Figures & data
Figure 1. TRB3 expression is increased in HG-treated hRPE cells in a concentration-dependent manner. (a) RT-qPCR and (b) western blot tested TRB3 expression in hRPE cells exposed to different concentrations of HG. **P < 0.01, ***P < 0.001. TRB3, Tribbles homolog 3.
Figure 2. TRB3 depletion alleviates HG-triggered hRPE cell impairment. The transfection efficiency of shRNA-TRB3-1/2 plasmids was tested by (a) RT-qPCR and (b) western blot. (c) The impacts of TRB3 silencing on the viability of HG-treated hRPE cells were appraised by CCK-8 assay. **P < 0.01, ***P < 0.001. TRB3, Tribbles homolog 3. MA, mannitol. HG, high glucose.
Figure 3. TRB3 inhibition protects HG-insulted hRPE cells against inflammation and oxidative stress. (a) TNF-α, IL-1β and IL-6 levels were determined by RT-qPCR. (b) ROS accumulation was detected by DCFH-DA assay. (c) MDA and SOD levels were confirmed by corresponding kits. ***P < 0.001. TRB3, Tribbles homolog 3. MA, mannitol. HG, high glucose. TNF-α, tumor necrosis factor alpha. IL-1β, interleukin-1beta. IL-6, interleukin-6. MDA, malondialdehyde. SOD, superoxidase dismutase.
Figure 4. TRB3 interference impedes HG-evoked apoptosis in hRPE cells. (a) TUNEL assay estimated the influence of TRB3 depletion on the apoptosis of HG-stimulated hRPE cells and (b) the quantification. (c) The protein levels of apoptosis-related factors were analyzed by western blot. **P < 0.01, ***P < 0.001. TRB3, Tribbles homolog 3. MA, mannitol. HG, high glucose. Bcl-2, B cell lymphoma-2. Bax, BCL-2 associated X.
Figure 5. TRB3 interacts with GRB2 in hRPE cells under HG conditions. (a) RT-qPCR and (b) western blot tested GRB2 expression in hRPE cells exposed to different concentrations of HG. (c-d) Co-IP assay testified the binding to TRB3 to GRB2. (e) RT-qPCR and (f) western blot tested GRB2 expression when TRB3 was down-regulated. **P < 0.01, ***P < 0.001. n.s., not significant. TRB3, Tribbles homolog 3. MA, mannitol. HG, high glucose. GRB2, growth factor receptor-bound 2.
Figure 6. TRB3 modulates HG-triggered hRPE cell damage through binding to GRB2. The transfection efficiency of Ov-GRB2 plasmid was tested by (a) RT-qPCR and (b) western blot. (c) CCK-8 assay evaluated cell viability. *P < 0.05, **P < 0.01, ***P < 0.001. TRB3, Tribbles homolog 3. MA, mannitol. HG, high glucose. GRB2, growth factor receptor-bound 2.
Figure 7. TRB3 has impacts on HG-provoked inflammation and oxidative stress in hRPE cells via interaction with GRB2. (a) TNF-α, IL-1β and IL-6 levels were determined by RT-qPCR. (b) ROS accumulation was detected by DCFH-DA assay. (c) MDA and SOD levels were confirmed by corresponding kits. *P < 0.05, **P < 0.01, ***P < 0.001. TRB3, Tribbles homolog 3. MA, mannitol. HG, high glucose. GRB2, growth factor receptor-bound 2. TNF-α, tumor necrosis factor alpha. IL-1β, interleukin-1beta. IL-6, interleukin-6. MDA, malondialdehyde. SOD, superoxidase dismutase.
Figure 8. TRB3 affects HG-mediated hRPE cell apoptosis by binding with GRB2. (a) TUNEL assay estimated the apoptosis of HG-stimulated hRPE cells and (b) the quantification. (c) The protein levels of apoptosis-related factors were analyzed by western blot. ***P < 0.001. TRB3, Tribbles homolog 3. MA, mannitol. HG, high glucose. GRB2, growth factor receptor-bound 2. Bcl-2, B cell lymphoma-2. Bax, BCL-2 associated X.
