by Keyword: Heterogeneity

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Mas, S., Torro, A., Fernández, L., Bec, N., Gongora, C., Larroque, C., Martineau, P., de Juan, A., Marco, S., (2020). MALDI imaging mass spectrometry and chemometric tools to discriminate highly similar colorectal cancer tissues Talanta 208, 120455

Intratumour heterogeneity due to cancer cell clonal evolution and microenvironment composition and tumor differences due to genetic variations between patients suffering of the same cancer pathology play a crucial role in patient response to therapies. This study is oriented to show that matrix-assisted laser-desorption ionization-Mass spectrometry imaging (MALDI-MSI), combined with an advanced multivariate data processing pipeline can be used to discriminate subtle variations between highly similar colorectal tumors. To this aim, experimental tumors reproducing the emergence of drug-resistant clones were generated in athymic mice using subcutaneous injection of different mixes of two isogenic cell lines, the irinotecan-resistant HCT116-SN50 (R) and its sibling human colon adenocarcinoma sensitive cell line HCT116 (S). Because irinotecan-resistant and irinotecan-sensitive are derived from the same original parental HCT116 cell line, their genetic characteristics and molecular compositions are closely related. The multivariate data processing pipeline proposed relies on three steps: (a) multiset multivariate curve resolution (MCR) to separate biological contributions from background; (b) multiset K-means segmentation using MCR scores of the biological contributions to separate between tumor and necrotic parts of the tissues; and (c) partial-least squares discriminant analysis (PLS-DA) applied to tumor pixel spectra to discriminate between R and S tumor populations. High levels of correct classification rates (0.85), sensitivity (0.92) and specificity (0.77) for the PLS-DA classification model were obtained. If previously labelled tissue is available, the multistep modeling strategy proposed constitutes a good approach for the identification and characterization of highly similar phenotypic tumor subpopulations that could be potentially applicable to any kind of cancer tissue that exhibits substantial heterogeneity. © 2019 Elsevier B.V.

Keywords: Chemometrics, Colorectal cancer, MALDI imaging, Multivariate analysis, Tumor heterogeneity

Feiner-Gracia, Natalia, Beck, Michaela, Pujals, Sílvia, Tosi, Sébastien, Mandal, Tamoghna, Buske, Christian, Linden, Mika, Albertazzi, Lorenzo, (2017). Super-resolution microscopy unveils dynamic heterogeneities in nanoparticle protein corona Small 13, (41), 1701631

The adsorption of serum proteins, leading to the formation of a biomolecular corona, is a key determinant of the biological identity of nanoparticles in vivo. Therefore, gaining knowledge on the formation, composition, and temporal evolution of the corona is of utmost importance for the development of nanoparticle-based therapies. Here, it is shown that the use of super-resolution optical microscopy enables the imaging of the protein corona on mesoporous silica nanoparticles with single protein sensitivity. Particle-by-particle quantification reveals a significant heterogeneity in protein absorption under native conditions. Moreover, the diversity of the corona evolves over time depending on the surface chemistry and degradability of the particles. This paper investigates the consequences of protein adsorption for specific cell targeting by antibody-functionalized nanoparticles providing a detailed understanding of corona-activity relations. The methodology is widely applicable to a variety of nanostructures and complements the existing ensemble approaches for protein corona study.

Keywords: Heterogeneity, Mesoporous silica nanoparticles, Protein corona, Super-resolution imaging, Targeting

Bueno, M., Paganetti, H., Duch, M. A., Schuemann, J., (2013). An algorithm to assess the need for clinical Monte Carlo dose calculation for small proton therapy fields based on quantification of tissue heterogeneity Medical Physics , 40, (8), 081704

Purpose: In proton therapy, complex density heterogeneities within the beam path constitute a challenge to dose calculation algorithms. This might question the reliability of dose distributions predicted by treatment planning systems based on analytical dose calculation. For cases in which substantial dose errors are expected, resorting to Monte Carlo dose calculations might be essential to ensure a successful treatment outcome and therefore the benefit is worth a presumably long computation time. The aim of this study was to define an indicator for the accuracy of dose delivery based on analytical dose calculations in treatment planning systems for small proton therapy fields to identify those patients for which Monte Carlo dose calculation is warranted. Methods: Fourteen patients treated at our facility with small passively scattered proton beams (apertures diameters below 7 cm) were selected. Plans were generated in the XiO treatment planning system in combination with a pencil beam algorithm developed at the Massachusetts General Hospital and compared to Monte Carlo dose calculations. Differences in the dose to the 50% of the gross tumor volume (D50, GTV) were assessed in a field-by-field basis. A simple and fast methodology was developed to quantify the inhomogeneity of the tissue traversed by a single small proton beam using a heterogeneity index (HI) - a concept presented by Plugfelder [Med. Phys. 34, 1506-1513 (2007)10.1118/1. 2710329] for scanned proton beams. Finally, the potential correlation between the error made by the pencil beam based treatment planning algorithm for each field and the level of tissue heterogeneity traversed by the proton beam given by the HI was evaluated. Results: Discrepancies up to 5.4% were found in D50 for single fields, although dose differences were within clinical tolerance levels (<3%) when combining all of the fields involved in the treatment. The discrepancies found for each field exhibited a strong correlation to their associated HI-values (Spearman's ρ = 0.8, p < 0.0001); the higher the level of tissue inhomogeneities for a particular field, the larger the error by the analytical algorithm. With the established correlation a threshold for HI can be set by choosing a tolerance level of 2-3% - commonly accepted in radiotherapy. Conclusions: The HI is a good indicator for the accuracy of proton field delivery in terms of GTV prescription dose coverage when small fields are delivered. Each HI-value was obtained from the CT image in less than 3 min on a computer with 2 GHz CPU allowing implementation of this methodology in clinical routine. For HI-values exceeding the threshold, either a change in beam direction (if feasible) or a recalculation of the dose with Monte Carlo would be highly recommended.

Keywords: Heterogeneities, Heterogeneity index, Monte Carlo, Proton therapy, Small fields

van Zanten, T. S., Gomez, J., Manzo, C., Cambi, A., Buceta, J., Reigada, R., Garcia-Parajo, M. F., (2010). Direct mapping of nanoscale compositional connectivity on intact cell membranes Proceedings of the National Academy of Sciences of the United States of America 107, (35), 15437-15442

Lateral segregation of cell membranes is accepted as a primary mechanism for cells to regulate a diversity of cellular functions. In this context, lipid rafts have been conceptualized as organizing principle of biological membranes where underlying cholesterol-mediated selective connectivity must exist even at the resting state. However, such a level of nanoscale compositional connectivity has been challenging to prove. Here we used single-molecule near-field scanning optical microscopy to visualize the nanolandscape of raft ganglioside GM1 after tightening by its ligand cholera toxin (CTxB) on intact cell membranes. We show that CTxB tightening of GM1 is sufficient to initiate a minimal raft coalescence unit, resulting in the formation of cholesterol-dependent GM1 nanodomains <120 nm in size. This particular arrangement appeared independent of cell type and GM1 expression level on the membrane. Simultaneous dual color high-resolution images revealed that GPI anchored and certain transmembrane proteins were recruited to regions proximal (<150 nm) to CTxB-GM1 nanodomains without physical intermixing. Together with in silico experiments, our high-resolution data conclusively demonstrate the existence of raft-based interconnectivity at the nanoscale. Such a linked state on resting cell membranes constitutes thus an obligatory step toward the hierarchical evolution of large-scale raft coalescence upon cell activation.

Keywords: Cholera toxin, Membrane heterogeneity, Near-field scanning optical microscopy, Raft ganglioside GM1, Single-molecule detection

Park, C. Y., Tambe, D., Alencar, A. M., Trepat, X., Zhou, E. H., Millet, E., Butler, J. P., Fredberg, J. J., (2010). Mapping the cytoskeletal prestress The American Journal of Physiology - Cell Physiology , 298, (5), C1245-C1252

Park CY, Tambe D, Alencar AM, Trepat X, Zhou EH, Millet E, Butler JP, Fredberg JJ. Mapping the cytoskeletal prestress. Am J Physiol Cell Physiol 298: C1245-C1252, 2010. First published February 17, 2010; doi: 10.1152/ajpcell.00417.2009.-Cell mechanical properties on a whole cell basis have been widely studied, whereas local intracellular variations have been less well characterized and are poorly understood. To fill this gap, here we provide detailed intracellular maps of regional cytoskeleton (CSK) stiffness, loss tangent, and rate of structural rearrangements, as well as their relationships to the underlying regional F-actin density and the local cytoskeletal prestress. In the human airway smooth muscle cell, we used micropatterning to minimize geometric variation. We measured the local cell stiffness and loss tangent with optical magnetic twisting cytometry and the local rate of CSK remodeling with spontaneous displacements of a CSK-bound bead. We also measured traction distributions with traction microscopy and cell geometry with atomic force microscopy. On the basis of these experimental observations, we used finite element methods to map for the first time the regional distribution of intracellular prestress. Compared with the cell center or edges, cell corners were systematically stiffer and more fluidlike and supported higher traction forces, and at the same time had slower remodeling dynamics. Local remodeling dynamics had a close inverse relationship with local cell stiffness. The principal finding, however, is that systematic regional variations of CSK stiffness correlated only poorly with regional F-actin density but strongly and linearly with the regional prestress. Taken together, these findings in the intact cell comprise the most comprehensive characterization to date of regional variations of cytoskeletal mechanical properties and their determinants.

Keywords: Cell mechanics, Stiffness, Remodeling, Heterogeneity