Caracterização da hemodinâmica uterina de éguas durante o ciclo estral

Pós-graduação em Medicina Veterinária - FMVZ

Caracterização do infiltrado inflamatório e avaliação dos marcadores de prognóstico Ki-67, p53, receptor de estrógeno e progesterona no tumor mamário maligno de cadelas

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Avaliação in vivo do potencial estrogênico de amostra de água para consumo humano produzida por estação de tratamento do estado de São Paulo

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Interação entre estradiol e receptores 5 - H T 1A no Núcleo Mediano da Rafe em ratas ovariectomizadas: na aquisição da informação aversiva e associação ao contexto

Pós-graduação em Biociências - FCLAS

Abordagem metodológica para a valuação do petencial desregulador endócrino de água de beber: estudo com amostras reais

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Effects of estrogen deficiency combined with chronic alcohol consumption on rat mandibular condyle

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Efeitos do acetato de deslorelina sobre a produção in vivo e in vitro de embriões de gatas domésticas

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Efeitos do acetato de deslorelina sobre a produção in vivo e in vitro de embriões de gatas domésticas

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Thyroid hormone profile in breast cancer patients in postmenopause

Objective: The aim of this study was to determine thyroid hormone (TH) profile in postmenopausal patients with breast cancer (BC). Subjects and methods: 12 CaM patients stages I or II, without interventions that could interfere with tumor progression were selected, as well as and a control group with 18 postmenopausal women without CaM. We measured serum anti-thyroperoxidase antibody (TPOAB), thyroid-stimulating hormone (TSH), free thyroxine (T4L), estradiol (E2), follicle-stimulating hormone (FSH), and luteinizing hormone (LH), before and after surgery, besides immunohistochemistry for estrogen (ER) and progesterone (PR) receptors. Results: Four patients with CaM showed changes in thyroid hormone profile: two had hyperthyroidism, one hypothyroidism, and one was positive for TPO-AB. All of them positive for ER and PR.TSH levels in breast cancer patients were not different from levels found in the control group (1.89 +/- 1.56 vs. 2.86 +/- 3.12 mIU/mL), but the levels of T4L in patients with CaM were statistically higher than those of the control group (1.83 +/- 0.57 vs. 1.10 +/- 0.20 ng/dL). Conclusion: These results reinforce the need for assessment of thyroid status in CaM patients, since in the absence of E2, changes in clinical HTs can act in E2-controlled processes. Arq Bras Endocrinol Metab. 2012;56(4):238-43

Myocardial Chemokine Expression and Intensity of Myocarditis in Chagas Cardiomyopathy Are Controlled by Polymorphisms in CXCL9 and CXCL10

Background: Chronic Chagas cardiomyopathy (CCC), a life-threatening inflammatory dilated cardiomyopathy, affects 30% of the approximately 8 million patients infected by Trypanosoma cruzi. Even though the Th1 T cell-rich myocarditis plays a pivotal role in CCC pathogenesis, little is known about the factors controlling inflammatory cell migration to CCC myocardium. Methods and Results: Using confocal immunofluorescence and quantitative PCR, we studied cell surface staining and gene expression of the CXCR3, CCR4, CCR5, CCR7, CCR8 receptors and their chemokine ligands in myocardial samples from end-stage CCC patients. CCR5+, CXCR3+, CCR4+, CCL5+ and CXCL9+ mononuclear cells were observed in CCC myocardium. mRNA expression of the chemokines CCL5, CXCL9, CXCL10, CCL17, CCL19 and their receptors was upregulated in CCC myocardium. CXCL9 mRNA expression directly correlated with the intensity of myocarditis, as well as with mRNA expression of CXCR3, CCR4, CCR5, CCR7, CCR8 and their ligands. We also analyzed single-nucleotide polymorphisms for genes encoding the most highly expressed chemokines and receptors in a cohort of Chagas disease patients. CCC patients with ventricular dysfunction displayed reduced genotypic frequencies of CXCL9 rs10336 CC, CXCL10 rs3921 GG, and increased CCR5 rs1799988CC as compared to those without dysfunction. Significantly, myocardial samples from CCC patients carrying the CXCL9/CXCL10 genotypes associated to a lower risk displayed a 2-6 fold reduction in mRNA expression of CXCL9, CXCL10, and other chemokines and receptors, along with reduced intensity of myocarditis, as compared to those with other CXCL9/CXCL10 genotypes. Conclusions: Results may indicate that genotypes associated to reduced risk in closely linked CXCL9 and CXCL10 genes may modulate local expression of the chemokines themselves, and simultaneously affect myocardial expression of other key chemokines as well as intensity of myocarditis. Taken together our results may suggest that CXCL9 and CXCL10 are master regulators of myocardial inflammatory cell migration, perhaps affecting clinical progression to the life-threatening form of CCC.

Prenatal retinoic acid improves lung vascularization and VEGF expression in CDH rat

OBJECTIVE: We sought to investigate the effects of antenatal retinoic acid on the pulmonary vasculature and vascular endothelial growth factor (VEGF) and VEGF receptors (VEGFR) expression in a nitrofen-induced congenital diaphragmatic hernia (CDH) model. STUDY DESIGN: Rat fetuses were exposed to nitrofen at gestational day 9.5 and/or all-trans retinoic acid (ATRA) at gestational days 18.5-20.5. We assessed lung growth, airway, and vascular morphometry. VEGF, VEGFR1, and VEGFR2 expression was analyzed by Western blotting and immunohistochemistry. Continuous data were analyzed by analysis of variance and Kruskal-Wallis test. RESULTS: CDH decreased lung to body weight ratio, increased mean linear intercept and mean transection length/airspace, and decreased mean airspace cord length. ATRA did not affect lung growth or morphometry. CDH increased proportional medial wall thickness of arterioles while ATRA reduced it. ATRA recovered expression of VEGF and receptors, which were reduced in CDH. CONCLUSION: Retinoic acid and VEGF may provide pathways for preventing pulmonary hypertension in CDH.

Motor recovery and synaptic preservation after ventral root avulsion and repair with a fibrin sealant derived from snake venom

Background: Ventral root avulsion is an experimental model of proximal axonal injury at the central/peripheral nervous system interface that results in paralysis and poor clinical outcome after restorative surgery. Root reimplantation may decrease neuronal degeneration in such cases. We describe the use of a snake venom-derived fibrin sealant during surgical reconnection of avulsed roots at the spinal cord surface. The present work investigates the effects of this fibrin sealant on functional recovery, neuronal survival, synaptic plasticity, and glial reaction in the spinal motoneuron microenvironment after ventral root reimplantation. Methodology/Principal Findings: Female Lewis rats (7 weeks old) were subjected to VRA and root replantation. The animals were divided into two groups: 1) avulsion only and 2) replanted roots with fibrin sealant derived from snake venom. Post-surgical motor performance was evaluated using the CatWalk system twice a week for 12 weeks. The rats were sacrificed 12 weeks after surgery, and their lumbar intumescences were processed for motoneuron counting and immunohistochemistry (GFAP, Iba-1 and synaptophysin antisera). Array based qRT-PCR was used to evaluate gene regulation of several neurotrophic factors and receptors as well as inflammatory related molecules. The results indicated that the root reimplantation with fibrin sealant enhanced motor recovery, preserved the synaptic covering of the motoneurons and improved neuronal survival. The replanted group did not show significant changes in microglial response compared to VRA-only. However, the astroglial reaction was significantly reduced in this group. Conclusions/Significance: In conclusion, the present data suggest that the repair of avulsed roots with snake venom fibrin glue at the exact point of detachment results in neuroprotection and preservation of the synaptic network at the microenvironment of the lesioned motoneurons. Also such procedure reduced the astroglial reaction and increased mRNA levels to neurotrophins and anti-inflammatory cytokines that may in turn, contribute to improving recovery of motor function.

Decreased expression of ADAMTS-1 in human breast tumors stimulates migration and invasion

Abstract Background ADAMTS-1 (a disintegrin and metalloprotease with thrombospondin motifs) is a member of the ADAMTS family of metalloproteases. Here, we investigated mRNA and protein levels of ADAMTS-1 in normal and neoplastic tissues using qPCR, immunohistochemistry and immunoblot analyses, and we addressed the role of ADAMTS-1 in regulating migration, invasion and invadopodia formation in breast tumor cell lines. Results In a series of primary breast tumors, we observed variable levels of ADAMTS-1 mRNA expression but lower levels of ADAMTS-1 protein expression in human breast cancers as compared to normal tissue, with a striking decrease observed in high-malignancy cases (triple-negative for estrogen, progesterone and Her-2). This result prompted us to analyze the effect of ADAMTS-1 knockdown in breast cancer cells in vitro. MDA-MB-231 cells with depleted ADAMTS-1 expression demonstrated increased migration, invasion and invadopodia formation. The regulatory mechanisms underlying the effects of ADAMTS-1 may be related to VEGF, a growth factor involved in migration and invasion. MDA-MB-231 cells with depleted ADAMTS-1 showed increased VEGF concentrations in conditioned medium capable of inducing human endothelial cells (HUVEC) tubulogenesis. Furthermore, expression of the VEGF receptor (VEGFR2) was increased in MDA-MB-231 cells as compared to MCF7 cells. To further determine the relationship between ADAMTS-1 and VEGF regulating breast cancer cells, MDA-MB-231 cells with reduced expression of ADAMTS-1 were pretreated with a function-blocking antibody against VEGF and then tested in migration and invasion assays; both were partially rescued to control levels. Conclusions ADAMTS-1 expression was decreased in human breast tumors, and ADAMTS-1 knockdown stimulated migration, invasion and invadopodia formation in breast cancer cells in vitro. Therefore, this series of experiments suggests that VEGF is involved in the effects mediated by ADAMTS-1 in breast cancer cells.

De Novo Design of secondary structures: Synthesis and conformational studies

Chemists have long sought to extrapolate the power of biological catalysis and recognition to synthetic systems. These efforts have focused largely on low molecular weight catalysts and receptors; however, biological systems themselves rely almost exclusively on polymers, proteins and RNA, to perform complex chemical functions. Proteins and RNA are unique in their ability to adopt compact, well-ordered conformations, and specific folding provides precise spatial orientation of the functional groups that comprise the “active site”. These features suggest that identification of new polymer backbones with discrete and predictable folding propensities (“foldamers”) will provide a basis for design of molecular machines with unique capabilities. The foldamer approach complements current efforts to design unnatural properties into polypeptides and polynucleotides. The aim of this thesis is the synthesis and conformational studies of new classes of foldamers, using a peptidomimetic approach. Moreover their attitude to be utilized as ionophores, catalysts, and nanobiomaterials were analyzed in solution and in the solid state. This thesis is divided in thematically chapters that are reported below. It begins with a very general introduction (page 4) which is useful, but not strictly necessary, to the expert reader. It is worth mentioning that paragraph I.3 (page 22) is the starting point of this work and paragraph I.5 (page 32) isrequired to better understand the results of chapters 4 and 5. In chapter 1 (page 39) is reported the synthesis and conformational analysis of a novel class of foldamers containing (S)-β3-homophenylglycine [(S)-β3-hPhg] and D- 4-carboxy-oxazolidin-2-one (D-Oxd) residues in alternate order is reported. The experimental conformational analysis performed in solution by IR, 1HNMR, and CD spectroscopy unambiguously proved that these oligomers fold into ordered structures with increasing sequence length. Theoretical calculations employing ab initio MO theory suggest a helix with 11-membered hydrogenbonded rings as the preferred secondary structure type. The novel structures enrich the field of peptidic foldamers and might be useful in the mimicry of native peptides. In chapter 2 cyclo-(L-Ala-D-Oxd)3 and cyclo-(L-Ala-DOxd) 4 were prepared in the liquid phase with good overall yields and were utilized for bivalent ions chelation (Ca2+, Mg2+, Cu2+, Zn2+ and Hg2+); their chelation skill was analyzed with ESI-MS, CD and 1HNMR techniques and the best results were obtained with cyclo-(L-Ala-D-Oxd)3 and Mg2+ or Ca2+. Chapter 3 describes an application of oligopeptides as catalysts for aldol reactions. Paragraph 3.1 concerns the use of prolinamides as catalysts of the cross aldol addition of hydroxyacetone to aromatic aldeydes, whereas paragraphs 3.2 and 3.3 are about the catalyzed aldol addition of acetone to isatins. By means of DFT and AIM calculations, the steric and stereoelectronic effects that control the enantioselectivity in the cross-aldol addition of acetone to isatin catalysed by L-proline have been studied, also in the presence of small quantities of water. In chapter 4 is reported the synthesis and the analysis of a new fiber-like material, obtained from the selfaggregation of the dipeptide Boc-L-Phe-D-Oxd-OBn, which spontaneously forms uniform fibers consisting of parallel infinite linear chains arising from singleintermolecular N-H···O=C hydrogen bonds. This is the absolute borderline case of a parallel β-sheet structure. Longer oligomers of the same series with general formula Boc-(L-Phe-D-Oxd)n-OBn (where n = 2-5), are described in chapter 5. Their properties in solution and in the solid state were analyzed, in correlation with their attitude to form intramolecular hydrogen bond. In chapter 6 is reported the synthesis of imidazolidin-2- one-4-carboxylate and (tetrahydro)-pyrimidin-2-one-5- carboxylate, via an efficient modification of the Hofmann rearrangement. The reaction affords the desired compounds from protected asparagine or glutamine in good to high yield, using PhI(OAc)2 as source of iodine(III).

Role of αvβ3 – Integrin and TLR2 in the innate response to Herpes Simplex Virus infection and Delivery of retargeted oncolytic Herpes Simplex via carrier cells

In the first part of my thesis I studied the mechanism of initiation of the innate response to HSV-1. Innate immune response is the first line of defense set up by the cell to counteract pathogens infection and it is elicited by the activation of a number of membrane or intracellular receptors and sensors, collectively indicated as PRRs, Patter Recognition Receptors. We reported that the HSV pathogen-associated molecular patterns (PAMP) that activate Toll-like receptor 2 (TLR2) and lead to the initiation of innate response are the virion glycoproteins gH/gL and gB, which constitute the conserved fusion core apparatus across the Herpesvirus. Specifically gH/gL is sufficient to initiate a signaling cascade which leads to NF-κB activation. Then, by gain and loss-of-function approaches, we found that αvβ3-integrin is a sensor of and plays a crucial role in the innate defense against HSV-1. We showed that αvβ3-integrin signals through a pathway that concurs with TLR2, affects activation/induction of interferons type 1, NF-κB, and a polarized set of cytokines and receptors. Thus, we demonstrated that gH/gL is sufficient to induce IFN1 and NF-κB via this pathway. From these data, we proposed that αvβ3-integrin is considered a class of non-TLR pattern recognition receptors. In the second part of my thesis I studied the capacity of human mesenchymal stromal cells isolated by fetal membranes (FM-hMSCs) to be used as carrier cells for the delivery of retargeted R-LM249 virus. The use of systemically administrated carrier cells to deliver oncolytic viruses to tumoral targets is a promising strategy in oncolytic virotherapy. We observed that FM-hMSCs can be infected by R-LM249 and we optimized the infection condition; then we demonstrate that stromal cells sustain the replication of retargeted R-LM249 and spread it to target tumoral cells. From these preliminary data FM-hMSCs resulted suitable to be used as carrier cells