High Glucose Exacerbates TNF-α-Induced Proliferative Inhibition in Human Periodontal Ligament Stem Cells through Upregulation and Activation of TNF Receptor 1

Abstract EN

Objective.

This research is aimed at investigating how high glucose affects the proliferation and apoptosis in periodontal ligament stem cells (PDLSCs) in the presence of TNF-α.

Methods.

PDLSCs obtained from periodontal healthy permanent teeth were treated under either high-glucose condition (30 mmol/L, G30 group) or normal glucose condition (5.6 mmol/L, G5.6 group) in the presence or absence of TNF-α (10 ng/ml) for 2 to 6 days.

Cell proliferation and cell cycle were evaluated by CCK-8, EdU incorporation assay, and flow cytometry.

Cell apoptosis was assessed by annexin V/PI staining.

Protein expression was detected by western blotting.

Cellular ROS expression was evaluated by CellROX labeling and flow cytometry.

Specific antibodies targeting TNFR1 and TNFR2 were used to block TNF-α signaling.

Vitamin C was also used to verify if the blockage of ROS can rescue PDLSCs in the presence of high glucose and TNF-α.

Results.

CCK-8 assay showed that high glucose exacerbated TNF-α-induced cell viability inhibition (57.0%, 85.2%, and 100% for the G30+TNF-α group, G5.6+TNF-α group, and control group, respectively) on day 6.

High glucose increased protein expression of TNFR1 compared with the control group on day 2 (1.24-fold) and day 6 (1.26-fold).

Blocking TNFR1 totally reversed the proliferative inhibition in G30+TNF-α group.

The addition of vitamin C or TNFR1 antibody totally reversed the elevation of intracellular ROS expression caused by high glucose and TNF-α.

Vitamin C partially restored cell proliferation in the presence of high glucose and TNF-α.

Conclusion.

High glucose exacerbates TNF-α-induced proliferative inhibition in human periodontal ligament stem cells through the upregulation and activation of TNF receptor 1.

Inhibition of intracellular ROS expression by vitamin C partially rescues PDLSCs in terms of cell proliferation.