(130d) Image-Based Modeling of Fibroblasts Modifying PDGF-BB Gradient Explains Cells’ Alternated Directional Decision during Chemotaxis in a Microfluidic Maze

Directed
fibroblast migration is central to highly proliferative processes in
regenerative medicine and developmental biology, such as wound healing and
embryogenesis. However, the mechanisms by which single fibroblasts affect each
other’s directional decisions, while chemotaxing in microscopic tissue pores,
are not well understood. Here we use image-based
multiscale modeling in order to explain an experimentally observed
contradiction to the classical chemotaxis theory:  fibroblasts
alternating their directional decisions in microscopic tissue-sized
confinement, instead of following the steepest chemotactic gradient. The
chemotaxis experiment was performed in a microfluidic maze (see Figure 1A),
which contains micron-sized paths (i.e. a long path and a short path) that the
cells preferred depending on the sequence in which they entered the maze. The
maze had two ports, one entrance and one exit. The entrance was connected to a
cell reservoir, while the exit was connected to a chemoattractant source of 50
ng/ml of PDGF-BB. NIH-3T3 fibroblasts were allowed to migrate along the PDGF-BB
gradient, towards the exit of the maze.  Real-time imaging of the migration was
performed using an automated Olympus IX83 microscope. Experimental data, such
as directional decisions of the cells, was manually extracted from time-lapse
images acquired at 15-min intervals. An image-based multiscale COMSOL model was
performed in order to visualize the gradient of PDGF-BB inside of the
microfluidic maze.  The macroscopic scale of the model simulated the global
chemoattractant gradient established in the maze, while the microscopic scale
was used to represent the localized gradient disturbances caused by the
presence of the cells.  In the former case, we used a transient two-dimensional
transport-of-diluted-species physics with two constant concentration boundary
conditions applied at the maze ports.  Moreover, the model took into account
depletion of the PDGF-BB via natural decay. On the microscopic level, on the
other hand, we modeled endocytosis of the PDGF-BB by the individual cells. The
simulation results showed that the reason why the fibroblasts alternate the
paths that they take through the maze can be explained by the localized PDGF-BB
consumption by the preceding cell.  Namely, the leading cell follows the
shortest path (i.e. the steepest gradient), as expected from classical
chemotaxis theory (see Figure 1B). The subsequent cell, however, sees a
gradient altered by the leading cell ahead of it (see Figure 1C), and
gets steered into the long path.  Overall, these finding supports the
conclusion that the presence of the individual cells in microconfinement can
modify the external chemotactic cues via self-generation of local gradients. 
Consequently, accounting for such effects could lead to a better understanding
of tissue generation in vivo, and result in more advanced engineered tissue
products in vitro.

Figure
1. Image-based
modeling of localized fibroblast effects on the PDGF-BB distribution within the
maze.  (A) Demonstration of the image-based concept.  Upper half corresponds to
the COMSOL simulation, with the PDGF-BB concentration scaled by the exit
boundary condition concentration; while the lower half is the phase microscopy
image of the fibroblasts (labeled in green) chemotaxing through the maze.  (B)
& (C) Simulation results of real-time modifications to the PDGF-BB gradient
overlaid on the experimental microscopy images, showing the first and second
cells’ directional decisions at the maze bifurcation, respectively.  Note that
although the gradient is scaled between 0 and 100%, the actual value
corresponding to the latter is chosen differently between the two timeframes in
order to enhance visibility.  Dashed white circle highlights the decision-making
cell, while white arrow points to the higher chemoattractant gradient path
chosen by the cells.