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Research Papers

# Microstructure and Mechanical Property of Glutaraldehyde-Treated Porcine Pulmonary Ligament

[+] Author and Article Information
Huan Chen, Xuefeng Zhao

California Medical Innovations Institute,
San Diego, CA 92121

Zachary C. Berwick

3DT Holdings, LLC,
San Diego, CA 92121

Joshua F. Krieger, Sean Chambers

Cook Medical, Inc.,
Bloomington, IN 47402

Ghassan S. Kassab

California Medical Innovations Institute,
San Diego, CA 92121
e-mail: gkassab@calmi2.org

1H. Chen and X. Zhao contributed equally to this work.

2Corresponding author.

Manuscript received June 22, 2015; final manuscript received March 21, 2016; published online April 27, 2016. Assoc. Editor: Hai-Chao Han.

J Biomech Eng 138(6), 061003 (Apr 27, 2016) (9 pages) Paper No: BIO-15-1308; doi: 10.1115/1.4033300 History: Received June 22, 2015; Revised March 21, 2016

## Abstract

There is a significant need for fixed biological tissues with desired structural and material constituents for tissue engineering applications. Here, we introduce the lung ligament as a fixed biological material that may have clinical utility for tissue engineering. To characterize the lung tissue for potential clinical applications, we studied glutaraldehyde-treated porcine pulmonary ligament (n = 11) with multiphoton microscopy (MPM) and conducted biaxial planar experiments to characterize the mechanical property of the tissue. The MPM imaging revealed that there are generally two families of collagen fibers distributed in two distinct layers: The first family largely aligns along the longitudinal direction with a mean angle of θ = 10.7 ± 9.3 deg, while the second one exhibits a random distribution with a mean θ = 36.6 ± 27.4. Elastin fibers appear in some intermediate sublayers with a random orientation distribution with a mean θ = 39.6 ± 23 deg. Based on the microstructural observation, a microstructure-based constitutive law was proposed to model the elastic property of the tissue. The material parameters were identified by fitting the model to the biaxial stress–strain data of specimens, and good fitting quality was achieved. The parameter $e0$ (which denotes the strain beyond which the collagen can withstand tension) of glutaraldehyde-treated tissues demonstrated low variability implying a relatively consistent collagen undulation in different samples, while the stiffness parameters for elastin and collagen fibers showed relatively greater variability. The fixed tissues presented a smaller $e0$ than that of fresh specimen, confirming that glutaraldehyde crosslinking increases the mechanical strength of collagen-based biomaterials. The present study sheds light on the biomechanics of glutaraldehyde-treated porcine pulmonary ligament that may be a candidate for tissue engineering.

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## Figures

Fig. 1

A schematic figure of biaxial testing setup

Fig. 2

Representative SHG and TPEF images demonstrating collagen and elastin structure at different depths of an unloaded pulmonary ligament specimen. (a, d) TPEF images for elastin, (b, e) SHG images for collagen, and (c, f) merged images. Row-wise: Z = 22.1 and 58.1 μm. columnwise: elastin, collagen, merged.

Fig. 3

Representative SHG and TPEF images demonstrating fiber arrangement at different depths of a specimen under equibiaxial stretch (E11=E22=0.6). Row-wise: elastin, collagen, merged; columnwise: Z = 9.9, 21.9, 37.9, and 69.9 μm.

Fig. 4

Quantitative measurement of fiber orientation angle based on SHG and TPEF images. (a) and (b) Fiber orientation angle measured based on images in Fig. 3. (c) The mean orientation angle at ten different depths of pulmonary ligament specimens. Error bar indicates standard deviation.

Fig. 5

The stress–strain curves of fresh and glutaraldehyde-treated pulmonary ligaments, respectively, based on equibiaxial testing data. Symbols denote experimental mean stress components, and solid lines denote model predicted stresses. Error bar indicates standard error.

Fig. 6

The stress–strain curves of a representative glutaraldehyde-treated sample under three biaxial protocols E11:E22  = 1.3:2, 2:1.3, 2:2. Symbols and solid lines denote experimental data and model predictions, respectively.

Fig. 7

The stress–strain curves of a representative glutaraldehyde-treated sample under all biaxial protocols. Symbols and solid lines denote experimental data and model predictions, respectively.

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