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- Lower melanoma pulmonary metastatic burden in obese mice: role of FGF-21Publication . Fonseca, Magda; Soares, Raquel; Coelho, PedroObesity is a risk factor for malignant melanoma. The lungs are main target organs for metastization and their immune response is a key modulator of this mechanism. The concept that the metastatic potential of some types of cancer is reduced or inhibited by obesity, known as the obesity paradox, drives major concerns on the prognosis of metastasized patients. The aim of this study was to investigate how high-fat diet (HFD)-induced obesity affects melanoma metastization. C57Bl6/J mice were fed with HFD or standard diet for 180 days and inoculated intravenously with B16F10 melanoma cells. Upon 21 days of inoculation, lung tissue of overweight and lean mice was assessed for histology and immunohistochemistry assays. Adipokine antibody arrays were performed in mice serum. In vitro RAW 264.7 macrophage cultures were established and incubated with FGF-21 and/or lipopolysaccharide (LPS). Conditioned media was added to B16F10 cells for viability quantification. HFD-fed mice presented a reduced number of metastases with lower proliferative rates. The high content of inflammatory foci observed in noninoculated obese mice was significantly decreased upon B16F10 inoculation, concurrent with a slight fibrosis reduction. Plasma levels of fibroblast growth factor-21 (FGF-21), an endocrine regulator, were elevated in noninoculated HFD mice and the expression of FGF receptor 1 (FGFR-1) was significantly upregulated after inoculation. FGF-21 reduced melanoma viability in LPS-stimulated macrophages. Altogether, these findings suggest that higher amounts of FGF-21 are able to counterbalance the proinflammatory effects associated with obesity, protecting the lungs from melanoma metastization.
- Adipocyte-released factors enhance melanocyte’s proliferation and motilityPublication . Fernandes, Rúben; Coelho, Pedro; Almeida, Joana; Prudêncio, Cristina; Soares, RaquelObesity, favored by the modern lifestyle, acquired epidemic proportions nowadays. Obesity has been associated with various major causes of death and morbidity including malignant neoplasms. Cutaneous melanoma incidence rates have also been increasing uring the last four decades in several countries. Obesity involvement in melanoma etiology has been recognized, but the implicated mechanisms remain unclear. We propose to address the above relationship and investigate the mechanism interplaying between obesity and an increased risk of melanoma onset.
- Quinoxaline-1, 4-dioxide derivatives inhibitory action in melanoma and brain tumor cellsPublication . Silva, Liliana; Coelho, Pedro; Soares, Raquel; Prudêncio, Cristina; Vieira, MónicaQuinoxaline-1,4-dioxide derivatives are synthetic heterocyclic compounds with multiple biological and pharmacological effects. In this study, we investigated the bioactivity of five quinoxaline-1,4-di-N-oxides derivatives in different animal cell lines.
- Effect of umbilical cord mesenchymal stem cells secretome in melanomaPublication . Accioly, Gustavo; Aguiar, Gonçalo; Areal, Lara; Costa, Pedro; Coelho, Pedro; Gomes, AndreiaMelanoma of the skin is one of the most prominent and fastest growing malignancies. More than 300 thousand diagnosed cases and 57 thousand deaths in 2020 worldwide and roughly 517 thousand cases of were registered during the 2015-2020 period. Melanoma is generally regarded as an aggressive and unpredictable cancer whose conventional therapies, such as local excision, chemotherapy and immunotherapy, have encountered difficulties to prevent larger scale-tumours and metastasis, as well as overcome recurrence and development of drug resistance. Stem cellbased therapies have been studied as interesting therapeutical approaches for cancer whenever conventional therapy fails to impede its progression. That is owing to the anti-proliferative and immunomodulatory capacity of some SC, being one of the major examples, Mesenchymal Stem Cells. Due to its high abundance, well defined extraction and expansion protocols as well as documented anti-tumorigenic characteristics, Umbilical Cord derived Mesenchymal Stem Cells (UC-MSC) have been observed as a promising candidate for Melanoma treatment, specially through acellular therapy using its secretome. MSC secretome is defined as the set of MSCs-derived bioactive factors available extracellularly and is responsible for the major therapeutic effects of MSCs, namely in oncological pathologies. In this study we hypothesize the ability of UC-MSC’s secretome to inhibit Melanoma growth in vitro. UC-MSC secretome, in the form of conditioned medium (CM), was obtained by extraction from selected umbilical cords and expansion of while murine melanoma cell line B16-F10 culture was established. After treating melanoma cells with different concentrations of CM (100%, 50% and 25%), common cancer hallmarks such as cell viability, motility, colony formation and cell interactions were assessed through MTT, Wound Healing and Colony formation and Hanging-Drop assays, respectively. General analysis of viability and motility showed no statistically significant difference between treated and control groups as well as no concentration-dependent effect whereas formation of cellular aggregates follows an inhibition trend on the treated groups. These results put into perspective the effect of secretome of UC-MSCs. Moreover, further larger scale studies are needed for deeper understanding of MSC secretome mechanisms of action, therefore enabling their use in acellular therapies against melanoma in the future.
- Secretome of umbilical cord mesenchymal stem cells: Potential effects on melanomaPublication . Fernandes, Pablo ; Gomes, Andreia ; Coelho, PedroMelanoma, a tumor resulting from the malignant transformation of melanocytes, is characterized by its aggressive nature and propensity to metastasize. Current treatment options for advanced melanoma are limited and mostly ineffective, highlighting the need for novel therapeutic approaches. Mesenchymal stem cells (MSCs) have increased considerable attention due to their anti-cancer and immunomodulatory properties. In particular, human umbilical cord MSCs (hUCMSCs) have shown promise in various therapeutic applications.
- Melanoma and obesity: Should antioxidant vitamins be addressed?Publication . Oliveira, Sofia; Coelho, Pedro; Prudêncio, Cristina; Vieira, Mónica; Soares, Raquel; Guerreiro, Susana G.; Fernandes, RúbenMelanoma is an aggressive form of skin cancer refractory to conventional therapies. Obesity has reached epidemic dimensions acting as a risk factor for several cancer types, such as melanoma. Several reactive species of oxygen are also involved in melanoma initiation and progression. Low levels of antioxidant content and/or activity in lightly pigmented cells could expose them to an extremely oxidative environment and rise the susceptibility to oxidative damage and consequently loss of cell homeostasis. Despite the knowledge about melanoma biology, pathogenesis and developed therapies, is extremely important to understand the antioxidant modulation of melanoma under an environment of obesity, especially the effect of some natural compounds of the diet, such as antioxidant vitamins A, C and E and selenium in order to establish alternatives to conventional therapies, which are known to be ineffective against melanoma.
- Effect of Adipocyte Secretome in Melanoma Progression and Vasculogenic MimicryPublication . Coelho, Pedro; Almeida, Joana; Prudêncio, Cristina; Fernandes, Rúben; Soares, RaquelObesity, favored by the modern lifestyle, acquired epidemic proportions nowadays. Obesity has been associated with various major causes of death and morbidity including malignant neoplasms. This increased prevalence has been accompanied by a worldwide increase in cutaneous melanoma incidence rates during the last decades. Obesity involvement in melanoma aetiology has been recognized, but the implicated mechanisms remain unclear. In the present study, we address this relationship and investigate the influence of adipocytes secretome on B16-F10 and MeWo melanoma cell lines. Using the 3T3-L1 adipocyte cell line, as well as ex vivo subcutaneous (SAT) and visceral (VAT) adipose tissue conditioned medium, we were able to show that adipocyte-released factors play a dual role in increasing melanoma cell overall survival, both by enhancing proliferation and decreasing apoptosis. B16-F10 cell migration and cell-cell and cell-matrix adhesion capacity were predominantly enhanced in the presence of SAT and VAT released factors. Melanocytes morphology and melanin content were also altered by exposure to adipocyte conditioned medium disclosing a more dedifferentiated phenotype of melanocytes. In addition, exposure to adipocyte-secreted molecules induced melanocytes to rearrange, on 3D cultures, into vessel-like structures, and generate characteristic vasculogenic mimicry patterns. These findings are corroborated by the released factors profile of 3T3-L1, SAT, and VAT assessed by microarrays, and led us to highlight the mechanisms by which adipose secretome from sub-cutaneous or visceral depots promote melanoma progression.
- Oxidative Stress Modulation and Radiosensitizing Effect of Quinoxaline-1,4-Dioxides DerivativesPublication . Silva, Liliana; Coelho, Pedro; Teixeira, Dulce; Monteiro, Armanda; Pinto, Gabriela; Soares, Raquel; Prudêncio, Cristina; Vieira, MónicaQuinoxaline-1,4-dioxide (QNX) derivatives are synthetic heterocyclic compounds with multiple biological and pharmacological effects. In this study, we investigated the oxidative status of quinoxaline-1,4-dioxides derivatives in modulating melanoma and glioma cell lines, based on previous results from the research group and their capability to promote cell damage by the production of Reactive Oxygen Species (ROS).
- Development of a stable melanoma dual reporter cell line expressing Luciferase and GFPPublication . Aguiar, Gonçalo; Torres, Sílvia; Prudêncio, Cristina; Soares, Raquel; Coelho, Pedro; Prudêncio, Cristina; Coelho, PedroMelanoma is the most aggressive and lethal form of skin cancer, with a high risk of metastatic spread. Obesity is recognized as a risk factor for various types of cancer. However, regarding melanoma, this association remains controversial. Obesity might act as a double-edged sword in melanoma, promoting primary tumour growth but at the same time limiting metastatic spread - the "obesity paradox”. Herein, we aimed to create a stable murine B16F10 melanoma cell line expressing both firefly luciferase (Luc) and green fluorescent protein (GFP), which will later be engrafted into diet induced-obesity animal model for future in vivo studies. B16F10-Luc-GFP cells were generated by transfection with premade lentiviral particles, featuring a construct with Luc and GFP under a cytomegalovirus promoter and mediated by a F2A element. The antibiotic selection marker (puromycin) is expressed under a Rous sarcoma virus promoter. Afterwards, the transfected cells were selected with 1 μg/ml of puromycin. The clones with the highest levels of GFP-positive cells and GFP fluorescence were purified by two rounds of cell sorting and submitted to fluorescence and bioluminescence quantification, morphology, injury, BrdU incorporation, 7-AAD, and PI cell cycle assays and compared to the parental cell line. B16F10-Luc-GFP were successfully generated, and both GFP fluorescence and D-luciferin bioluminescence are present and proportional to cell density. As expected, the parental cell line didn’t display GFP or Luc activities. Moreover, transduced cells exhibit similar morphology, motility, proliferation, viability, and cell cycle progression as B16F10 cells. Conclusions: Altogether, the future engraftment of B16F10-LucGFP in obese mice, will improve melanoma research models, enabling the in vivo and ex vivo visualization of primary tumours and metastasis, providing a better understanding of the underlying molecular mechanisms, to clarify the “obesity paradox” in melanoma.