Disruption of the interaction between PMCA2 and calcineurin triggers apoptosis and enhances paclitaxel-induced cytotoxicity in breast cancer cells

Cancer is caused by defects in the signalling mechanisms that govern cell proliferation and apoptosis. It is well known that calcium-dependent signalling pathways play a critical role in cell regulation. A tight control of calcium homeostasis by transporters and channel proteins is required to assure a proper functioning of the calcium-sensitive signal transduction pathways that regulate cell growth and apoptosis. The Plasma Membrane Calcium ATPase 2 (PMCA2) has been recently identified as a negative regulator of apoptosis that can play a significant role in cancer progression by conferring cells resistance to apoptosis. We have previously reported an inhibitory interaction between PMCA2 and the calcium-activated signalling molecule calcineurin in breast cancer cells. Here we demonstrate that disruption of the PMCA2/calcineurin interaction in a variety of human breast cancer cells results in activation of the calcineurin/NFAT pathway, up-regulation in the expression of the pro-apoptotic protein Fas Ligand, and in a concomitant loss of cell viability. Reduction in cell viability is the consequence of an increase in cell apoptosis. Impairment of the PMCA2/calcineurin interaction enhances paclitaxel-mediated cytotoxicity of breast tumoral cells. Our results suggest that therapeutic modulation of the PMCA2/calcineurin interaction might have important clinical applications to improve current treatments for breast cancer patients.

Induction of protein degradation in skeletal muscle by a phorbol ester involves upregulation of the ubiquitin-proteasome proteolytic pathway

Although muscle atrophy is common to a number of disease states there is incomplete knowledge of the cellular mechanisms involved. In this study murine myotubes were treated with the phorbol ester 12-0-tetradecanoylphorbol-13-acetate (TPA) to evaluate the role of protein kinase C (PKC) as an upstream intermediate in protein degradation. TPA showed a parabolic dose-response curve for the induction of total protein degradation, with an optimal effect at a concentration of 25 nM, and an optimal incubation time of 3 h. Protein degradation was attenuated by co-incubation with the proteasome inhibitor lactacystin (5 μM), suggesting that it was mediated through the ubiquitin-proteasome proteolytic pathway. TPA induced an increased expression and activity of the ubiquitin-proteasome pathway, as evidenced by an increased functional activity, and increased expression of the 20S proteasome α-subunits, the 19S subunits MSS1 and p42, as well as the ubiquitin conjugating enzyme E214k, also with a maximal effect at a concentration of 25 nM and with a 3 h incubation time. There was also a reciprocal decrease in the cellular content of the myofibrillar protein myosin. TPA induced activation of PKC maximally at a concentration of 25 nM and this effect was attenuated by the PKC inhibitor calphostin C (300 nM), as was also total protein degradation. These results suggest that stimulation of PKC in muscle cells initiates protein degradation through the ubiquitin-proteasome pathway. TPA also induced degradation of the inhibitory protein, I-κBα, and increased nuclear accumulation of nuclear factor-κB (NF-κB) at the same time and concentrations as those inducing proteasome expression. In addition inhibition of NF-κB activation by resveratrol (30 μM) attenuated protein degradation induced by TPA. These results suggest that the induction of proteasome expression by TPA may involve the transcription factor NF-κB. © 2005 Elsevier Inc. All rights reserved.

Increased expression of proteasome subunits in skeletal muscle of cancer patients with weight loss

Atrophy of skeletal muscle is common in patients with cancer and results in increased morbidity and mortality. In order to design effective therapy the mechanism by which this occurs needs to be elucidated. Most studies suggest that the ubiquitin-proteasome proteolytic pathway is most important in intracellular proteolysis, although there have been no reports on the activity of this pathway in patients with different extents of weight loss. In this report the expression of the ubiquitin-proteasome pathway in rectus abdominis muscle has been determined in cancer patients with weight loss of 0-34% using a competitive reverse transcriptase polymerase chain reaction to measure expression of mRNA for proteasome subunits C2 and C5, while protein expression has been determined by western blotting. Overall, both C2 and C5 gene expression was increased by about three-fold in skeletal muscle of cachectic cancer patients (average weight loss 14.5 ± 2.5%), compared with that in patients without weight loss, with or without cancer. The level of gene expression was dependent on the amount of weight loss, increasing maximally for both proteasome subunits in patients with weight loss of 12-19%. Further increases in weight loss reduced expression of mRNA for both proteasome subunits, although it was still elevated in comparison with patients with no weight loss. There was no evidence for an increase in expression at weight losses less than 10%. There was a good correlation between expression of proteasome 20Sα subunits, detected by western blotting, and C2 and C5 mRNA, showing that increased gene expression resulted in increased protein synthesis. Expression of the ubiquitin conjugating enzyme, E214k, with weight loss followed a similar pattern to that of proteasome subunits. These results suggest variations in the expression of key components of the ubiquitin-proteasome pathway with weight loss of cancer patients, and suggest that another mechanism of protein degradation must be operative for patients with weight loss less than 10%. © 2004 Elsevier Ltd. All rights reserved.

The effect of male -male competition and its underlying regulatory mechanisms on the electric signal of the gymnotiform fish Brachyhypopomus gauderio

Sexually-selected communication signals can be used by competing males to settle contests without incurring the costs of fighting. The ability to dynamically regulate the signal in a context-dependent manner can further minimize the costs of male aggressive interactions. Such is the case in the gymnotiform fish Brachyhypopomus gauderio, which, by coupling its electric organ discharge (EOD) waveform to endocrine systems with circadian, seasonal, and behavioral drivers, can regulate its signal to derive the greatest reproductive benefit. My dissertation research examined the functional role of the EOD plasticity observed in male B. gauderio and the physiological mechanisms that regulate the enhanced male EOD. To evaluate whether social competition drives the EOD changes observed during male-male interactions, I manipulated the number of males in breeding groups to create conditions that exemplified low and high competition and measured their EOD and steroid hormone levels. My results showed that social competition drives the enhancement of the EOD amplitude of male B. gauderio. In addition, changes in the EOD of males due to changes in their social environment were paralleled by changes in the levels of androgens and cortisol. I also examined the relationship between body size asymmetry, EOD waveform parameters, and aggressive physical behaviors during male-male interactions in B. gauderio, in order to understand more fully the role of EOD waveforms as reliable signals. While body size was the best determinant of dominance in male B. gauderio, EOD amplitude reliably predicted body condition, a composite of length and weight, for fish in good body condition. To further characterize the mechanisms underlying the relationship between male-male interactions and EOD plasticity, I identified the expression of the serotonin receptor 1A, a key player in the regulation of aggressive behavior, in the brains of B. gauderio. I also identified putative regulatory regions in this receptor in B. gauderio and other teleost fish, highlighting the presence of additional plasticity. In conclusion, male-male competition seems to be a strong selective driver in the evolution of the male EOD plasticity in B. gauderio via the regulatory control of steroid hormones and the serotonergic system.

The Effect of male-male competition and its Underlying Regulatory Mechanisms on the Electric Signal of the Gymnotiform fish Brachyhypopomus gauderio

Sexually-selected communication signals can be used by competing males to settle contests without incurring the costs of fighting. The ability to dynamically regulate the signal in a context-dependent manner can further minimize the costs of male aggressive interactions. Such is the case in the gymnotiform fish Brachyhypopomus gauderio, which, by coupling its electric organ discharge (EOD) waveform to endocrine systems with circadian, seasonal, and behavioral drivers, can regulate its signal to derive the greatest reproductive benefit. My dissertation research examined the functional role of the EOD plasticity observed in male B. gauderio and the physiological mechanisms that regulate the enhanced male EOD. To evaluate whether social competition drives the EOD changes observed during male-male interactions, I manipulated the number of males in breeding groups to create conditions that exemplified low and high competition and measured their EOD and steroid hormone levels. My results showed that social competition drives the enhancement of the EOD amplitude of male B. gauderio. In addition, changes in the EOD of males due to changes in their social environment were paralleled by changes in the levels of androgens and cortisol. I also examined the relationship between body size asymmetry, EOD waveform parameters, and aggressive physical behaviors during male-male interactions in B. gauderio, in order to understand more fully the role of EOD waveforms as reliable signals. While body size was the best determinant of dominance in male B. gauderio, EOD amplitude reliably predicted body condition, a composite of length and weight, for fish in good body condition. To further characterize the mechanisms underlying the relationship between male-male interactions and EOD plasticity, I identified the expression of the serotonin receptor 1A, a key player in the regulation of aggressive behavior, in the brains of B. gauderio. I also identified putative regulatory regions in this receptor in B. gauderio and other teleost fish, highlighting the presence of additional plasticity. In conclusion, male-male competition seems to be a strong selective driver in the evolution of the male EOD plasticity in B. gauderio via the regulatory control of steroid hormones and the serotonergic system.

An investigation of the molecular interactions between statins and bacterial pathogens, and their combined impact on the human immune system

Statins are a class of drug that inhibits cholesterol biosynthesis, and are used to treat patients with high serum cholesterol levels. They exert this function by competitively binding to the enzyme 3-hydroxy-3-methylglutaryl-CoenzymeA reductase (HMGR), which catalyses the formation of mevalonate, a rate-limiting step in cholesterol biosynthesis. In addition, statins have what are called “pleiotropic effects”, which include the reduction of inflammation, immunomodulation, and antimicrobial effects. Statins can also improve survival of patients with sepsis and pneumonia. Cystic fibrosis (CF) is the most common recessive inherited disease in the Caucasian population, which is characterised by factors including, but not limited to, excessive lung inflammation and increased susceptibility to infection. Therefore, the overall objective of this study was to examine the effects of statins on CFassociated bacterial pathogens and the host response. In this work, the prevalence of HMGR was examined in respiratory pathogens, and several CF-associated pathogens were found to possess homologues of this enzyme. HMGR homology was analysed in Staphylococcus aureus, Burkholderia cenocepacia and Streptococcus pneumoniae, and the HMGR of B. cenocepacia was found to have significant conservation to that of Pseudomonas mevalonii, which is the most widely-characterised bacterial HMGR. However, in silico analysis revealed that, unlike S. aureus and S. pneumoniae, B. cenocepacia did not possess homologues of other mevalonate pathway proteins, and that the HMGR of B. cenocepacia appeared to be involved in an alternative metabolic pathway. The effect of simvastatin was subsequently tested on the growth and virulence of S. aureus, B. cenocepacia and S. pneumoniae. Simvastatin inhibited the growth of all 3 species in a dose-dependent manner. In addition, statin treatment also attenuated biofilm formation of all 3 species, and reduced in vitro motility of S. aureus. Interestingly, simvastatin also increased the potency of the aminoglycoside antibiotic gentamicin against B. cenocepacia. The impact of statins was subsequently tested on the predominant CF-associated pathogen Pseudomonas aeruginosa, which does not possess a HMGR homologue. Mevastatin, lovastatin and simvastatin did not influence the growth of this species. However, sub-inhibitory statin concentrations reduced the swarming motility and biofilm formation of P. aeruginosa. The influence of statins was also examined on Type 3 toxin secretion, quorum sensing and chemotaxis, and no statin effect was observed on any of these phenotypes. Statins did not appear to have a characteristic effect on the P. aeruginosa transcriptome. However, a mutant library screen revealed that the effect of statins on P. aeruginosa biofilm was mediated through the PvrR regulator and the Cup fimbrial biosynthesis genes. Furthermore, proteomic analysis demonstrated that 6 proteins were reproducibly induced by simvastatin in the P. aeruginosa swarming cells. The effect of statins on the regulation of the host-P. aeruginosa immune response was also investigated. Statin treatment increased expression of the pro-inflammatory cytokines IL-8 and CCL20 in lung epithelial cells, but did not attenuate P. aeruginosa-mediated inflammatory gene induction. In fact, simvastatin and P. aeruginosa caused a synergistic effect on CCL20 expression. The expression of the transcriptional regulators KLF2 and KLF6 was also increased by statins and P. aeruginosa, with the induction of KLF6 by simvastatin proving to be a novel effect. Interestingly, both statins and P. aeruginosa were capable of inducing alternative splicing of KLF6. P. aeruginosa was found to induce KLF6 alternative splicing by way of the type 3 secreted toxin ExoS. In addition, a mechanistic role was elucidated for KLF6 in the lung, as it was determined that statin-mediated induction of this protein was responsible for the induction of the host response genes CCL20 and iNOS. Moreover, statin treatment caused a slight increase in infection-related cytotoxicity, and increased bacterial adhesion to cells. Taken together, these data demonstrate that statins can reduce the virulence of CFassociated bacterial pathogens and alter host response effectors. Furthermore, novel statin effectors were identified in both bacterial and host cells.

Structural and Functional Analysis of the Caspase –dependent and –independent Domains of the X-linked Inhibitor of Apoptosis Protein in Inflammatory Breast Cancer Tumor Biology

Inflammatory breast cancer (IBC) is an extremely rare but highly aggressive form of breast cancer characterized by the rapid development of therapeutic resistance leading to particularly poor survival. Our previous work focused on the elucidation of factors that mediate therapeutic resistance in IBC and identified increased expression of the anti-apoptotic protein, X-linked inhibitor of apoptosis protein (XIAP), to correlate with the development of resistance to chemotherapeutics. Although XIAP is classically thought of as an inhibitor of caspase activation, multiple studies have revealed that XIAP can also function as a signaling intermediate in numerous pathways. Based on preliminary evidence revealing high expression of XIAP in pre-treatment IBC cells rather than only subsequent to the development of resistance, we hypothesized that XIAP could play an important signaling role in IBC pathobiology outside of its heavily published apoptotic inhibition function. Further, based on our discovery of inhibition of chemotherapeutic efficacy, we postulated that XIAP overexpression might also play a role in resistance to other forms of therapy, such as immunotherapy. Finally, we posited that targeting of specific redox adaptive mechanisms, which are observed to be a significant barrier to successful treatment of IBC, could overcome therapeutic resistance and enhance the efficacy of chemo-, radio-, and immuno- therapies. To address these hypotheses our objectives were: 1. to determine a role for XIAP in IBC pathobiology and to elucidate the upstream regulators and downstream effectors of XIAP; 2. to evaluate and describe a role for XIAP in the inhibition of immunotherapy; and 3. to develop and characterize novel redox modulatory strategies that target identified mechanisms to prevent or reverse therapeutic resistance.

Using various genomic and proteomic approaches, combined with analysis of cellular viability, proliferation, and growth parameters both in vitro and in vivo, we demonstrate that XIAP plays a central role in both IBC pathobiology in a manner mostly independent of its role as a caspase-binding protein. Modulation of XIAP expression in cells derived from patients prior to any therapeutic intervention significantly altered key aspects IBC biology including, but not limited to: IBC-specific gene signatures; the tumorigenic capacity of tumor cells; and the metastatic phenotype of IBC, all of which are revealed to functionally hinge on XIAP-mediated NFκB activation, a robust molecular determinant of IBC. Identification of the mechanism of XIAP-mediated NFκB activation led to the characterization of novel peptide-based antagonist which was further used to identify that increased NFκB activation was responsible for redox adaptation previously observed in therapy-resistant IBC cells. Lastly, we describe the targeting of this XIAP-NFκB-ROS axis using a novel redox modulatory strategy both in vitro and in vivo. Together, the data presented here characterize a novel and crucial role for XIAP both in therapeutic resistance and the pathobiology of IBC; these results confirm our previous work in acquired therapeutic resistance and establish the feasibility of targeting XIAP-NFκB and the redox adaptive phenotype of IBC as a means to enhance survival of patients.

Mineralogical (bulk rock and clay fraction) and stable isotope composition (d13C, d18O) of Aptian deposits of the Bir Oum Ali succession, central Tunisia

Alternations between siliciclastic, carbonate and evaporitic sedimentary systems, as recorded in the Aptian mixed succession of southern Tunisia, reflect profound palaeoceanographic and palaeoclimatic changes in this area of the southern Tethyan margin. The evolution from Urgonian-type carbonates (Berrani Formation, lower Aptian) at the base of the series, to intervals dominated by gypsum or detrital deposits in the remainder of the Aptian is thought to result from the interplay between climate change and tectonic activity that affected North Africa. Based on the evolution of clay mineral assemblages, the early Aptian is interpreted as having been dominated by slightly humid conditions, since smectitic minerals are observed. Near the early to late Aptian boundary, the onset of a gypsiferous sedimentation is associated with the appearance of palygorskite and sepiolite, which supports the installation of arid conditions in this area of the southern Tethyan margin. The evaporitic sedimentation may have also been promoted by the peculiar tectonic setting of the Bir Oum Ali area during the Aptian, where local subsidence may have been tectonically enhanced linked to the opening of northern and central Atlantic. Stress associated with the west and central African rift systems may have triggered the development of NW-SE, hemi-graben structures. Uplifted areas may have constituted potential new sources for clastic material that has been subsequently deposited during the late Aptian. Chemostratigraphic (d13C) correlation of the Bir Oum Ali succession with other peri-Tethyan regions complements biostratigraphic findings, and indicates that a potential expression of the Oceanic Anoxic Event (OAE) 1a may be preserved in this area of Tunisia. Although the characteristic negative spike at the base of this event is not recognized in the present study, a subsequent, large positive excursion with d13C values is of similar amplitude and absolute values to that reported from other peri-Tethyan regions, thus supporting the identification of isotopic segments C4-C7 of the OAE1a. The absence of the negative spike may be linked to either non preservation or non deposition: the OAE1a occurred in a global transgressive context, and since the Bir Oum Ali region was located in the innermost part of the southern Tethyan margin during most of the Aptian, stratigraphic hiatuses may have been longer than in other regions of the Tethys. This emphasizes the importance of integrating several stratigraphic disciplines (bio-, chemo- and sequence stratigraphy) when performing long-distance correlation.

Palynomorphs from the Lateglacial and Holocene of the Mt-Athos Basin, Aegean Sea

To unravel the climatic and environmental dynamics in the borderlands of the Aegean Sea during the early and middle Holocene, and notably for the interval of sapropel S1 (S1) formation, we have analysed terrestrial palynomorphs from a marine core in the northern Aegean Sea. The qualitative results were complemented by quantitative pollen-based climate reconstructions. A land-sea correlation was established based on pollen data and sediment lightness measurements from the same core, and previously published benthic foraminifer data from a nearby core. The borderlands of the Aegean Sea underwent a transition from an open vegetation to oak-dominated woodlands between ~10.4 and ~9.5 ka cal BP. A coeval increase in winter precipitation suggests that moisture availability was the main factor controlling Holocene reforestation. The ~50% higher winter precipitation during S1 formation relative to "pre-sapropelic" conditions suggests a strong contribution from the borderlands of the Aegean Sea to the freshwater surplus during S1 formation. The humid and mild winter conditions during S1 formation were repeatedly punctuated by short-term climatic events that caused a partial deforestation and a reorganisation within the broad-leaved arboreal vegetation. In the marine realm, these events are documented by improved benthic oxygenation. The strongest event represents the regional expression of the 8.2 ka cold event and led to an interruption in S1 formation. Except for the interval of S1 formation, the pollen-derived winter temperatures correlate with the smoothed GISP2 K+ series. They support the previously published, marine-based concept that the intensity of the Siberian High strongly controlled the winter climate in the Aegean region. During S1 formation in the Aegean Sea, however, climate conditions in the borderlands were more strongly affected by the monsoonally influenced climate system of the lower latitudes.

Suppression of the Insulin Receptors in Adult Schistosoma japonicum Impacts on Parasite Growth and Development: Further Evidence of Vaccine Potential

To further investigate the importance of insulin signaling in the growth, development, sexual maturation and egg production of adult schistosomes, we have focused attention on the insulin receptors (SjIRs) of Schistosoma japonicum, which we have previously cloned and partially characterised. We now show, by Biolayer Interferometry, that human insulin can bind the L1 subdomain (insulin binding domain) of recombinant (r)SjIR1 and rSjIR2 (designated SjLD1 and SjLD2) produced using the Drosophila S2 protein expression system. We have then used RNA interference (RNAi) to knock down the expression of the SjIRs in adult S. japonicum in vitro and show that, in addition to their reduced transcription, the transcript levels of other important downstream genes within the insulin pathway, associated with glucose metabolism and schistosome fecundity, were also impacted substantially. Further, a significant decrease in glucose uptake was observed in the SjIR-knockdown worms compared with luciferase controls. In vaccine/challenge experiments, we found that rSjLD1 and rSjLD2 depressed female growth, intestinal granuloma density and faecal egg production in S. japonicum in mice presented with a low dose challenge infection. These data re-emphasize the potential of the SjIRs as veterinary transmission blocking vaccine candidates against zoonotic schistosomiasis japonica in China and the Philippines.

Negative regulation of the hepatic fibrogenic response by suppressor of cytokine signaling 1 (SOCS1)

Abstract: Suppressor of cytokine signaling 1 (SOCS1) is an indispensable regulator of IFN-γ signaling and has been implicated in the regulation of liver fibrosis. However, it is not known whether SOCS1 mediates its anti-fibrotic functions in the liver directly, or via modulating IFN-γ, which has been implicated in attenuating hepatic fibrosis. Additionally, it is possible that SOCS1 controls liver fibrosis by regulating hepatic stellate cells (HSC), a key player in fibrogenic response. While the activation pathways of HSCs have been well characterized, the regulatory mechanisms are not yet clear. The goals of this study were to dissociate IFN-γ-dependent and SOCS1-mediated regulation of hepatic fibrogenic response, and to elucidate the regulatory functions of SOCS1 in H SC activation. Liver fibrosis was induced in Socs1[superscript -/-]Ifng[superscript -/-] mice with dimethylnitrosamine or carbon tetrachloride. Ifng[superscript -/-] and C57BL/6 mice served as controls. Following fibrogenic treatments, Socs1[superscript -/-]Ifng[superscript -/-] mice showed elevated serum ALT levels and increased liver fibrosis com-pared to mice Ifng[superscript -/-]. The latter group showed higher alanine aminotransferase (ALT) levels and fibrosis than C57BL/6 controls. The livers of Socs1-deficient mice showed bridging fibrosis, which was associated with increased accumulation of myofibroblasts and abundant collagen deposition. Socs1-deficient livers showed increased expression of genes coding for smooth muscle actin, collagen, and enzymes involved in remodeling the extracellular matrix, namely matrix metalloproteinases and tissue inhibitor of metalloproteinases. Primary HSCs from Socs1-deficient mice showed increased proliferation in response to growth factors such as HGF, EGF and PDGF, and the fibrotic livers of Socs1-deficient mice showed increased expression of the Pdgfb gene. Taken together, these data indicate that SOCS1 controls liver fibrosis independently of IFN-γ and that part of this regulation may occur via regulating HSC proliferation and limiting growth factor availability.

Tectonic expression of an active slab tear from high-resolution seismic and bathymetric data offshore Sicily (Ionian Sea)

Subduction of a narrow slab of oceanic lithosphere beneath a tightly curved orogenic arc requires the presence of at least one lithospheric scale tear fault. While the Calabrian subduction beneath southern Italy is considered to be the type example of this geodynamic setting, the geometry, kinematics and surface expression of the associated lateral, slab tear fault offshore eastern Sicily remain controversial. Results from a new marine geophysical survey conducted in the Ionian Sea, using high-resolution bathymetry and seismic profiling reveal active faulting at the seafloor within a 140 km long, two-branched fault system near Alfeo Seamount. The previously unidentified 60 km long NW trending North Alfeo Fault system shows primarily strike-slip kinematics as indicated by the morphology and steep-dipping transpressional and transtensional faults. Available earthquake focal mechanisms indicate dextral strike-slip motion along this fault segment. The 80 km long SSE trending South Alfeo fault system is expressed by one or two steeply dipping normal faults, bounding the western side of a 500+ m thick, 5 km wide, elongate, syntectonic Plio-Quaternary sedimentary basin. Both branches of the fault system are mechanically capable of generating magnitude 6-7 earthquakes like those that struck eastern Sicily in 1169, 1542, and 1693.

Recombinant expression of Aryl Hydrocarbon Receptor for quantitative ligand-binding analysis

Recombinant expression of the Aryl Hydrocarbon Receptor (AhR) yields small amounts of ligand- binding competent AhR. Therefore, Spodoptera frugiperda (Sf9) cells and baculovirus have been evaluated for high level and functional expression of AhR. Rat and human AhR were expressed as soluble protein in significant amounts. Expression of ligand-binding competent AhR was sensitive to the protein concentration of Sf9 extract, and co-expression of the chaperone p23 failed to affect the yield of functional ligand-binding AhR. The expression system yielded high levels of functional protein, with the ligand-binding capacity (Bmax) typically 20- fold higher than that obtained with rat liver cytosol. Quantitative estimates of the ligand-binding affinity of human and rat AhR were obtained; the Kd for recombinant rat AhR was indistinguishable from that of native rat AhR, thereby validating the expression system as a faithful model for native AhR. The human AhR bound TCDD with significantly lower affinity than the rat AhR. These findings demonstrate high-level expression of ligand-binding competent AhR, and sufficient AhR for quantitative analysis of ligand-binding.

Enhancing bioactive fatty acids of the meat from lambs reared in intensive systems through nutritional modulation

Tese de Doutoramento em Ciências Veterinárias, especialidade de Produção Animal

Characterization of bone pathologies in the ins2+/akita mouse, a new model for diabetes insulindependent: contribution for a better understanding of the disease in humans

Tese de Doutoramento, Ciências Biomédicas, Departamento de Ciências Biomédicas e Medicina, Universidade do Algarve, 2016