Annals of Biomedical Engineering

, Volume 39, Issue 1, pp 205–222 | Cite as

Three-Dimensional Quantitative Micromorphology of Pre- and Post-Implanted Engineered Heart Valve Tissues

  • Chad E. Eckert
  • Brandon T. Mikulis
  • Danielle Gottlieb
  • Dane Gerneke
  • Ian LeGrice
  • Robert F. Padera
  • John E. Mayer
  • Frederick J. Schoen
  • Michael S. Sacks


There is a significant gap in our knowledge of engineered heart valve tissue (EHVT) development regarding detailed three-dimensional (3D) tissue formation and remodeling from the point of in vitro culturing to full in vivo function. As a step toward understanding the complexities of EHVT formation and remodeling, a novel serial confocal microscopy technique was employed to obtain 3D microstructural information of pre-implant (PRI) and post-implant for 12 weeks (POI) EHVT fabricated from PGA:PLLA scaffolds and seeded with ovine bone-marrow-derived mesenchymal stem cells. Custom scaffold fiber tracking software was developed to quantify scaffold fiber architectural features such as length, tortuosity, and minimum scaffold fiber–fiber separation distance and scaffold fiber orientation was quantified utilizing a 3D fabric tensor. In addition, collagen and cellular density of ovine pulmonary valve leaflet tissue were also analyzed for baseline comparisons. Results indicated that in the unseeded state, scaffold fibers formed a continuous, oriented network. In the PRI state, the scaffold showed some fragmentation with a scaffold volume fraction of 7.79%. In the POI specimen, the scaffold became highly fragmented, forming a randomly distributed short fibrous network (volume fraction of 2.03%) within a contiguous, dense collagenous matrix. Both PGA and PLLA scaffold fibers were observed in the PRI and POI specimens. Collagen density remained similar in both PRI and POI specimens (74.2 and 71.5%, respectively), though the distributions in the transmural direction appeared slightly more homogenous in the POI specimen. Finally, to guide future 2D histological studies for large-scale studies (since acquisition of high-resolution volumetric data is not practical for all specimens), we investigated changes in relevant collagen and scaffold metrics (collagen density and scaffold fiber orientation) with varying section spacing. It was found that a sectioning spacing up to 25 μm (for scaffold morphology) and 50 μm (for collagen density) in both PRI and POI tissues did not result in loss of information fidelity, and that sectioning in the circumferential or radial direction provides the greatest preservation of information. This is the first known work to investigate EHVT microstructure over a large volume with high resolution and to investigate time evolving in vivo EHVT morphology. The important scaffold fiber structural changes observed provide morphological information crucial for guiding future structurally based constitutive modeling efforts focused on better understanding EHVT tissue formation and remodeling.


Engineered heart valve tissue Heart valves Tissue engineering Structural mechanical modeling Quantitative morphology 



The authors would like to thank Dr. Bruce Smaill for his invaluable input and assistance in utilizing the extended-volume scanning laser confocal microscope and Helen Shing, M.S., for her expert technical assistance with histology. This work was supported by the National Science Foundation’s East Asia and Pacific Island Summer Institute research program (CEE and BTM) and National Institute of Health grants R01 HL-068816 (MSS) and R01 HL-089750 (MSS).


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Copyright information

© Biomedical Engineering Society 2010

Authors and Affiliations

  • Chad E. Eckert
    • 1
  • Brandon T. Mikulis
    • 1
  • Danielle Gottlieb
    • 2
  • Dane Gerneke
    • 3
  • Ian LeGrice
    • 3
  • Robert F. Padera
    • 4
  • John E. Mayer
    • 2
  • Frederick J. Schoen
    • 4
  • Michael S. Sacks
    • 1
  1. 1.Department of Bioengineering, Swanson School of EngineeringMcGowan Institute for Regenerative Medicine, University of PittsburghPittsburghUSA
  2. 2.Department of Cardiology and Cardiovascular SurgeryChildren’s Hospital BostonBostonUSA
  3. 3.Auckland Bioengineering InstituteUniversity of AucklandAucklandNew Zealand
  4. 4.Department of PathologyBrigham and Women’s Hospital and Harvard Medical SchoolBostonUSA

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