Interactive Effects of Copper-Doped Urological Implants with Tissue in the Urinary Tract for the Inhibition of Cell Adhesion and Encrustation in the Animal Model Rat

At a Glance

Metadata	Details
Publication Date	2022-08-16
Journal	Polymers
Authors	Wolfgang Kram, Henrike Rebl, Julia E. de la Cruz, Antonia Haag, Jürgen Renner
Institutions	Centro de Cirugía de Mínima Invasión Jesús Usón, University of Rostock
Citations	7
Analysis	Full AI Review Included

Executive Summary

This research investigated the interactive effects of copper-doped amorphous hydrogenated carbon (a-C:H/Cu-multilayer) coatings on Elastollan^® implants to inhibit bacterial adhesion and encrustation in the urinary tract.

Therapeutic Window Established: In vitro studies determined a copper ion concentration window (0.5-1.0 mM CuCl₂) that effectively eliminated E. coli viability (to less than 1%) while maintaining excellent human urothelial cell viability (greater than 80%).
Biocompatibility Confirmed: The Elastollan^® base material, including 25% Barium Sulphate (BaSO₄) contrast agent, and the a-C:H/Cu-multilayer coating showed very good biocompatibility with urothelial cells.
Non-Toxic Release: In the rat animal model, copper concentrations in the urinary bladder tissue and blood serum remained within the non-toxic range (tissue: < 250 µg/g dry weight; serum: < 1.69 mg/L).
Delayed Release Mechanism: The formation of organic protein layers on the implants resulted in a delayed and reduced copper release, particularly in the Cu-coated glass bead group.
Encrustation Failure: The a-C:H/Cu-multilayer coated implants exhibited significantly higher encrustation (mean 1.994 g) compared to the uncoated reference (mean 1.117 g).
Causal Factor Identified: The increased encrustation is attributed primarily to the higher surface roughness resulting from the morphology of the amorphous carbon layer combined with copper doping.
Encrustation Composition: FTIR analysis showed the encrustation consisted mainly of struvite (80%), whewellite (10%), and protein (10%).

Technical Specifications

Parameter	Value	Unit	Context
Base Material	Elastollan^® 1185 A 10 FC	N/A	Polyurethane used for stent prototypes.
X-ray Contrast Agent	25	%	Barium Sulphate (BaSO₄) concentration in bulk material.
Single Cu Layer Thickness	200	nm	Thickness of copper layers in the a-C:H/Cu multilayer stack.
PECVD Gas Flow (Ar)	7	sccm	Process 1 (a-C:H deposition).
PECVD Gas Flow (C₂H₂)	50	sccm	Process 1 (a-C:H deposition).
PECVD Pressure	2.3 x 10^-2	Mbar	Process 1 (a-C:H deposition).
PECVD RF Power	250	W	Process 1 (a-C:H deposition).
PVD Gas Flow (Ar)	50	sccm	Process 2 (Cu sputtering).
PVD Pressure	3.8 x 10^-2	Mbar	Process 2 (Cu sputtering).
PVD RF Power	150	W	Process 2 (Cu sputtering).
Therapeutic Cu Window	0.5 - 1.0	mM	Concentration range lethal to E. coli but safe for urothelium.
Non-Toxic Cu Threshold (Tissue)	< 250	µg/g dry tissue	Reference limit for copper in liver tissue.
Mean Encrustation (Coated)	1.994	g	Group B (a-C:H/Cu-multilayer) after 21 days.
Mean Encrustation (Reference)	1.117	g	Group A (Uncoated Elastollan) after 21 days.
Encrustation Composition (FTIR)	80/10/10	%	Struvite/Whewellite/Protein ratio.

Key Methodologies

The study utilized a hybrid PVD-PECVD process for coating and a rat model simulating tissue injury and lithogenic conditions for in vivo testing.

Coating and Material Preparation

Hybrid Deposition: a-C:H/Cu-multilayers were deposited on Elastollan^® platelets using alternating Plasma Enhanced Chemical Vapor Deposition (PECVD) for the a-C:H layers and Physical Vapor Deposition (PVD) magnetron sputtering for the Cu layers.
Multilayer Structure: The process alternated 4 cycles of PECVD (Process 1) and 4 cycles of PVD (Process 2) to achieve time-dependent copper release.
Implant Types: Three types of implants were tested in the bladder tissue interaction study: uncoated borosilicate glass beads (Group 1), copper-coated glass beads (Group 2, Cu), and solid copper beads (Group 3, Cu+).

In Vitro Testing

Biocompatibility: MTS assay was performed on human urothelial HUC-1 cells incubated with materials and CuCl₂ solutions for 24 hours to quantify cell viability.
Antibacterial Efficacy: Escherichia coli HB 101 was incubated in synthetic urine with varying CuCl₂ concentrations (0.5 mM to 3 mM) to determine the minimum concentration required for bacterial elimination (CFU counting).

In Vivo Animal Model (Rat)

Model: Male Sprague-Dawley rats were used with an implant retention time of 21 days.
Tissue Injury Simulation: The urothelial barrier was reversibly disrupted via intraoperative microinjection of protamine sulphate (10 mg/mL) into the urinary bladder.
Encrustation Induction: Ethylene glycol (oxalate precursor) was added to the drinking water to promote urinary crystal formation (calcium oxalate).
Copper Quantification: Copper levels in bladder tissue were measured by F-AAS (detection limit 8 µg/g dry tissue). Serum and urine copper levels were measured by ICP-MS.
Histology: Bladder tissue was analyzed using Hematoxylin-Eosin (HE) staining and rhodanine staining to detect free or associated copper accumulation.

Post-Explantation Analysis

Encrustation Weight: Implants were weighed to quantify encrustation mass (Group A vs. Group B).
Compositional Analysis: Encrustations were analyzed using EDX mapping (for elemental distribution of Ca, P, Mg) and FTIR spectrometry (for molecular composition, confirming struvite, whewellite, and protein).

Commercial Applications

The development of copper-doped DLC coatings for controlled antimicrobial release targets several high-value medical and industrial sectors:

Urological Implants:
- Anti-encrustation and antibacterial coatings for ureteral stents (Double-J stents) to extend indwelling time and reduce morbidity.
- Coatings for urinary catheters and drainage tubes to combat biofilm formation and catheter-associated urinary tract infections (CAUTI).
General Medical Devices:
- Antimicrobial surfaces for temporary orthopedic implants (e.g., wires, pins) where localized infection prevention is critical.
- Biomaterials requiring high mechanical hardness and low friction (provided by DLC) combined with intrinsic antimicrobial properties (provided by Cu).
Controlled Release Systems:
- Multilayer thin-film technology for defined, time-dependent release of therapeutic metal ions or drugs in physiological environments.
Surface Engineering:
- Development of highly durable, biocompatible amorphous carbon coatings (a-C:H) for internal use, focusing on optimizing surface roughness to minimize protein adhesion and crystal nucleation.

View Original Abstract

The insertion of a ureteral stent provides acute care by restoring urine flow and alleviating urinary retention or dysfunction. The problems of encrustation, bacterial colonization and biofilm formation become increasingly important when ureteral stents are left in place for a longer period of time. One way to reduce encrustation and bacterial adherence is to modify the stent surface with a diamond-like carbon coating, in combination with copper doping. The biocompatibilities of the Elastollan® base material and the a-C:H/Cu-mulitilayer coating were tested in synthetic urine. The copper content in bladder tissue was determined by atomic absorption spectroscopy and in blood and in urine by inductively coupled plasma mass spectrometry. Encrustations on the materials were analyzed by scanning electron microscopy, energy dispersive X-ray spectroscopy and Fourier transform infrared spectroscopy. A therapeutic window for copper ions of 0.5-1.0 mM was determined to kill bacteria without affecting human urothelial cells. In the rat animal model, it was found that copper release did not reach toxic concentrations in the affecting tissue of the urinary tract or in the blood. The encrustation behavior of the surfaces showed that the roughness of the amorphous carbon layer with the copper doping is probably the causal factor for the higher encrustation.

Tech Support

Original Source

DOI: https://doi.org/10.3390/polym14163324

References

2001 - Study on concretions developed around urinary catheters and mechanisms of renal calculi development [Crossref]
2006 - The role of biofilm infection in urology [Crossref]
2007 - Biofilms—A microbial life perspective: A critical review [Crossref]
2000 - Biofilms and their role in infections in urology [Crossref]
2019 - Innovations in Ureteral Stent Technology [Crossref]
1998 - The effect of urease inhibitors on the encrustation of urethral catheters [Crossref]
2017 - Efficacy of silver/hydrophilic poly(p-xylylene) on preventing bacterial growth and biofilm formation in urinary catheters [Crossref]