Polyphenols delivery by polymeric materials: challenges in cancer treatment

AbstractNanotechnology can offer different solutions for enhancing the therapeutic efficiency of polyphenols, a class of natural products widely explored for a potential applicability for the treatment of different diseases including cancer. While possessing interesting anticancer properties, polyphenols suffer from low stability and unfavorable pharmacokinetics, and thus suitable carriers are required when planning a therapeutic protocol. In the present review, an overview of the different strategies based on polymeric materials is presented, with the aim to highlight the strengths and the weaknesses of each approach and offer a platform of ideas for researchers working in the field

Stretching and heating cells with light-nonlinear photothermal cell rheology

Stretching and heating are everyday experiences for skin and tissue cells. They are also standard procedures to reduce the risk for injuries in physical exercise and to relieve muscle spasms in physiotherapy. Here, we ask which immediate and long-term mechanical effects of such treatments are quantitatively detectable on the level of individual living cells. Combining versatile optical stretcher techniques with a well-tested mathematical model for viscoelastic polymer networks, we investigate the thermomechanical properties of suspended cells with a photothermal rheometric protocol that can disentangle fast transient and slow 'inelastic' components in the nonlinear mechanical response. We find that a certain minimum strength and duration of combined stretching and heating is required to induce long-lived alterations of the mechanical state of the cells, which then respond qualitatively differently to mechanical tests than after weaker/shorter treatments or merely mechanical preconditioning alone. Our results suggest a viable protocol to search for intracellular biomolecular signatures of the mathematically detected dissimilar mechanical response modes

Investigating the effect of cell substrate on cancer cell stiffness by optical tweezers

The mechanical properties of cells are influenced by their microenvironment. Here we report cell stiffness alteration by changing the cell substrate stiffness for isolated cells and cells in contact with other cells. Polydimethylsiloxane (PDMS) is used to prepare soft substrates with three different stiffness values (173, 88 and 17 kPa respectively). Breast cancer cells lines, namely HBL-100, MCF-7 and MDA-MB-231 with different level of aggressiveness are cultured on these substrates and their local elasticity is investigated by vertical indentation of the cell membrane. Our preliminary results show an unforeseen behavior of the MDA-MB-231 cells. When cultured on glass substrate as isolated cells, they are less stiff than the other two types of cells, in agreement with the general statement that more aggressive and metastatic cells are softer. However, when connected to other cells the stiffness of MDA-MB-231 cells becomes similar to the other two cell lines. Moreover, the stiffness of MDA-MB-231 cells cultured on soft PDMS substrates is significantly higher than the stiffness of the other cell types, demonstrating thus the strong influence of the environmental conditions on the mechanical properties of the cells

Astrocytes-derived extracellular vesicles in motion at the neuron surface: Involvement of the prion protein

Astrocytes-derived extracellular vesicles (EVs) are key players in glia-neuron communication. However, whether EVs interact with neurons at preferential sites and how EVs reach these sites on neurons remains elusive. Using optical manipulation to study single EV-neuron dynamics, we here show that large EVs scan the neuron surface and use neuronal processes as highways to move extracellularly. Large EV motion on neurites is driven by the binding of EV to a surface receptor that slides on neuronal membrane, thanks to actin cytoskeleton rearrangements. The use of prion protein (PrP)-coated synthetic beads and PrP knock out EVs/neurons points at vesicular PrP and its receptor(s) on neurons in the control of EV motion. Surprisingly, a fraction of large EVs contains actin filaments and has an independent capacity to move in an actin-mediated way, through intermittent contacts with the plasma membrane. Our results unveil, for the first time, a dual mechanism exploited by astrocytic large EVs to passively/actively reach target sites on neurons moving on the neuron surface

PNIPAAm microgels with defined network architecture as temperature sensors in optical stretchers

Stretching individual living cells with light is a standard method to assess their mechanical properties. Yet, heat introduced by the laser light of optical stretchers may unwittingly change the mechanical properties of cells therein. To estimate the temperature induced by an optical trap, we introduce cell-sized, elastic poly(N-isopropylacrylamide) (PNIPAAm) microgels that relate temperature changes to hydrogel swelling. For their usage as a standardized calibration tool, we analyze the effect of free-radical chain-growth gelation (FCG) and polymer-analogous photogelation (PAG) on hydrogel network heterogeneity, micromechanics, and temperature response by Brillouin microscopy and optical diffraction tomography. Using a combination of tailor-made PNIPAAm macromers, PAG, and microfluidic processing, we obtain microgels with homogeneous network architecture. With that, we expand the capability of standardized microgels in calibrating and validating cell mechanics analysis, not only considering cell and microgel elasticity but also providing stimuli-responsiveness to consider dynamic changes that cells may undergo during characterization

Neuronal chemotaxis by optically manipulated liposomes

We probe chemotaxis of single neurons, induced by signalling molecules which were optically delivered from liposomes in the neighbourhood of the cells. We implemented an optical tweezers setup combined with a micro-dissection system on an inverted microscope platform. Molecules of Netrin-1 protein were encapsulated into micron-sized liposomes and manipulated to micrometric distances from a specific growth cone of a hippocampal neuron by the IR optical tweezers. The molecules were then released broken the liposomes with UV laser pulses. Chemotaxis induced by the delivered molecules was confirmed by the migration of the growth cone toward the liposome position. Since the delivery can be manipulated with high temporal and spatial resolution and the number of molecules released can be controlled quite precisely by tuning the liposome size and the solution concentration, this technique opens new opportunities to investigate the effect of physiological active compounds as Netrin-1 to neuronal signalling and guidance, which represents an important issue in neurobiology

Correlative all-optical quantification of mass density and mechanics of subcellular compartments with fluorescence specificity

Quantitative measurements of physical parameters become increasingly important for understanding biological processes. Brillouin microscopy (BM) has recently emerged as one technique providing the 3D distribution of viscoelastic properties inside biological samples − so far relying on the implicit assumption that refractive index (RI) and density can be neglected. Here, we present a novel method (FOB microscopy) combining BM with optical diffraction tomography and epifluorescence imaging for explicitly measuring the Brillouin shift, RI, and absolute density with specificity to fluorescently labeled structures. We show that neglecting the RI and density might lead to erroneous conclusions. Investigating the nucleoplasm of wild-type HeLa cells, we find that it has lower density but higher longitudinal modulus than the cytoplasm. Thus, the longitudinal modulus is not merely sensitive to the water content of the sample − a postulate vividly discussed in the field. We demonstrate the further utility of FOB on various biological systems including adipocytes and intracellular membraneless compartments. FOB microscopy can provide unexpected scientific discoveries and shed quantitative light on processes such as phase separation and transition inside living cells

Combined fluorescence, optical diffraction tomography and Brillouin microscopy

Quantitative measurements of physical parameters become increasingly important for understanding biological processes. Brillouin microscopy (BM) has recently emerged as one technique providing the 3D distribution of viscoelastic properties inside biological samples — so far relying on the implicit assumption that refractive index (RI) and density can be neglected. Here, we present a novel method (FOB microscopy) combining BM with optical diffraction tomography and epi-fluorescence imaging for explicitly measuring the Brillouin shift, RI and absolute density with molecular specificity. We show that neglecting the RI and density might lead to erroneous conclusions. Investigating the cell nucleus, we find that it has lower density but higher longitudinal modulus. Thus, the longitudinal modulus is not merely sensitive to the water content of the sample — a postulate vividly discussed in the field. We demonstrate the further utility of FOB on various biological systems including adipocytes and intracellular membraneless compartments. FOB microscopy can provide unexpected scientific discoveries and shed quantitative light on processes such as phase separation and transition inside living cells

Mechanical deformation induces depolarization of neutrophils

The transition of neutrophils from a resting state to a primed state is an essential requirement for their function as competent immune cells. This transition can be caused not only by chemical signals but also by mechanical perturbation. After cessation of either, these cells gradually revert to a quiescent state over 40 to 120 min. We use two biophysical tools, an optical stretcher and a novel microcirculation mimetic, to effect physiologically relevant mechanical deformations of single nonadherent human neutrophils. We establish quantitative morphological analysis and mechanical phenotyping as label-free markers of neutrophil priming. We show that continued mechanical deformation of primed cells can cause active depolarization, which occurs two orders of magnitude faster than by spontaneous depriming. This work provides a cellular-level mechanism that potentially explains recent clinical studies demonstrating the potential importance, and physiological role, of neutrophil depriming in vivo and the pathophysiological implications when this deactivation is impaired, especially in disorders such as acute lung injury.We acknowledge financial support by the Cambridge Commonwealth Trust (to A.E.E.), the European Research Council (Starting Grant “Light Touch” to J.G.), and the National Institute for Health Research Cambridge Biomedical Research Centre (to E.R.C.). C.S. is a Wellcome Trust Postdoctoral Clinical Research Fellow (101692MA), and C.F. is a Medical Research Council Clinical Training Fellow

Investigation into local cell mechanics by atomic force microscopy mapping and optical tweezer vertical indentation

Investigating the mechanical properties of cells could reveal a potential source of label-free markers of cancer progression, based on measurable viscoelastic parameters. The Young’s modulus has proved to be the most thoroughly studied so far, however, even for the same cell type, the elastic modulus reported in different studies spans a wide range of values, mainly due to the application of different experimental conditions. This complicates the reliable use of elasticity for the mechanical phenotyping of cells. Here we combine two complementary techniques, atomic force microscopy (AFM) and optical tweezer microscopy (OTM), providing a comprehensive mechanical comparison of three human breast cell lines: normal myoepithelial (HBL-100), luminal breast cancer (MCF-7) and basal breast cancer (MDA-MB-231) cells. The elastic modulus was measured locally by AFM and OTM on single cells, using similar indentation approaches but different measurement parameters. Peak force tapping AFM was employed at nanonewton forces and high loading rates to draw a viscoelastic map of each cell and the results indicated that the region on top of the nucleus provided the most meaningful results. OTM was employed at those locations at piconewton forces and low loading rates, to measure the elastic modulus in a real elastic regime and rule out the contribution of viscous forces typical of AFM. When measured by either AFM or OTM, the cell lines’ elasticity trend was similar for the aggressive MDA-MB-231 cells, which were found to be significantly softer than the other two cell types in both measurements. However, when comparing HBL-100 and MCF-7 cells, we found significant differences only when using OTM