Figure 4

Inhibiting TRAF4 expression using Lv-TRAF4 abolished the difference in LPS-induced autophagy between HDMSCs and ASMSCs. (a) GFP-positive cells were observed under a confocal microscope (× 50, scale bar=200 μm), and the transduction efficiencies were equivalent between HDMSCs (n=30) and ASMSCs (n=30). (b) The expression levels of TRAF4, LC3-II and P62 in ASMSCs (n=30) and HDMSCs (n=30) were determined. Lv-TRAF4 effectively decreased the expression of TRAF4 in both ASMSCs and HDMSCs compared with that in the control and Lv-NC treatments before and after LPS stimulation. Lv-TRAF4 transduction led to increased LC3-II expression but decreased P62 expression in both ASMSCs and HDMSCs after LPS stimulation. In addition, silencing TRAF4 abolished the differences in LC3-II and P62 expression between HDMSCs and ASMSCs after LPS stimulation. The figures in b are typical examples of MSCs from one healthy donor and one AS subject. (b) are presented as the means±s.d.; *P<0.05. The control group consisted of MSCs not transduced with lentivirus. The Lv-NC group consisted of MSCs transduced with lentivirus encoding a negative control shRNA sequence. The Lv-TRAF4 group consisted of MSCs transduced with lentivirus encoding shRNA targeting TRAF4. AS, ankylosing spondylitis; ASMSCs, ankylosing spondylitis mesenchymal stem cells; HDMSCs, healthy donors mesenchymal stem cells; LPS, lipopolysaccharide; MSCs, mesenchymal stem cells.