Neutrophils Respond Selectively to Physical Cues: Roughness Modulates Its Granule Release, and NETosis

Our study examined how different titanium alloy Ti6Al4V (Ti64) and zirconia (ZrO2) surfaces, ranging from rough to very smooth, affect the expression of elastase (NE), matrix metalloproteinase (MMP)-8, MMP-9, and extracellular traps (NETs) by neutrophils. Discs of Ti64 and ZrO2, 10 mm in diameter and 1.5 mm thick, were created using diamond-impregnated polishing burs and paste to produce rough (Ra > 3 µm), smooth (Ra ≥ 1 to 1.5 µm), and very smooth (Ra < 0.1 µm) surfaces. Neutrophils from Wistar rats were cultured on these surfaces, and the culture supernatants were then examined for NE, MMP-8, and MMP-9 using ELISA. At the same time, NET formation was demonstrated immunohistochemically by staining neutrophils with CD16b and DNA with DAPI. Overall, the expressions of NE and MMP-8 were significantly higher from neutrophil culture on Ti64 and ZrO2 rough surfaces compared to the very smooth surface (R > S > VS) after 2 h and 4 h of culture. The expression of MMP-9 also increased with culture time; however, no significant surface effects on expression were observed. Similarly, rough Ti64 and ZrO2 surfaces (R & S) also showed significantly larger NET formation compared to the very smooth surface (VS) after 4 h and 8 h cultures. Our findings suggest that increasing surface roughness on Ti64 and ZrO2 triggers higher NE, MMP-8, and NET formation secretion.