Padronização da pesquisa de linfonodos sentinelas em estômago por métodos combinados

Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas, Pós-Graduação em Ciências da Cirurgia, Campinas, 2012.Introdução - Com os estudos de Gould et al. (1960), Cabanas (1977) e Morton et al. (1992), estabeleceu-se o conceito da pesquisa do linfonodo sentinela. Esse se baseia na teoria de que ao identificar a presença ou ausência de metástase no primeiro linfonodo que recebe a drenagem linfática a partir do tumor (sentinela), poderia representar o estado de acometimento dos outros linfonodos. Isto evitaria a realização desnecessária de linfadenectomias. Com o passar dos anos, foi consagrada para ser aplicada em casos de melanoma e câncer de mama. Nesta última década, tenta-se estender os princípios da utilização da pesquisa de linfonodo sentinela para os cânceres do aparelho digestivo. Entretanto, no caso do estômago, existem algumas dificuldades, como: presença de sistema de drenagem linfática multidirecional, ocorrência de metástases saltatórias e identificação de mais de um linfonodo sentinela por indivíduo. Objetivo - Criar e padronizar um modelo animal para o treinamento de pesquisa de linfonodos sentinelas em estômago. Método - Trinta e dois coelhos, saudáveis, foram submetidos à anestesia exclusivamente intramuscular. Por meio de laparotomia, foi injetado na subserosa da parede anterior do corpo gástrico, 0,1 ml de fitato marcado com tecnécio-99m (0,2 mCi), em seguida pelo mesmo orifício, de 0,2 ml de Azul Patente V® 2,5%. A cavidade abdominal foi avaliada, in vivo , para pesquisa de suspeitas de linfonodos azuis (corados em azul) e com detector manual de radiação gamma aos 5, 10 e 20 minutos para detecção de suspeitas de linfonodos radioativos (radioatividade identificada superior a 10X o valor apresentado pelo fundo). Após 20 minutos, realizou-se ressecção e exérese total do estômago, baço e suspeitas de linfonodos, para posterior avaliação da radioatividade ex vivo . A seguir, encaminharam-se as suspeitas de linfonodos para estudo histológico para identificação de tecido linfóide. Resultados - Foram identificados linfonodos em 30 coelhos (93,75%) com média de 2,2 por animal. Das 90 suspeitas de linfonodos detectadas, em 70 casos (77,77%) obteve-se confirmação histológica para tecido linfóide. Dessas, a maioria foi identificada e localizada na região entre o esôfago e o fundo gástrico durante a avaliação in vivo aos 5 minutos. Dois coelhos faleceram durante os experimentos (Taxa de mortalidade = 6,25%). Conclusão - O modelo experimental em coelhos para pesquisa de linfonodos sentinelas em estômago por métodos combinados foi factível, de fácil execução e baixa mortalidade, podendo ser usado para treinamento.Abstract : Introduction - The concept of sentinel lymph node was established by the studies of Gould et al. (1960), Cabanas (1977) and Morton et al. (1992). It is based on the theory that, whenever the presence or absence of metastasis is identified in the first lymph node that receives the lymphatic drainage from the tumor (sentinel) the status of involvement of other lymph nodes might be infered. This could avoid the performance of unnecessary lymphadenectomies. Over the years, its use was consecrated by its application in melanoma and breast cancer. In the last decade, attempts have been made to extend the principles of sentinel lymph node investigation to cancers of the digestive tract. In the case of stomach cancer, additional difficulties were found, such as multiple and aberrant lymphatic routes, the occurrence of skip metastasis and the possible identification of more than one sentinel lymph node in the same patient. Aim - To develop and evaluate an animal model for training sentinel lymph node navigation in the stomach. Methods - Thirtytwo healthy rabbits, were prepped and given intramuscular anesthesia. Through a formal laparotomy, they received a subserosal injection of 0.1 ml of phytate labeled with technetium-99m (0.2 mCi) in the anterior wall of the gastric corpus, followed by 0.2 ml of Blue Patent ® V 2.5%, through the same puncture site. Suspicious lymph nodes were searched in-vivo at 5, 10 and 20 minutes, both visually (Blue Patent stained lymph nodes) and with a manual gamma radiation detector (to detect suspected radioactive lymph nodes, displaying radioactivity levels over 10X the value displayed by the background). En-block resection of the stomach, spleen, visible limph nodes and local fat tissue was then performed and the specimen was assessed "ex vivo" for radioactivity. Suspected lymph nodes were sent for histological study to evaluate the presence of lymphoid tissue. Results Radiolabeled or stained lymph nodes were identified in 30 rabbits (93.75%) with an average of 2.2 specimens per animal; of the 90 suspicious lymph nodes detected, histology confirmed lymphoid tissue in 70 cases (77.77%). Most lymph nodes were identified at the 5-minute in-vivo evaluation and their most common location was found to be in the region between the esophagus and the gastric fundus. Two rabbits died during the procedure resulting in a 6.25% mortality. Conclusion - The rabbit model proved adequate for training in sentinel node navigation in the stomach by combined methods (dye and radiocolloid) being easy to execute and associated with low mortality

Quantum Yield Regeneration: Influence of Neutral Ligand Binding on Photophysical Properties in Colloidal Core/Shell Quantum Dots

This article describes an experiment designed to identify the role of specific molecular ligands in maintaining the high photoluminescence (PL) quantum yield (QY) observed in as-synthesized CdSe/CdZnS and CdSe/CdS quantum dots (QDs). Although it has been possible for many years to prepare core/shell quantum dots with near-unity quantum yield through high-temperature colloidal synthesis, purification of such colloidal particles is frequently accompanied by a reduction in quantum yield. Here, a recently established gel permeation chromatography (GPC) technique is used to remove weakly associated ligands without a change in solvent: a decrease in ensemble QY and average PL lifetime is observed. Minor components of the initial mixture that were removed by GPC are then added separately to purified QD samples to determine whether reintroduction of these components can restore the photophysical properties of the initial sample. We show that among these putative ligands trioctylphosphine and cadmium oleate can regenerate the initial high QY of all samples, but only the “L-type” ligands (trioctyphosphine and oleylamine) can restore the QY without changing the shapes of the optical spectra. On the basis of the PL decay analysis, we confirm that quenching in GPC-purified samples and regeneration in ligand-introduced samples are associated chiefly with changes in the relative population fraction of QDs with different decay rates. The reversibility of the QY regeneration process has also been studied; the introduction and removal of trioctylphosphine and oleylamine tend to be reversible, while cadmium oleate is not. Finally, isothermal titration calorimetry has been used to study the relationship between the binding strength of the neutral ligands to the surface and photophysical property changes in QD samples to which they are added

Probing Surface Saturation Conditions in Alternating Layer Growth of CdSe/CdS Core/Shell Quantum Dots

We monitor effective band gap energy shifts and free reagent concentration during the formation of CdS shells on CdSe nanocrystals to test the hypothesis that alternating addition of stoichiometric doses of precursors can effectively saturate surface sites and thereby enforce conformal shell growth. The selective ionic layer addition and reaction (SILAR) mechanism has been proposed to describe growth under such conditions, and the method is attractive because of the opportunity to (1) avoid cross-reaction of precursors in growing binary films in solution and (2) enforce conformal growth, rather than regioselective growth, by saturating all available surface sites in a self-limiting manner in each half-cycle. The strong red shift that takes place when CdS shells are grown on CdSe cores provides a convenient process monitoring tool that complements scanning transmission electron microscopy imaging and analytical measurements of free reagent concentration. We find that, under commonly used conditions, a cadmium oleate precursor reacts incompletely at chalcogenide-saturated nanocrystal surfaces. Although approximately spherical particles are obtained, the growth does not proceed via saturating cycles, as described in the SILAR mechanism. This has implications for the rational control of conformal and regioselective growth of epilayers on nanocrystal quantum dots and higher-dimensional chalcogenide semiconductor nanostructures via solution processes

Discrete Miktoarm Star Block Copolymers with Tailored Molecular Architecture

Molecular architecture is a critical factor in regulating phase behaviors of the block copolymer and prompting the formation of unconventional nanostructures. This work meticulously designed a library of isomeric miktoarm star polymers with an architectural evolution from the linear-branched block copolymer to the miktoarm star block copolymer and to the star-like block copolymer (i.e., 3AB → 3(AB1)B2 → 3(AB)). All of the polymers have precise chemical composition and uniform chain length, eliminating inherent molecular uncertainties such as chain length distribution or architectural defects. The self-assembly behaviors were systematically studied and compared. Gradually increasing the relative length of the branched B1 block regulates the ratio between the bridge and loop configuration and effectively releases packing frustration in the formation of the spherical or cylindrical structures, leading to a substantial deflection of phase boundaries. Complex structures, such as Frank–Kasper phases, were captured at a surprisingly higher volume fraction. Rationally regulating the molecular architecture offers rich possibilities to tune the packing symmetry of block copolymers

Novel Organic–Inorganic Hybrid Electrolyte to Enable LiFePO₄ Quasi-Solid-State Li-Ion Batteries Performed Highly around Room Temperature

A novel type of organic–inorganic hybrid polymer electrolytes with high electrochemical performances around room temperature is formed by hybrid of nanofillers, Y-type oligomer, polyoxyethylene and Li-salt (PBA-Li), of which the <i>T</i>_g and <i>T</i>_m are significantly lowered by blended heterogeneous polyethers and embedded nanofillers with benefit of the dipole modification to achieve the high Li-ion migration due to more free-volume space. The quasi-solid-state Li-ion batteries based on the LiFePO₄/15PBA-Li/Li-metal cells present remarkable reversible capacities (133 and 165 mAh g^–1 @0.2 C at 30 and 45 °C, respectively), good rate ability and stable cycle performance (141.9 mAh g^–1 @0.2 C at 30 °C after 150 cycles)

Purification of Quantum Dots by Gel Permeation Chromatography and the Effect of Excess Ligands on Shell Growth and Ligand Exchange

This article describes the use of gel permeation chromatography (GPC) as a means to separate natively capped colloidal CdSe and CdSe/Cd_<i>x</i>Zn_1–<i>x</i>S quantum dots (QDs) from small-molecule impurities in hydrophobic solvents. A range of analysis techniques, including ¹H NMR, diffusion-ordered NMR analysis (DOSY), and thermogravimetric analysis (TGA) have been used to compare the nature and quantities of ligands adsorbed on the QDs after GPC and after alternative purification methods. We show that the GPC purified samples display lower ligand-to-QD ratio (135 oleate substituents per nanocrystal for CdSe QDs with lowest-energy absorption peak at 534 nm) than what we can achieve by the multiple precipitation/redissolution method, and the GPC purified samples are stable at both room temperature and high temperature (180–200 °C for CdSe QDs). The achievement of an efficient and highly reproducible method for the preparation of clean QD samples allowed us to test whether impurities that reside in samples prepared by standard purification methods have a significant effect on further surface modification reactions. We found that the reactivity of CdSe QDs toward precursors for CdS shell growth was profoundly affected by the presence of excess ligands in impure QD samples prepared by multiple precipitations and that the removal of excess ligands and impurities significantly improved the speed and reliability by which water-soluble CdSe/Cd_<i>x</i>Zn_1–<i>x</i>S QDs could be prepared by ligand exchange with cysteine. GPC purification provides a preparative-scale, consistent, size-based purification of QDs without perturbing the solvent environment and as such could serve as the basis for advanced syntheses and enable detailed measurements of QD surface chemical properties

Copolymerization and Synthesis of Multiply Binding Histamine Ligands for the Robust Functionalization of Quantum Dots

The polymeric functionalization of quantum dots via ligand exchange is a robust method for the preparation of stable fluorescent particles with high quantum yields. For most biological applications of quantum dots, water solubility is a key requirement; to achieve biocompatibility, polymeric ligand systems that can provide water solubility as well as effective anchoring groups are advantageous. In this work, histamine functional polymers bearing poly­(ethylene glycol) (PEG) side chains were prepared using RAFT polymerization. A versatile postmodification strategy using activated ester units of <i>N</i>-methacryloxysuccinimide (NMS) and poly­(ethylene glycol) methacrylate in the polymer chain afforded copolymers ranging from 6K to 50K with low polydispersities, along with tailored composition of each monomer along the copolymer chain. By controlling the monomer ratio, PEGMA molecular weight, time, and temperature, the composition could be tuned to study its effect on quantum dot functionalization. Representative oleate-capped CdSe/Cd_<i>x</i>Zn_1–<i>x</i>S QDs purified by a recently established gel permeation chromatography (GPC) method were used to test the effectiveness of the histamine-bearing polymers for preparation of water-soluble QDs. Successful ligand exchange of the QDs was characterized by good dispersions in water, lack of aggregation between QDs, and good quantum yields in water. Overall, the synthetic method demonstrates a facile and robust postmodification strategy for the formation of multiply binding, histamine-bearing copolymers, which can be applied to nanomaterials for fundamental investigations and bioimaging/biodistribution studies

Antineoplastic Agents. 585. Isolation of <i>Bridelia ferruginea</i> Anticancer Podophyllotoxins and Synthesis of 4‑Aza-podophyllotoxin Structural Modifications

Cytotoxic constituents of the terrestrial plant <i>Bridelia ferruginea</i> were isolated using bioactivity-guided fractionation, which revealed the presence of the previously known deoxypodophyllotoxin (<b>1</b>), isopicrodeoxypodophyllotoxin (<b>2</b>), β-peltatin (<b>3</b>), β-peltatin-5-<i>O</i>-β-d-glucopyranoside (<b>3a</b>), and the indole neoechinulin (<b>4</b>). As an extension of previous podophyllotoxin research, SAR studies were undertaken focused on 4-aza-podophyllotoxin structural modifications. A number of such derivatives were synthesized following modifications to the A and E rings. Such structural modifications with alkyl and 4-fluorobenzyl substituents at the 4-aza position provided the most potent cancer cell growth inhibitory activity (GI₅₀ 0.1 to <0.03 μg/mL) against a panel of six human cancer cell lines and one murine cancer cell line. Several compounds corresponding to 4′-demethylated modifications were also synthesized and found to be significantly less potent

Composition and antimicrobial activity of the essential oil from the branches of <i>Jacaranda cuspidifolia</i> Mart. growing in Sichuan, China

<p>This study represents the first report on the chemical composition and antimicrobial activity of the essential oil from the branches of <i>Jacaranda cuspidifolia</i> Mart. Thirty-three compounds were identified by Gas Chromatography-Mass Spectrometry (GC-MS) and the major constituents of the essential oil were Palmitic acid (31.36%), (<i>Z</i>) − 9,17-Octadecadienal (12.06%), Ethyl palmitate (3.81%), Perhydrofarnesyl acetone (2.07%), <i>γ</i>-Maaliene (1.88%), and Cedro (1.42%) and 9,12-Octadecadienoic acid ethyl ester (1.42%). The <i>in vitro</i> antimicrobial activities of the essential oil were evaluated by the disc diffusion method, and the inhibition zones against <i>Escherichia coli</i>, <i>Staphylococcus aureus</i> and <i>Candida albicans</i> were 7.10, 8.20 and 7.25 mm, respectively. The oil showed moderate activities against <i>E. coli</i>, <i>S. aureus</i> and <i>C. albicans</i> with minimum inhibition concentration (MIC) values of 17.3 mg/mL, 12.9 mg/mL and 16.0 mg/mL, respectively.</p

Fe-Cluster Pushing Electrons to N‑Doped Graphitic Layers with Fe₃C(Fe) Hybrid Nanostructure to Enhance O₂ Reduction Catalysis of Zn-Air Batteries

Non-noble metal catalysts with catalytic activity toward oxygen reduction reaction (ORR) comparable or even superior to that of Pt/C are extremely important for the wide application of metal–air batteries and fuel cells. Here, we develop a simple and controllable strategy to synthesize Fe-cluster embedded in Fe₃C nanoparticles (designated as Fe₃C­(Fe)) encased in nitrogen-doped graphitic layers (NDGLs) with graphitic shells as a novel hybrid nanostructure as an effective ORR catalyst by directly pyrolyzing a mixture of Prussian blue (PB) and glucose. The pyrolysis temperature was found to be the key parameter for obtaining a stable Fe₃C­(Fe)@NDGL core–shell nanostructure with an optimized content of nitrogen. The optimized Fe₃C­(Fe)@NDGL catalyst showed high catalytic performance of ORR comparable to that of the Pt/C (20 wt %) catalyst and better stability than that of the Pt/C catalyst in alkaline electrolyte. According to the experimental results and first principle calculation, the high activity of the Fe₃C­(Fe)@NDGL catalyst can be ascribed to the synergistic effect of an adequate content of nitrogen doping in graphitic carbon shells and Fe-cluster pushing electrons to NDGL. A zinc–air battery utilizing the Fe₃C­(Fe)@NDGL catalyst demonstrated a maximum power density of 186 mW cm^–2, which is slightly higher than that of a zinc–air battery utilizing the commercial Pt/C catalyst (167 mW cm^–2), mostly because of the large surface area of the N-doped graphitic carbon shells. Theoretical calculation verified that O₂ molecules can spontaneously adsorb on both pristine and nitrogen doped graphene surfaces and then quickly diffuse to the catalytically active nitrogen sites. Our catalyst can potentially become a promising replacement for Pt catalysts in metal-air batteries and fuel cells