Nano-bio lab course: microvasculature scaffolds

Editor’s note: Over the next several days, we will share the student impressions of some of the techniques learned in INBT’s nano-bio laboratory course (670.621). These reports demonstrate the wide variety of techniques students trained at the Johns Hopkins Institute for NanoBioTechnology are expected to understand. Each technique is taught in a different affiliated faculty lab. More lab techniques to come.

Tiny scaffolds build microvessels

In the Sharon Gerecht lab led by her PhD candidate Sebastian Barreto, we learned about the protocol they use for constructing microvasculature using a 250-micron diameter fibrin fiber scaffold. The scaffold has endothelial colony forming cells seeded on the surface that rapidly proliferate and excrete ESM to form a cell-based structure around the fibrin fiber scaffold.

Lab6 Figure DRW

A) Confocal microscopy image of collagen IV from endothelial colony forming cells seeded on fibrin fibers of various diameters. (B) Scatter plot of angle of collagen IV deposition showing significance between the 445 um fiber and 105-370 um fibers. The significance of the finding is that the endothelial cells are unable to sense the curvature of the fiber above 445 um.

The addition of plasmin to the media allows the fibrin scaffold to be rapidly degraded leaving the cell construct in the shape of a micro-vessel, after which media can be perfused through the vessel with a pulsatile pump to simulate heart powered blood flow and stimulate the cells within the construct. The method of creating the cell construct also allows for the seeding of perivascular cells on the surface of the endothelial cells to study the interaction between the two cell types.

Confocal microscopy following fixing and staining of the cell construct allows the orientation of the ECM and cell proliferation to be analyzed. As a tool for studying endothelial blood vessel formation this platform technology is very interesting. Of particular interest is the fact that above a certain diameter of the fibrin fiber the endothelial cells do not rearrange, indicating that they have a mechanism for sensing the effective radius of curvature of the vessel.

Reference: Barreto-Ortiz SF, Zhang S, Davenport M, Fradkin J, Ginn B, et al. (2013) A Novel In Vitro Model for Microvasculature Reveals Regulation of Circumferential ECM Organization by Curvature. PLoS ONE 8(11): e81061. doi:10.1371/journal.pone.0081061

About the author: David Wilson is a first year PhD student in biomedical engineering working in the drug delivery laboratory of Jordan Green.

For all press inquiries regarding INBT, its faculty and programs, contact Mary Spiro, or 410-516-4802.

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