Conductive bioimprint for regenerative medicine : synthesis and characterisation
Thesis DisciplineBiomedical Engineering
Degree GrantorUniversity of Canterbury
Degree NameDoctor of Philosophy
The ability to manufacture the cell culture platforms that guide biological cells to differentiate in controlled manners and in situ monitoring their viability and response to stimulate is important in a number of applications such as lab-on-chip, tissue engineering and biosensing.
This research presents the development of conductive bioimprinting based on conducting polymer poly(3,4–ethylenedioxythiophene):polystyrene sulfonate (PEDOT:PSS) for cell features replication and monitoring platform. Conductive bioimprinting is a new technology that integrates hydrogel and glycerol into PEDOT: PSS to imitate cell-like features using a soft lithography method. Adding functionality to the bioimprint process by enabling not only high resolution replica but also monitoring cell growth/viability through changes in their conductivity, an approach that has not been investigated before. Moreover, the use of hydrogel, glycerol and crosslinker demonstrates enhancement in biocompatibility, low degradation/water solubility and mechanical properties of the conductive substrate.
Six different concentrations of PEDOT:PSS (0%, 2%, 3%, 4% and 6% w/w) were studied in the synthesis of the conductive hydrogel via chemical method. The success in the synthesis and optimisation of the conductive hydrogel resulted in a very high replication fidelity with more than 90% accuracy when used for replicating myoblast C2C12 cellular features down to micro- and nano-sized details. The high-resolution conductive bioimprint replicas were utilised as a secondary cell culture substrate to investigate the cells response to cell–like topography as compared to featureless flat surfaces.
The synthesised conductive hydrogel underwent chemical and surface modifications to optimise the electrical conductivity, swelling behaviour, mechanical properties, biocompatibility and cell response on different topography. Conductive hydrogel films that were chemically crosslinked with microbial Transglutaminase (mTg) showed significant increase in the electrical conductivity from 10-6 to 1 Scm-1, in addition to improve water solubility. Moreover, cultured myoblast C2C12 cells on the crosslinked conductive substrates showed that they attached, grew and proliferated on all surfaces after 24 hours of cell seeding. In addition, conventional patterning of the conductive hydrogel substrates were employed in vitro to culture myoblast C2C12 cells and observe the cell–material interactions. Crystal violet stained cells revealed that cells are very responsive to their micro environment and they prefer to attach on a patterned surface as compared to a flat surface under the same cell culture media conditions.
In investigating the physicochemical properties of crosslinked conductive hydrogel films with different concentrations, several techniques have been employed for the analysis and testing of the synthesised material. Wettability, biocompatibility, biodegradability, mechanical analysis and some chemical analysis were performed. Results showed that the conductive hydrogel films have a good wettability and excellent hydrophilic properties. These material features acted favourably for the myoblast cell adhesion onto the conductive hydrogel substrate as compared to glass, gold and PDMS substrates. In vitro biodegradation tests when conducted in a culture medium at temperature of 37 ℃ for 15 days revealed that the degradation of the conductive hydrogel films was slowed down with increasing PEDOT: PSS concentrations. The tensile testing showed that the conductive hydrogel exhibited excellent stretchability properties: the elasticity modulus was decreased from 77.87 ± 1.12 MPa for the 2% films to 1.11 ± 0.10 MPa for the 6% films. Furthermore, from the chemical analysis: ultraviolet visible spectroscopy (UV-Vis), X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FTIR) and Thermogravimetric analysis (TGA) confirmed that the appropriate and expected chemical elements were present in the synthesised conductive hydrogel polymer films.
The electrical cell-substrate impedance spectroscopy (ECIS) measurements were performed on myoblasts C2C12 cells. The myoblast cell’s impedance changed during adhesion, growth, proliferation and toxin addition. The sensitivity of the conductive hydrogel biochip during cell culture experiments were measured and analysed. These results demonstrate a new method of using cell impedance spectroscopy on conductive hydrogel to study the cell behaviour and would help researchers’ better understanding on cellular responses/ events in real–time.