Effect of incorporation of titanium dioxide nanotubes to convencional glass ionomer cement on the expression pattern of immunomarkers in fibroblasts stimulated by lipopolysaccharide

Abstract

Titanium dioxide (n-TiO2) nanotubes have been suggested as promising materials for several applications. In order to determine the impact of the addition of n-TiO2 on the physicochemical properties of glass ionomer cement (GIC), our research group carried out a series of in vitro studies (ARP FAPESP #2016/13786-0; 2019/14078-8). In general, the results obtained demonstrate that the presence of n-TiO2 in the composition of CIVs produces an increase of GIC hardness without affecting surface roughness, dentin micro shear-bond-strengh, and cell morphology of fibroblasts grown on CIV prepared with different concentrations of nanotubes. Despite the significant advance in this area, it is not known whether the presence of n-TiO2 in the composition of CIV can modulate the pattern of inflammatory cytokine expression. Thus, the present study aims to determine whether the presence of n-TiO2 in the composition of the CIV modifies the pattern of expression of inflammatory cytokines in fibroblast culture (NIH/3T3 cells) with the presence of the total extract of Fusobacterium nucleatum (Fn). Different concentrations of n-TiO2 (3%; 5%; 7% by weight) synthesized by the alkaline method (E20 nm) will be incorporated into the CIV [Ketac Molar EasyMix - (KM)]. Fibroblasts will be cultured on CIV discs with and without nanotubes, and with the presence of the total extract of Fusobacterium nucleatum (Fn) (concentration to be determined by the cell viability test) and the following tests will be performed: 1. Cell proliferation/viability assay (Trypan blue and mitochondrial-MTT activity) (n=6; 1, 3 and 4 days); 2. Morphology of fibroblasts (Confocal Laser Microscopy (Confocal) (n=6; 1, 3 and 4 days); 3. Protein expression - determination of cytokine levels (IL-1², IL-6, IL-10, and TNF -±) using multiplex methodology (n=6; 12 and 18 h) and 4. Gene expression - determination of IL-1², IL-6, IL-10, and TNF-± levels) (n=6; 3, 6, and 18h) using real-time PCR. Data will be submitted to statistical analysis, considering the significance level of 5%.(AU)