(C-120) Controlled phase separation between conjugated and zwitterionic polymers increases conductivity and suppresses the foreign body response

Implantable medical devices (IMDs) are playing an increasingly important role due to an aging population and the associated increasing prevalence of chronic diseases. However, the foreign body response (FBR) has limited IMDs from realizing their full potential. The FBR is characterized by inflammatory and fibrotic processes that surround the implanted material. Fibrotic encapsulation is particularly detrimental for biosensors and electrophysiological devices because it would impede the diffusion of analytes and ions. A class of materials with promising FBR suppressing properties are zwitterionic polymers, due to their antifouling properties. Previous work has shown that zwitterionic hydrogel implants can nearly eliminate the FBR for at least three months in mice. As electronic materials for IMDs, conjugated polymers are promising candidates due to their low mechanical moduli, broad chemical design space, and mixed electronic/ionic conducting properties. Specifically, poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate) (PEDOT:PSS) is of interest because it is chemically stable, highly conductive, highly processable, and non-cytotoxic. Nonetheless, previous work addressing the FBR against PEDOT:PSS did not place their focus on long-term solutions or did not carry out FBR-specific characterization methods. We hypothesize that by combining PEDOT:PSS and a zwitterionic polymer into a double-network hydrogel and carefully tuning its phase separation morphology, the conductivity can be increased and the FBR can be effectively suppressed. In addition, the porosity of the hydrogel enables the diffusion of analytes and ions. In this work, we demonstrate a process for inducing PEDOT:PSS network formation in situ in a zwitterionic hydrogel matrix, which significantly improves its conductivity and reduces FBR-associated fibrosis.

Zwitterionic PEDOT:PSS hydrogels (ZIPH) were prepared by encapsulating PEDOT:PSS nanoparticles in a zwitterionic poly(sulfobetaine methacrylate) hydrogel matrix. Thereafter, a novel post-treatment process (undisclosed) was applied to the hydrogel to generate ZIPH-3c. Electrical conductivity was determined via standard four-point probe method, and electrochemical properties were determined via electrochemical impedance spectroscopy. Tensile tests were carried out for mechanical characterization. The material structure was characterized via Raman mapping, SEM, conductive AFM, cryo-TEM, and grazing-incidence wide-angle X-ray scattering (GIWAXS), and the results were correlated with the observed bulk properties. Extensive fouling experiments were carried out using a modified CBQCA assay.

To assess the level of foreign body response and to gain insight into the mechanisms at play, hydrogels were implanted subcutaneously into the backs of mice. Mice were sacrificed 1-, 4-, and 12-weeks post-implantation. Masson’ trichrome stain was used to determine the collagen distribution around the implants. Flow cytometry was utilized to quantify macrophage polarization. Monocytes, T cells, neutrophils, fibroblasts, and myofibroblasts were also quantified using flow cytometry. Immunofluorescence microscopy was used to determine the spatial distribution of M1 and M2 macrophages, myofibroblasts, and new blood vessels. A custom made LegendPlex kit was used to quantify the cytokine concentrations in the granuloma. C3a complement fragments were quantified by ELISA. Finally, NanoString RNA analysis was conducted to determine the key genes associated with the tissues surrounding the implants.

The electrical conductivity of the hydrogel that has undergone the post-treatment process (ZIPH-3c) was over three orders of magnitude higher than the hydrogel that has not undergone the post-treatment process (ZIPH-u) (Fig. 1). The post-treatment process (undisclosed) also increased the Young’s modulus. To gain insight into the reason behind the change in bulk properties, a series of physical characterizations were conducted. GIWAXS showed that no change in crystallinity occurred, and the hydrogels stayed amorphous after the post-treatment process. Conductive AFM and cryo-TEM revealed that before the post-treatment process, PEDOT:PSS existed as spheroidal nanoparticles in the zwitterionic matrix, but after the post-treatment process, the PEDOT:PSS nanoparticles transitioned to an interconnected fibrillar structure. It was concluded that the increase in conductivity and the Young’s modulus are caused by the interconnected fibrillar structure formed by PEDOT:PSS (Fig. 2).

Fouling experiments demonstrated that both ZIPH hydrogels were no more antifouling than PEDOT:PSS itself, but in vivo experiments revealed a disconnect between the fouling properties and the FBR outcome. It was found that the tissue around ZIPH-3c had the lowest collagen density 4-weeks post-implantation, followed by ZIPH-u. The PEDOT:PSS hydrogel control had the highest collagen density (Fig. 3). To further understand the observations, immunofluorescence microscopy and flow cytometry were conducted. We observed that ZIPH-3c was associated with the greatest number of macrophages with CD206 expression, a marker for M2 polarization. Furthermore, it was found that ZIPH-3c was associated with the highest concentration of MCP-1 and VEGF after 1 week. Since MCP-1 and VEGF are associated with arteriogenesis and angiogenesis, respectively, immunofluorescence microscopy was conducted on CD31- and αSMA-stained tissue sections. The results suggest that neovascularization is occurring in tissues surrounding ZIPH-3c.

The results indicate that the novel post-treatment process significantly alters the properties of ZIPH. The changes in bulk physical properties can be explained by the data. Since all other factors were held mostly equal, the differences in phase separation and the morphology of the PEDOT:PSS nanoparticles could explain the extent of FBR-associated fibrosis. We believe that our findings could make significant contributions to the advancement and understanding of the design principles for implantable electronics.