Génie biologique | EPFL Graph Search

vignette|Biologiste en pleine étude Le génie biologique désigne l'application des concepts et méthodes de la biologie (et accessoirement de la physique, chimie, mathématiques et informatique) afin de résoudre les problèmes relatifs aux sciences du vivant, en utilisant les méthodes analytiques et de synthèses propres à l'ingénierie ainsi que son expérience quant au coût et à la faisabilité d'une solution. Dans cette optique, alors que l'ingénierie traditionnelle applique les sciences physiques et mathématiques afin d'analyser, concevoir et fabriquer des outils, des structures et des processus non vivants, le génie biologique utilise principalement le domaine des connaissances de la biologie moléculaire afin d'étudier et de promouvoir son application aux organismes vivants. L'une de ses applications majeures est l'analyse et la résolution économique des problèmes liés à la santé humaine. Cependant, son champ d'application est encore bien plus étendu, on peut citer par exemple le biomimé

Field-based Strategies for Solid-Liquid Separation

Thierry Chappuis

The ever increasing demand for large quantities of bio-engineered pharmaceuticals and the tendency for doses to become larger stimulate the development of more productive and reliable mammalian cell culture technologies. In this context, perfusion bioreactors appear an efficient means to achieve high volumetric productivity and high product concentration. Since large-scale implementation of perfusion is limited by the reliability and scale-up potential of the device used to achieve cell retention, the present work proposes to tackle the problem bottom-up from the fundamentals of physics in order to develop, characterize and model novel particle retention strategies able to overcome the limitations of state-of-the-art systems. The first part of the present thesis focuses on the identification of physical force fields chosen for their ability to drive particles or cells into highly localized regions of a suspension. The ability of ultrasonic, dielectrophoretic and magnetophoretic interactions to make cells levitate over a surface was identified as an innovative way of implementing a Boycott effect (similar to that occurring in inclined sedimentation channels) while maintaining the flow channel in a vertical position. Therefore, inclined settlers were chosen as a reference system and the Ponder-Nakamura-Kuroda theory describing their performance was extended to take into account ultrasonic, dielectrophoretic and magnetic effects. The second part of the work provides deep modeling insight into the mechanisms underlying mammalian cell retention in these new filter implementations. Potentials and limitations are discussed in details. In particular, coupling dielectrophoresis with the Boycott effect proved to be successful in separating polystyrene particles from their moderately electrically conductive suspending medium. However, in highly conductive media, this process was shown to be very sensitive to order-disruptive electrokinetic effects such as ac electroosmosis and electrothermal fluid flow. Heat dissipation is however not specific to dielectrophoresis. This phenomenon is also a known limitation of the ultrasonic or magnetophoretic implementations, in certain conditions. The last part focused its work on the development of magnetophoretically-enhanced sedimentation, which was proposed as a means to partly overcome the heat dissipation problem observed with dielectrophoresis in highly conductive culture media. For such a magnetic approach to work correctly with intrinsically nonmagnetic materials, a paramagnetic suspending medium has to be used. The current implementation proposes to prepare media with fair quantities of gadolinium(III) or dysprosium(III) salts dissolved in them. Using such a paramagnetic buffer, the feasibility of magnetically retaining polystyrene particles could be verified in a vertical separation channel, hence providing first demonstration of the principle. The use of paramagnetic substances in mammalian cell culture media is naturally a matter of concern. The present work addresses this question as well and highlights the feasibility of growing Chinese Hamster Ovary cells in Gd(III)- or Dy(III)-rich environments. In contrast, attempts to develop low electrically conductive culture media failed and replacement of the NaCl content by D-mannitol or glycerol demonstrated severe growth inhibition effects.

EPFL2010

Bioengineering human bone marrow models to elucidate the triggers of the stem cell niche

Queralt Vallmajó Martín

The bone marrow (BM) microenvironment, or niche, significantly affects behaviors of its resident stem cell populations. Disruptions in the BM niche contribute to a number of severe clinical pathologies. Discovery of novel therapeutic strategies for BM-related diseases necessitates development of superior preclinical models to study parameters affecting BM homeostasis. To address this, a poly(ethylene glycol) hydrogel featuring a particular crosslinking chemistry based on the enzyme transglutaminase factor XIII, dubbed TG-PEG, was utilized to engineer humanized bone marrow models. Its modular nature was exploited throughout this work to systematically evaluate contributions of biophysical, cellular, and biochemical BM niche components to stem cell biology. Stiffness of TG-PEG hydrogels was optimized for each cell type and individual application explored. For bone formation by bone marrow stromal cells (BMSCs), an intermediate stiffness was optimal and significantly improved total bone volume over softer TG-PEG gels or natural materials. Results indicated that stability of the scaffold was more important than their chemical and biological properties. Alternatively, for hematopoietic stem and progenitor cells (HSPCs), softer gels were optimal for preserving multipotency while promoting cell proliferation. A number of BM-specific stem and progenitor cell types with richly diverse functions were explored. Skeletal stem cells (SSCs) encapsulated in TG-PEG hydrogels demonstrated their ability to form de novo bone in healing critically sized calvaria defects in mice. Next, subcutaneous implantation of BMSC-laden gels revealed their significant contribution to formation of ossicles. For some BMSC donors, ossicles could be formed in absence of bone morphogenetic protein (BMP)-2 offering a novel tool to evaluate their intrinsic capacity. Later, titration of BMSC seeding densities and BMP-2 doses yielded optimal conditions for robust BM formation, regardless of donor, enabling reproducibility for follow-on studies involving BM models. In humanized mice, homing of HSPCs to BMSC-laden TG-PEG gels was observed. Finally, humanized xenograft models were employed to determine vulnerability of the BM niche to subversion by cancer cells simulating osteotropism for both leukemia and breast cancer metastasis. Molecular factors known to trigger particular cell reactions within the niche were also investigated. Including RGD cell-adhesion ligands, as well as matrix metalloproteinase (MMP)-susceptible crosslinks to render the gels degradable, were critical for facilitating migration of recruited BM cells from murine hosts in xenograft studies. Hyaluronic acid (HA), when added to the hydrogel backbone, conferred superior engraftment to transplanted BMSCs and afforded reduced immunogenicity. Incorporation of the Notch-signaling ligands Jagged1 or DLL-4 into soft TG-PEG gels during in vitro HSPC expansion substantially stimulated proliferation, but recovered cells could no longer ensure long-term reconstitution. Aging, trauma, or cancer can cause disruptions of the tightly controlled balance between cells and their microenvironment in the BM niche. This body of work presents a number of bioengineered tools optimized for studying how niche components are affected by or contribute to such pathologies. Further work with these or similar models will no doubt highlight key clinically relevant targets and inform novel therapeutic approaches.

EPFL2019

Strategic Insights into Engineering Parameters Affecting Cell Type-Specific Uptake of DNA-Based Nanomaterials

Maartje Martina Cornelia Bastings, Alice Comberlato, Marianna Mainardi Koga, Hugo José Rodriguez Franco

DNA-based nanomaterials are gaining popularity as uniform and programmable bioengineering tools as a result of recent solutions to their weak stability under biological conditions. The DNA nanotechnology platform uniquely allows decoupling of engineering parameters to comprehensively study the effect of each upon cellular encounter. We here present a systematic analysis of the effect of surface parameters of DNA-based nanoparticles on uptake in three different cell models: tumor cells, macrophages, and dendritic cells. The influence of surface charge, stabilizing coating, fluorophore types, functionalization technique, and particle concentration employed is found to cause significant differences in material uptake among these cell types. We therefore provide new insights into the large variance in cell type-specific uptake, highlighting the necessity of proper engineering and careful assay development when DNA-based materials are used as tools in bioengineering and as future nanotherapeutic agents.