Korean Circ J.  2017 Jan;47(1):123-131. 10.4070/kcj.2016.0203.

The Control of Drug Release and Vascular Endothelialization after Hyaluronic Acid-Coated Paclitaxel Multi-Layer Coating Stent Implantation in Porcine Coronary Restenosis Model

• Bae IH
• Jeong MH
• Kim JH
• Park YH
• Lim KS
• Park DS
• Shim JW
• Kim JH
• Ahn Y
• Hong YJ
• Sim DS

• KMID: 2365291
• DOI: http://doi.org/10.4070/kcj.2016.0203

Abstract

BACKGROUND AND OBJECTIVES
Hyaluronic acid (HA) is highly biocompatible with cells and the extracellular matrix. In contrast to degradation products of a synthetic polymer, degradation products of HA do not acidify the local environment. The aim of this study was to fabricate an HA-coated paclitaxel (PTX)-eluting stent via simple ionic interactions and to evaluate its effects in vitro and in vivo.
MATERIALS AND METHODS
HA and catechol were conjugated by means of an activation agent, and then the stent was immersed in this solution (resulting in a HA-coated stent). After that, PTX was immobilized on the HA-coated stent (resulting in a hyaluronic acid-coated paclitaxel-eluting stent [H-PTX stent]). Study groups were divided into 4 groups: bare metal stent (BMS), HA, H-PTX, and poly (L-lactide)-coated paclitaxel-eluting stent (P-PTX). Stents were randomly implanted in a porcine coronary artery. After 4 weeks, vessels surrounding the stents were isolated and subjected to various analyses.
RESULTS
Smoothness of the surface was maintained after expansion of the stent. In contrast to a previous study on a PTX-eluting stent, in this study, the PTX was effectively released up to 14 days (a half amount of PTX in 4 days). The proliferation of smooth muscle cells was successfully inhibited (by 80.5±12.11% at 7 days of culture as compared to the control) by PTX released from the stent. Animal experiments showed that the H-PTX stent does not induce an obvious inflammatory response. Nevertheless, restenosis was clearly decreased in the H-PTX stent group (9.8±3.25%) compared to the bare-metal stent group (29.7±8.11%).
CONCLUSION
A stent was stably coated with PTX via simple ionic interactions with HA. Restenosis was decreased in the H-PTX group. These results suggest that HA, a natural polymer, is suitable for fabrication of drug-eluting stents (without inflammation) as an alternative to a synthetic polymer.

Keyword

Stents; Hyaluronic acid; Paclitaxel; Preclinical drug evaluation; Coronary restenosis

MeSH Terms

Animal Experimentation
Coronary Restenosis*
Coronary Vessels
Drug Evaluation, Preclinical
Drug Liberation*
Drug-Eluting Stents
Extracellular Matrix
Hyaluronic Acid
In Vitro Techniques
Myocytes, Smooth Muscle
Paclitaxel*
Polymers
Stents*
Hyaluronic Acid
Paclitaxel
Polymers

Figure

• 

  Fig. 1 Schematic illustration of the coating approaches. To prepare and immobilize PTX NPs, HSA and HA were used. Ionic interactions between PTX NPs and HA were induced. BMS: bare-metal stent, HA: hyaluronic acid, H-PTX: hyaluronic-acid-coated paclitaxel-eluting stent, P-PTX: poly(L-lactide)-coated paclitaxel-eluting stent, HSA: human serum albumin, PTX: paclitaxel, PLA: polylactide, PTX NPs: paclitaxel nanoparticles.

• 

  Fig. 2 In vitro cumulative kinetics of the PTX release from the stents. As described in the Materials and Methods, the stents were placed in tubing, and then PBS was circulated in the tubes. The amount of PTX released into the medium was measured at certain time points using a spectrophotometer. The data are shown as mean±standard (n=10). PTX: paclitaxel, H-PTX: hyaluronic-acid-coated paclitaxel-eluting stent, P-PTX: poly(L-lactide)-coated paclitaxel-eluting stent, PBS: phosphate-buffered saline.

• 

  Fig. 3 Inhibitory effects of PTX on SMC proliferation. To assess the effects of HA and PTX on cell proliferation, a 2-dimensional version of a stent was immersed in the cell culture medium. Cells were seeded in the stent-containing culture dish and incubated. Proliferation was assessed by an XTT assay. The results are shown as a mean±standard as compared to the control, n=10. SMC: smooth muscle cell, BMS: bare-metal stent, HA: hyaluronic acid, H-PTX: hyaluronic-acid-coated paclitaxel-eluting stent, P-PTX: poly(L-lactide)-coated paclitaxel-eluting stent, PTX: paclitaxel.

• 

  Fig. 4 Histopathological analysis of the porcine coronary model. Twenty-eight days postimplantation, the vessels containing the stents were isolated and subjected to a histopathological analysis. Typical optical photographs (25×) of the cross-sectional slices. BMS: bare-metal stent, HA: hyaluronic acid, H-PTX: hyaluronic-acid-coated paclitaxel-eluting stent, P-PTX: poly(L-lactide)-coated paclitaxel-eluting stent.

• 

  Fig. 5 Histomorphometric analysis of the stented arteries. The data are presented as mean±standard (n=10), *p<0.05. NS: not statistically significant, BMS: bare-metal stent, HA: hyaluronic acid, H-PTX: hyaluronic-acid-coated paclitaxel-eluting stent, P-PTX: poly(L-lactide)-coated paclitaxel-eluting stent.

• 

  Fig. 6 Immunofluorescence analysis of the stents. Stent sections after coronary implantation were paraffin-embedded and stained for CD68. Red staining by secondary antibodies was Alexa fluor 568. DAPI staining was performed to visualize the cell nuclei (40×). BMS: bare-metal stent, HA: hyaluronic acid, H-PTX: hyaluronic-acid-coated paclitaxel-eluting stent, P-PTX: poly(L-lactide)-coated paclitaxel-eluting stent, DAPI: 4',6-diamidino-2-phenylindole.

• 

  Fig. 7 The strut coverage rate. (A) Representative SEM images of the luminal area of the stents. The latter were harvested at 4 weeks postimplantation. They were bisected longitudinally and subjected to SEM examination. (B) Histomorphometric analysis of strut coverage rate ntified by ntified by special software (Visus 2000 Visual Image Analysis System; IMT Tech., San Diego, CA, USA). (Visus 2000 Visual Image Analysis System; IMT Tech., San Diego, CA, USA). special software. BMS: bare-metal stent, HA: hyaluronic acid, H-PTX: hyaluronic-acid-coated paclitaxel-eluting stent, P-PTX: poly(L-lactide)-coated paclitaxel-eluting stent, SEM: scanning electron microscope.

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