Fibroblastos/efeitos dos fármacos , Camundongos , Tamanho da Partícula , Pele/lesõesRESUMO
Nine compounds bearing pyridinyl (or piperidinyl, benzimidazolyl, benzotriazolyl) groups bound to an azelayl moiety through an amide bond were synthesized. The structural analogy with some histone deacetylase inhibitors inspired their syntheses, seeking new selective histone deacetylase inhibitors (HDACi). The azelayl moiety recalls part of 9-hydroxystearic acid, a cellular lipid showing antiproliferative activity toward cancer cells with HDAC as a molecular target. Azelayl derivatives bound to a benzothiazolyl moiety further proved to be active as HDACi. The novel compounds were tested on a panel of both normal and tumor cell lines. Non-specific induction of cytotoxicity was observed in the normal cell line, while three of them induced a biological effect only on the osteosarcoma (U2OS) cell line. One of them induced a change in nuclear shape and size. Cell-cycle alterations are associated with post-transcriptional modification of both H2/H3 and H4 histones. In line with recent studies, revealing unexpected HDAC7 function in osteoclasts, molecular docking studies on the active molecules predicted their proneness to interact with HDAC7. By reducing side effects associated with the action of the first-generation inhibitors, the herein reported compounds, thus, sound promising as selective HDACi.
Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Compostos Aza/química , Ácidos Dicarboxílicos/química , Compostos Heterocíclicos/química , Neoplasias Ósseas , Linhagem Celular Tumoral , Técnicas de Química Sintética , Relação Dose-Resposta a Droga , Humanos , Ligação de Hidrogênio , Modelos Moleculares , Conformação Molecular , Estrutura Molecular , Osteossarcoma , Relação Estrutura-AtividadeRESUMO
The first detailed analysis of FLIM applications for Mg cell imaging is presented. We employed the Mg-sensitive fluorescent dye named DCHQ5, a derivative of diaza-18-crown-6 ethers appended with two 8-hydroxyquinoline groups, to perform fluorescence lifetime imaging in control and Mg deprived SaOS-2 live cells, which contain different concentrations of magnesium. We found that the lifetime maps are almost uniform all over the cells and, most relevantly, we showed that the ratio of the amplitude terms is related to the magnesium intracellular concentration.
Assuntos
Neoplasias Ósseas/metabolismo , Magnésio/metabolismo , Imagem Óptica/métodos , Osteossarcoma/metabolismo , Espectrometria de Fluorescência/métodos , Humanos , Magnésio/análise , Células Tumorais CultivadasRESUMO
Magnesium (Mg) is crucial for bone health. Low concentrations of Mg inhibit the activity of osteoblasts while promoting that of osteoclasts, with the final result of inducing osteopenia. Conversely, little is known about the effects of high concentrations of extracellular Mg on osteoclasts and osteoblasts. Since the differentiation and activation of these cells is coordinated by vitamin D3 (VD3), we investigated the effects of high extracellular Mg, as well as its impact on VD3 activity, in these cells. U937 cells were induced to osteoclastic differentiation by VD3 in the presence of supra-physiological concentrations (>1 mM) of extracellular Mg. The effect of high Mg concentrations was also studied in human bone-marrow-derived mesenchymal stem cells (bMSCs) induced to differentiate into osteoblasts by VD3. We demonstrate that high extra-cellular Mg levels potentiate VD3-induced osteoclastic differentiation, while decreasing osteoblastogenesis. We hypothesize that Mg might reprogram VD3 activity on bone remodeling, causing an unbalanced activation of osteoclasts and osteoblasts.
Assuntos
Diferenciação Celular , Colecalciferol/metabolismo , Magnésio/metabolismo , Osteoblastos/citologia , Osteoblastos/metabolismo , Osteoclastos/citologia , Osteoclastos/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Colecalciferol/farmacologia , Perfilação da Expressão Gênica , Humanos , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Monócitos/citologia , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Osteoblastos/efeitos dos fármacos , Osteoclastos/efeitos dos fármacos , Células U937RESUMO
The quantification of elemental concentration in cells is usually performed by analytical assays on large populations missing peculiar but important rare cells. The present article aims at comparing the elemental quantification in single cells and cell population in three different cell types using a new approach for single cells elemental analysis performed at sub-micrometer scale combining X-ray fluorescence microscopy and atomic force microscopy. The attention is focused on the light element Mg, exploiting the opportunity to compare the single cell quantification to the cell population analysis carried out by a highly Mg-selective fluorescent chemosensor. The results show that the single cell analysis reveals the same Mg differences found in large population of the different cell strains studied. However, in one of the cell strains, single cell analysis reveals two cells with an exceptionally high intracellular Mg content compared with the other cells of the same strain. The single cell analysis allows mapping Mg and other light elements in whole cells at sub-micrometer scale. A detailed intensity correlation analysis on the two cells with the highest Mg content reveals that Mg subcellular localization correlates with oxygen in a different fashion with respect the other sister cells of the same strain. Graphical abstract Single cells or large population analysis this is the question!
Assuntos
Corantes Fluorescentes/química , Magnésio/análise , Microscopia de Fluorescência/métodos , Imagem Óptica/métodos , Análise de Célula Única/métodos , Contagem de Células , Linhagem Celular Tumoral , Células Endoteliais da Veia Umbilical Humana , Humanos , Síncrotrons , Raios XRESUMO
Magnesium plays a pivotal role in energy metabolism and in the control of cell growth. While magnesium deprivation clearly shapes the behavior of normal and neoplastic cells, little is known on the role of this element in cell differentiation. Here we show that magnesium deficiency increases the transcription of multipotency markers and tissue-specific transcription factors in human adipose-derived mesenchymal stem cells exposed to a mixture of natural molecules, i.e., hyaluronic, butyric and retinoid acids, which tunes differentiation. We also demonstrate that magnesium deficiency accelerates the osteogenic differentiation of human bone marrow-derived mesenchymal stem cells. We argue that magnesium deprivation generates a stressful condition that modulates stem cell plasticity and differentiation potential. These studies indicate that it is possible to remodel transcription in mesenchymal stem cells by lowering extracellular magnesium without the need for genetic manipulation, thus offering new hints for regenerative medicine applications.
Assuntos
Magnésio/metabolismo , Células-Tronco Mesenquimais/metabolismo , Transcrição Gênica , Tecido Adiposo/citologia , Adulto , Células da Medula Óssea/citologia , Ciclo Celular/genética , Diferenciação Celular/genética , Feminino , Regulação da Expressão Gênica , Humanos , Osteogênese/genética , Espécies Reativas de Oxigênio/metabolismoRESUMO
The biological function of a chemical element in cells not only requires the determination of its intracellular quantity, but also the spatial distribution of its concentration. Different strategies can be employed to quantify and map the intracellular concentration of elements in single cells. The assessment of the intracellular elemental concentration, which is the relevant information, requires the measurement of cell volume. This challenging and demanding task requires combining different techniques allowing gathering of both morphological and compositional information on the same cell. Moreover, the need to analyse samples more similar to their natural state requires complex hardware equipment, and supplementary efforts in preparation protocols. Nevertheless, the response to the question: "where is it and how much?" is worth all these efforts. This review aims at providing an insight into the recent and most advanced techniques and strategies for quantifying and mapping chemical elements in single cells. We describe and discuss indirect detection techniques (label based) which make use of fluorescent dyes, and direct ones (label free), such as particle induced X-ray emission, proton backscattering spectrometry, scanning transmission ion spectrometry, nano-secondary ion mass spectrometry, X-ray fluorescence microscopy, complemented by X-ray imaging.