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Fucoxanthin (Fx) is a carotenoid derived from marine organisms that exhibits anticancer activities. However, its role as a potential drug adjuvant in breast cancer (BC) treatment is still poorly explored. Firstly, this study investigated the cytotoxic effects of Fx alone and combined with doxorubicin (Dox) and cisplatin (Cis) on a panel of 2D-cultured BC cell lines (MCF7, SKBR3 and MDA-MB-231) and one non-tumoral cell line (MCF12A). Fucoxanthin induced cytotoxicity against all the cell lines and potentiated Dox cytotoxic effects towards the SKBR3 and MDA-MB-231 cells. The combination triggering the highest cytotoxicity (Fx 10 µM + Dox 1 µM in MDA-MB-231) additionally showed significant induction of cell death and genotoxic effects, relative to control. In sequence, the same combination was tested on 3D cultures using a multi-endpoint approach involving bioactivity assays and microscopy techniques. Similar to 2D cultures, the combination of Fx and Dox showed higher cytotoxic effects on 3D cultures compared to the isolated compounds. Furthermore, this combination increased the number of apoptotic cells, decreased cell proliferation, and caused structural and ultrastructural damages on the 3D models. Overall, our findings suggest Fx has potential to become an adjuvant for Dox chemotherapy regimens in BC treatment.
Fernanda Malhão; Ana Macedo; Carla Costa; Eduardo Rocha; Alice Ramos. Fucoxanthin Holds Potential to Become a Drug Adjuvant in Breast Cancer Treatment: Evidence from 2D and 3D Cell Cultures. Molecules 2021, 26, 4288 .
AMA StyleFernanda Malhão, Ana Macedo, Carla Costa, Eduardo Rocha, Alice Ramos. Fucoxanthin Holds Potential to Become a Drug Adjuvant in Breast Cancer Treatment: Evidence from 2D and 3D Cell Cultures. Molecules. 2021; 26 (14):4288.
Chicago/Turabian StyleFernanda Malhão; Ana Macedo; Carla Costa; Eduardo Rocha; Alice Ramos. 2021. "Fucoxanthin Holds Potential to Become a Drug Adjuvant in Breast Cancer Treatment: Evidence from 2D and 3D Cell Cultures." Molecules 26, no. 14: 4288.
Brown trout is an environmental freshwater sentinel species and is economically important for recreational fishing and aquaculture. Despite that, there is limited knowledge regarding morpho-physiological variations in adults throughout the reproductive cycle. Thus, this study aimed to analyze the fitness and gonadal maturation of cultured adult brown trout in four reproductive phases (spawning capable—December, regressing—March, regenerating—July, and developing—November). The systematic evaluation of males and females was based on biometric, biochemical, and hormonal parameters, along with a histomorphological grading of gonads and the immunophenotype location of key steroidogenic enzymes. The total weight and lengths reached the lowest levels in December. Gonad weights were higher in December and November, while the opposite pattern was found for liver weights. The lowest levels of cholesterol and total protein were also noted during those stages. The 11-ketotestosterone (11-KT) and testosterone (T) for males, and estradiol (E2) and T for females, mostly explained the hormonal variations. The immunohistochemistry of cytochrome P450c17 (CYP17-I), aromatase (CYP19), and 17β-hydroxysteroid dehydrogenase (17β-HSD) showed sex and site-specific patterns in the distinct reproductive phases. The sex- and season-specific changes generated discriminative multi-parameter profiles, serving as a tool for environmental and aquaculture surveys.
Diana Santos; Eduardo Rocha; Fernanda Malhão; Célia Lopes; José Gonçalves; Tânia Madureira. Multi-Parametric Portfolio to Assess the Fitness and Gonadal Maturation in Four Key Reproductive Phases of Brown Trout. Animals 2021, 11, 1290 .
AMA StyleDiana Santos, Eduardo Rocha, Fernanda Malhão, Célia Lopes, José Gonçalves, Tânia Madureira. Multi-Parametric Portfolio to Assess the Fitness and Gonadal Maturation in Four Key Reproductive Phases of Brown Trout. Animals. 2021; 11 (5):1290.
Chicago/Turabian StyleDiana Santos; Eduardo Rocha; Fernanda Malhão; Célia Lopes; José Gonçalves; Tânia Madureira. 2021. "Multi-Parametric Portfolio to Assess the Fitness and Gonadal Maturation in Four Key Reproductive Phases of Brown Trout." Animals 11, no. 5: 1290.
Herein, sixteen priority sixteen priority polycyclic aromatic hydrocarbons (PAHs) and seven polychlorinated biphenyls (PCBs), considered seawater quality indicators, were quantified in four beaches of the Iberian (Portuguese) northwest coastline, including one ornithological reserve. These seashores are close to an important industrial region and a harbour. Water samples were extracted for PAHs and PCBs in their aqueous phase (DAP) and suspended particulate matter (SPM) and analysed by GC-MS/MS. Annual average concentrations reached ∼8 ng/L (DAP) and ∼100 ng/g dw (SPM) for PAHs and ∼3 ng/L (DAP) and ∼19 ng/g dw (SPM) for PCBs. PAHs and PCBs had fluctuations suggestive of seasonality. The possible origins of PAHs point to pyrogenic (forest fires) and petrogenic activities linked to maritime transports. The source(s) of PCBs appear to be related with leaks from waste sorting centres located in the vicinity of the sampling sites, together with the burning of contaminated materials. Toxic equivalents (TEQs) suggested a low carcinogenic potential for PAHs in seawater samples (∼0.3 ng/L and ∼4 ng/g dw, for DAP and SPM). Evaluation of risk coefficients for Σ16 PAHs revealed "Low-risk" of both matrices in all sampling areas. PCBs thyroid toxicity equivalents (TEQs-TH), suggested a low impact on biota (∼7.0E-05 ng/L and ∼5.2E-04 ng/g dw, for DAP and SPM). However, in vivo acute assays with Artemia salina exposed to environmental concentrations of PAHs, PCBs, and mixtures, confirmed the theoretical approach, showing that this coastline is not "Risk-free". This fact calls for further toxicological approaches to fully understand the risks posed by these compounds locally.
Maria João Rocha; Ana Beatriz Ribeiro; Diana Campos; Eduardo Rocha. Temporal-spatial survey of PAHs and PCBs in the Atlantic Iberian northwest coastline, and evaluation of their sources and risks for both humans and aquatic organisms. Chemosphere 2021, 279, 130506 .
AMA StyleMaria João Rocha, Ana Beatriz Ribeiro, Diana Campos, Eduardo Rocha. Temporal-spatial survey of PAHs and PCBs in the Atlantic Iberian northwest coastline, and evaluation of their sources and risks for both humans and aquatic organisms. Chemosphere. 2021; 279 ():130506.
Chicago/Turabian StyleMaria João Rocha; Ana Beatriz Ribeiro; Diana Campos; Eduardo Rocha. 2021. "Temporal-spatial survey of PAHs and PCBs in the Atlantic Iberian northwest coastline, and evaluation of their sources and risks for both humans and aquatic organisms." Chemosphere 279, no. : 130506.
Purpose: The aims of this work were a) to describe the histology of the lacrimal gland (LG) and cornea induced by an adenovirus (Ad) vector encoding the human erythropoietin (Epo) gene delivered to the LG and b) to evaluate the therapeutic potential of this strategy to prevent benzalkonium chloride (BAK) corneal toxicity. Methods: Structure and function of male Wistar rats LG were compared in the groups: 1) naïve control and 2) Ad-hEpo in the right LG (RLG). The protective response against BAK eye drops was compared among the groups 1) naïve control, 2) BAK in the right eye, 3) Ad-hEpo RLG + BAK and 4) Ad-hEpo in the right salivary gland (RSG)+BAK. Ad-hEpo groups received an injection of AdLTR2EF1a-hEPO (25 ul, 1010 particles/ml) in the right LG or SG (positive control). The BAK groups received 0.2% BAK in the right cornea twice a day. The tests applied after 7 days, included tear secretion, hEPO mRNA detection by qRT-PCR, LG and cornea histology, LG ELISA for cytokines and hematocrit. Results: hEPO mRNA was present in the Ad-hEpo RLG and RSG, but not kidney or liver samples (negative controls). TNF-α and IL-1β increased in the LG exposed to Ad-hEpo compared to naïve control (p = .0115 and p = .0397, respectively). BAK reduced tear secretion, but this reduction was prevented by Ad-hEpo RLG+BAK and Ad-hEpo RSG+BAK (p = .017). The corneal epithelia were thinner in the BAK-treated groups independent of Ad-hEpo (p = .0009). Hematocrit increased only in the Ad-hEpo RSG group (p = .01). Conclusions: Ad-hEpo infection of rat LG and SG induces local, but only the SG infection induced systemic changes in rats. Importantly, Ad-hEpo attenuated the BAK-mediated toxic reduction in tear flow. Future studies must consider viral vector tissue tropism, biodistribution and effective therapeutic gene products for ocular surface diseases.
Lara Cristina Dias; Changyu Zheng; Adriana De Andrade Batista Murashima; Ana Carolina Dias; Marina Zilio Fantucci; Luiz Fernando Nominato; Lilian Eslaine Costa Mendes da Silva; Eduardo Melani Rocha. Lacrimal Gland as a Target Organ for Adenovirus Gene Therapy Encoding Erythropoietin for Dry Eye Induced by Benzalkonium Chloride. Current Eye Research 2021, 46, 1314 -1319.
AMA StyleLara Cristina Dias, Changyu Zheng, Adriana De Andrade Batista Murashima, Ana Carolina Dias, Marina Zilio Fantucci, Luiz Fernando Nominato, Lilian Eslaine Costa Mendes da Silva, Eduardo Melani Rocha. Lacrimal Gland as a Target Organ for Adenovirus Gene Therapy Encoding Erythropoietin for Dry Eye Induced by Benzalkonium Chloride. Current Eye Research. 2021; 46 (9):1314-1319.
Chicago/Turabian StyleLara Cristina Dias; Changyu Zheng; Adriana De Andrade Batista Murashima; Ana Carolina Dias; Marina Zilio Fantucci; Luiz Fernando Nominato; Lilian Eslaine Costa Mendes da Silva; Eduardo Melani Rocha. 2021. "Lacrimal Gland as a Target Organ for Adenovirus Gene Therapy Encoding Erythropoietin for Dry Eye Induced by Benzalkonium Chloride." Current Eye Research 46, no. 9: 1314-1319.
Despite of physiological and toxicological relevance, the potential of androgens to influence fish lipid metabolism remains poorly explored. Here, brown trout primary hepatocytes were exposed to six concentrations (1 nM to 100 μM) of dihydrotestosterone (DHT) and testosterone (T), to assess changes in the mRNA levels of genes covering diverse lipid metabolic pathways. Acsl1, essential for fatty acid activation, was up-regulated by T and DHT, whereas the lipogenic enzymes FAS and ACC were up-regulated by the highest (100 μM) concentration of T and DHT, respectively. ApoA1, the major component of high-density lipoprotein (HDL), was down-regulated by both androgens. PPARγ, linked to adipogenesis and peroxisomal β-oxidation, was down-regulated by T and DHT, while Acox1–3I, rate-limiting in peroxisomal β-oxidation, was down-regulated by T. Fabp1, StAR and LPL were not altered. Our findings suggest that androgens may impact on lipid transport, adipogenesis and fatty acid β-oxidation and promote lipogenesis in fish liver.
Célia Lopes; Eduardo Rocha; Inês L. Pereira; Tânia V. Madureira. Deciphering influences of testosterone and dihydrotestosterone on lipid metabolism genes using brown trout primary hepatocytes. Aquatic Toxicology 2021, 235, 105819 .
AMA StyleCélia Lopes, Eduardo Rocha, Inês L. Pereira, Tânia V. Madureira. Deciphering influences of testosterone and dihydrotestosterone on lipid metabolism genes using brown trout primary hepatocytes. Aquatic Toxicology. 2021; 235 ():105819.
Chicago/Turabian StyleCélia Lopes; Eduardo Rocha; Inês L. Pereira; Tânia V. Madureira. 2021. "Deciphering influences of testosterone and dihydrotestosterone on lipid metabolism genes using brown trout primary hepatocytes." Aquatic Toxicology 235, no. : 105819.
Anthropogenic activities can introduce polycyclic aromatic hydrocarbons (PAHs) in coastal waters. Here, 16 priority PAHs were quantified by gas chromatography-mass spectroscopy (GC–MS) in seawater samples from three Iberian Atlantic habitats. Results showed global concentrations of ≅9 ng/L in the aqueous phase (DAP) and ≅94 ng/g, dw in suspended particulate matter (SPM). The identified sources were both petrogenic and pyrogenic. Nonetheless, the application of several quantitative approaches, including Principal Component Analysis, point to distinct PAHs inputs amongst the sampling areas and demonstrate that sources in S1 (fire forests/sea harbor) are different from those at S2 and S3 (oil refinery/sea harbor). The calculated carcinogenicity potential in both DAP and SPM was low for humans (high percentage of Group 3 PAHs). However, both toxic equivalent factors (TEQs) and environmental risks coefficients (RQs) reveal that the evaluated areas are not risk-free, as confirmed by the practical Artemia salina acute-test assay.
Maria João Rocha; Eduardo Rocha. Concentrations, sources and risks of PAHs in dissolved and suspended material particulate fractions from the Northwest Atlantic Coast of the Iberian Peninsula. Marine Pollution Bulletin 2021, 165, 112143 .
AMA StyleMaria João Rocha, Eduardo Rocha. Concentrations, sources and risks of PAHs in dissolved and suspended material particulate fractions from the Northwest Atlantic Coast of the Iberian Peninsula. Marine Pollution Bulletin. 2021; 165 ():112143.
Chicago/Turabian StyleMaria João Rocha; Eduardo Rocha. 2021. "Concentrations, sources and risks of PAHs in dissolved and suspended material particulate fractions from the Northwest Atlantic Coast of the Iberian Peninsula." Marine Pollution Bulletin 165, no. : 112143.
Seaweed bioactive compounds have shown anticancer activities in in vitro and in vivo studies. However, tests remain limited, with conflicting results, and effects in combination with anticancer drugs are even scarcer. Here, the cytotoxic effects of five seaweed compounds (astaxanthin, fucoidan, fucosterol, laminarin, and phloroglucinol) were tested alone and in combination with anticancer drugs (cisplatin—Cis; and doxorubicin—Dox), in breast cell lines (three breast cancer (BC) subtypes and one non-tumoral). The combinations revealed situations where seaweed compounds presented potentiation or inhibition of the drugs’ cytotoxicity, without a specific pattern, varying according to the cell line, concentration used for the combination, and drug. Fucosterol was the most promising compound, since: (i) it alone had the highest cytotoxicity at low concentrations against the BC lines without affecting the non-tumoral line; and (ii) in combination (at non-cytotoxic concentration), it potentiated Dox cytotoxicity in the triple-negative BC cell line. Using a comparative approach, monolayer versus 3D cultures, further investigation assessed effects on cell viability and proliferation, morphology, and immunocytochemistry targets. The cytotoxic and antiproliferative effects in monolayer were not observed in 3D, corroborating that cells in 3D culture are more resistant to treatments, and reinforcing the use of more complex models for drug screening and a multi-approach that should include histological and ICC analysis.
Fernanda Malhão; Alice Ramos; Ana Macedo; Eduardo Rocha. Cytotoxicity of Seaweed Compounds, Alone or Combined to Reference Drugs, against Breast Cell Lines Cultured in 2D and 3D. Toxics 2021, 9, 24 .
AMA StyleFernanda Malhão, Alice Ramos, Ana Macedo, Eduardo Rocha. Cytotoxicity of Seaweed Compounds, Alone or Combined to Reference Drugs, against Breast Cell Lines Cultured in 2D and 3D. Toxics. 2021; 9 (2):24.
Chicago/Turabian StyleFernanda Malhão; Alice Ramos; Ana Macedo; Eduardo Rocha. 2021. "Cytotoxicity of Seaweed Compounds, Alone or Combined to Reference Drugs, against Breast Cell Lines Cultured in 2D and 3D." Toxics 9, no. 2: 24.
Triple-negative breast cancer (TNBC) has the poorest BC prognosis, chemotherapy being the mainstream treatment. Prior studies revealed potential synergistic effects of natural compounds with drugs, such as doxorubicin (Dox), frequently used for TNBC. Fucosterol (Fct), a phytosterol of brown seaweeds, because of its antioxidant and antitumor effects, is one of these promising compounds. Using a TNBC cell line (MDA-MB-231), we aimed to test the effects of Fct alone and in combination with Dox on cell viability and proliferation, in monolayer and three-dimensional (3D) cultures. At this stage of the research, data demonstrated that Fct (5 µM) alone did not affect cell viability and proliferation. In monolayer, Dox (≥1 µM) decreased cell viability and proliferation, while in 3D only cell viability was affected at Dox 5 µM. The combination of Fct/Dox (5/0.1 µM), in monolayer, significantly decreased cell viability and proliferation, differing from the control and both compounds alone; hence, this suggests that Fct enhanced the Dox effect. These promising Fct effects in monolayer were not observed in 3D. We suggest that Fct may increase Dox effects, under certain conditions. Our results corroborate other studies reporting more treatment resistance of cells in 3D culture, reinforcing the need to use more complex models for more realistic drug screening.
Fernanda Malhão; Alice Ramos; Eduardo Rocha. Cytotoxic and Anti-Proliferative Effects of Fucosterol, Alone and in Combination with Doxorubicin, in 2D and 3D Cultures of Triple-Negative Breast Cancer Cells. Medical Sciences Forum 2020, 2, 14 .
AMA StyleFernanda Malhão, Alice Ramos, Eduardo Rocha. Cytotoxic and Anti-Proliferative Effects of Fucosterol, Alone and in Combination with Doxorubicin, in 2D and 3D Cultures of Triple-Negative Breast Cancer Cells. Medical Sciences Forum. 2020; 2 (1):14.
Chicago/Turabian StyleFernanda Malhão; Alice Ramos; Eduardo Rocha. 2020. "Cytotoxic and Anti-Proliferative Effects of Fucosterol, Alone and in Combination with Doxorubicin, in 2D and 3D Cultures of Triple-Negative Breast Cancer Cells." Medical Sciences Forum 2, no. 1: 14.
Estrogenic effects triggered by androgens have been previously shown in a few studies. Aromatization and direct binding to estrogen receptors (ERs) are the most proposed mechanisms. For example, previously, a modulation of vitellogenin A (VtgA) by testosterone (T), an aromatizable androgen, was reported in brown trout primary hepatocytes. The effect was reversed by an ER antagonist. In this study, using the same model the disruption caused by T and by the non-aromatizable androgen - dihydrotestosterone (DHT), was assessed in selected estrogenic targets. Hepatocytes were exposed (96 h) to six concentrations of each androgen. The estrogenic targets were VtgA, ERα, ERβ1 and two zona pellucida genes, ZP2.5 and ZP3a.2. The aromatase CYP19a1 gene and the androgen receptor (AR) were also included. Modulation of estrogenic targets was studied by quantitative real-time PCR and immunohistochemistry, using an HScore system. VtgA and ERα were up-regulated by DHT (1, 10, 100 μM) and T (10, 100 μM). In contrast, ERβ1 was down-regulated by DHT (10, 100 μM), and T (100 μM). ZP2.5 mRNA levels were increased by DHT and T (1, 10, 100 μM), while ZP3a.2 was up-regulated by DHT (100 μM) and T (10, 100 μM). Positive correlations were found between VtgA and ERα mRNA levels and ZPs and ERα, after exposure to both androgens. The mRNA levels of CYP19a1 were not changed, while AR expression tended to increase after micromolar DHT exposures. HScores for Vtg and ZPs corroborated the molecular findings. Both androgens triggered estrogen signaling through direct binding to ERs, most probably ERα.
Célia Lopes; Tânia V. Madureira; José F. Gonçalves; Eduardo Rocha. Disruption of classical estrogenic targets in brown trout primary hepatocytes by the model androgens testosterone and dihydrotestosterone. Aquatic Toxicology 2020, 227, 105586 .
AMA StyleCélia Lopes, Tânia V. Madureira, José F. Gonçalves, Eduardo Rocha. Disruption of classical estrogenic targets in brown trout primary hepatocytes by the model androgens testosterone and dihydrotestosterone. Aquatic Toxicology. 2020; 227 ():105586.
Chicago/Turabian StyleCélia Lopes; Tânia V. Madureira; José F. Gonçalves; Eduardo Rocha. 2020. "Disruption of classical estrogenic targets in brown trout primary hepatocytes by the model androgens testosterone and dihydrotestosterone." Aquatic Toxicology 227, no. : 105586.
Strong evidence has suggested an important role of telomeres in meiosis, fertilization, and embryo development. To determine if sperm telomere length (STL) in sperm purified by differential gradient centrifugation followed by swim-up (selected STL) is correlated with sperm quality and clinical outcomes. Relative selected STL was assessed by quantitative polymerase chain reaction (Q-PCR) in 78 consecutive assisted reproductive technology (ART) treatments during 2017. Statistical analyses were performed in the totality of patients, and in normozoospermic and non-normozoospermic patients. These included correlations between selected STL and sperm quality parameters, embryological parameters (multivariable linear regression), and clinical parameters (multivariable logistic regression). No significant correlations were found between selected STL and sperm quality in the total population. However, selected STL was significantly correlated with total sperm count (r = 0.361; P = 0.039) and sperm DNA fragmentation-post-acrosomal region pattern (r = − 0.464; P = 0.030) in normozoospermic patients. No relation was observed between selected STL and clinical outcomes in any clinical group. As the correlations observed in normozoospermic patients were not representative of the whole heterogeneous population, differences in the sperm characteristics of the study population may lead to discrepant results when evaluating the association of STL with sperm quality. Since the total population selected STL was not related with sperm quality and with clinical outcomes, results do not support the use of selected STL measurement to evaluate the reproductive potential of the male patient or to predict the success rates of ART treatments.
Ana Catarina Lopes; Pedro Fontes Oliveira; Soraia Pinto; Carolina Almeida; Maria João Pinho; Rosália Sá; Eduardo Rocha; Alberto Barros; Mário Sousa. Discordance between human sperm quality and telomere length following differential gradient separation/swim-up. Journal of Assisted Reproduction and Genetics 2020, 37, 2581 -2603.
AMA StyleAna Catarina Lopes, Pedro Fontes Oliveira, Soraia Pinto, Carolina Almeida, Maria João Pinho, Rosália Sá, Eduardo Rocha, Alberto Barros, Mário Sousa. Discordance between human sperm quality and telomere length following differential gradient separation/swim-up. Journal of Assisted Reproduction and Genetics. 2020; 37 (10):2581-2603.
Chicago/Turabian StyleAna Catarina Lopes; Pedro Fontes Oliveira; Soraia Pinto; Carolina Almeida; Maria João Pinho; Rosália Sá; Eduardo Rocha; Alberto Barros; Mário Sousa. 2020. "Discordance between human sperm quality and telomere length following differential gradient separation/swim-up." Journal of Assisted Reproduction and Genetics 37, no. 10: 2581-2603.
The main aim of this study was to histochemically estimate the relative copper content in the liver during the breeding cycle of female Ohrid trout, a wild population endemic to the Ohrid Lake. The histology of the oogenetic cycle in this species was documented for the first time. Histochemistry with semi-quantification revealed positive correlations between the gonadosomatic indices and copper load. Uptake of the metal increases in the liver as the ovaries increase in size, with the highest hepatic copper load occurring postspawning. The plasmatic levels of 17β-oestradiol were also quantified, but no correlations to copper load were found, so there is no direct estrogenic control on the uptake and release of copper from the bloodstream to either organ. Therefore, when interpreting field studies or assays on hepatic copper pollution and toxicity, breeding-related seasonal changes in Ohrid trout must be taken into account.
Maja Jordanova; Katerina Rebok; Eduardo Rocha; Maria João Rocha. Changes in copper load in hepatocytes of Ohrid trout in relation to the ovarian maturation cycle. Toxicological & Environmental Chemistry 2020, 102, 272 -283.
AMA StyleMaja Jordanova, Katerina Rebok, Eduardo Rocha, Maria João Rocha. Changes in copper load in hepatocytes of Ohrid trout in relation to the ovarian maturation cycle. Toxicological & Environmental Chemistry. 2020; 102 (5-6):272-283.
Chicago/Turabian StyleMaja Jordanova; Katerina Rebok; Eduardo Rocha; Maria João Rocha. 2020. "Changes in copper load in hepatocytes of Ohrid trout in relation to the ovarian maturation cycle." Toxicological & Environmental Chemistry 102, no. 5-6: 272-283.
The goldfish is a model organism showing great potential for research, particularly in comparative endocrinology concerning the neuroendocrine signalling and regulation of vertebrate reproduction. Furthermore, this teleost is increasingly stressed as a relevant alternative to more common fish model organisms, namely zebrafish. However, quality descriptions and illustrations of the complete goldfish gonadal histology are surprisingly scarce, but needed, to support research using this fish. Therefore, the main aim of this work is to describe in detail and adequately illustrate the goldfish oogenesis, from oogonia to late maturation, by applying routine stains (haematoxylin–eosin) and special procedures (periodic acid–Schiff and Goldner's trichrome). We hypothesized that the combined strategies would enable not only to observe the most general features but also to perceive some poorly described details of oocytes better. We describe the details of the following maturation stages: oogonia proliferation, chromatin–nucleolus, primary growth (one nucleolus step, multiple nucleoli step, perinucleolar step, cortical alveoli step) and secondary growth (early secondary growth step, late secondary growth step). Additionally, we report aspects of early and late follicular atresia. The study allowed comparisons with other species and showed that the Goldner's trichrome has the best discriminative power and should be the preferred stain, despite more time‐consuming.
Maria Leonor Ferrão; Maria João Rocha; Eduardo Rocha. Histological characterization of the maturation stages of the ovarian follicles of the goldfish Carassius auratus (Linnaeus, 1758). Anatomia, Histologia, Embryologia 2020, 49, 749 -762.
AMA StyleMaria Leonor Ferrão, Maria João Rocha, Eduardo Rocha. Histological characterization of the maturation stages of the ovarian follicles of the goldfish Carassius auratus (Linnaeus, 1758). Anatomia, Histologia, Embryologia. 2020; 49 (6):749-762.
Chicago/Turabian StyleMaria Leonor Ferrão; Maria João Rocha; Eduardo Rocha. 2020. "Histological characterization of the maturation stages of the ovarian follicles of the goldfish Carassius auratus (Linnaeus, 1758)." Anatomia, Histologia, Embryologia 49, no. 6: 749-762.
: Chronic Myeloid Leukemia (CML) represents 15-20% of all new cases of leukemia and is characterized by an uncontrolled proliferation of abnormal myeloid cells. Currently, the first-line of treatment involves Tyrosine Kinase Inhibitors (TKIs), which specifically inhibits the activity of the fusion protein BCR-ABL. However, resistance, mainly due to mutations, can occur. In the attempt to find more effective and less toxic therapies, several approaches are taken into consideration such as research of new anti-leukemic drugs and “combination chemotherapy” where different drugs that act by different mechanisms are used. Here, we reviewed the molecular mechanisms of CML, the main mechanisms of drug resistance and current strategies to enhance the therapeutic effect of TKIs in CML. Despite major advances in CML treatment, new, more potent anticancer drugs and with fewer side effects are needed. Marine organisms, and particularly seaweed, have a high diversity of bioactive compounds with some of them having anticancer activity in several in vitro and in vivo models. The state-of-art suggests that their use during cancer treatment may improve the outcome. We reviewed here the yet few data supporting anti-leukemic activity of some carotenoids and phlorotannins in some leukemia models. Also, strategies to overcome drug resistance are discussed, particularly the combination of conventional drugs with natural compounds.
Tania P. Almeida; Alice A. Ramos; Joana Ferreira; Amaya Azqueta; Eduardo Rocha. Bioactive Compounds from Seaweed with Anti-Leukemic Activity: A Mini-Review on Carotenoids and Phlorotannins. Mini-Reviews in Medicinal Chemistry 2020, 20, 39 -53.
AMA StyleTania P. Almeida, Alice A. Ramos, Joana Ferreira, Amaya Azqueta, Eduardo Rocha. Bioactive Compounds from Seaweed with Anti-Leukemic Activity: A Mini-Review on Carotenoids and Phlorotannins. Mini-Reviews in Medicinal Chemistry. 2020; 20 (1):39-53.
Chicago/Turabian StyleTania P. Almeida; Alice A. Ramos; Joana Ferreira; Amaya Azqueta; Eduardo Rocha. 2020. "Bioactive Compounds from Seaweed with Anti-Leukemic Activity: A Mini-Review on Carotenoids and Phlorotannins." Mini-Reviews in Medicinal Chemistry 20, no. 1: 39-53.
The liver hosts numerous vital functions, such as biotransformation and excretion of xenobiotics. Synthetic oestrogens influence liver structure and function, leading to adaptations or to dysfunctions/injury. They are often stated to induce increases in fish liver weight, but there is controversy regarding how: if by changes in hepatocyte size (hypertrophy) and/or number (hyperplasia). Using platyfish as the experimental model, our primary aim was to assess if/how hepatocytes reacted to a sub-acute oestrogenic exposure. A complementary aim was to generate fundamental structural data for the liver of that model organism. Adult males were injected intramuscularly with 17α-ethinylestradiol (EE2) (25 μg/g), every 72 h for two weeks. Control fish were given solvent only. Body and liver morphometry were registered, and hepatocytes examined through histology and stereology at light microscopy. Immunohistochemistry evaluated hepatocytic vitellogenin (VTG) content. Treated and control fish did not differ as to quantitative parameters. Nevertheless, exposed fish were sensitive to EE2. VTG tagging was positive in their hepatocytes and these tended to be more basophilic, though not fully oestrogenized. We hypothesise that the platyfish liver is not particularly sensitive to the disrupting action of EE2 because of its reproductive mode; with no production peaks of VTG and no huge changes in endogenous sex-steroids. The fish may have had no evolutionary pressure for hepatocytes to be particularly reactive to oestradiol (E2). In the end, this study offers the first unbiased estimation of the liver cellularity in the platyfish, as well of the hepatocytic volume, serving now as a baseline reference.
Madalena V.F. Real; Maria J. Rocha; José F. Gonçalves; Eduardo Rocha. Histology and design-based estimation of hepatocellularity and volumes of hepatocytes in control and ethynylestradiol exposed males of platyfish (Xiphophorus maculatus). Tissue and Cell 2019, 63, 101327 .
AMA StyleMadalena V.F. Real, Maria J. Rocha, José F. Gonçalves, Eduardo Rocha. Histology and design-based estimation of hepatocellularity and volumes of hepatocytes in control and ethynylestradiol exposed males of platyfish (Xiphophorus maculatus). Tissue and Cell. 2019; 63 ():101327.
Chicago/Turabian StyleMadalena V.F. Real; Maria J. Rocha; José F. Gonçalves; Eduardo Rocha. 2019. "Histology and design-based estimation of hepatocellularity and volumes of hepatocytes in control and ethynylestradiol exposed males of platyfish (Xiphophorus maculatus)." Tissue and Cell 63, no. : 101327.
Doxorubicin (Dox) is one of the most successful anticancer drugs in use. However, chemoresistance is one of the main limitations that patients face. Therefore, development of new strategies to improve the efficacy of Dox is needed. Marine‐derived fungi are especially promising sources of new anticancer compounds. In this work, antitumor activity of crude ethyl extract of the cultures of the marine‐derived fungus Neosartorya siamensis KUFA 0017 (NS), combined with Dox, was evaluated in six cancer cell lines. To evaluate possible mechanisms involved in the eventual improvement of Dox's cytotoxicity by NS extract, effects on DNA damage, cell death, ultrastructural modifications, and intracellular accumulation of Dox were assessed. The NS extract demonstrated a significant enhancement of Dox's cytotoxic activity in A549 cells, inducing DNA damage, cell death, and intracellular accumulation of Dox. Additionally, the cytotoxic effect of eight compounds, isolated from this extract, that is, 2,4‐dihydroxy‐3‐methylacetophenone‐(C1), nortryptoquivaline‐(C2), chevalone C‐(C3), tryptoquivaline H‐(C4), fiscalin A‐(C5), epi‐fiscalin‐C (C6), epi‐neofiscalin A‐(C7), and epi‐fiscalin A‐(C8), alone and combined with Dox was also evaluated in lung cancer cells. The cytotoxic effect of Dox was potentiated by all the isolated compounds (except C1) in A549 cells. Therefore, we concluded that NS extract potentiated cytotoxicity by inhibiting cell proliferation, increasing intracellular accumulation of Dox, and inducing cell death (possibly by an autophagic process). The isolated compounds also enhanced the activity of Dox, supporting the potential of this sort of combination. These data call for further studies to characterize drug interactions and underlying mechanisms.
Alice Ramos; Bruno Castro‐Carvalho; Maria Prata‐Sena; Fernanda Malhão; Suradet Buttachon; Tida Dethoup; Anake Kijjoa; Eduardo Rocha. Can marine‐derived fungus Neosartorya siamensis KUFA 0017 extract and its secondary metabolites enhance antitumor activity of doxorubicin? An in vitro survey unveils interactions against lung cancer cells. Environmental Toxicology 2019, 35, 507 -517.
AMA StyleAlice Ramos, Bruno Castro‐Carvalho, Maria Prata‐Sena, Fernanda Malhão, Suradet Buttachon, Tida Dethoup, Anake Kijjoa, Eduardo Rocha. Can marine‐derived fungus Neosartorya siamensis KUFA 0017 extract and its secondary metabolites enhance antitumor activity of doxorubicin? An in vitro survey unveils interactions against lung cancer cells. Environmental Toxicology. 2019; 35 (4):507-517.
Chicago/Turabian StyleAlice Ramos; Bruno Castro‐Carvalho; Maria Prata‐Sena; Fernanda Malhão; Suradet Buttachon; Tida Dethoup; Anake Kijjoa; Eduardo Rocha. 2019. "Can marine‐derived fungus Neosartorya siamensis KUFA 0017 extract and its secondary metabolites enhance antitumor activity of doxorubicin? An in vitro survey unveils interactions against lung cancer cells." Environmental Toxicology 35, no. 4: 507-517.
Breast cancer (BC) accounts for the most cancer-related deaths amongst women worldwide, implying an urgent need of finding new drugs more effective than those currently in use [1, 2]. Fucoxanthin (Fx) is a marine carotenoid derived from brown seaweed that has been showing antitumor effects on different cancer cell lines, mainly in 2D models [3]. However, 3D culture models have a better predictive capacity of in vivo cellular responses against cytotoxic compounds [4]. This study aimed to evaluate the potential anticancer effects of Fx versus Doxorubicin (Dox) (a conventional anticancer drug) in a panel of three BC cell lines, representative of different molecular subtypes (MCF-7, SKBR3 and MDA-MB-231), and in a non-tumoral BC cell line (MCF-12A), cultured under 2D and 3D conditions. Effects on cell viability, death, proliferation and DNA damage were targeted. Results from the 2D cultures showed that Dox and Fx caused cytotoxicity on all the cell lines. Cytotoxic activity of Dox involved mechanisms of cell death induction (in MCF-12A cells) and genotoxicity (in MDA-MB-231 cells), contrarily to Fx, where neither cell death nor genotoxic mechanisms were detected. Under 3D conditions, cells were less responsive to Dox and Fx, compared to 2D cultures. Antiproliferative effects were solely registered in SKBR3 cells exposed to Dox and Fx, and in MCF-12A cells exposed to Dox. The in vitro data revealed that the Fx may be a potential anticancer agent against BC cells, with differential effects according to the cell subtype. The data warrants further studies on the underlying anticancer mechanisms. Acknowledgments: We thank the fundamental financial support of the ICBAS of the UPorto, namely via its Laboratory of Histology and Embryology and Master Program in Environmental Contamination and Toxicology. This research was also partially supported by the Strategic Funding UID/Multi/04423/2019 through national funds provided by FCT and ERDF, in the framework of the program PT2020. [1] DeSantis, C.E., F. Bray, J. Ferlay, J. Lortet-Tieulent, B.O. Anderson, and A. Jemal, International Variation in Female Breast Cancer Incidence and Mortality Rates. Cancer Epidemiol Biomarkers Prev, 2015. 24(10): p. 1495-506. [2] Pádua, D., E. Rocha, D. Gargiulo, and A.A. Ramos, Bioactive compounds from brown seaweeds: Phloroglucinol, fucoxanthin and fucoidan as promising therapeutic agents against breast cancer. Phytochem Lett, 2015. 14: p. 91-98. [3] Kumar, S.R., M. Hosokawa, and K. Miyashita, Fucoxanthin: a marine carotenoid exerting anti-cancer effects by affecting multiple mechanisms. Mar Drugs, 2013. 11(12): p. 5130-47. [4] Santo, V.E., S.P. Rebelo, M.F. Estrada, P.M. Alves, E. Boghaert, and C. Brito, Drug screening in 3D in vitro tumor models: overcoming current pitfalls of efficacy read-outs. Biotechnol J, 2017. 12(1): p. 1600505.
Ana Catarina Macedo; Fernanda Malhão; Eduardo Rocha; Alice Ramos. Anticancer activity of the seaweed compound fucoxanthin in breast cancer cell lines cultured as 2D and 3D models. Proceedings of 5th International Electronic Conference on Medicinal Chemistry 2019, 1 .
AMA StyleAna Catarina Macedo, Fernanda Malhão, Eduardo Rocha, Alice Ramos. Anticancer activity of the seaweed compound fucoxanthin in breast cancer cell lines cultured as 2D and 3D models. Proceedings of 5th International Electronic Conference on Medicinal Chemistry. 2019; ():1.
Chicago/Turabian StyleAna Catarina Macedo; Fernanda Malhão; Eduardo Rocha; Alice Ramos. 2019. "Anticancer activity of the seaweed compound fucoxanthin in breast cancer cell lines cultured as 2D and 3D models." Proceedings of 5th International Electronic Conference on Medicinal Chemistry , no. : 1.
Preussin, a hydroxyl pyrrolidine derivative isolated from the marine sponge-associated fungus Aspergillus candidus KUFA 0062, displayed anticancer effects in some cancer cell lines, including MCF7. Preussin was investigated for its cytotoxic and antiproliferative effects in breast cancer cell lines (MCF7, SKBR3, and MDA-MB-231), representatives of major breast cancers subtypes, and in a non-tumor cell line (MCF12A). Preussin was first tested in 2D (monolayer), and then in 3D (multicellular aggregates), cultures, using a multi-endpoint approach for cytotoxicity (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), resazurin and lactate dehydrogenase (LDH)) and proliferative (5-bromo-2′-deoxyuridine (BrdU)) assays, as well as the analysis of cell morphology by optical/electron microscopy and immunocytochemistry for caspase-3 and ki67. Preussin affected cell viability and proliferation in 2D and 3D cultures in all cell lines tested. The results in the 3D culture showed the same tendency as in the 2D culture, however, cells in the 3D culture were less responsive. The effects were observed at different concentrations of preussin, depending on the cell line and assay method. Morphological study of preussin-exposed cells revealed cell death, which was confirmed by caspase-3 immunostaining. In view of the data, we recommend a multi-endpoint approach, including histological evaluation, in future assays with the tested 3D models. Our data showed cytotoxic and antiproliferative activities of preussin in breast cancer cell lines in 2D and 3D cultures, warranting further studies for its anticancer potential.
Fernanda Malhão; Alice A. Ramos; Suradet Buttachon; Tida Dethoup; Anake Kijjoa; Eduardo Rocha. Cytotoxic and Antiproliferative Effects of Preussin, a Hydroxypyrrolidine Derivative from the Marine Sponge-Associated Fungus Aspergillus candidus KUFA 0062, in a Panel of Breast Cancer Cell Lines and Using 2D and 3D Cultures. Marine Drugs 2019, 17, 448 .
AMA StyleFernanda Malhão, Alice A. Ramos, Suradet Buttachon, Tida Dethoup, Anake Kijjoa, Eduardo Rocha. Cytotoxic and Antiproliferative Effects of Preussin, a Hydroxypyrrolidine Derivative from the Marine Sponge-Associated Fungus Aspergillus candidus KUFA 0062, in a Panel of Breast Cancer Cell Lines and Using 2D and 3D Cultures. Marine Drugs. 2019; 17 (8):448.
Chicago/Turabian StyleFernanda Malhão; Alice A. Ramos; Suradet Buttachon; Tida Dethoup; Anake Kijjoa; Eduardo Rocha. 2019. "Cytotoxic and Antiproliferative Effects of Preussin, a Hydroxypyrrolidine Derivative from the Marine Sponge-Associated Fungus Aspergillus candidus KUFA 0062, in a Panel of Breast Cancer Cell Lines and Using 2D and 3D Cultures." Marine Drugs 17, no. 8: 448.
Estrone, 17β-estradiol, 17α-ethynylestradiol, alkylphenols and alkylphenol ethoxylates, are endocrine disruptor compounds. In Portugal, the report of male fish feminization in Minho, Ave and Mondego River estuaries made pertinent their evaluation in surface water samples. Data, got by specific enzyme-linked immunosorbent assays, showed that estrogens concentrations were lower in Minho than in Ave or Mondego estuaries: [Estrone] > [17β-estradiol] > [17α-ethynylestradiol] (p < 0.05). Converting estrogen concentrations in 17α-ethynylestradiol equivalents, the contribution of estrogens was 1.3 ng/L, 3.5 ng/L, and 2.4 ng/L, respectively, for Minho, Ave, and Mondego estuaries stressing out a high risk for local aquatic species. The concentrations of alkylphenols and alkylphenol ethoxylates were lower at both Minho and Mondego estuaries ≈600 ng/L and 2700 ng/L, respectively than at the Ave, ≈1070 ng/L and 4855 ng/L (p < 0.05). In conclusion, this study reports the first systematic data about the presence of the targeted endocrine disruptor compounds in the Minho River estuary and, in parallel, provides new information about their concentrations in two other habitats previously monitored. This study also shows that in Ave and Mondego estuaries the concentrations of industrial estrogens were still excessive, whereas the load of the pure estrogens has decreased significantly facing previous surveys.
Maria João Rocha; Tânia V. Madureira; Carla Sofia Venade; Irene Martins; Joana Campos; Eduardo Rocha. Presence of estrogenic endocrine disruptors in three European estuaries in Northwest Iberian Peninsula (Portugal). Toxicological & Environmental Chemistry 2019, 101, 244 -264.
AMA StyleMaria João Rocha, Tânia V. Madureira, Carla Sofia Venade, Irene Martins, Joana Campos, Eduardo Rocha. Presence of estrogenic endocrine disruptors in three European estuaries in Northwest Iberian Peninsula (Portugal). Toxicological & Environmental Chemistry. 2019; 101 (3-6):244-264.
Chicago/Turabian StyleMaria João Rocha; Tânia V. Madureira; Carla Sofia Venade; Irene Martins; Joana Campos; Eduardo Rocha. 2019. "Presence of estrogenic endocrine disruptors in three European estuaries in Northwest Iberian Peninsula (Portugal)." Toxicological & Environmental Chemistry 101, no. 3-6: 244-264.
Background Sjögren's syndrome (SS) induces difficulty in chewing and swallowing due to low salivary flow. However, these symptoms may be associated with other factors, such as orofacial myofunctional disorders and temporomandibular disorder (TMD), which have not been comprehensively assessed in this population. Objective The aims of this study were to investigate orofacial muscles and functions as well as the presence of TMD in patients with SS compared with a group without SS and analyze whether the patients’ experience of limitations in orofacial functioning is associated with the orofacial functional status and muscle pain related to TMD. Methods Women with SS based on the 2002 American‐European Criteria and volunteers paired by age and sex were compared. The exams included the orofacial myofunctional evaluation with scores protocol (OMES), tongue and lip strength measures and electromyography of the masticatory muscles. TMD investigations included clinical examination, self‐report of symptoms and jaw functional limitation scale. Results Patients with Sjögren's Syndrome present impaired muscle and orofacial functions based on lower scores of all categories of orofacial myofunctional evaluation (P< 0.0001), tongue strength (P= 0.0003‐0.0004), and masticatory muscle activity (P= 0.0002‐0.007) as well as worse TMD signs and symptoms (P< 0.05) and jaw functional limitation (P< 0.0001‐0.0003). The patients’ experiences with limitation in mastication and swallowing were associated with orofacial myofunctional status and muscle pain related to TMD. Those disorders should be monitored along with disease control and must be addressed in the clinical evaluation to prevent nutritional and metabolic comorbidities in SS patients. This article is protected by copyright. All rights reserved.
Mariana Cristina Zanin; Denny Marcos Garcia; Eduardo Melani Rocha; Cláudia Maria De Felício. Orofacial Motor Functions and Temporomandibular Disorders in Patients With Sjögren's Syndrome. Arthritis Care & Research 2019, 72, 1057 -1065.
AMA StyleMariana Cristina Zanin, Denny Marcos Garcia, Eduardo Melani Rocha, Cláudia Maria De Felício. Orofacial Motor Functions and Temporomandibular Disorders in Patients With Sjögren's Syndrome. Arthritis Care & Research. 2019; 72 (8):1057-1065.
Chicago/Turabian StyleMariana Cristina Zanin; Denny Marcos Garcia; Eduardo Melani Rocha; Cláudia Maria De Felício. 2019. "Orofacial Motor Functions and Temporomandibular Disorders in Patients With Sjögren's Syndrome." Arthritis Care & Research 72, no. 8: 1057-1065.
Climate change and pharmaceuticals contamination constitute two of the most relevant stressors on the aquatic ecosystems, however, there is a huge lack of information regarding the interactive effects of both stressors. For that, a mesocosm experiment was implemented where adult zebrafish were exposed to combined temperature and the progestin levonorgestrel (LNG) for 21 days. Considering that the liver is one of the organs where there is a greater metabolization and accumulation of toxicants, the main objective of this work was to assess the effects of both stressors on the female zebrafish hepatocytes morphology and functioning, through stereological and immunohistochemical techniques. Our results revealed an increase of coefficient of variation of the number distribution of hepatocytes volume (CVN(υ)) for individuals exposed to LNG, which denotes an increase of the hepatocytes size variability and is suggestive of functional impacts. This was corroborated by the signs of increased glycogen content with the exposure to increased LNG concentrations and temperature, indicating modified hepatocyte glycogen metabolism. Such disturbances can be considered indicators that the fish had to deal with impacts caused by the stress factors. Regarding the immunoreactivity, from the four proteins selected (catalase, CYP1A, HSP90 and Vtg), just in two of them (catalase and Vtg) were observed some responses to both stressors. For catalase there was a hormetic response, in which exposure to lower LNG concentrations caused a significant higher positive immunostaining than under higher LNG concentrations. While, for Vtg, significant effects of temperature and LNG existed, in which a decline in Vtg immunostaining was observed with exposure to higher temperature and lower LNG concentrations. These results should be seen as a warning sign about fine impacts of multiple stressors, such as temperature and progestogens, on the structure and functioning of zebrafish liver and potentially in other aquatic organisms, and on their health implications.
P.G. Cardoso; R. Resende-De-Oliveira; Eduardo Rocha. Combined effects of increased temperature and levonorgestrel exposure on zebrafish female liver, using stereology and immunohistochemistry against catalase, CYP1A, HSP90 and vitellogenin. Environmental Pollution 2019, 252, 1059 -1067.
AMA StyleP.G. Cardoso, R. Resende-De-Oliveira, Eduardo Rocha. Combined effects of increased temperature and levonorgestrel exposure on zebrafish female liver, using stereology and immunohistochemistry against catalase, CYP1A, HSP90 and vitellogenin. Environmental Pollution. 2019; 252 ():1059-1067.
Chicago/Turabian StyleP.G. Cardoso; R. Resende-De-Oliveira; Eduardo Rocha. 2019. "Combined effects of increased temperature and levonorgestrel exposure on zebrafish female liver, using stereology and immunohistochemistry against catalase, CYP1A, HSP90 and vitellogenin." Environmental Pollution 252, no. : 1059-1067.