# Enhancing Biocompatibility and Biophysical Properties of Three-Dimensional Collagen Scaffolds Using Nonthermal Plasma Treatment

## TL;DR

Nonthermal plasma treatment improves the mechanical strength and biocompatibility of collagen scaffolds, making them better for bone tissue engineering.

## Contribution

NTP treatment is shown to enhance 3D collagen scaffolds without altering their chemical composition.

## Key findings

- NTP treatment increased scaffold porosity and hydrophilicity without changing chemical composition.
- Mechanical stiffness improved by 14.5-16.7% after NTP treatment.
- NTP-treated scaffolds supported higher cell numbers and enhanced osteogenic marker expression.

## Abstract

Collagen-glycosaminoglycan
(CG) scaffolds are extensively
utilized
in tissue engineering for their excellent biocompatibility and low
immunogenicity; however, their poor mechanical stiffness typically
requires further physical or chemical modifications to enhance their
structural integrity for clinical applications. We investigate the
effects of nonthermal plasma (NTP) treatment; an emerging technology
commonly used in the biomedical field for surface modifications, sterilization,
and wound healing. A comprehensive analysis is conducted to evaluate
the surface characteristics, biophysical properties, and biocompatibility
of the 3D CG scaffolds treated with NTP for 2 and 5 min, compared
with untreated controls. Histological and SEM analyses demonstrated
thickening of the scaffold pore struts and an increase in porosity,
while Energy Dispersive X-ray Spectroscopy (EDS) and Fourier transform
infrared spectroscopy (FTIR) confirmed that the native chemical composition
of the scaffolds remained intact and unchanged following NTP exposure.
Post-treatment, the scaffolds exhibited increased hydrophilicity demonstrated
by a reduced contact angle. Mechanical testing showed significant
improvements in the scaffold’s compression modulus, with increases
of approximately 16.7% and 14.5% for 2 min and 5 min treatments, respectively
(p < 0.05). In vitro biocompatibility
assays indicated increased metabolic rates and significantly higher
cell numbers in the ADSC-seeded on NTP-treated scaffolds (p = 0.001; p = 0.02, respectively). Following
21 day osteogenic conditions, both 2 min and 5 min NTP-treated scaffolds
exhibited significantly elevated expression of key osteogenic markers,
with RUNX2 showing a 9-fold increase at 2 min and an 11-fold increase
at 5 min (p < 0.001), and Osteocalcin demonstrating
increases of 2.5-fold and 2.3-fold, respectively (p < 0.01), compared to untreated controls. The enhanced biocompatibility
and ability to serve as a supportive matrix that promotes osteogenic
lineage commitment observed in the NTP-treated scaffolds suggest that
these materials could be effectively utilized as allogenic osteocyte-loaded
biomaterials for bone regeneration. Collectively, these results demonstrate
that NTP treatment significantly improves the functional performance
and mechanical strength of the 3D CG scaffolds, establishing it as
an effective approach for enhancing scaffold performance in regenerative
medicine applications.