RESUMEN
Phenotypic heterogeneity is widely observed in cancer cell populations. Here, to probe this heterogeneity, we developed an image-guided genomics technique termed spatiotemporal genomic and cellular analysis (SaGA) that allows for precise selection and amplification of living and rare cells. SaGA was used on collectively invading 3D cancer cell packs to create purified leader and follower cell lines. The leader cell cultures are phenotypically stable and highly invasive in contrast to follower cultures, which show phenotypic plasticity over time and minimally invade in a sheet-like pattern. Genomic and molecular interrogation reveals an atypical VEGF-based vasculogenesis signalling that facilitates recruitment of follower cells but not for leader cell motility itself, which instead utilizes focal adhesion kinase-fibronectin signalling. While leader cells provide an escape mechanism for followers, follower cells in turn provide leaders with increased growth and survival. These data support a symbiotic model of collective invasion where phenotypically distinct cell types cooperate to promote their escape.
Asunto(s)
Movimiento Celular/genética , Heterogeneidad Genética , Genómica/métodos , Esferoides Celulares/metabolismo , Comunicación Celular/genética , Línea Celular Tumoral , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Invasividad Neoplásica , Neoplasias/irrigación sanguínea , Neoplasias/genética , Neoplasias/patología , Fenotipo , Esferoides Celulares/patología , Microambiente Tumoral/genética , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Upon activation, the epidermal growth factor (EGF) receptor becomes phosphorylated and triggers a vast signaling network that has profound effects on cell growth. The EGF receptor is observed to assemble into clusters after ligand binding and tyrosine kinase autophosphorylation, but the role of these assemblies in the receptor signaling pathway remains unclear. To address this question, we measured the phosphorylation of EGFR when the EGF ligand was anchored onto laterally mobile and immobile surfaces. We found that cells generated clusters of ligand-receptor complex on mobile EGF surfaces, and displayed a lower ratio of phosphorylated EGFR to EGF when compared to immobilized EGF that is unable to cluster. This result was verified by tuning the lateral assembly of ligand-receptor complexes on the surface of living cells using patterned supported lipid bilayers. Nanoscale metal lines fabricated into the supported membrane constrained lipid diffusion and EGF receptor assembly into micron and sub-micron scale corrals. Single cell analysis indicated that clustering impacts EGF receptor activation, and larger clusters (>1 µm(2)) of ligand-receptor complex generated lower EGF receptor phosphorylation per ligand than smaller assemblies (<1 µm(2)) in HCC1143 cells that were engaged to ligand-functionalized surfaces. We investigated the mechanism of EGFR clustering by treating cells with compounds that disrupt the cytoskeleton (Latrunculin B), clathrin-mediated endocytosis (Pitstop2), and inhibit EGFR activation (Gefitinib). These results help elucidate the nature of large-scale EGFR clustering, thus underscoring the general significance of receptor spatial organization in tuning biochemical function.
Asunto(s)
Materiales Biocompatibles/química , Neoplasias de la Mama/metabolismo , Factor de Crecimiento Epidérmico/química , Factor de Crecimiento Epidérmico/metabolismo , Receptores ErbB/metabolismo , Difusión , Femenino , Humanos , Fosforilación , Unión Proteica , Transducción de Señal , Propiedades de Superficie , Células Tumorales Cultivadas , Regulación hacia ArribaRESUMEN
A random sample of ten community-based Vita-Stat automatic blood pressure recorders (ABPR) was evaluated for accuracy and repeatability. Each of 518 adult volunteers had two randomly ordered systolic and diastolic Vita-Stat blood pressure measurements compared with two corresponding measurements obtained by a trained observer using a Random-Zero sphygmomanometer. Eight of the ten Vita-Stat recorders underestimated systolic blood pressure. In contrast, diastolic blood pressure was overestimated by nine of the ten machines. While the overall differences in average blood pressure were small (mean +/- SEM = -2.4 +/- 0.6 mm Hg for systolic blood pressure and +2.3 +/- 0.4 mm Hg for diastolic blood pressure), the magnitude of the average discrepancy varied considerably by machine (+4.7 to -13.8 mm Hg for systolic and +5.0 to -2.0 for diastolic blood pressure). At every level of systolic and diastolic blood pressure the Vita-Stat ABPR provided a less accurate method of classifying blood pressure among individuals than the human observer. Additional analyses exploring the ability of the Vita-Stat machine to measure an individual's blood pressure within 2, 4, or 6 mm Hg of the corresponding Random-Zero value again suggested that the Vita-Stat ABPR was less accurate than the human observer. Duplicate Vita-Stat readings were less repeatable than corresponding Random-Zero measurements. Based on the findings of this study, the Vita-Stat ABPR appears to be an unsatisfactory tool for community-based self-measurement of blood pressure.
Asunto(s)
Monitores de Presión Sanguínea/normas , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Envejecimiento/fisiología , Presión Sanguínea , Determinación de la Presión Sanguínea/instrumentación , Estudios de Evaluación como Asunto , Humanos , Hipertensión/prevención & control , Tamizaje Masivo , Persona de Mediana Edad , Análisis de Regresión , Reproducibilidad de los ResultadosRESUMEN
Hypercholesterolemia is an important determinant of progressive atherosclerosis in both vein grafts and native vessels after coronary artery bypass surgery. We demonstrated that nearly half of all patients admitted for coronary bypass have elevated cholesterol levels before surgery. Of these, half are unaware of the elevations in cholesterol level, and few are undergoing any treatment. The results suggest that lipids should be measured before coronary bypass surgery to optimize postoperative risk reduction education.