Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 5 de 5
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Eur Urol ; 52(6): 1700-8, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17646044

RESUMO

OBJECTIVES: With conventional methodology, sentinel lymph node (SLN) mapping of invasive urinary bladder cancer is technically challenging. This study was performed to determine the utility of invisible, near-infrared fluorescent (NIRF) light for patient-specific SLN mapping, in real time under complete image guidance. METHODS: Lymphatic tracers, injection volume, NIRF excitation fluence rate, light collection of emitted fluorescence, and degree of bladder distension were systematically optimized in normal dogs and pigs. SLN mapping was then performed in pet dogs with naturally occurring invasive transitional cell carcinoma (InvTCC) of the urinary bladder, which closely mimics the human disease. RESULTS: NIRF albumin (hydrodynamic diameter [HD], 7.4 nm) and NIRF quantum dots (15-20 nm HD) injected into the bladder wall resulted in identification of draining lymph nodes (LNs) in under 3 min. In both species, considerable variability in the lymphatic drainage was observed among individuals. Optimal SLN mapping was achieved with the use of superficial, serosal injection of NIRF tracer, with the bladder distended to an intraluminal pressure of 20-40 cm H(2)O. In dogs with InvTCC, NIRF tracers identified SLNs that were confirmed histologically to harbor metastases. CONCLUSIONS: The use of invisible NIRF light permits real-time, patient-specific identification of SLNs that drain bladder cancer. Intraluminal bladder pressure is a key parameter that needs to be controlled for optimal results.


Assuntos
Modelos Animais de Doenças , Biópsia de Linfonodo Sentinela , Neoplasias da Bexiga Urinária/patologia , Animais , Carcinoma de Células de Transição/patologia , Cães , Feminino , Masculino , Invasividade Neoplásica , Suínos
2.
Ann Surg Oncol ; 14(2): 286-98, 2007 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17096058

RESUMO

BACKGROUND: Understanding lymph drainage patterns of the peritoneum could assist in staging and treatment of gastrointestinal and ovarian malignancies. Sentinel lymph nodes (SLNs) have been identified for solid organs and the pleural space. Our purpose was to determine whether the peritoneal space has a predictable lymph node drainage pattern. METHODS: Rats received intraperitoneal injections of near-infrared (NIR) fluorescent tracers: namely, quantum dots (designed for retention in SLNs) or human serum albumin conjugated with IRDye800 (HSA800; designed for lymphatic flow beyond the SLN). A custom imaging system detected NIR fluorescence at 10 and 20 minutes and 1, 4, and 24 hours after injection. To determine the contribution of viscera to peritoneal lymphatic flow, additional cohorts received bowel resection before NIR tracer injection. Associations with appropriate controls were assessed with the chi(2) test. RESULTS: Quantum dots drained to the celiac, superior mesenteric, and periportal lymph node groups. HSA800 drained to these same groups at early time points but continued flowing to the mediastinal lymph nodes via the thoracic duct. After bowel resection, both tracers were found in the thoracic, not abdominal, lymph node groups. Additionally, HSA800 was no longer found in the thoracic duct but in the anterior chest wall and diaphragmatic lymphatics. CONCLUSIONS: The peritoneal space drains to the celiac, superior mesenteric, and periportal lymph node groups first. Lymph continues via the thoracic duct to the mediastinal lymph nodes. Bowel lymphatics are a key determinant of peritoneal lymph flow, because bowel resection shifts lymph flow directly to the intrathoracic lymph nodes via chest wall lymphatics.


Assuntos
Sistema Linfático/fisiologia , Sistema Linfático/fisiopatologia , Cavidade Peritoneal/fisiologia , Cavidade Peritoneal/fisiopatologia , Animais , Diafragma/fisiologia , Procedimentos Cirúrgicos do Sistema Digestório , Intestinos/fisiologia , Intestinos/fisiopatologia , Intestinos/cirurgia , Linfonodos/fisiologia , Linfonodos/fisiopatologia , Vasos Linfáticos/fisiologia , Vasos Linfáticos/fisiopatologia , Masculino , Modelos Animais , Pontos Quânticos , Ratos , Ratos Sprague-Dawley , Soroalbumina Radioiodada , Espectroscopia de Luz Próxima ao Infravermelho , Ducto Torácico/fisiologia , Parede Torácica/fisiologia
3.
Ann Thorac Surg ; 79(1): 269-77; discussion 269-77, 2005 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-15620956

RESUMO

BACKGROUND: The presence of lymph node metastases is an important prognostic marker with regard to non-small-cell lung cancer (NSCLC). Assessment of the sentinel lymph node (SLN) for the presence of tumor may improve staging. Our objective was to develop an optical noninvasive imaging tool that would permit intraoperative SLN mapping and provide real-time visual feedback for image-guided localization and resection. METHODS: Invisible near-infrared (NIR) light penetrates relatively deeply into tissue and background autofluorescence is low. We have developed a NIR fluorescence imaging system that simultaneously displays color video and NIR images of the surgical field. We recently engineered 15 nm nonradioactive NIR fluorescent quantum dots (QDs) as optimal lymphotrophic optical probes. The introduction of these QDs into lung tissue allows real-time visualization of draining lymphatic channels and nodes. RESULTS: In 12 Yorkshire pigs (mean weight 35 kg) we demonstrated that 200 pmol of NIR QDs injected into lobar parenchyma accurately maps lymphatic drainage and the SLN. All SLNs were strongly fluorescent and easily visualized within 5 minutes of injection. In 14 separate injections QDs localized to a mediastinal node, whereas in 2 injections QDs localized to a hilar intraparenchymal node. Histologic analysis in all cases confirmed the presence of nodal tissue. CONCLUSIONS: We report a highly sensitive rapid technique for SLN mapping of the lung. This technique permits precise real-time imaging and therefore overcomes many limitations of currently available techniques.


Assuntos
Corantes Fluorescentes , Fluorometria , Pulmão/patologia , Linfonodos/patologia , Sistema Linfático/patologia , Pontos Quânticos , Biópsia de Linfonodo Sentinela/métodos , Espectroscopia de Luz Próxima ao Infravermelho , Cirurgia Vídeoassistida/métodos , Animais , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/secundário , Corantes/farmacocinética , Sistemas Computacionais , Estudos de Viabilidade , Retroalimentação Psicológica , Feminino , Corantes Fluorescentes/farmacocinética , Cuidados Intraoperatórios , Neoplasias Pulmonares/patologia , Metástase Linfática/patologia , Masculino , Corantes de Rosanilina/farmacocinética , Sensibilidade e Especificidade , Sus scrofa , Cirurgia Vídeoassistida/instrumentação
4.
Mol Cell Biol ; 24(7): 2698-709, 2004 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15024060

RESUMO

The recruitment of corepressors by DNA-bound repressors is likely to be a critical rate-limiting step in the transcriptional regulation of many genes. An excellent paradigm for such an interaction is the association of the basic helix-loop-helix zipper protein Mad1 with the corepressor mSin3A. When bound together, the Sin3 interaction domain (SID) of Mad1 forms extensive hydrophobic contacts with the four-helix bundle formed by the paired amphipathic helix 2 (PAH2) domain of mSin3A. Using the costructure to predict the principle residues required for binding, we have carried out an extensive mutational analysis to examine the Mad1 SID-mSin3A PAH2 interaction in vitro and in vivo. Bulky hydrophobic residues in the alpha1 (I308 and V311) and alpha2 (L329 and L332) helices of the PAH2 domain are necessary to accommodate the precise arrangement of bulky (L12) and short (A15 and A16) hydrophobic residues in the amphipathic Mad1 SID. We have also used phage display to derive an optimal SID, which shows an essentially identical arrangement of key residues. By manipulating these key residues, we have generated altered-specificity Mad1 SID mutants that bind only to a PAH2 domain with a reciprocal mutation, permitting us to demonstrate for the first time that these domains interact directly in vivo. We have also found that the integrity of the PAH1 domain affects the Mad1 SID-PAH2 interaction. It is conceivable that cross talk between different PAH domains and their binding partners helps to determine the subunit composition and order of assembly of mSin3A complexes.


Assuntos
Fosfoproteínas/metabolismo , Proteínas Repressoras/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , Sequência de Aminoácidos , Animais , Proteínas de Ciclo Celular , Linhagem Celular , Genes Reporter , Humanos , Modelos Moleculares , Mutação , Proteínas Nucleares/química , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fosfoproteínas/química , Fosfoproteínas/genética , Ligação Proteica , Conformação Proteica , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Repressoras/química , Proteínas Repressoras/genética , Complexo Correpressor Histona Desacetilase e Sin3 , Fatores de Transcrição/química , Fatores de Transcrição/genética
5.
Anal Biochem ; 308(2): 364-72, 2002 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-12419351

RESUMO

Peptide phage display is a method by which large numbers of diverse peptides can be screened for binding to a target of interest. Even when successful, the rate-limiting step is usually validation of peptide bioactivity using living cells. In this paper, we describe an integrated system of vectors that expedites both the screening and the characterization processes. Library construction and screening is performed using an optimized type 3 phage display vector, mJ(1), which is shown to accept peptide libraries of at least 23 amino acids in length. Peptide coding sequences are shuttled from mJ(1) into one of three families of mammalian expression vectors for cell physiological studies. The vector pAL(1) expresses phage display-derived peptides as Gal4 DNA binding domain fusion proteins for transcriptional activation studies. The vectors pG(1), pG(1)N, and pG(1)C express phage display-derived peptides as green fluorescent protein fusions targeted to the entire cell, nucleus, or cytoplasm, respectively. The vector pAP(1) expresses phage display-derived peptides as fusions to secreted placental alkaline phosphatase. Such enzyme fusions can be used as highly sensitive affinity reagents for high-throughput assays and for cloning of peptide-binding cell surface receptors. Taken together, this system of vectors should facilitate the development of phage display-derived peptides into useful biomolecules.


Assuntos
Fosfatase Alcalina/metabolismo , Vetores Genéticos , Proteínas Luminescentes/metabolismo , Fragmentos de Peptídeos/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Fatores de Transcrição/metabolismo , Fosfatase Alcalina/genética , Sequência de Aminoácidos , Animais , Bacteriófagos , Sequência de Bases , Clonagem Molecular , Primers do DNA/química , DNA Recombinante , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Fluorescência Verde , Humanos , Proteínas Luminescentes/genética , Melanoma Experimental/genética , Melanoma Experimental/metabolismo , Camundongos , Dados de Sequência Molecular , Proteínas Associadas a Pancreatite , Fragmentos de Peptídeos/genética , Biblioteca de Peptídeos , Reação em Cadeia da Polimerase , Proteínas Recombinantes de Fusão/isolamento & purificação , Proteínas de Saccharomyces cerevisiae/genética , Fatores de Transcrição/genética , Células Tumorais Cultivadas , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...