Microfluidic devices for the study of cell-cell, cell-particle and particle-particle interactions

Microfluidics and microtechnologies are of great interest for biological applications. This interest is linked to the fact that microtechnologies enable the study of single cells at the cellular and sub-cellular level. One of many applications of such single cell assays is cell-cell interaction probing. Current cell-cell interaction tools used by biologists rely on manual cell manipulation. But in the last decade new microfluidic technologies have arisen to tackle this biological challenge. Despite this growing need, the developped microfluidic devices still force biologists to compromise between the throughput of the assay (number of cell pairs interrogated) and the control over the interaction parameters (contact force and contact time). This thesis proposes an engineering point of view of microfluidic tools for single cell and in particular cell-cell interaction studies. The developed microfluidic devices enable an advancement of the microfluidic technologies to leverage new tools for cell-cell interaction interrogation in a controlled manner and at a high throughput. In this work we report the development of a novel microfluidic device based on a “roll-over” mechanism. This new chip design enables the multiplexing of cell-cell interactions. This multiplexing is achieved by forcing into contact all cells from a sample of a first cell population with all cells from a sample of a second cell population. Using such a device, we characterized the interaction of cells expressing olfactory receptors. This chip enables the maximization of cell-cell interaction interrogation and thus is ideally suited for rare and nonredundant cell populations. A droplet microfluidic chip for controlled and reliable cell co-encapsulation was designed and implemented. Common droplet microfluidic devices rely either on random cell co-encapsulation in droplets, thus leading to high cell loss, or use multiple microfluidic devices sequentially to perform droplet manipulation steps and achieve cell co-encapsulation. Both cases lead to cell sample loss, first from the random cell co-encapsulation process and second from the possible droplet loss between different microfluidic devices. Therefore, the implementation of a single fully integrated device enabling controlled cell co-encapsulation in droplets is of high interest for the study of precious, large cell libraries interactions for example in the case of cancer immunotherapy. In this thesis we also developed new concepts, using already existing microfluidic designs. By reusing the microfabrication technologies and microfluidic principles of the previously designed devices we could develop a novel microfluidic chip for another biological application. The third device studied in this thesis enables to look further into cell membrane receptors by isolating plasma membrane fragments. This tool enables an in-depth study of receptors involved in cell-cell interactions. It can also be used for any other cell membrane receptor study as for example G protein coupled receptor screening for therapeutic drug discovery. The tools developed in this thesis set the grounds for the development of new generations of cell-cell interaction microfluidic devices, enabling high throughput and control over the cell-cell interaction parameters. The technological advancements presented in this work were validated for various biological applications

Feedback-free microfluidic oscillator with impinging jets

The present paper describes a microfluidic oscillator based on facing impinging jets and operating in laminar flow conditions. Using appropriate microchannel configurations, pulsatile liquid flows are generated at the microscale from steady and equal inlet flow conditions and without moving parts or external stimuli. An experimental campaign has been carried out, using oscillator structures manufactured in silicon using conventional microfabrication techniques. This allowed us to study in detail the impact of the main geometric parameters of these structures on the oscillation frequency. The observed range of regular oscillations was found to depend on the geometry of the output channels, with highly regular oscillations occurring over a very large range of Reynolds numbers (Re) when an expansion of the output channel is added. The evolution of the self-oscillating frequency was shown to be dependent on the distance separating the impinging jets and on the average speed of the jets. Direct numerical simulations have been performed using a spectral element method. The computed dye concentration fields and nondimensional self-oscillation frequencies compare well with the experiments. The simulations enable a detailed characterization of the self-oscillation phenomenon in terms of pressure and velocity fields

Biotechnologies to tackle the challenge of neoantigen identification

Among immune correlates of clinical responses, tumor-specificneoantigens took the spotlight as relevant targets for cancerimmunotherapy. The implementation of pipelines forpersonalized cancer therapy remains challenging due to theprivacy, that is patient-specificity, of neoantigens and the low-frequency of neoantigen-specific T cells in blood and tumorsamples. To overcome these obstacles, recent developmentsin the field of biotechnology have allowed the multiplexedidentification of neoepitope-specific T cells. This reviewaddresses the pros and cons of conventional neoantigenscreening methodologies and highlights the current as well asthe prospective biotechnological opportunities in the field

Bioprinting the Cancer Microenvironment

Cancer is intrinsically complex, comprising both heterogeneous cellular compositions and microenvironmental cues. During the various stages of cancer initiation, development, and metastasis, cell-cell interactions (involving vascular and immune cells besides cancerous cells) as well as cell-extracellular matrix (ECM) interactions (e.g., alteration in stiffness and composition of the surrounding matrix) play major roles. Conventional cancer models both two- and three-dimensional (2D and 3D) present numerous limitations as they lack good vascularization and cannot mimic the complexity of tumors, thereby restricting their use as biomimetic models for applications such as drug screening and fundamental cancer biology studies. Bioprinting as an emerging biofabrication platform enables the creation of high-resolution 3D structures and has been extensively used in the past decade to model multiple organs and diseases. More recently, this versatile technique has further found its application in studying cancer genesis, growth, metastasis, and drug responses through creation of accurate models that recreate the complexity of the cancer microenvironment. In this review we will focus first on cancer biology and limitations with current cancer models. We then detail the current bioprinting strategies including the selection of bioinks for capturing the properties of the tumor matrices, after which we discuss bioprinting of vascular structures that are critical toward construction of complex 3D cancer organoids. We finally conclude with current literature on bioprinted cancer models and propose future perspectives

Global biosphere primary productivity changes during the past eight glacial cycles

Global biosphere productivity is the largest uptake flux of atmospheric carbon dioxide (CO2), and it plays an important role in past and future carbon cycles. However, global estimation of biosphere productivity remains a challenge. Using the ancient air enclosed in polar ice cores, we present the first 800,000-year record of triple isotopic ratios of atmospheric oxygen, which reflects past global biosphere productivity. We observe that global biosphere productivity in the past eight glacial intervals was lower than that in the preindustrial era and that, in most cases, it starts to increase millennia before deglaciations. Both variations occur concomitantly with CO2 changes, implying a dominant control of CO2 on global biosphere productivity that supports a pervasive negative feedback under the glacial climate

Triple isotopic composition of atmospheric oxygen (Δ17O of O2) over 58.0-150.0, 233.2-238.1, and 445.6-796.3 ka from EPICA Dome C ice core

Here we present the triple isotopic composition of atmospheric oxygen (Δ17O of O2) measured from EPICA Dome C (EDC) ice core, Antarctica, over 58.0-150.0, 233.2-238.1, and 445.6-796.3 ka, in order to reconstruct the past global biosphere productivity (Luz et al., 1999). The data were produced between Feb. 2019 and Oct. 2020 at Laboratoire des Sciences du Climat et de l'Environnement (LSCE), Gif-sur-Yvette, France. The air samples trapped in EDC ice cores were extracted and purified by a semi-automated system of LSCE, following the identical principles of Barkan and Luz (2005)

Global biosphere primary productivity changes during the past eight glacial cycles

International audienceGlobal biosphere productivity is the largest uptake flux of atmospheric carbon dioxide (CO$_2$), and it plays an important role in past and future carbon cycles. However, global estimation of biosphere productivity remains a challenge. Using the ancient air enclosed in polar ice cores, we present the first 800,000-year record of triple isotopic ratios of atmospheric oxygen, which reflects past global biosphere productivity. We observe that global biosphere productivity in the past eight glacial intervals was lower than that in the preindustrial era and that, in most cases, it starts to increase millennia before deglaciations. Both variations occur concomitantly with CO$_2$ changes, implying a dominant control of CO$_2$ on global biosphere productivity that supports a pervasive negative feedback under the glacial climate

Microfluidic device performing on flow study of serial cell–cell interactions of two cell populations

In this study we present a novel microfluidic hydrodynamic trapping device to probe the cell-cell interaction between all cell samples of two distinct populations. We have exploited an hydrodynamic trapping method using microfluidics to immobilize a batch of cells from the first population at specific locations, then relied on hydrodynamic filtering principles, the flowing cells from the second cell population are placed in contact with the trapped ones, through a roll-over mechanism. The rolling cells interact with the serially trapped cells one after the other. The proposed microfluidic phenomenon was characterized with beads. We have shown the validity of our method by detecting the capacity of olfactory receptors to induce adhesion of cell doublets overexpressing these receptors. We report here the first controlled on-flow single cell resolution cell-cell interaction assay in a microfluidic device for future application in cell-cell interactions-based cell library screenings

Toward Microfluidic Label-Free Isolation and Enumeration of Circulating Tumor Cells from Blood Samples

The isolation, analysis, and enumeration of circulating tumor cells (CTCs) from cancer patient blood samples are a paradigm shift for cancer patient diagnosis, prognosis, and treatment monitoring. Most methods used to isolate and enumerate these target cells rely on the expression of cell surface markers, which varies between patients, cancer types, tumors, and stages. Here, we propose a label-free high-throughput platform to isolate, enumerate, and size CTCs on two coupled microfluidic devices. Cancer cells were purified through a Vortex chip and subsequently flowed in-line to an impedance chip, where a pair of electrodes measured fluctuations of an applied electric field generated by cells passing through. A proof-of-concept of the coupling of those two devices was demonstrated with beads and cells. First, the impedance chip was tested as a stand-alone device: (1) with beads (mean counting error of 1.0%, sizing information clearly separated three clusters for 8, 15, and 20 um beads, respectively) as well as (2) with cancer cells (mean counting error of 3.5%). Second, the combined setup was tested with beads, then with cells in phosphate-buffered saline, and finally with cancer cells spiked in healthy blood. Experiments demonstrated that the Vortex HT chip enriched the cancer cells, which then could be counted and differentiated from smaller blood cells by the impedance chip based on size information. Further discrimination was shown with dual high-frequency measurements using electric opacity, highlighting the potential application of this combined setup for a fully integrated label-free isolation and enumeration of CTCs from cancer patient samples. (c) 2019 International Society for Advancement of Cytometr