Plasma-Activated Polydimethylsiloxane Microstructured Pattern with Collagen for Improved Myoblast Cell Guidance

Abstract

We focused on polydimethylsiloxane (PDMS) as a substrate for replication, micropatterning, and construction of biologically active surfaces. The novelty of this study is based on the combination of the argon plasma exposure of a micropatterned PDMS scaffold, where the plasma served as a strong tool for subsequent grafting of collagen coatings and their application as cell growth scaffolds, where the standard was significantly exceeded. As part of the scaffold design, templates with a patterned microstructure of different dimensions (50 × 50, 50 × 20, and 30 × 30 μm²) were created by photolithography followed by pattern replication on a PDMS polymer substrate. Subsequently, the prepared microstructured PDMS replicas were coated with a type I collagen layer. The sample preparation was followed by the characterization of material surface properties using various analytical techniques, including scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), and X-ray photoelectron spectroscopy (XPS). To evaluate the biocompatibility of the produced samples, we conducted studies on the interactions between selected polymer replicas and micro- and nanostructures and mammalian cells. Specifically, we utilized mouse myoblasts (C2C12), and our results demonstrate that we achieved excellent cell alignment in conjunction with the development of a cytocompatible surface. Consequently, the outcomes of this research contribute to an enhanced comprehension of surface properties and interactions between structured polymers and mammalian cells. The use of periodic microstructures has the potential to advance the creation of novel materials and scaffolds in tissue engineering. These materials exhibit exceptional biocompatibility and possess the capacity to promote cell adhesion and growth.

Keywords: PDMS; coating; collagen type I; cytocompatibility; microstructure; myoblast cell; nanostructured pattern; replication; soft lithography.

Figure 1

SEM images of PDMS-negative replicas: (a) 30 × 30 μm and (b) 50 × 50 μm photoresist pattern. Detailed SEM image of a PDMS-negative replica (30 × 30 μm) demonstrating pattern edges.

Figure 2

Detailed SEM images of PDMS-negative replicas coated with a type I collagen layer after lyophilization. (a) Pattern size 30 × 30 μm and (b) 50 × 50 μm.

Figure 3

Chemical composition of PDMS surfaces (30 × 30 μm, 50 × 50 μm) before lyophilization and after collagen coating; EDS analysis, measured area size of 30 × 30 μm².

Figure 4

XPS spectra of PDMS surfaces (30 × 30 μm) before lyophilization (A) and after collagen coating (B), acquired from XPS analysis, and XPS spectra of PDMS surfaces (50 × 50 μm) before lyophilization (C) and after collagen coating (D), acquired from XPS analysis.

Figure 5

Deconvoluted C1s XPS spectra of PDMS surfaces (30 × 30 μm) before lyophilization (A) and after collagen coating (B), acquired from XPS analysis, and XPS spectra of PDMS surfaces (50 × 50 μm) before lyophilization (C) and after collagen coating (D), acquired from XPS analysis. The particular description of characterictic peaks are introduced in Figure 5B, the colours corresponds on all figures (A–D) to this description. The figures show the line positions as measured, without calibration to remove drift due to charging.

Figure 6

The number of adhered (day 1) and proliferated (day 3) C2C12 cells cultured on PDMS surfaces (30 × 30 μm, 50 × 50 μm) after collagen coating; tissue polystyrene (TCPS) for comparison is also shown. PDMS surface (line 50 × 50 μm) after collagen coating—fluorescence microscopy images of C2C12 cells with labeled cytoskeletons (in green) and nuclei (in blue) are also shown. White line represents 50 micron.

Figure 7

Images from fluorescence microscopy of C2C12 with phalloidin (Atto 488)-stained cells on PDMS surfaces (30 × 30 μm, 50 × 50 μm) after collagen coating and tissue polystyrene (TCPS). The results for the 1st day, 3rd day, and 6th day from seeding are presented.