Fig. 2: Resveratrol (RSV) rescued the cell aggregate formation and osteogenic differentiation of N-PDLSCs under inflammatory cytokine tumor necrosis factor alpha (TNF-α) treatment.

a–c Histological analysis of PDLSC aggregates (a) showed that N-PDLSCs under TNF-α treatment formed thinner aggregates (b) with less collagen deposition (c), which could be rescued by RSV application. Bars: 50 μm. d Osteogenesis of PDLSC aggregates demonstrated by ALP (top) and alizarin red staining (bottom). Bars: 5 mm (top) and 50 μm (bottom). e, f RSV treatment restored the ALP activity (e) and mineralizing activity (f) of N-PDLSC aggregates under TNF-α application. g Quantitative real-time polymerase chain reaction (qRT-PCR) analysis of the osteogenic marker genes Alp, Runt-related transcription factor 2 (Runx2), Osteocalcin (Ocn), and Collagen I (Col1) in PDLSC aggregates after osteogenic induction. The data showed that RSV rescued the osteogenic potential of N-PDLSC aggregates under TNF-α treatment. h Western blot analysis of the extracellular matrix (ECM) and osteogenic marker genes Col1, Periostin, and Runx2 in PDLSC aggregates. i Western blot analysis of molecular targets related to key signaling pathways in PDLSC aggregates: the Sirtuin 1 (Sirt1)-peroxisome proliferator-activated receptor gamma coactivator-1 alpha (Pgc1α) pathway; the nuclear factor kappaB (NFκB) pathway; and the adenosine monophosphate-activated protein kinase (AMPK) pathway. j Western blot analysis showed that RSV improved osteogenesis and ECM deposition in N-PDLSC aggregates under TNF-α treatment, possibly by inhibiting the NFκB pathway and stimulating the Pgc1α and AMPK pathways. TNF-α was applied at 10 ng/ml, and RSV was applied at 10 nM in all the experiments. n = 6 per group (a–f) and n = 3 per group (g–j). The data represent the mean ± SD. *P < 0.05