Remote sensing data and numerical simulation are important tools to rebuild any oil spill accident letting to identify its source and trajectory. Through these tools was identified an oil spill that ...affected Oaxacan coast in October 2022. The SAR images were processed with a standard method included in SNAP software, and the numerical simulation was made using Lagrangian transport model included in GNOME software. With the combining of these tools was possible to discriminate the look-alikes from true oil slicks; which are the main issue when satellite images are used. Obtained results showed that 4.3m3 of crude oil were released into the ocean from a punctual point of oil pollution. This oil spill was classified such as a small oil spill. The marine currents and weathering processes were the main drivers that controlled the crude oil displacement and its dispersion. It was estimated in GNOME that 1.6 m3 of crude oil was floating on the sea (37.2 %), 2.4 m3 was evaporated into the atmosphere (55.8 %) and 0.3 m3 reached the coast of Oaxaca (7 %). This event affected 82 km of coastline, but the most important touristic areas as well as turtle nesting zones were not affected by this small crude oil spill. Results indicated that the marine-gas-pump number 3 in Salina Cruz, Oaxaca, is a punctual point of oil pollution in the Southern Mexican Pacific Ocean. Further work is needed to assess the economic and ecological damage to Oaxacan coast caused by this small oil spill.
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•Remote sensing data and numerical simulation let to rebuild an oil spill accident.•Look-alikes were identified using SAR images and numerical simulation.•Marine currents were the main driver in oil transportation to shoreline.•4.3 m3 of oil was released in the Southern Mexican Pacific Ocean.•82 km of shoreline was affected by 0.3 m3 of oil.
Despite the fundamental importance of the redox metabolism of mitochondria under normal and pathological conditions, our knowledge regarding the transport of vitamin C across mitochondrial membranes ...remains far from complete. We report here that human HEK-293 cells express a mitochondrial low-affinity ascorbic acid transporter that molecularly corresponds to SVCT2, a member of the sodium-coupled ascorbic acid transporter family 2. The transporter SVCT1 is absent from HEK-293 cells. Confocal colocalization experiments with anti-SVCT2 and anti-organelle protein markers revealed that most of the SVCT2 immunoreactivity was associated with mitochondria, with minor colocalization at the endoplasmic reticulum and very low immunoreactivity at the plasma membrane. Immunoblotting of proteins extracted from highly purified mitochondrial fractions confirmed that SVCT2 protein was associated with mitochondria, and transport analysis revealed a sigmoidal ascorbic acid concentration curve with an apparent ascorbic acid transport Km of 0.6mM. Use of SVCT2 siRNA for silencing SVCT2 expression produced a major decrease in mitochondrial SVCT2 immunoreactivity, and immunoblotting revealed decreased SVCT2 protein expression by approximately 75%. Most importantly, the decreased protein expression was accompanied by a concomitant decrease in the mitochondrial ascorbic acid transport rate. Further studies using HEK-293 cells overexpressing SVCT2 at the plasma membrane revealed that the altered kinetic properties of mitochondrial SVCT2 are due to the ionic intracellular microenvironment (low in sodium and high in potassium), with potassium acting as a concentration-dependent inhibitor of SVCT2. We discarded the participation of two glucose transporters previously described as mitochondrial dehydroascorbic acid transporters; GLUT1 is absent from mitochondria and GLUT10 is not expressed in HEK-293 cells. Overall, our data indicate that intracellular SVCT2 is localized in mitochondria, is sensitive to an intracellular microenvironment low in sodium and high in potassium, and functions as a low-affinity ascorbic acid transporter. We propose that the mitochondrial localization of SVCT2 is a property shared across cells, tissues, and species.
•We describe a general mechanism for the mitochondrial acquisition of ascorbic acid.•Mitochondria express a low-affinity form of the ascorbic acid transporter SVCT2.•SVCT2 transport kinetics is altered by the intracellular ionic microenvironment.•Mitochondrial SVCT2 shows an apparent transport Km of 0.6mM.•Mitochondrial SVCT2 is fully functional under normal high ascorbate concentrations.
The potential role of vitamin C in cancer prevention and treatment remains controversial. While normal human cells obtain vitamin C as ascorbic acid, the prevalent form of vitamin C in vivo, the ...uptake mechanisms by which cancer cells acquire vitamin C has remained unclear. The aim of this study is to characterize how breast cancer cells acquire vitamin C. For this, we determined the expression of vitamin C transporters in normal and breast cancer tissue samples, and in ZR-75, MCF-7, MDA-231 and MDA-468 breast cancer cell lines. At the same time, reduced (AA) and oxidized (DHA) forms of vitamin C uptake experiments were performed in all cell lines.
We show here that human breast cancer tissues differentially express a form of SVCT2 transporter, that is systematically absent in normal breast tissues and it is increased in breast tumors. In fact, estrogen receptor negative breast cancer tissue, exhibit the most elevated SVCT2 expression levels. Despite this, our analysis in breast cancer cell lines showed that these cells are not able to uptake ascorbic acid and depend on glucose transporter for the acquisition of vitamin C by a bystander effect. This is consistent with our observations that this form of SVCT2 is completely absent from the plasma membrane and is overexpressed in mitochondria of breast cancer cells, where it mediates ascorbic acid transport. This work shows that breast cancer cells acquire vitamin C in its oxidized form and are capable of accumulated high concentrations of the reduced form. Augmented expression of an SVCT2 mitochondrial form appears to be a common hallmark across all human cancers and might have implications in cancer cells survival capacity against pro-oxidant environments.
A final model is proposed identifying three potentially regulatable steps: i) The local extracellular generation of DHA, ii) DHA transport dependent on plasmamembrane glucose transporters (GLUT), and iii) Mitochondrial uptake of accumulated AA by a SVCT2 form that is overexpressed in breast cancer. This mechanism migth be implicated in mitochondrial oxidative stress reduction. Display omitted
•Breast cancer tissues exhibit high levels of SVCT2 expression, an AA transporter, that is absent in normal tissue.•However, breast cancer cells are unable to acquire vitamin C as AA and they only have the capacity to acquire it as DHA.•SVCT2 is located at the inner membrane of mitochondria of breast cancer cells, where it mediates AA transport.•Mitochondrial SVCT2 overexpression seems to be a common hallmark across all human cancers.
GLUT12 is a member of the facilitative family of glucose transporters. The goal of this study was to characterize the functional properties of GLUT12, expressed in Xenopus laevis oocytes, using ...radiotracer and electrophysiological methods. Our results showed that GLUT12 is a facilitative sugar transporter with substrate selectivity: d-glucose ≥ α-methyl-d-glucopyranoside (α-MG) > 2-deoxy-d-glucose(2-DOG) > d-fructose = d-galactose. α-MG is a characteristic substrate of the Na(+)/glucose (SGLT) family and has not been shown to be a substrate of any of the GLUTs. In the absence of sugar, (22)Na(+) was transported through GLUT12 at a higher rate (40%) than noninjected oocytes, indicating that there is a Na(+) leak through GLUT12. Genistein, an inhibitor of GLUT1, also inhibited sugar uptake by GLUT12. Glucose uptake was increased by the PKA activator 8-bromoadenosine 3',5'-cyclic monophosphate (8-Br-cAMP) but not by the PKC activator phorbol-12-myristate-13-acetate (PMA). In high K(+) concentrations, glucose uptake was blocked. Addition of glucose to the external solution induced an inward current with a reversal potential of approximately -15 mV and was blocked by Cl(-) channel blockers, indicating the current was carried by Cl(-) ions. The sugar-activated Cl(-) currents were unaffected by genistein. In high external K(+) concentrations, sugar-activated Cl(-) currents were also blocked, indicating that GLUT12 activity is voltage dependent. Furthermore, glucose-induced current was increased by the PKA activator 8-Br-cAMP but not by the PKC activator PMA. These new features of GLUT12 are very different from those described for other GLUTs, indicating that GLUT12 must have a specific physiological role within glucose homeostasis, still to be discovered.
Resveratrol acts as a chemopreventive agent for cancer and as a potential antiobesity and antidiabetic compound, by leading to reduced body fat and improved glucose homeostasis. The exact mechanisms ...involved in improving hyperglycemic state are not known, but most of the glucose uptake into mammalian cells is facilitated by the GLUT hexose transporters. Resveratrol is structurally similar to isoflavones such as genistein, which inhibit the glucose uptake facilitated by the GLUT1 hexose transporter. Here we examined the direct effects of resveratrol on glucose uptake and accumulation in HL-60 and U-937 leukemic cell lines, which express mainly GLUT1, under conditions that discriminate transport from the intracellular substrate phosphorylation/accumulation. Resveratrol blocks GLUT1-mediated hexose uptake and thereby affects substrate accumulation on these cells. Consequently, we characterized the mechanism involved in inhibition of glucose uptake in human red cells. Resveratrol inhibits glucose exit in human red cells, and the displacement of previously bound cytochalasin B revealed the direct interaction of resveratrol with GLUT1. Resveratrol behaves as a competitive blocker of glucose uptake under zero-trans exit and exchange kinetic assays, but it becomes a mixed noncompetitive blocker when zero-trans entry transport was assayed, suggesting that the binding site for resveratrol lies on the endofacial face of the transporter. We propose that resveratrol interacts directly with the human GLUT1 hexose transporter by binding to an endofacial site and that this interaction inhibits the transport of hexoses across the plasma membrane. This inhibition is distinct from the effect of resveratrol on the intracellular phosphorylation/accumulation of glucose.
The facilitative hexose transporter GLUT1 activity is blocked by tyrosine kinase inhibitors that include natural products such as flavones and isoflavones and synthetic compounds such as tyrphostins, ...molecules that are structurally unrelated to the transported substrates Vera, et al. (2001) Biochemistry, 40, 777–790. Here we analyzed the interaction of GLUT1 with quercetin (a flavone), genistein (an isoflavone), and tyrphostin A47 and B46 to evaluate if they share one common or have several binding sites on the protein. Kinetic assays showed that genistein, quercetin, and tyrphostin B46 behave as competitive inhibitors of equilibrium exchange and zero-trans uptake transport and noncompetitive inhibitors of net sugar exit out of human red cells, suggesting that they interact with the external surface of the GLUT1 molecule. In contrast, tyrphostin A47 was a competitive inhibitor of equilibrium exchange and zero-trans exit transport and a noncompetitive inhibitor of net sugar entry into red cells, suggesting that it interacts with the cytoplasmic surface of the transporter. Genistein protected GLUT1 against iodide-elicited fluorescence quenching and also decreased the affinity of d-glucose for its external binding site, while quercetin and tyrphostins B46 and A47 promoted fluorescence quenching and did not affect the external d-glucose binding site. These findings are explained by a carrier that presents at least three binding sites for tyrosine kinase inhibitors, in which (i) genistein interacts with the transporter in a conformation that binds glucose on the external surface (outward-facing conformation), in a site which overlaps with the external binding site for d-glucose, (ii) quercetin and tyrphostin B46 interact with the GLUT1 conformation which binds glucose by the internal side of the membrane (inward-facing conformation), but to a site accessible from the external surface of the protein, and (iii) the binding site for tyrphostin A47 is accessible from the inner surface of GLUT1 by binding to the inward-facing conformation of the transporter. These data provide groundwork for a molecular understanding of how the tyrosine kinase inhibitors directly affect glucose transport in animal cells.
To determine the feasibility of performing neoadjuvant chemotherapy (NCH) followed by radical surgery in patients with locally advanced squamous cell carcinoma of the vulva.
Prospective and ...multicenter trial. Thirty-five patients with a diagnosis of previously untreated locally advanced squamous cell carcinoma of the vulva were given 4 schemes of cisplatin-based NCH and 1 NCH regimen with single bleomycin. Then, they underwent radical surgery of the vulva if clinical response was 50% or more. Age, NCH schemes used, toxicity, response to treatment, type of radical surgery performed, and clinical outcome were evaluated.
Thirty-three patients completed the proposed schemes, and 30 were assessed for radical surgery. Finally, 27 patients underwent radical surgery (radical vulvectomy or radical local excision plus bilateral inguinofemoral lymphadenectomy). In 2 cases of persistent rectal involvement, posterior pelvic exenteration was performed. Moreover, 24 of 27 patients remain with no evidence of disease to date. Toxicity was acceptable. Median age was 62 years (range, 54-72 years). Median follow-up was 49 months (range, 4-155 months).
The use of NCH in selected groups may increase surgical feasibility in initially inoperable patients, thus favoring organ preservation and less extensive resections. Adverse reactions were acceptable, and vulvoperineal deleterious effects that may occur after radiotherapy were consequently avoided.
Glucose transporter (GLUT)1 has become an attractive target to block glucose uptake in malignant cells since most cancer cells overexpress GLUT1 and are sensitive to glucose deprivation. ...Methylxanthines are natural compounds that inhibit glucose uptake; however, the mechanism of inhibition remains unknown. Here, we used a combination of binding and glucose transport kinetic assays to analyze in detail the effects of caffeine, pentoxifylline, and theophylline on hexose transport in human erythrocytes. The displacement of previously bound cytochalasin B revealed a direct interaction between the methylxanthines and GLUT1. Methylxanthines behave as noncompetitive blockers (inhibition constant values of 2-3 mM) in exchange and zero-trans efflux assays, whereas mixed inhibition with a notable uncompetitive component is observed in zero-trans influx assays (inhibition constant values of 5-12 mM). These results indicate that methylxanthines do not bind to either exofacial or endofacial d-glucose-binding sites but instead interact at a different site accessible by the external face of the transporter. Additionally, infinite-cis exit assays (Sen-Widdas assays) showed that only pentoxifylline disturbed d-glucose for binding to the exofacial substrate site. Interestingly, coinhibition assays showed that methylxanthines bind to a common site on the transporter. We concluded that there is a methylxanthine regulatory site on the external surface of the transporter, which is close but distinguishable from the d-glucose external site. Therefore, the methylxanthine moiety may become an attractive framework for the design of novel specific noncompetitive facilitative GLUT inhibitors.
The data presented in this article are related to the research paper entitled “Increased expression of mitochondrial sodium-coupled ascorbic acid transporter-2 (mitSVCT2) as a central feature in ...breast cancer”, available in Free Radical Biology and Medicine Journal 1. In this article, we examined the SVCT2 transporter expression in various breast cancer cell lines using RT-PCR and Western blot assays. In addition, we analyzed the subcellular localization of SVCT2 by immunofluorescence colocalization assays and cellular fractionation experiments. Finally, an analysis of different cancer tissue microarrays immunostained for SVCT2 and imaged by The Human Protein Atlas (https://www.proteinatlas.org) is presented.
We characterized the human Na⁺-ascorbic acid transporter SVCT2 and developed a basic model for the transport cycle that challenges the current view that it functions as a Na⁺-dependent transporter. ...The properties of SVCT2 are modulated by Ca²⁺/Mg²⁺ and a reciprocal functional interaction between Na⁺ and ascorbic acid that defines the substrate binding order and the transport stoichiometry. Na⁺ increased the ascorbic acid transport rate in a cooperative manner, decreasing the transport Km without affecting the Vmax, thus converting a low affinity form of the transporter into a high affinity transporter. Inversely, ascorbic acid affected in a bimodal and concentration-dependent manner the Na⁺ cooperativity, with absence of cooperativity at low and high ascorbic acid concentrations. Our data are consistent with a transport cycle characterized by a Na⁺:ascorbic acid stoichiometry of 2:1 and a substrate binding order of the type Na⁺:ascorbic acid:Na⁺. However, SVCT2 is not electrogenic. SVCT2 showed an absolute requirement for Ca²⁺/Mg²⁺ for function, with both cations switching the transporter from an inactive into an active conformation by increasing the transport Vmax without affecting the transport Km or the Na⁺ cooperativity. Our data indicate that SVCT2 may switch between a number of states with characteristic properties, including an inactive conformation in the absence of Ca²⁺/Mg²⁺. At least three active states can be envisioned, including a low affinity conformation at Na⁺ concentrations below 20 mM and two high affinity conformations at elevated Na⁺ concentrations whose Na⁺ cooperativity is modulated by ascorbic acid. Thus, SVCT2 is a Ca²⁺/Mg²⁺-dependent transporter.
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