Preferential location of prilocaine and etidocaine in phospholipid bilayers: A molecular dynamics study

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

Preferential location of lidocaine and etidocaine in lecithin bilayers as determined by EPR, fluorescence and H-2 NMR

We have examined the effect of the uncharged species of lidocaine (LDC) and etidocaine (EDC) on the acyl chain moiety of egg phosphatidylcholine liposomes. Changes in membrane organization caused by both anesthetics were detected through the use of EPR spin labels (5, 7 and 12 doxyl stearic acid methyl ester) or fluorescence probes (4, 6, 10, 16 pyrene-fatty acids). The disturbance caused by the LA was greater when the probes were inserted in more external positions of the acyl chain and decreased towards the hydrophobic core of the membrane. The results indicate a preferential insertion of LDC at the polar interface of the bilayer and in the first half of the acyl chain, for EDC. Additionally, 2 H NMR spectra of multilamellar liposomes composed by acyl chain-perdeutero DMPC and EPC (1:4 mol%) allowed the determination of the segmental order (S-mol) and dynamics (T-1) of the acyl chain region. In accordance to the fluorescence and EPR results, changes in molecular orientation and dynamics are more prominent if the LA preferential location is more superficial, as for LDC while EDC seems to organize the acyl chain region between carbons 2-8, which is indicative of its positioning. We propose that the preferential location of LDC and EDC inside the bilayers creates a "transient site", which is related to the anesthetic potency since it could modulate the access of these molecules to their binding site(s) in the voltage-gated sodium channel. (C) 2007 Elsevier B.V. All rights reserved.

Preferential infection sites of Cysticercus bovis in cattle experimentally infected with Taenia saginata eggs

The preferential sites of infection of Cysticercus bovis were evaluated in the skeletal muscle and entrails of 25 cattle that were experimentally infected with Taenia saginata (2 x 10(4) eggs). Two other animals were not inoculated (control). Ninety days after inoculation, all the cattle were euthanized. The carcasses were deboned and dissected into 26 anatomical sections (masseter muscles, brain, tongue, esophagus, heart, diaphragm, lungs, liver, kidneys, spleen, top sirloin butt, bottom sirloin butt, outside round, top (inside) round, transversus abdominus, top sirloin cap, strip loin, full tenderloin, eye of round, knuckle, shoulder clod, foreshank, shank, chuck, back ribs, and tail muscles). The dissected tissues were sliced into 5 mm sections. From the 25 cattle, 9258 C. bovis (cysticerci) were recovered; 75.02% (6946) of these were recovered from skeletal muscles and 24.98% (2312) from the entrails. A high parasitism level was found in the shoulder clod (12.55%), heart (11.02%), liver (9.48%), masseter muscles (8.51%), chuck (8.25%), strip loin and full tenderloin (7.26%), knuckle (6.63%), and back ribs (5.53%), totaling 69.23% (5738) of all of the detected cysticerci. on the other hand, there was a low C. bovis parasitism level in the brain, spleen, tail muscles, kidneys, esophagus, and diaphragm, representing just 3.9% of the total number of cysticerci. Given these results, we conclude that specific skeletal musculature regions, such as the shoulder blade, chuck, strip loin and full tenderloin, knuckle, back ribs and top round, which are not officially examined in many countries, are effective sites to efficiently screen C. bovis infection. To date, these regions have not been considered as preferential sites of C. bovis infection. Based on our work, however, these regions deserve greater attention from health inspectors because they contained a greater number of Cysticercus than the other regions of carcasses that are parasitized by T. saginata larvae. (c) 2010 Elsevier Ltd. All rights reserved.

Interactions of Paracoccidioides brasiliensis with host cells: recent advances

Host-fungal interactions are inherently complex and dynamic. In order to identify new microbial targets and develop more effective anti-fungal therapies, it is important to understand the cellular and molecular mechanisms of disease. Paracoccidioidomycosis provokes a variety of clinical symptoms, and Paracoccidioides brasiliensis can reach many tissues, but primarily attacks the lungs. The ability of the pathogen to interact with the host surface structures is essential to further colonization, invasion, and growth. Epithelial cells may represent the first host barrier or the preferential site of entry of the fungus. For this reason, interactions between P. brasiliensis and Vero/A549 epithelial cells were evaluated, with an emphasis on the adherence, induction of cytoskeletal alterations, and differential signaling activity of the various surface molecules. The adhesion to and invasion of epithelial cells by P. brasiliensis may represent strategies employed to thwart the initial host immune response, and may help in the subsequent dissemination of the pathogen throughout the body.

Invasion of epithelial mammalian cells by Paracoccidioides brasiliensis leads to cytoskeletal rearrangement and apoptosis of the host cell

Paracoccidioides brasiliensis (Pb) yeast cells can enter mammalian cells and probably manipulate the host cell environment to favor their own growth and survival. We studied the uptake of strain Pb 18 into A549 lung and Vero epithelial cells, with an emphasis on the repercussions in the cytoskeleton and the apoptosis of host cells. Cytoskeleton components of the host cells, such as actin and tubulin, were involved in the P. brasiliensis invasion process. Cytochalasin D and colchicine treatment substantially reduced invasion, indicating the functional participation of microfilaments (MFs) and microtubules (MTs) in this mechanism. Cytokeratin could also play a role in the P. brasiliensis interaction with the host. Gp43 was recognized by anti-actin and anti-cytokeratin antibodies, but not by anti-tubulin. The apoptosis induced by this fungus in infected epithelial cells was demonstrated by various techniques: TUNEL, DNA fragmentation and Bak and Bcl-2 immunocytochemical expression. DNA fragmentation was observed in infected cells but not in uninfected ones, by both TUNEL and gel electrophoresis methods. Moreover, Bcl-2 and Bak did not show any differences until 24 h after infection of cells, suggesting a competitive mechanism that allows persistence of infection. Overexpression of Bak was observed after 48 h, indicating the loss of competition between death and survival signals. In conclusion, the mechanisms of invasion of host cells, persistence within them, and the subsequent induction of apoptosis of such cells may explain the efficient dissemination of P. brasiliensis. (C) 2004 Published by Elsevier SAS.

The self-polarization effect in Pb(Zr0.50Ti0.50)O-3 thin films with no preferential orientation

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

Amniotic Fluid Interleukin-1 Beta and Interleukin-6, but not Interleukin-8 Correlate with Microbial Invasion of the Amniotic Cavity in Preterm Labor

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

Perineural Invasion in Aggressive Skin Carcinomas of the Head and Neck

Introduction: Perineural invasion is a well-recognized form of cancer dissemination. However, it has been reported only in few papers concerning cutaneous carcinomas ( basal cell, BCC, and squamous cell, SCC). Moreover, the incidence is considered to be very low. Niazi and Lambert [Br J Plast Surg 1993; 46: 156-157] reported only 0.18% of perineural invasion among 3,355 BCCs. It is associated with high-risk subtypes, as morphea-like, as well as with an increased risk of local recurrence. No paper was found in the literature looking for perineural invasion in very aggressive skin cancers with skull base extension, with immunohistochemical analysis. Methods: This is a retrospective review, including 35 very advanced skin carcinomas with skull base invasion (24 BCCs and 11 SCCs, operated on at a single institution from 1982 to 2000). Representative slides were immunohistochemically evaluated with antiprotein S-100, in order to enhance nerve fibers and to detect perineural invasion. The results were compared to 34 controls with tumors with a good outcome, treated in the same time frame at the same Institution. Results: Twelve (50.0%) of the BCCs with skull base invasion had proven perineural invasion, as opposed to only 1 (4.6%) of the controls, and this difference was statistically significant (p < 0.001). Regarding SCCs, 7 aggressive tumors (63.6%) showed perineural invasion compared to only 1 (10.0%) of the controls, but this difference did not reach significance (p=0.08), due to the small number of cases. Conclusions: In this series, it was demonstrated that immunohistochemically detected perineural invasion was very prevalent in advanced skin carcinomas. In addition, it was statistically associated with extremely aggressive BCCs with skull base invasion. Copyright (c) 2008 S. Karger AG, Basel

Cortactin is associated with perineural invasion in the deep invasive front area of laryngeal carcinomas

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

Implication of RNA-Binding Protein La in Proliferation, Migration and Invasion of Lymph Node-Metastasized Hypopharyngeal SCC Cells

The 5-year survival rate for oral cavity cancer is poorer than for breast, colon or prostate cancer, and has improved only slightly in the last three decades. Hence, new therapeutic strategies are urgently needed. Here we demonstrate by tissue micro array analysis for the first time that RNA-binding protein La is significantly overexpressed in oral squamous cell carcinoma (SCC). Within this study we therefore addressed the question whether siRNA-mediated depletion of the La protein may interfere with known tumor-promoting characteristics of head and neck SCC cells. Our studies demonstrate that the La protein promotes cell proliferation, migration and invasion of lymph node-metastasized hypopharyngeal SCC cells. We also reveal that La is required for the expression of beta-catenin as well as matrix metalloproteinase type 2 (MMP-2) within these cells. Taken together these data suggest a so far unknown function of the RNA-binding protein La in promoting tumor progression of head and neck SCC.

Loss of Expression and Function of SOCS3 Is an Early Event in HNSCC: Altered Subcellular Localization as a Possible Mechanism Involved in Proliferation, Migration and Invasion

Background: Suppressor of cytokine signaling 3 (SOCS3) is an inducible endogenous negative regulator of signal transduction and activator of transcription 3 (STAT3). Epigenetic silencing of SOCS3 has been shown in head and neck squamous cell carcinoma (HNSCC), which is associated with increased activation of STAT3. There is scarce information on the functional role of the reduction of SOCS3 expression and no information on altered subcellular localization of SOCS3 in HNSCC.Methodology/Principal Findings: We assessed endogenous SOCS3 expression in different HNSCC cell lines by RT-qPCR and western blot. Immunofluorescence and western blot were used to study the subcellular localization of endogenous SOCS3 induced by IL-6. Overexpression of SOCS3 by CMV-driven plasmids and siRNA-mediated inhibition of endogenous SOCS3 were used to verify the role of SOCS3 on tumor cell proliferation, viability, invasion and migration in vitro. In vivo relevance of SOCS3 expression in HNSCC was studied by quantitative immunohistochemistry of commercially-available tissue microarrays. Endogenous expression of SOCS3 was heterogeneous in four HNSCC cell lines and surprisingly preserved in most of these cell lines. Subcellular localization of endogenous SOCS3 in the HNSCC cell lines was predominantly nuclear as opposed to cytoplasmic in non-neoplasic epithelial cells. Overexpression of SOCS3 produced a relative increase of the protein in the cytoplasmic compartment and significantly inhibited proliferation, migration and invasion, whereas inhibition of endogenous nuclear SOCS3 did not affect these events. Analysis of tissue microarrays indicated that loss of SOCS3 is an early event in HNSCC and was correlated with tumor size and histological grade of dysplasia, but a considerable proportion of cases presented detectable expression of SOCS3.Conclusion: Our data support a role for SOCS3 as a tumor suppressor gene in HNSCC with relevance on proliferation and invasion processes and suggests that abnormal subcellular localization impairs SOCS3 function in HNSCC cells.

Conquering Brazil: the invasion by the exotic gekkonid lizard Hemidactylus mabouia (Squamata) in Brazilian natural environments

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

The invasion of five alien species in the Delta do Parnaiba Environmental Protection Area, Northeastern Brazil

Marine biological invasions have been regarded as one of the major causes of native biodiversity loss, with shipping and aquaculture being the leading contributors for the introductions of alien species in aquatic ecosystems. In the present study, five aquatic alien species (one mollusk, three crustaceans and one fish species) were detected during dives, shore searches and from the fisheries on the coast of the Delta do Parnaiba Environmental Protection Area, in the States of Piaui and Maranhao, Northeastern Brazil. The species were the bicolor purse-oyster Isognomon bicolor, the whiteleg shrimp Litopenaeus vannamei, the giant river prawn Macrobrachium rosenbergii, the Indo-Pacific swimming crab Charybdis hellerii and, the muzzled blenny Omobranchus punctatus. Ballast water (I. bicolor, C. hellerii, and O. punctatus) and aquaculture activities (L. vannamei and M. rosenbergii) in adjacent areas are the most likely vectors of introduction. All exotic species found have potential impact risks to the environment because they are able to compete against native species for resources (food and habitat). Isognomon bicolor share the same habitat and food items with the native bivalve species of mussels and barnacles. Litopenaeus vannamei share the same habitat and food items with the native penaeids such as the pinkspot shrimp Farfantepenaeus brasiliensis, the Southern brown shrimp Farfantepenaeus subtilis, and the Southern white shrimp Litopenaeus schmitti, and in the past few years L. vannamei was responsible for a viral epidemics in the cultivation tanks that could be transmitted to native penaeid shrimps. Charybdis hellerii is also able to cause impacts on the local fisheries as the species can decrease the populations of native portunid crabs which are commercialized in the studied region. Macrobrachium rosenbergii may be sharing natural resources with the Amazon River prawn Macrobrachium amazonicum. Omobranchus punctatus shares habit with the native redlip blenny Ophioblennius atlanticus and other fishes, such as the frillfin goby Bathigobius soporator. Some immediate remedial measures to prevent further introductions from ballast water and shrimp farm ponds should be: (i) to prevent the release of ballast water by ship/vessels in the region; (ii) to reroute all effluent waters from shrimp rearing facilities through an underground or above-ground dry well; (iii) to install adequate sand and gravel filter which will allow passage of water but not livestock; (iv) outdoor shrimp pounds located on floodable land should be diked, and; (v) to promote environmental awareness of those directly involved with ballast water (crews of ship/vessels) and shrimp farms in the region. Rev. Biol. Trop. 58 (3): 909-923. Epub 2010 September 01.

RECK-Mediated Inhibition of Glioma Migration and Invasion

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