Bo Cao

This PhD thesis investigates the intricate interplay between titanium alloy surface characteristics, macrophage immune responses and novel therapeutic strategies to enhance the resistance of orthopedic implants to infection. The studies described provide new insights into the role of surface morphology in regulating macrophage function and explores the mechanisms of transcriptional regulation as a potential immunomodulatory target.

Chapter 1 describes the clinical and biological challenges associated with orthopedic implant infections, with particular emphasis on the enduring threat posed by bacterial biofilms and the urgent need for more in-depth host-pathogen-material studies to find a new approach to the treatment of infections in orthopedic implant materials. It also reviews the latest methods employed in infection biology and implantation studies, including advanced imaging, transcriptomics and real-time in vitro infection modelling. It acts as a foundation for the mechanistic exploration of host-pathogen-material interactions.

Chapter 2 investigated the effect of infected titanium surface characteristics on macrophages, revealing that a moderately rough surface promotes macrophage adhesion and polarization towards both M1 and M2 phenotypes. The M1 phenotype is dominant which is critical for early antimicrobial activity, particularly in the context of foreign body-associated infections. Proteomics revealed that key protein and NFATc1 immune pathways are influenced by the titanium disk surface roughness, suggesting that tailored surface engineering may provide a effective approach to the critical early treatment phase of orthopedic implant infections.

Chapter 3 extends this inquiry utilizing a more complex biological scenario, examining macrophage-bacteria interactions on different titanium surfaces. By constructing GFP-expressing S. aureus strains using electron microscopy, laser confocal, and proteomics techniques, it was revealed that the combined effects of surface roughness and S. aureus co-cultures can significantly reduce macrophage bacterial clearance by inducing macrophage M1 polarization, adhesion, improved phagocytic activity, and reactive oxygen species (ROS) generation. Altering the roughness of orthopedic implant materials opens new frontiers in the design of bio interfacesfor anti-infective implants. Proteomics proved that surface roughness modulates both NFATc1 macrophage polarization signaling pathway and key metabolic pathways that regulate immune responses.

Chapter 4 shifts towards molecular processes, describing the cooperative DNA binding kinetics of the transcription factors NFATc1 and c-Jun. Bases on the SPR data, itcan be concluded that although each factor binds DNA independently, their cooperative binding significantly enhances binding kinetics, suggesting a synergistic mechanism of gene regulation. This mechanistic insight provides a conceptual basis for immune-targeted immune interventions that go beyond traditional antimicrobial strategies.

Finally, chapter 5 builds on these observations, introducing a rationally designed peptide, MBB, which structurally mimics c-Jun and selectively inhibits NFATc1/c-Jun DNA co-binding. Functional tests demonstrated that MBB enhanced S. aureus phagocytosis and ROS generation by macrophages. This represents a novel class of immunotherapeutic agents for implant infections, as this peptide selectively interferes with NFATc1/c-Jun DNA co-binding and enhances macrophage clearance of the pathogenic bacterium S. aureus in the case of implant infections without relying on antibiotics.