RESUMO
BACKGROUND: Pheochromocytomas and paragangliomas (PCPG) are rare catecholamine-secreting endocrine tumors deriving from chromaffin cells of the embryonic neural crest. Although distinct molecular PCPG subtypes have been elucidated, certain characteristics of these tumors have yet to be fully examined, namely the tumor microenvironment (TME). To further understand tumor-stromal interactions in PCPG subtypes, the present study deconvoluted bulk tumor gene expression to examine ligand-receptor interactions. METHODS: RNA-sequencing data primary solid PCPG tumors were derived from The Cancer Genome Atlas (TCGA). Tumor purity was estimated using two robust algorithms. The tumor purity estimates and bulk tumor expression values allowed for non-negative linear regression to predict the average expression of each gene in the stromal and tumor compartments for each PCPG molecular subtype. The predicted expression values were then used in conjunction with a previously curated ligand-receptor database and scoring system to evaluate top ligand-receptor interactions. RESULTS: Across all PCPG subtypes compared to normal samples, tumor-to-tumor signaling between bone morphogenic proteins 7 (BMP7) and 15 (BMP15) and cognate receptors ACVR2B and BMPR1B was increased. In addition, tumor-to-stroma signaling was enriched for interactions between predicted tumor-originating delta-like ligand 3 (DLL3) and predicted stromal NOTCH receptors. Stroma-to-tumor signaling was enriched for interactions between ephrins A1 and A4 with ephrin receptors EphA5, EphA7, and EphA8. Pseudohypoxia subtype tumors displayed increased predicted stromal expression of genes related to immune-exhausted T-cell response, including those for inhibitory receptors HAVCR2 and CTLA4. CONCLUSION: The current exploratory study predicted stromal and tumor through compartmental deconvolution and yielded previously unrecognized interactions and putative biomarkers in PCPG.
Assuntos
Neoplasias das Glândulas Suprarrenais , Paraganglioma , Feocromocitoma , Neoplasias das Glândulas Suprarrenais/patologia , Biomarcadores Tumorais/genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Ligantes , Proteínas de Membrana/genética , Paraganglioma/genética , Paraganglioma/patologia , Feocromocitoma/genética , Feocromocitoma/patologia , Transcriptoma , Microambiente Tumoral/genéticaRESUMO
Developmental genes are often regulated by multiple elements with overlapping activity. Yet, in most cases, the relative function of those elements and their contribution to endogenous gene expression remain poorly characterized. An example of this phenomenon is that distinct sets of enhancers have been proposed to direct Fgf8 in the limb apical ectodermal ridge and the midbrain-hindbrain boundary. Using in vivo CRISPR/Cas9 genome engineering, we functionally dissect this complex regulatory ensemble and demonstrate two distinct regulatory logics. In the apical ectodermal ridge, the control of Fgf8 expression appears distributed between different enhancers. In contrast, we find that in the midbrain-hindbrain boundary, one of the three active enhancers is essential while the other two are dispensable. We further dissect the essential midbrain-hindbrain boundary enhancer to reveal that it is also composed by a mixture of essential and dispensable modules. Cross-species transgenic analysis of this enhancer suggests that its composition may have changed in the vertebrate lineage.
Assuntos
Desenvolvimento Embrionário/genética , Elementos Facilitadores Genéticos/genética , Fator 8 de Crescimento de Fibroblasto/genética , Regulação da Expressão Gênica no Desenvolvimento , Engenharia Genética/métodos , Animais , Sistemas CRISPR-Cas/genética , Ectoderma/embriologia , Embrião de Mamíferos , Extremidades/embriologia , Estudos de Viabilidade , Feminino , Fator 8 de Crescimento de Fibroblasto/metabolismo , Redes Reguladoras de Genes , Masculino , Mesencéfalo/embriologia , Camundongos , Camundongos Transgênicos , Rombencéfalo/embriologiaRESUMO
BACKGROUND AND PURPOSE: Current treatment guidelines for gallbladder cancer range from simple cholecystectomy to regional hepatic resection. Treatment patterns for radical resection and adjuvant chemotherapy vary. We aim to determine if there is any disparity in treatment or difference in survival between academic versus community treatment centers. METHODS: The National Cancer Database (NCDB) was queried from 2004 to 2014 for gallbladder carcinoma. Cases were stratified into treatment sites as "Community Cancer Center" (CCC) or "Academic Cancer Center" (ACC). Propensity score matching was performed for patient demographics, TNM stage, resection type, and administration of adjuvant chemotherapy. The primary outcome included 30-day mortality, 90-day mortality, and overall survival. RESULTS: There are similar frequencies of radical versus simple resection and administration of adjuvant chemotherapy between ACC and CCC. When propensity-matched for resection type, cases treated at ACC have lower 30-day mortality (4.1% vs. 6.9%) and 90-day mortality (13.2% vs. 18.5%) and increased 5-year overall survival (26.2% vs. 22.4%) (p < 0.01). After propensity matching for adjuvant chemotherapy, cases at ACC have lower 30-day mortality (4.12% vs. 7.71%) and 90-day mortality (13.22% vs. 19.19%) and increased overall survival (13.6% vs. 11.0%) (p < 0.01). DISCUSSION AND CONCLUSIONS: While treatment patterns for gallbladder cancer at ACC and CCC were similar, there was a decrease in 30-day and 90-day mortality and improved overall survival associated with patients treated at ACC. Treatment site may have an impact in the surgical outcomes of gallbladder cancer patients. This disparity warrants further research.
Assuntos
Neoplasias da Vesícula Biliar , Quimioterapia Adjuvante , Colecistectomia , Neoplasias da Vesícula Biliar/patologia , Neoplasias da Vesícula Biliar/cirurgia , Humanos , Estadiamento de Neoplasias , Pontuação de Propensão , Resultado do TratamentoRESUMO
BACKGROUND: The MELD score has been demonstrated to be predictive of hepatectomy outcomes in mixed patient samples of primary and secondary liver cancers. Because MELD is a measure of hepatic dysfunction, prior conclusions may rely on the high prevalence of cirrhosis observed with primary lesions. This study aims to evaluate MELD score as a predictor of mortality and develop a risk prediction model for patients specifically undergoing hepatic metastasectomy. METHODS: ACS-NSQIP 2005-2013 was analyzed to select patients who had undergone liver resections for metastases. A receiver operating characteristic (ROC) analysis determined the MELD score most associated with 30-day mortality. A literature review identified variables that impact hepatectomy outcomes. Significant factors were included in a multivariable analysis (MVA). A risk calculator was derived from the final multivariable model. RESULTS: Among the 14,919 patients assessed, the mortality rate was 2.7%, and the median MELD was 7.3 (range = 34.4). A MELD of 7.24 was identified by ROC (sensitivity = 81%, specificity = 51%, c-statistic = 0.71). Of all patients above this threshold, 4.4% died at 30 days vs. 1.1% in the group ≤7.24. This faction represented 50.1% of the population but accounted for 80.3% of all deaths (p < 0.001). The MVA revealed mortality to be increased 2.6-times (OR = 2.55, 95%CI 1.69-3.84, p < 0.001). A risk calculator was successfully developed and validated. CONCLUSIONS: MELD>7.24 is an important predictor of death following hepatectomy for metastasis and may prompt a detailed assessment with the provided risk calculator. Attention to MELD in the preoperative setting will improve treatment planning and patient education prior to oncologic liver resection.
Assuntos
Hepatectomia/mortalidade , Neoplasias Hepáticas/secundário , Neoplasias Hepáticas/cirurgia , Metastasectomia/mortalidade , Idoso , Creatinina/sangue , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de DoençaRESUMO
The ancestral role of the Hox gene family is specifying morphogenetic differences along the main body axis. In vertebrates, HoxD genes were also co-opted along with the emergence of novel structures such as limbs and genitalia. We propose that these functional recruitments relied on the appearance, or implementation, of regulatory sequences outside of the complex. Whereas transgenic human and murine HOXD clusters could function during axial patterning, in mice they were not expressed outside the trunk. Accordingly, deletion of the entire cluster abolished axial expression, whereas recently acquired regulatory controls were preserved.
Assuntos
Proteínas de Ligação a DNA , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/fisiologia , Camundongos Transgênicos , Proteínas de Neoplasias , Animais , Desenvolvimento Ósseo/genética , Embrião de Mamíferos/metabolismo , Evolução Molecular , Deleção de Genes , Regulação da Expressão Gênica no Desenvolvimento , Genes Reporter , Humanos , Hibridização In Situ , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Modelos Genéticos , Família Multigênica , Mutação , Fenótipo , Recombinação Genética , Fatores de Tempo , Fatores de Transcrição/genéticaRESUMO
This paper discusses a method for overcoming the problem of weak sinks representing wells that result from spatial discretization effects when using MODPATH, the particle-tracking postprocessor for the ground water flow model MODFLOW. Weak sink cells are model cells that represent a well that does not discharge at a sufficiently large rate to capture all of the flow entering the cell; therefore, flowpaths within these cells cannot be uniquely defined because it is impossible to know whether a given water particle discharges to the well or passes through the cell. Creating a submodel of the well cell by using the nested rediscretization method can eliminate this ambiguity by converting the weak sink cell into a strong sink cell. The method is designed to be run manually for each well and for steady-state conditions. Other advantages, disadvantages, technical considerations, and limitations of the method are presented. Software created for the method consists of five Fortran programs that are operated using a set of instructions. A practical application of the method is presented by using an example wellhead-protection problem that demonstrates that nested rediscretization can provide more accurate particle-tracking results than those obtained by using a coarsely discretized model alone.
Assuntos
Modelos Teóricos , Movimentos da Água , Abastecimento de Água , SoloRESUMO
Most patients succumbing to colorectal cancer fail with liver-predominant metastases. To make a clinical impact in this disease, a systemic or whole-liver therapy may be required, whereas most cancer gene therapy approaches are limited in their ability to treat beyond local disease. As a preclinical model for cancer gene therapy, recombinant adenovirus containing the human IFN-beta (hIFN-beta) cDNA was delivered systemically in nude mouse xenograft models of human colorectal cancer liver metastases. The vector targeted hepatocytes that produced high levels of hIFN-beta in the liver, resulting in a profound apoptotic response in the tumors and significant tumor regression. hIFN-beta gene therapy not only resulted in improved survival and long-term cure in a micrometastatic model, but provided similar benefits in a clinically relevant gross disease model. A similar recombinant adenovirus containing the murine IFN-beta (mIFN-beta) cDNA also resulted in a therapeutic response and improved survival in syngeneic mouse models of colorectal cancer liver metastases. Depletion studies demonstrate a contribution of natural killer cells to this therapeutic response. The toxicity of an adenoviral vector expressing murine IFN-beta in a syngeneic model is also presented. These encouraging results warrant further investigation of the use of cancer gene therapy for targeting metastatic disease.
Assuntos
Adenocarcinoma/secundário , Adenoviridae/genética , Neoplasias Colorretais/patologia , DNA Complementar/uso terapêutico , Terapia Genética , Vetores Genéticos/uso terapêutico , Interferon beta/uso terapêutico , Neoplasias Hepáticas/secundário , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/terapia , Animais , Apoptose , Citomegalovirus/genética , DNA Complementar/administração & dosagem , DNA Complementar/genética , DNA Complementar/toxicidade , Feminino , Genes Sintéticos , Vetores Genéticos/administração & dosagem , Vetores Genéticos/genética , Vetores Genéticos/toxicidade , Hepatócitos/metabolismo , Humanos , Injeções Intraperitoneais , Injeções Intravenosas , Interferon beta/administração & dosagem , Interferon beta/genética , Interferon beta/toxicidade , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/terapia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Camundongos SCID , Transplante de Neoplasias , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/terapia , Regiões Promotoras Genéticas , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/fisiologia , Proteínas Recombinantes de Fusão/uso terapêutico , Proteínas Recombinantes de Fusão/toxicidade , Células Tumorais Cultivadas/transplante , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Hepatic artery infusion of adenoviral vectors has been shown to increase transduction of certain hepatocellular malignancies in preclinical studies. In addition, clinical trials have begun evaluating the efficacy of gene transfer of cytotoxic genes to metastatic colorectal tumors through hepatic artery infusion. Here we evaluate the extent of gene expression and therapeutic effect following various routes of administration of recombinant adenovirus in a rat model of metastatic colorectal carcinoma. We administered adenovirus (AdCMVlacZ) to rats with established colorectal metastases through infusion into the hepatic artery, intravenous infusion, or direct injection into a tumor. Intravenous administration resulted in transduction of hepatocytes, but not tumor cells. Hepatic arterial administration failed to substantially increase transduction of tumor cells. In addition, ligation of the hepatic artery following infusion of adenovirus or the addition of lipiodol infusion had no effect on the transduction of tumor cells. We administered AdCMVp53 by direct injection into tumors, intravenous administration, or hepatic artery infusion to evaluate the delivery of a therapeutic gene. Direct injection of AdCMVp53 into established hepatic colorectal metastases resulted in a therapeutic response in comparison with both hepatic arterial and intravenous infusion of vector. These preclinical studies fail to support a strategy of infusion through the hepatic artery of recombinant adenovirus targeting tumor cells in the treatment of colorectal cancer liver metastases.
Assuntos
Neoplasias Colorretais/genética , Técnicas de Transferência de Genes , Vetores Genéticos/administração & dosagem , Adenoviridae/genética , Animais , Feminino , Expressão Gênica , Terapia Genética , Artéria Hepática , Humanos , Infusões Intra-Arteriais , Infusões Intravenosas , Metástase Neoplásica , Ratos , Ratos Nus , Transdução Genética , Células Tumorais CultivadasRESUMO
Gene therapy remains a new and exciting therapy that holds the potential to impact the care of many diseases. Cancer gene therapy strategies encompass a major part of this developing field. Initial preclinical and phase I clinical trials have demonstrated the ability to transfer genetic material to cells in vitro and in vivo with resultant expression of biologically active protein. Most of these studies have involved direct injection or local installation of vector. A majority of patients succumbing to cancer do so because of metastatic disease. Clearly, to broaden the impact of cancer gene therapy on these patients' outcome, new strategies for targeting regional or systemic disease are required. This article offers a review of current vectors and therapeutic strategies along with the application of these in human cancers.
Assuntos
Terapia Genética/métodos , Metástase Neoplásica/terapia , Adjuvantes Imunológicos/uso terapêutico , Previsões , Terapia Genética/normas , Terapia Genética/tendências , Vetores Genéticos/classificação , Vetores Genéticos/uso terapêutico , Humanos , Metástase Neoplásica/genética , Resultado do TratamentoRESUMO
INTRODUCTION: Sarcomatosis is the disseminated intraperitoneal spread of sarcoma. It is a condition for which there is no effective treatment. Photodynamic therapy (PDT) is a cancer treatment modality that uses a photosensitizing agent and laser light to kill cells. We report our preliminary Phase II clinical trial experience using PDT for the treatment of intraperitoneal sarcomatosis. METHODS: From May 1997 to December 1998 eleven patients received twelve PDT treatments for intraperitoneal sarcomatosis. Photofrin (PF) 2.5 mg/kg was administered intravenously 48 hours before surgical debulking to a maximum residual tumor size of less than 5 mm. Light therapy was administered at a fluence of 2.5 J/cm2 of 532 nm green light to the mesentery and serosa of the small bowel and colon; 5 J/cm2 of 630 nm red light to the stomach and duodenum; 7.5 J/cm2 of red light to the surface of the liver, spleen, and diaphragms; and 10 J/cm2 of red light to the retroperitoneal gutters and pelvis. Light fluence was measured with an on-line light dosimetry system. Response to treatment was evaluated by abdominal CT scan at 3 and 6 months, diagnostic laparoscopy at 3 to 6 months, and clinical examination every 3 months. RESULTS: Adequate tumor debulking required an omentectomy in eight patients (73%), small bowel resection in seven patients (64%), colon resection in four patients (36%), splenectomy in one patient (9%), and a left spermatic cord resection in one patient. Five patients (45%) have no evidence of disease at follow-up (range, 1.7-17.3 months), including patients at 13.8 and 17.3 months examined by CT. Two patients (18%) died from disease progression. Four patients (36%) are alive with disease progression. Toxicities related to PDT included substantial postoperative fluid shifts with volume overload, transient thrombocytopenia, and elevated liver function tests. One patient suffered a postoperative pulmonary embolism complicated by adult respiratory distress syndrome (ARDS). CONCLUSIONS: Debulking surgery with intraperitoneal PDT for sarcomatosis is feasible. Preliminary response data suggest prolonged relapse-free survival in some patients. Additional follow-up with more patients will be necessary for full evaluation of the added benefit of PDT and aggressive surgical debulking in these patients.
Assuntos
Fotorradiação com Hematoporfirina/métodos , Neoplasias Peritoneais/tratamento farmacológico , Sarcoma/tratamento farmacológico , Adulto , Terapia Combinada , Éter de Diematoporfirina/efeitos adversos , Éter de Diematoporfirina/uso terapêutico , Intervalo Livre de Doença , Feminino , Seguimentos , Humanos , Laparotomia , Masculino , Pessoa de Meia-Idade , Pennsylvania/epidemiologia , Neoplasias Peritoneais/mortalidade , Neoplasias Peritoneais/cirurgia , Sarcoma/mortalidade , Sarcoma/cirurgia , Taxa de SobrevidaAssuntos
Articulações/embriologia , Mesoderma/citologia , Proteínas/genética , Proteínas/metabolismo , Animais , Desenvolvimento Ósseo , Cartilagem/citologia , Cartilagem/embriologia , Contagem de Células , Diferenciação Celular , Embrião de Galinha , Condrócitos/citologia , Técnicas de Cultura , Expressão Gênica , Cápsula Articular/metabolismo , Articulações/citologia , Articulações/metabolismo , Botões de Extremidades , Mesoderma/metabolismo , Transdução de Sinais , Células-Tronco/citologia , Membrana Sinovial/metabolismoRESUMO
BACKGROUND: Photodynamic therapy (PDT) combines photosensitizer drug, oxygen, and laser light to kill tumor cells on surfaces. This is the initial report of our phase II trial, designed to evaluate the effectiveness of surgical debulking and PDT in carcinomatosis and sarcomatosis. METHODS: Fifty-six patients were enrolled between April 1997 and January 2000. Patients were given Photofrin (2.5 mg/kg) intravenously 2 days before tumor-debulking surgery. Laser light was delivered to all peritoneal surfaces. Patients were followed with CT scans and laparoscopy to evaluate responses to treatment. RESULTS: Forty-two patients were adequately debulked at surgery; these comprise the treatment group. There were 14 GI malignancies, 12 ovarian cancers and 15 sarcomas. Actuarial median survival was 21 months. Median time to recurrence was 3 months (range, 1-21 months). The most common serious toxicities were anemia (38%), liver function test (LFT) abnormalities (26%), and gastrointestinal toxicities (19%), and one patient died. CONCLUSIONS: Photofrin PDT for carcinomatosis has been successfully administered to 42 patients, with acceptable toxicity. The median survival of 21 months exceeds our expectations; however, the relative contribution of surgical resection versus PDT is unknown. Deficiencies in photosensitizer delivery, tissue oxygenation, or laser light distribution leading to recurrences may be addressed through the future use of new photosensitizers.
Assuntos
Antineoplásicos/uso terapêutico , Carcinoma/cirurgia , Éter de Diematoporfirina/uso terapêutico , Neoplasias Peritoneais/cirurgia , Fotoquimioterapia , Sarcoma/cirurgia , Adulto , Idoso , Carcinoma/tratamento farmacológico , Terapia Combinada , Feminino , Seguimentos , Humanos , Infusões Intravenosas , Masculino , Pessoa de Meia-Idade , Neoplasias Peritoneais/tratamento farmacológico , Sarcoma/tratamento farmacológico , Taxa de Sobrevida , Resultado do TratamentoRESUMO
Yeast two-hybrid techniques were used to identify possible effectors for the heterotrimeric G protein G(z) in human bone marrow cells. Eya2, a human homologue of the Drosophila Eya transcription co-activator, was identified. Eya2 interacts with activated Galpha(z) and at least one other member of the Galpha(i) family, Galpha(i2). Interactions were confirmed in mammalian two-hybrid and glutathione S-transferase fusion protein pull-down assays. Regions of Eya2-mediating interaction were mapped to the C-terminal Eya consensus domain. Eya2 is an intrinsically cytosolic protein that is translocated to the nucleus by members of the Six homeodomain-containing family of proteins. Activated Galpha(z) and Galpha(i2) prevent Eya2 translocation and inhibit Six/Eya2-mediated activation of a reporter gene controlled through the MEF3/TATA promoter. Although G proteins are known to regulate the activity of numerous transcription factors, this regulation is normally achieved indirectly via one or more intermediates. We show here a novel functional regulation of a co-activator directly by G protein subunits.
Assuntos
Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/metabolismo , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Proteínas de Homeodomínio/metabolismo , Transativadores/metabolismo , Animais , Células da Medula Óssea/metabolismo , Núcleo Celular/metabolismo , Sequência Consenso , Citosol/metabolismo , Imunofluorescência , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/genética , Genes Reporter/genética , Células HeLa , Proteínas Heterotriméricas de Ligação ao GTP/genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Proteínas do Tecido Nervoso/metabolismo , Proteínas Nucleares , Regiões Promotoras Genéticas/genética , Ligação Proteica , Subunidades Proteicas , Transporte Proteico , Proteínas Tirosina Fosfatases , Transativadores/antagonistas & inibidores , Transativadores/química , Transativadores/genética , Técnicas do Sistema de Duplo-HíbridoRESUMO
BACKGROUND: Patients with thin primary melanomas (< or = 1 mm) generally have an excellent prognosis. However, the presence of a vertical growth phase (VGP) adversely impacts the survival rate. We report on the rate of occurrence of nodal metastasis in patients with thin primary melanomas with a VGP who are offered sentinel lymph node (SLN) biopsy. METHODS: Among 235 patients with clinically localized cutaneous melanomas who underwent successful SLN biopsy, 71 had lesions 1 mm or smaller with a VGP. The SLN was localized by using blue dye and a radiotracer. If negative for tumor by using hematoxylin and eosin staining, the SLN was further examined by immunohistochemistry. RESULTS: The rate of occurrence of SLN metastasis was 15.2% in patients with melanomas deeper than 1 mm and 5.6% in patients with thin melanomas. Three patients with thin melanomas and a positive SLN had low-risk lesions, based on a highly accurate six-variable multivariate logistic regression model for predicting 8-year survival in stage I/II melanomas. The fourth patient had a low- to intermediate-risk lesion based on this model. At the time of the lymphadenectomy, one patient had two additional nodes with metastasis. CONCLUSIONS: VGP in a melanoma 1 mm or smaller seems to be a risk factor for nodal metastasis. The risk of nodal disease may not be accurately predicted by the use of a multivariate logistic regression model that incorporates thickness, mitotic rate, regression, tumor-infiltrating lymphocytes, sex, and anatomical site. Patients with thin lesions having VGP should be evaluated for SLN biopsy and trials of adjuvant therapy when stage III disease is found.
Assuntos
Metástase Linfática , Melanoma/patologia , Neoplasias Cutâneas/patologia , Adolescente , Adulto , Idoso , Biópsia , Feminino , Humanos , Modelos Logísticos , Excisão de Linfonodo , Linfonodos/diagnóstico por imagem , Linfonodos/patologia , Metástase Linfática/diagnóstico , Masculino , Melanoma/cirurgia , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Cintilografia , Neoplasias Cutâneas/cirurgiaRESUMO
Overexpression of wild-type p53 in cancer cells by adenovirus-mediated p53 gene transfer can result in the induction of apoptosis. To identify the potential mediators of this p53-induced apoptosis, we examined apoptotic protein levels in human lung cancer cells after Adp53 gene transfer. We observed up-regulation of Bax and Bak protein levels 18-36 h after transduction with Adp53 in H1299, H358, and H322 lung cancer cells. Contrary to expected observations, no changes in Bcl-2 and Bcl-X(L) protein levels were observed. Morphological cell changes and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining showed evidence of apoptosis in all cell lines 48 h after transduction with Adp53. These results indicate that the induction of apoptosis by adenovirus-mediated p53 transfer may be mediated by the induction of proapoptotic mechanisms rather than suppression of antiapoptotic mechanisms.
Assuntos
Técnicas de Transferência de Genes , Neoplasias Pulmonares/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteína Supressora de Tumor p53/genética , Adenoviridae/genética , Apoptose/genética , Western Blotting , DNA Recombinante/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/metabolismo , Regulação para Cima , Proteína Killer-Antagonista Homóloga a bcl-2 , Proteína X Associada a bcl-2 , Proteína bcl-XRESUMO
We introduced a functional p16 cDNA into non-small cell lung cancer (NSCLC) cell lines expressing different combinations of normal and mutated p16, p53, and Rb genes via a recombinant adenovirus to determine the effect of exogenous p16 expression on cell growth. Analysis of p16-deficient cells infected with Adv/p16 identified growth arrest of the cells in the G0 - G1 phase early on. Apoptosis was identified to occur by the 5th day after infection which corresponded with increased p16 expression, reduced Rb expression, and increased Rb hypophosphorylation, but only occurred in cells expressing functional p53. Further analysis indicated that the expression of the anti-apoptotic protein bcl-2 was greatly reduced in the NSCLC cell lines H460 and A549 (both -p16, +p53, +Rb), again only by the 5th day after Adv/p16 infection, but no affect on Bax expression was observed. H1299 cells (-p16, -p53, +Rb) infected with Adv/p16 only exhibited apoptosis by an additional infection with Adv/p53 which also corresponded with a down-regulation of bcl-2. In addition, the infection of A549 cells with Adv/p16 followed by a subsequent infection with Adv/Rb lead to a significant decrease in apoptosis which correlated with an increase in bcl-2 expression. These studies suggest that p16 is capable of mediating apoptosis in NSCLC cell lines expressing wild-type p53, through a direct down-regulation of Rb and an indirect down-regulation of the anti-apoptotic protein bcl-2.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Proteínas de Transporte/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Adenoviridae/genética , Apoptose , Western Blotting , Carcinoma Pulmonar de Células não Pequenas/virologia , Divisão Celular , Inibidor p16 de Quinase Dependente de Ciclina , Regulação para Baixo , Citometria de Fluxo , Humanos , Marcação In Situ das Extremidades Cortadas , Neoplasias Pulmonares/virologia , Proteína do Retinoblastoma/metabolismo , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/metabolismoRESUMO
Several lines of evidence seem to indicate that some neurocognitive measures could be phenotypic markers of predisposition to schizophrenia. The aim of this study was to investigate 21 patients with schizophrenia, 51 of their first-degree relatives and 46 nonpsychiatric controls, with a series of tests known to be sensitive to prefrontal cortical damage--the Trail Making Test, part B (TMT B), the Wisconsin Card Sorting Test (WCST) and a verbal fluency test (VFT)- and/or sensitive to temporo-hippocampic dysfunctions: verbal and visual memory and verbal learning tests from the Wechsler Memory Scale-Revised (Wechsler, 1987). Since parents and siblings share on average 50% of their genes with the schizophrenic proband, firstly we predicted that the first-degree relatives' performance would be at an intermediate level between patients and control subjects and secondly, we expected that a higher proportion of relatives than of control subjects would be impaired. The patients demonstrated deviant patterns of neuropsychological performance on the three tests sensitive to frontal dysfunctions and on most of the memory and learning tests. In the relative group, performance on the TMT B, VFT, immediate verbal recall and verbal learning was at an intermediate level between both other groups and significantly impaired compared to control subjects. However, the relative group did not differ from the control group on the WCST, immediate visual recall, and delayed verbal and visual recalls. Furthermore, compared to the control group, the percentages of patients and relatives who scored one standard deviation below the mean control group were significantly higher for the VFT and immediate verbal recall scores. Among all the tests studied, the verbal fluency and the immediate verbal recall appeared to be valuable phenotypic markers of schizophrenia since: (i) their mean scores were poorer in the patient and in the relative groups, (ii) the percentages of patients and relatives with poor performance were higher than the percentage of controls, (iii) these deficits were not due to poorer general intellectual abilities in the relative group, (iv) these deficits did not correlate with anxiety or depression scores.
Assuntos
Rememoração Mental , Reconhecimento Visual de Modelos , Esquizofrenia/genética , Psicologia do Esquizofrênico , Transtorno da Personalidade Esquizotípica/genética , Aprendizagem Verbal , Adulto , Feminino , Marcadores Genéticos/genética , Testes Genéticos , Humanos , Masculino , Memória de Curto Prazo , Pessoa de Meia-Idade , Testes Neuropsicológicos , Fenótipo , Retenção Psicológica , Esquizofrenia/diagnóstico , Transtorno da Personalidade Esquizotípica/diagnóstico , Transtorno da Personalidade Esquizotípica/psicologiaRESUMO
UNLABELLED: Sentinel lymph node (SLN) biopsy has emerged as a novel approach for identifying patients with melanoma and regional nodal micrometastasis who may benefit from full nodal basin resection. To identify the pattern of tumor lymphatic drainage and the SLN, lymphoscintigraphy has been performed using primarily 99mTc-sulfur colloid (SC). In this study, we compare the efficacy of SLN biopsy using 99mTc-human serum albumin (HSA) with SLN biopsy after SC-based lymphoscintigraphy. METHODS: One hundred and six patients with localized cutaneous melanoma were studied. Lymphoscintigraphy was performed after intradermal injection of HSA in 85 patients and SC in 21 patients. Four patients underwent lymphoscintigraphy twice, once with SC and once with HSA. Dynamic images were acquired for up to 1 h, followed by high-count images of the SLN in various projections so that the most likely site was marked on the skin for biopsy. Intraoperatively, blue dye was injected around the primary site. Twenty-four patients underwent SLN dissection directed by preoperative lymphoscintigraphy and vital blue dye mapping; in the remaining 80 patients, a gamma probe was added intraoperatively to the localization procedure. Two patients underwent mapping with gamma probe alone. RESULTS: Draining lymphatic basins and nodes were identified by lymphoscintigraphy in all patients. The SLN was identified in 95% of patients when both blue dye and intraoperative gamma probe were used. When 99mTc-HSA was used for imaging, 98% of the SLNs ultimately identified were radiolabeled, and 82% were both hot and blue. Of the SLN recovered with SC, all the nodes were radiolabeled; however, there was only 58% hot and blue concordance. Greater numbers of SLNs were removed in the SC group (median 2.0 versus 1.0, P = 0.02); however, the incidence of micrometastasis was statistically similar in both HSA and SC cohorts. In the 4 patients examined with both tracers, SLN mapping was similar. CONCLUSION: Although SC has been the radiotracer of choice for SLN mapping in melanoma, HSA appears to be a suitable alternative, with identical success rates. In fact, the higher concordance between hot and blue nodes using HSA suggests superiority of this tracer for this purpose.
Assuntos
Linfonodos/diagnóstico por imagem , Metástase Linfática/diagnóstico por imagem , Melanoma/patologia , Neoplasias Cutâneas/patologia , Agregado de Albumina Marcado com Tecnécio Tc 99m , Biópsia , Feminino , Humanos , Excisão de Linfonodo , Masculino , Melanoma/diagnóstico por imagem , Pessoa de Meia-Idade , Cintilografia , Compostos Radiofarmacêuticos , Corantes de Rosanilina , Neoplasias Cutâneas/diagnóstico por imagem , Coloide de Enxofre Marcado com Tecnécio Tc 99mRESUMO
During the post-natal period, skeletal muscles undergo important modifications leading to the appearance of different types of myofibers which exhibit distinct contractile and metabolic properties. This maturation process results from the activation of the expression of different sets of contractile proteins and metabolic enzymes, which are specific to the different types of myofibers. The muscle-specific promoter of the aldolase A gene (pM) is expressed mainly in fast-twitch glycolytic fibers in adult body muscles. We investigate here how pM is regulated during the post-natal development of different types of skeletal muscles (slow or fast-twitch muscles, head or body muscles). We show that pM is expressed preferentially in prospective fast-twitch muscles soon after birth; pM is up-regulated specifically in body muscles only later in development. This activation pattern is mimicked by a transgene which comprises only the 355 most proximal sequences of pM. Within this region, we identify a DNA element which is required for the up-regulation of the transgene during post-natal development in body muscles. Comparison of nuclear M1-binding proteins from young or adult body muscles show no qualitative differences. Distinct M1-binding proteins are present in both young and adult tongue nuclear extracts, compared to that present in gastrocnemius extracts.
Assuntos
Frutose-Bifosfato Aldolase/genética , Regulação da Expressão Gênica no Desenvolvimento , Músculo Esquelético/fisiologia , Regiões Promotoras Genéticas , Receptores Citoplasmáticos e Nucleares/metabolismo , Alitretinoína , Animais , Sítios de Ligação , Células Cultivadas , Galinhas , Camundongos , Camundongos Transgênicos , Desenvolvimento Muscular , Músculo Esquelético/embriologia , Músculo Esquelético/crescimento & desenvolvimento , Elementos de Resposta , Tretinoína/metabolismo , Regulação para CimaRESUMO
BACKGROUND: Sentinel lymph node (SLN) biopsy is being investigated as a staging procedure for breast carcinoma. The authors evaluated whether immunohistochemical (IHC) analysis improves the sensitivity of this procedure. METHODS: Forty-four women with breast carcinoma were recruited for SLN biopsy. Preoperative lymphoscintigraphy was followed by intraoperative localization using a handheld gamma probe and blue dye. After SLN identification, an immediate complete axillary lymph node dissection was performed in all patients. All lymph nodes were subjected to routine histology (hematoxylin and eosin [H&E]) and IHC using antibody to cytokeratins. RESULTS: The SLN was identified in 41 of 43 patients (95%). Successful SLN identification was independent of biopsy technique (open surgical [95%] vs. fine-needle aspiration/core needle biopsy [96%]). Twelve of 41 patients (29%) had evidence of lymph node metastasis in the SLN by routine histology. Of the twenty-nine patients with H&E negative SLN, 3 were found to have metastasis by IHC for a conversion rate of 10%. Fifteen of 41 patients (37%) had evidence of metastasis in SLN. All 26 patients with H&E and IHC negative SLN had negative nonsentinel lymph nodes by routine histology and IHC (100% negative predictive value). All patients with tumors < 2 cm and micrometastasis to the SLN had no additional lymph node disease, in contrast to patients with lesions > 2 cm or patients with macrometastasis to the SLN (P = 0.007). CONCLUSIONS: These results confirm that SLN biopsy is extremely accurate for patients with breast carcinoma, even after open surgical biopsy. IHC analysis or serial sectioning of SLN improves the sensitivity of this staging technique.