Engineering chemoattractant gradients using controlled release polysaccharide microspheres
Author(s)Wang, Yana, Ph. D. Massachusetts Institute of Technology
Massachusetts Institute of Technology. Dept. of Chemical Engineering.
Darrell J. Irvine.
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Chemoattractant gradients play important roles in the normal function of immune system, from lymphocyte homeostasis to mounting efficient immune responses against infection. Improved fundamental knowledge about the role of chemoattractant gradients developed around single source cells in controlling chemotaxis of "receiving" cells would not only greatly advance our understanding of the basic mechanisms of cell chemotaxis but also would inform strategies for modulating chemoattractant gradients in therapeutic applications, such as adjuvant materials for vaccines and cancer immunotherapy recruiting immune cells of interest. In this thesis, we first applied mathematical modeling to understand the key characteristics of chemoattractant gradients secreted from single source cells at physiological rates. During the transport of chemoattractants, we considered the diffusion of soluble attractants, binding to matrix and degradation by proteolytic enzymes. From the calculated chemoattractant concentration gradients, we predicted the characteristics of attractant receptor engagement on responding cells, and estimated the maximum stimulation distance effectively triggering chemotaxis of responding cells based on the threshold for receptor engagement gradients, a difference of ~10 ligand-engaged receptors between the front and back of responding cells. This characteristic maximum stimulation distance is a function of multiple parameters including secretion rate of the source cell, diffusion constant of the chemoattractant, interaction with matrix, degradation or clearance of chemoattractant in the tissues, and the density of source cells. In addition, chemokine receptor desensitization induced by chemoattractants could shorten the maximum stimulation distance. We then developed Artificial Secreting Cells (ASCs) to mimic real chemoattractant secreting cells using cell-sized polysaccharide-based hydrogel microspheres releasing chemoattractant in a controlled manner. These alginate hydrogel microspheres, ~30 [mu]m in size, were crosslinked with Ca2+ between gluronic acid units on alginate backbones and provided a natural and bioactive environment for chemokines. The chemokines could be loaded into these alginate microspheres by soaking them in concentrated chemokine solutions and released in a reversible manner. This approach was shown as a general strategy for several chemokines, such as CCL21, CCL19, CXCL10 and CXCL12. The loading and release properties of individual chemokines were highly correlated with the average charge density on protein surface. We have also demonstrated that the controlled gradients created by ASCs were similar to the modeled gradients developed around single source cells. Further we used 3D collagen hydrogels embedded with ASCs as an in vitro model to investigate single human T-cell and dendritic cell migration dynamics to CCL21 and CCL19 chemokine gradients. Individual T-cells exhibited a binary response to isolated attractant sources, migrating highly directionally or ignoring the gradient completely; the fraction of responding cells correlated with chemokine receptor occupancy induced by the gradient. In sustained gradients eliciting low receptor desensitization, attracted T-cells or dendritic cells swarmed around isolated ASCs for hours. With increasing ASC density, overlapping gradients and high attractant concentrations caused a transition from local swarming to transient "hopping" of cells bead to bead. Thus, diverse migration responses observed in vivo may be determined by chemoattractant source density and secretion rate, which govern receptor occupancy patterns in nearby cells.
Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemical Engineering, February 2012.Cataloged from PDF version of thesis.Includes bibliographical references (p. 111-122).
DepartmentMassachusetts Institute of Technology. Dept. of Chemical Engineering.; Massachusetts Institute of Technology. Department of Chemical Engineering
Massachusetts Institute of Technology