Ror β expression in activated macrophages and dental pulp stem cells

To assess the expression of Retinoic acid-related orphan receptor beta (Ror β) in human inflamed dental pulp stem cells (hI-DPSCs) and during macrophage phenotypic conversion.

Commercially procured THP-1 monocytes conversion to macrophages was judged by their morphology, the percentage of adherent cells and the expression of CD-14 surface marker. THP-1 macrophage cell viability following LPS, IFN-γ/IL-4, IL-13 stimulus was evaluated at 24 and 48h. The phenotypic conversion of macrophages to M1 and M2 was confirmed by flow cytometry and Western blot analysis. Cytokine release following polarization was estimated by the BD cytokine flex kit. The expression of Ror β in THP-1 macrophages and hI-DPSCs following LPS, IFN-γ/IL-4, IL-13 stimulus was assessed by Western blot analysis. Statistical significance was analysed using one-way Anova followed by Tukey's Post hoc test.

THP-1 monocytes pretreated with PMA (100 ng mL^-1 ) for 48 h followed by culturing in PMA-free media for another 48 h yielded cells with morphological characteristics similar to macrophages with a high percentage of adherence capability and CD-14 expression. Macrophages treated with LPS 100 ng mL^-1 and IFN-γ 20 ng mL^-1 or IL-4 20 ng mL^-1 had high expression of the respective M1 and M2 CD markers in flow cytometry and Western blot analysis. Cytokine release studies demonstrated the expression of IL-1β, TNF-α and IL-10 in the M1-polarized macrophages (P < 0.01), whilst TGF- β levels were seen in the M1 and M2-polarized macrophages. Ror β expression was upregulated when macrophages and hI-DPSCs were treated with anti-inflammatory cytokines.

Ror β was expressed in THP-1 macrophages and hI-DPSCs during their resting stage. Upregulated expression of Ror β occurred following an anti-inflammatory stimulus.