Name: nondestructive monitoring of degradable scaffold-based tissue-engineered blood vessel development using optical coherence tomography
Uploaded: 2018-10-03T14:00:00
Description: Watch this Scientific Journal Video about Nondestructive Monitoring of Degradable Scaffold-Based Tissue-Engineered Blood Vessel Development Using Optical Coherence Tomography at JoVE.com

We would like to acknowledge the Science and Technology Planning Project of the Guangdong Province of China (2016B070701007) for supporting this work.

1. Degradable PGA Scaffold based Tissue-engineered Vessels Culture
<ol>
	<li>PGA Scaffold Fabrication
	<ol>
		<li>Sew PGA mesh (19 mm diameter and 1 mm thick) around silicone tubing sterilized by ethylene oxide (17 cm length, 5.0 mm diameter, and 0.3 mm thick) using 5-0 suture.</li>
		<li>Sew polytetrafluoroethylene (ePTFE, 1cm length) with 4-0 suture onto each end of PGA mesh and overlapped by 2 mm.</li>
		<li>Dip PGA scaffolds with the hand in 1 mol/L NaOH for 1 min to adjust the spatial structure of the mesh and soa...

<ol>
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To generate engineered vessels with structural and mechanical properties similar to those of native blood vessels can lead to shorten the time for clinical use and is the ultimate goal of vascular engineering. Optical imaging techniques permit the visualization of tissue engineered vascular specific components, which cannot monitor individual constructs throughout culture and exposure grafts to a culture environment without compromising sterility7. In this article, the culture chamber is separated...

Tissue-engineered blood vessels (TEBVs) is of the most promising material as an ideal vascular graft1. In order to develop grafts to be clinically useful with similar structural and functional properties as native vessels, multiple techniques have been designed to maintain vascular function2,3. Although there have been engineered vessels with acceptable patency rates during implantation and in Phase III clinical study4, long-term culture and high cost also show the necessity of monitoring the development of TEBVs. Understanding of extracellular matrix(ECM) growth, remodeling, and adaptation processes in TEBVs in the biomimetic chemo-mechanical environment can provide crucial information for the development of vascular tissue engineering.
The ideal strategy to track the development of small-diameter engineered vessels5 should be nondestructive, sterile, longitudinal, three-dimensional and quantitative. TEBVs under different culture conditions could be assessed by this imaging modality, even including changes before and after vascular transplantation. Strategies to describe features of living engineered vessels are needed. Optical imaging techniques allow visualization and quantification of tissue deposition and biomaterials. Other advantages are the possibility to enable deep-tissue and label-free imaging with high resolution6,7. However, image-specific molecules and less easily accessible optical equipment for real-time monitoring is a significant practical obstacle, which has limited the extensive application of nonlinear optical microscopy. Optical coherence tomography (OCT) is an optical approach with intravascular imaging modality as a widely-used clinical tool to guide cardiac interventional therapy8. In the literature the method of OCT was reported as a way to assess the wall thickness of TEBVs9,10, coupled with affirmative imaging modalities for vascular tissue engineering research. Whereas, the dynamics of engineered vascular growth and remodeling was not observed.
In this manuscript, we detail the preparation of biodegradable polymeric scaffold-based TEBVs for four weeks culture. Human umbilical arteries vascular smooth muscle cells (HUASMCs) are expanded and seeded into a porous degradable polyglycolic acid (PGA) scaffolds in the bioreactor. Biodegradable polymers play the role in a temporary substrate for tissue engineering and have a certain degradation rate11. In order to ensure an appropriate match between scaffold degradation and neo-tissue formation, ECM and PGA scaffolds are crucial factors for effective vascular remodeling. The perfusion system simulates the biomechanical microenvironment of native vessels and maintains a consistent deformation under pressure stimulation. 
The aim of the presented protocol is to describe a relatively simple and nondestructive strategy for TEBVs imaging and long-term monitoring of culture. This protocol can be utilized for visualization of morphological changes and thickness measurements of engineered vessels under different culture conditions. Additionally, the analyses of polymer-based materials degradation in the tissue engineering scaffolds can be performed for the identification. By combining methods of scanning electron microscopic(SEM) and polarized microscope used in this protocol, correlation and quantification of extracellular matrix distribution and PGA degradation can be made, which can facilitate assessing scaffold degradation combined with OCT imaging.

<table border="0" cellpadding="0" cellspacing="0" style="width:797px;" width="797">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">PGA mesh</td>
			<td style="width:265px;">Synthecon</td>
			<td style="width:149px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">silicone tube</td>
			<td style="width:265px;">Cole Parmer</td>
			<td style="width:149px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">connector</td>
			<td style="width:265px;">Cole Parmer</td>
			<td style="width:149px;"></td>
			<td></td>
		</tr>
		<tr height="46">
			<td height="46" style="height:47px;width:216px;">intravascular OCT system</td>
			<td style="width:265px;">St. Jude Medical, Inc</td>
			<td style="width:149px;">ILUMIEN&#8482; OPTIS&#8482; SYSTEM</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">scanning electron microscopic</td>
			<td style="width:265px;">Philips&#160;</td>
			<td style="width:149px;">FEI Philips XL-30</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">polarized microscope</td>
			<td style="width:265px;">Olympus</td>
			<td style="width:149px;">Olympus BX51</td>
			<td></td>
		</tr>
		<tr height="30">
			<td height="30" style="height:31px;width:216px;">sutures</td>
			<td style="width:265px;">Johnson &#38; Johnson</td>
			<td style="width:149px;"></td>
			<td></td>
		</tr>
		<tr height="30">
			<td height="30" style="height:30px;width:216px;">pulsatile pump</td>
			<td style="width:265px;">Guangdong Cardiovascular Institute</td>
			<td style="width:149px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">LightLab Imaging software</td>
			<td style="width:265px;">St. Jude Medical, Inc</td>
			<td style="width:149px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

nondestructive monitoring of degradable scaffold-based tissue-engineered blood vessel development using optical coherence tomography

Engineered vascular grafts with structural and mechanical properties similar to natural blood vessels are expected to meet the growing demand for arterial bypass. Characterization of the growth dynamics and remodeling process of degradable polymer scaffold-based tissue-engineered blood vessels (TEBVs) with pulsatile stimulation is crucial for vascular tissue engineering. Optical imaging techniques stand out as powerful tools for monitoring vascularization of engineered tissue enabling high-resolution imaging in real-time culture. This paper demonstrates a nondestructive and fast real-time imaging strategy to monitor the growth and remodeling of TEBVs in long-term culture by using optical coherence tomography (OCT). Geometric morphology is evaluated, including vascular remodeling process, wall thickness, and comparison of TEBV thickness in different culture time points and presence of pulsatile stimulation. Finally, OCT provides practical possibilities for real-time observation of the degradation of polymer in the reconstructing tissues under pulsatile stimulation or not and in each vessel segment, by compared with the assessment of polymer degradation using scanning electron microscopic(SEM) and polarized microscope.

The three-dimensional culture system consisted of a culture chamber in the bioreactor and the perfusion system with a closed fluid cycle10,13 (Figure 1). The OCT imaging catheter was inserted into the distal end of the Y-junction and pulled back in the silicone tube for imaging. OCT imaging was first used to delineate the structural characterization of biodegradable polymeric scaffold-based TEBVs duri...

Watch this Scientific Journal Video about Nondestructive Monitoring of Degradable Scaffold-Based Tissue-Engineered Blood Vessel Development Using Optical Coherence Tomography at JoVE.com

Nondestructive Monitoring of Degradable Scaffold-Based Tissue-Engineered Blood Vessel Development Using Optical Coherence Tomography

A step by step protocol for nondestructive and long-period monitoring the process of vascular remodeling and scaffold degradation in real-time culture of biodegradable polymeric scaffold-based tissue-engineered blood vessels with pulsatile stimulation using optical coherence tomography is described here.

Engineered vascular grafts with structural and mechanical properties similar to natural blood vessels are expected to meet the growing demand for arterial ...

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