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An Atelocollagen Coating for Efficient Local Gene Silencing by Using Small Interfering RNA.

Olivia Koenig1, Dimitrios Nothdurft1, Nadja Perle1

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This study introduces a novel gene-eluting stent coating using atelocollagen (ATCOL) and small interfering RNA (siRNA) to improve vascular regeneration after stenting. The ATCOL-siRNA system effectively silenced ICAM-1, showing promise for enhanced artery wall repair.

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atelocollagenatherosclerosisgene knockdownlocal deliverysiICAM-1siRNA transfection

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Area of Science:

  • Biomaterials Science
  • Vascular Biology
  • Regenerative Medicine

Background:

  • Coronary artery stenting can lead to adverse effects, necessitating improved vascular regeneration strategies.
  • Drug-eluting stents represent an advancement, but gene-eluting stents offer a novel personalized approach for altering gene expression.
  • Intercellular adhesion molecule-1 (ICAM-1) on defective endothelial cells plays a role in vascular wall responses post-stenting.

Purpose of the Study:

  • To investigate a novel coating system comprising atelocollagen (ATCOL) and small interfering RNA (siRNA) targeting ICAM-1 for enhanced vascular regeneration.
  • To evaluate the biocompatibility, transfection efficiency, and gene-silencing capabilities of the ATCOL-siRNA coating.
  • To assess the suitability of this gene-eluting coating for potential in vivo applications in artery wall repair.

Main Methods:

  • Coating of endothelial cells (EA.hy926) with varying concentrations of ATCOL (0.008%, 0.032%) and immobilized siRNA (5 μg).
  • Cell viability assays, hemocompatibility testing, and transfection efficiency analysis using fluorescent-labeled siRNA.
  • Gene silencing assessment via flow cytometry for ICAM-1 knockdown and 5'-RNA ligase-mediated rapid amplification of cDNA ends PCR (RLM-RACE-PCR) for specificity.

Main Results:

  • High cell viability was observed for EA.hy926 cells cultured on ATCOL layers.
  • Hemocompatibility assays confirmed the biocompatibility of the ATCOL coating.
  • Optimal transfection efficiency was achieved with 5 μg siRNA in ATCOL within 2 days, with siRNA release lasting approximately 9 days.
  • Sustained ICAM-1 gene silencing up to 8 days was demonstrated with the 0.008% ATCOL and 5 μg siICAM-1 coating.
  • RLM-RACE-PCR confirmed the specificity of the gene-silencing effect.

Conclusions:

  • The developed ATCOL-based coating effectively delivers siRNA to endothelial cells, achieving specific and long-term gene silencing of ICAM-1.
  • The demonstrated biocompatibility and transfection efficiency highlight the potential of this ATCOL-siRNA system for vascular regeneration.
  • This gene-eluting coating is well-suited for further in vivo investigations aimed at improving artery wall regeneration after stent implantation.