The role of macrophage polarization on fibroblast behavior-an in vitro investigation on titanium surfaces

Abstract

Objectives

This study investigated the effect of smooth and rough titanium surface topographies on macrophage polarization and their influence on gingival fibroblast behavior cultured on titanium surfaces.

Materials and methods

RAW 264.7 macrophages were seeded on smooth (pickled titanium (PT)) and rough Sand-blasted with Large grit particles followed by Acid-etching (SLA) titanium surfaces and first investigated for macrophage polarization towards tissue-inflammatory M1 macrophages or wound-healing M2 macrophages. Thereafter, culture media collected from macrophages on both surfaces were cultured with gingival fibroblasts seeded on their respective topographies. All experiments were performed in triplicate with three independent experiments.

Results

Macrophages seeded on SLA surfaces polarized towards tissue-inflammatory M1 macrophages at early time points. Immunofluorescent staining and RT-PCR analysis demonstrated higher levels of iNOS and gene expression of IL-1β, IL-6, and TNF-alpha on SLA surfaces at 3 days when compared to both tissue culture plastic (TCP) and PT surfaces (p < 0.001). Very little differences were found between smooth PT surfaces and TCP. Interestingly, proliferation assay (CCK-8) suggested that conditioned media (CM) from macrophages seeded on SLA surfaces drastically inhibited gingival fibroblast proliferation at 3 and 5 days (p < 0.001). Meanwhile, CM from macrophages cultured on SLA surfaces also significantly reduced collagen 1 synthesis on SLA surfaces at 14 days as assessed by immunofluorescent staining (p < 0.001).

Conclusion

The results from this study demonstrate that the polarization of macrophages towards a pro-inflammatory (M1) phenotype on SLA surfaces may have a negative impact on gingival fibroblast behavior on titanium surfaces. Future strategies to better modulate macrophage polarization should be investigated to support a favorable immune response and encourage tissue integration.

Clinical relevance

As SLA surfaces have a potential to shift macrophages towards tissue-inflammatory M1 macrophages, this might be a negative impact for soft tissue healing. Therefore, SLA surfaces should be kept within the bone, as when in contact with soft tissue, they are prone to support a lack of soft tissue integration leading to inflammation.

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