In vitro response of THP-1 derived macrophages to antimicrobially effective PHMB-coated Ti6Al4V alloy implant material with and without contamination with S. epidermidis and P. aeruginosa

Abstract

Aim Periprosthetic joint infections are a devastating complication after arthroplasty, leading to rejection of the prosthesis. The prevention of septic loosening may be possible by an antimicrobial coating of the implant surface. Poly (hexamethylene) biguanide hydrochloride [PHMB] seems to be a suitable antiseptic agent for this purpose since previous studies revealed a low cytotoxicity and a long-lasting microbicidal effect of Ti6Al4V alloy coated with PHMB. To preclude an excessive activation of the immune system, possible inflammatory effects on macrophages upon contact with PHMB-coated surfaces alone and after killing of S. epidermidis and P. aeruginosa are analyzed. Methods THP-1 monocytes were differentiated to M0 macrophages by phorbol 12-myristate 13-acetate and seeded onto Ti6Al4V surfaces coated with various amounts of PHMB. Next to microscopic immunofluorescence analysis of labeled macrophages after adhesion on the coated surface, measurement of intracellular reactive oxygen species and analysis of cytokine secretion at different time points without and with previous bacterial contamination were conducted. Results No influence on morphology of macrophages and only slight increases in iROS generation were detected. The cytokine secretion pattern depends on the surface treatment procedure and the amount of adsorbed PHMB. The PHMB coating resulted in a high reduction of viable bacteria, resulting in no significant differences in cytokine secretion as reaction to coated surfaces with and without bacterial burden. Conclusion Ti6Al4V specimens after alkaline treatment followed by coating with 5–7 μg PHMB and specimens treated with H2O2 before PHMB-coating (4 μg) had the smallest influence on the macrophage phienotype and thus are considered as the surface with the best cytocompatibility to macrophages tested in the present study.

In vitro co-culture models for the assessment of orthopedic antibacterial biomaterials

“…In the case of the im-bac models ( Figure 4F ), the co-culture time did not exceed 72 h. For simultaneous seeding, the time varied between 1.5 and 24 h, while for bacteria-first seeding, the range extended from 1 to 72 h. In the case of the cell-first models, the primary culture duration spanned from 0.5 to 4 h. The shorter co-culture times in the cell-first studies can be explained by the focus of those studies on the immediate immunomodulatory effect of the biomaterial upon seeding. However, in the study by Zwicker and co-workers ( Zwicker et al, 2022 ), the co-culture time was extended to 72 h to observe a polarization shift in the macrophages. No such shift was, however, observed in the co-culture while it did occur in their monoculture experiments.…”

“…Only one im-bac study has directly focused on macrophage polarization to limit the foreign body response toward a contact-killing surface instead of focusing on enhanced phagocytosis ( Zwicker et al, 2022 ). The monoculture of THP-1 stimulated macrophages on Ti6Al4V, with and without Poly (hexamethylene) biguanide hydrochloride (PHMB) coating, showed that the layer did not induce a pro-inflammatory response.…”

Deletion of codY impairs Staphylococcus epidermidis biofilm formation, generation of viable but non-culturable cells and stimulates cytokine production in human macrophages

“…S. epidermidis has been shown to induce high levels of pro-inflammatory cytokines secretion, both in vitro and in vivo . Namely, a marked increase in IL-6, IL-8 and TNF-α secretion was detected upon in vitro stimulation of THP-1-derived macrophages [43, 44] and human peripheral blood mononuclear cells [33, 45] with different S. epidermidis strains. Our research team has also shown this pro-inflammatory profile in vivo , in a murine model of bloodstream infection [28].…”

Galangin Targets HSP90β to Alleviate Ulcerative Colitis by Controlling Fatty Acid Synthesis and Subsequent NLRP3 Inflammasome Activation

“…The cells were cultured in a humidified environment with 5% CO 2 at 37 °C. [ 14 ] To induce macrophage differentiation, they were incubated with PMA (50 ng mL −1 ) for 3 h, washed with phosphate‐buffered saline (PBS) for three times. Then, they were treated with LPS (2 µg mL −1 ) in the absence or presence of Gal for 3 h, then stimulated with ATP (5 mM) for 1 h.…”