Abstract
Purpose :
Sunitinib has anti-fibrotic effects in various organ fibrotic diseases. α-Tocotrienol is an antioxidant with reversible inhibit cell proliferation and anti-fibrotic effects. High concentrations of tinib drugs may have serious side effects on eye tissues. In this study, we evaluated the potential of sunitinib in combination with alpha-tocotrienol at non-toxic concentrations as an anti-fibrotic therapeutic drug combination.
Methods :
Human Tenon's capsule fibroblasts (HTFs) were treated with 10ng/ml TGF-β. Then, in mitomycin C (MMC), sunitinib, α-tocotrienol, sunitinib or MMC combined with α- tocotrienol treatment group, antiproliferative, and antifibrotic effects were observed. At 12, 24,48,73 hours, observe the cell morphology to evaluate the toxicity of the drug to the cells. MTT measures the effect of HTFs proliferation and cell viability. LDH kit detects the drug toxicity. Annexin V/PI double staining was detected by flow cytometry to determine the level of apoptosis. Western Blot was used to detect apoptosis-related proteins (caspase-3, -9 and PARP) and fibrosis-related proteins (α-SMA, Col-1 ) expression level.
Results :
MMC showed a strong anti-proliferation effect, and irreversible pro-apoptotic effect in the early stage of HTFs cell differentiation. α-tocotrienol regulates the activity of cell lysosomes and has a reversible inhibitory effect on cell proliferation. Sunitinib also has the effect of inhibiting cell proliferation. In the drug combination of α-tocotrienol and MMC, the living cells of HTFs were significantly improved. In the combination of α-tocotrienol and sunitinib, the anti-cell proliferation ability was enhanced. α-tocotrienol, sunitinib and MMC all have anti-fibrosis effects. After sunitinib treated HTFs, Western Blot showed that sunitinib activated caspase-3, -9 and PARP cleavage, and down-regulated the expression of Bcl-xL and Bax, suggesting that the apoptosis was mediated through the mitochondrial pathway. In addition, the results of Western blot experiments showed that the expression of α-SMA and Col-I decreased, suggesting that sunitinib inhibited TGF-β-induced cellular fibrosis.
Conclusions :
Compared with MMC, the combination of α-tocotrienol and sunitinib can significantly inhibit the proliferation of HTFs cells at a non-toxic concentration, and effectively inhibit the formation of fibrosis.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.