Molecular Engineering of Extrinsic and Intrinsic Cues to Control Stem Cell Function

David Schaffer, University of California, Berkeley

Stem cell microenvironments present complex repertoires of signals to regulate the processes self-renewal and differentiation.  There has been considerable progress in studying soluble signals that regulate stem cell function, but comparatively less work has focused on investigating the “solid phase” of the microenvironment, in large part due to experimental complexities in investigating and mimicking the complexity of the extracellular matrix (ECM), cell-cell interactions, and other components.  Recent work demonstrates that bioactive, synthetic materials can be harnessed to emulate and thereby study the effects of solid phase, biophysical cues on cell function.  By using a modular, bioactive material, we have found that that the matrix modulus or stiffness profoundly impacts neural stem cell self-renewal and differentiation, and mechanistic analysis implicates key mechanotransductive pathways in this process that are important in cell culture and in vivo.  Furthermore, immobilization of biochemical signals to the solid phase of a natural niche can lead to nanoscale organization of these signals, and nanostructured biological-polymeric conjugates likewise serve as potent effectors of neural stem and human embryonic stem cell function.  Finally, the combinatorial presentation of different matrix motifs from a material can generate synthetic systems capable of supporting the self-renewal and differentiation of both neural stem cells and human embryonic stem cells, thereby enabling the dissection or distillation of the ECM into key individual signals necessary to support stem cell function.  Biomimetic materials can thus be employed to study mechanisms by which the solid phase of a stem cell microenvironment regulates cell function, as well as offer safe, scaleable, and robust systems to control stem cells for biomedical application.

In addition to engineering the microenvironment, one can control a cell’s behavior by editing its genome.  Gene delivery vehicles based on viruses offer a number of advantageous properties, including the potential for safe and efficient gene delivery; however, they face a number of challenges including inefficient delivery to some therapeutically relevant cells such as stem cells.  Such shortcomings arguably arise from the fact that viruses did not naturally evolve to be utilized as human therapeutics, and we have thus been developing directed evolution – the iterative generation of large libraries genetic mutants and selection for enhanced properties – as an approach to create new viruses with useful properties.  For example, we have evolved adeno-associated virus for enhanced gene delivery to and gene targeting in neural stem cells and human embryonic stem cells.  Engineering both extrinsic and intrinsic cues thus provides strong capabilities to study natural mechanisms of stem cell fate regulation, as well as to control these choices for therapeutic applications.