RESUMO
AIM: To explore the effect of the Andrographis paniculata (A. paniculata) polysaccharide on the proliferation and apoptosis of human retinoblastoma (RB) Y79 cells and its mechanism. METHODS: The refined A. paniculata polysaccharide was obtained using techniques such as water extraction, ethanol precipitation, and decompression concentration. The inhibition effect of the A. paniculata polysaccharide on the proliferation of Y79 cells was detected by cell proliferation assay. Flow cytometry was used for the detection of cell apoptosis rate and cycle change. Real-time qunatitative polymerase chain reaction (RT qPCR)and Western blotting were used to detect the expression of cell apoptosis signal pathway-related factors (caspase-3, caspase-8, and caspase-9) and cell cycle signal pathway-related factors (CDK1 and cyclinB1) at the transcriptional and translational levels. RESULTS: Infrared and ultraviolet spectrum scanning showed that the extracted drug was a polysaccharide with high purity. After being treated with different concentrations of A. paniculata polysaccharide for different periods of time, the Y79 cells showed different degrees of proliferation inhibition. Flow cytometric observations showed that the cell apoptosis rate and the proportion of cells blocked in the G2/M phase were significantly increased after A. paniculata polysaccharide treatment. Further analysis revealed that the mRNA and protein expression of caspase-3, caspase-8, and caspase-9 in the A. paniculata polysaccharide treatment groups increased significantly compared with that in the control groups, while the expression of CDK1 and cyclinB1 decreased significantly. CONCLUSION: The A. paniculata polysaccharide could inhibit the proliferation and induce apoptosis of Y79 cells. Its possible mechanism is via the upregulation of caspase-3, caspase-8, and caspase-9 expression in the cell apoptotic signaling pathway and the downregulation of CDK1 and cyclinB1 expression in the cell cycle signaling pathway.
RESUMO
OBJECTIVE: To evaluate the wound healing mode of rabbit corneal epithelium after Epi-LASIK(epiplolic laser in-situ keratomileusis) and Flap-free Epi-LASIK(Flap-free epiploic laser in-situ keratomileusis). METHODS: Twenty-four New Zealand white rabbits (48 eyes) were randomly divided into four groups. There were six rabbits in every group, which were randomly treated with Epi-LASIK in one eye and Flap-free Epi-LASIK in the other by VISX20/20 excimer laser. Other three rabbits (6 eyes) without any treatment were used as control. The corneal epithelium wound healing mode was observed at 1, 2, 4, 7 days after surgery by light microscope and electron microscope. The number of keratocytes cells in the central anterior stroma were counted by hematoxylin-eosin-stained. The expression of basic fibroblast growth factor (bFGF) in corneal stroma were assessed by Immunohistochemistry staining. Cornea stromal cell apoptosis was evaluated by terminal deoxyribonuc-1-eotidyl transferase-mediated deoxynuridine triphosphate nick end labeling (TUNEL) assay.All the data was analysised by t-test to evaluate the different of wound healing mode after Epi-LASIK and Flap-free Epi-LASIK. RESULTS: The findings of light microscope and electron microscope investigation demonstrated that, at the first day after surgery, there was one layer of neogenesis of the corneal epithelial cells covering the area of surgery in Flap-free Epi-LASIK specimens. It would increase gradually after 7 days. At the first day after the surgery, the base membrane of the corneal epithelium was tightly attached to the stroma in Epi-LASIK specimens. At the second day after the surgery, there was edema observed on the epithelium flap in the Epi-LASIK specimens. There were more keratocytes cells found at 4 days after surgery in Flap-free Epi-LASIK group (33.85 +/- 3.39) than that in Epi-LASIK group (t = -2.315, P < 0.05). The expression of bFGF in Flap-free Epi-LASIK group at 2, 4 days after surgery were 101.75 +/- 8.07 and 110.56 +/- 9.00, which were more than Epi-LASIK (t = -2.339, -2.396, P < 0.05). There was less keratocyte apoptosis cells in Flap-free Epi-LASIK group than Epi-LASIK group at 2, 4 days after surgery (t = 3.332, 2.989, P < 0.05). CONCLUSIONS: Due to the specific way of the epithelium removal resulting in the more rapid epithelium healing, the Flap-free Epi-LASIK technique is advantageous in reducing the stimulative symptoms after the surgery. It's also beneficial for the corneal wound healing.
