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Human Saphenous Vein Response to Trans-wall Oxygen Gradients in a Novel Ex Vivo Conditioning Platform

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An Erratum to this article was published on 18 September 2015

Abstract

Autologous saphenous veins are commonly used for the coronary artery bypass grafting even if they are liable to progressive patency reduction, known as ‘vein graft disease’. Although several cellular and molecular causes for vein graft disease have been identified using in vivo models, the metabolic cues induced by sudden interruption of vasa vasorum blood supply have remained unexplored. In the present manuscript, we describe the design of an ex vivo culture system allowing the generation of an oxygen gradient between the luminal and the adventitial sides of the vein. This system featured a separation between the inner and the outer vessel culture circuits, and integrated a purpose-developed de-oxygenator module enabling the trans-wall oxygen distribution (high oxygen level at luminal side and low oxygen level at the adventitial side) existing in arterialized veins. Compared with standard cultures the bypass-specific conditions determined a significant increase in the proliferation of cells around adventitial vasa vasorum and an elevation in the length density of small and large caliber vasa vasorum. These results suggest, for the first time, a cause-effect relationship between the vein adventitial hypoxia and a neo-vascularization process, a factor known to predispose the arterialized vein conduits to restenosis.

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Acknowledgments

This work was supported by the Italian Ministry of Health research Project RF-2011-02346867. The authors would also like to thank Dr. Elena Bresciani for her support during the designing and manufacturing of the de-oxygenator module and Dr. Martina Malavasi for her support in functional assessment of the ex vivo culture system.

Conflict of interest

The authors declare no conflict of interest to disclose.

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Author notes

    Authors and Affiliations

    1. Dipartimento di Elettronica, Informazione e Bioingegneria, Politecnico di Milano, P.zza Leonardo da Vinci 32, 20133, Milan, Italy

      Marco Piola, Gianfranco Beniamino Fiore & Monica Soncini

    2. Unità di Ingegneria Tissutale, Centro Cardiologico Monzino-IRCCS, Via Parea 4, 20138, Milan, Italy

      Francesca Prandi & Maurizio Pesce

    3. Dipartimento di Scienze Cliniche e di Comunità, Università di Milano, Via Parea 4, 20138, Milan, Italy

      Marco Agrifoglio & Gianluca Polvani

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    1. Marco Piola
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    3. Gianfranco Beniamino Fiore
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    Corresponding author

    Correspondence to Marco Piola.

    Additional information

    Associate Editor Andreas Anayiotos oversaw the review of this article.

    Marco Piola and Francesca Prandi have contributed equally to this work.

    Electronic supplementary material

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    Figure S1

    A) Simplified scheme of the DC-EVCS consisting of the inner chamber and the de-oxygenator flow circuit. Physical model of the multiple layers inherent to the silicone tubing membrane system used as de-oxygenator. In the figure, C IN and C OUT are the oxygen concentration at the inlet and outlet of the inner culture chamber; In N 2 and Out N 2 , are the inlet and outlet port for the nitrogen (N 2 ) connection line, and Q is the recirculating flow rate. With regard to the tubing, J is the local oxygen transfer per unit length crossing the tubing wall by diffusion (green arrow), t is the tubing thickness, Ro and Ri are the outer and inner radius respectively; v is the fluid velocity through the tubing; pO 2 OUT Deoxy and pO 2 IN Deoxy represent the oxygen partial pressures at the inlet and at the outlet of a silicone tubing of length L, pO 2x and pO 2x+Δx are the oxygen partial pressures at the inlet and at the outlet of a generic infinitesimal tubing element of length Δx; pO 2GAS is the oxygen partial pressure within the de-oxygenator chamber. (B) Length of the silicone tubing as a function of the recirculating medium flow rate. The tubing length is parameterized for different values of the desired reduction of fluid oxygen partial pressure at the outlet of the silicone tubing. The oxygen reduction is defined as: pO 2 IN Deoxy - pO 2 OUT Deoxy . The pink area highlights the suitable solution region (silicone tubing length < 100 cm) in the length/flow rate graph. C) Oxygen concentration (C OUT ) trend as a function of time. The oxygen concentration is parameterized for different values of recirculation flow rate (Q), and for different silicone tubing length:20 cm, 40 cm, and 80 cm. c GAS is set equal to 0 mmol/ml and c 0 equal to 2.09×10-4 mmol/ml. (PNG 241 kb)

    Figure S2

    A) Scheme of the open loop setup for testing the de-oxygenator module without recirculation. The peristaltic pump draws the fluid from the reservoir 1 to the de-oxygenator module and finally to the reservoir 2. Silicone tubing are used for connecting the reservoir 1 to the de-oxygenator, while PVC tubing (lower permeability to oxygen) are used for connecting the de-oxygenator to the reservoir 2. The oxygen flow chamber sensor is placed between the de-oxygenator and reservoir 2. Pure nitrogen is used within the de-oxygenator for driving the extraction of oxygen from the silicone tubing. B) Scheme of the closed loop setup for studying the transient and steady state of the system. PVC tubing (lower permeability to oxygen) are used for connecting the de-oxygenator to the inner chamber. The oxygen flow chamber sensor is placed between the inner chamber reservoir (red box) and the de-oxygenator module. (PNG 112 kb)

    Figure S3

    Quantification of TUNEL+ cells in the SV wall. Arrows indicate TUNEL+ nuclei. Statistical comparison by paired t-test did not show significant differences (n=4 CABG samples; n=3 standard samples). (PNG 927 kb)

    Figure S4

    Schematic representation of the length density quantification. In figure, t is the thickness of the histological section, a i and b i are the major and minor axes of the elliptical print, α i is the angle between the axis of the cylinder and the surface, and l i is the length of the vessel. (PNG 26 kb)

    Supplementary material 5 (PDF 168 kb)

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    Piola, M., Prandi, F., Fiore, G.B. et al. Human Saphenous Vein Response to Trans-wall Oxygen Gradients in a Novel Ex Vivo Conditioning Platform. Ann Biomed Eng 44, 1449–1461 (2016). https://doi.org/10.1007/s10439-015-1434-0

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