Abstract:

Objective To discuss the inhibitory effect of Schisandrin B on the proliferation of pancreatic cancer Pan02 cells， and to clarify the mechanism. Methods CCK-8 method was used to detect the proliferation rates of the Pan02 cells after treated with different concentrations（0， 0.78， 1.56， 3.12， 6.25， 12.50， and 25.00 mg·L^－1 ） of Schisandrin B to select the optimal concentration and treatment time of Schisandrin B. The mouse pancreatic cancer Pan02 cells were divided into control group （0 mg·L^－1 Schisandrin B）， 2.5 mg·L^－1 Schisandrin B group， 5.0 mg·L^－1 Schisandrin B group， and 10.0 mg·L^－1 Schisandrin B group. The morpholoy of Pan02 cells invarious groups was observed with light microscope； 5-ethynyl-2'-deoxyuridine （EdU） staining assay was used to detect the positive expression rates of the Pan02 cells in various groups； flow cytometry was used to detect the percentages of the Pan02 cells at different cell cycles and the apoptotic rates of the cells in various groups； Western blotting method was used to detect the expression levels of cell cycle and apoptosis-related proteins in the cells in various groups. Results The CCK-8 method results showed that after treated with Schisandrin B for 48 and 72 h， compared with 0 mg·L^－1 Schisandrin B， the proliferation rates of the Pan02 cells after treated with different concentrations of Schisandrin B were decreased （P<0.01）， especially at 72 h. 0.25， 5.0， and 10.0 mg·L^－1 Schisandrin B were selected to treat the Pan02 cells， and 72 h was the treatment time. In control group， the Pan02 cells had a spindle shape， with good condition， and grew closely adhered to the wall with normal organelles and cytoplasm， in 2.5 and 5.0 mg·L^－1 Schisandrin B groups， the cell volume was decreased， the intercellular adhesion was disappeared， and the cell membrane was intact but more permeable； the cytoplasm shrank and vacuolar structures appeared inside the cells， with some fragmented and floating on the surface of the solution； in 10.0 mg·L^－1 Schisandrin B group， the Pan02 cells exhibited notable apoptotic bodies， indicating an apoptotic state. The EdU staining results showed that compared with control group， the rates of EdU positive cells in 2.5， 5.0， and 10.0 mg·L^－1 Schisandrin B groups were significantly decreased （P<0.01）. The flow cytometry results showed that compared with control group， the percentages of the cells at S phase in 2.5， 5.0， and 10.0 mg·L^－1 Schisandrin B groups were significantly increased （P<0.01）， while the percentages of the cells at G₂/M phase were significantly decreased （P<0.01）， and the percentages of the cells at G₀/G₁ phase in 5.0 amd 1.0 mg·L^－1 Schisandrin groups were decreased （P<0.01）； compared with control group， the apoptotic rates of the cells in 2.5， 5.0， and 10.0 mg·L^－1 Schisandrin B groups were significantly increased （P<0.01）. The Western blotting results showed that compared with control group， the expression levels of p27， B-cell lymphoma 2 （Bcl-2） associated X protein （Bax）， cleaved cysteine aspartic acid protease-3 （cleaved Caspase-3）， and cleaved poly adenosine diphosphate（ADP） ribose polymerase （cleaved PARP） proteins in the cells in 2.5 mg·L^－1 Schisandrin B group were significantly increased （P<0.05 or P<0.01）， the expression levels of cyclin A2， cyclin E2， and Bcl-2 proteins in the cells in 5.0 and 10.0 mg·L^－1 Schisandrin B groups were significantly decreased （P<0.05 or P<0.01）， while the expression levels of p27， Bax， cleaved Caspase-3， and cleaved PARP proteins in the cells in 5.0 and 10.0 mg·L^－1 Schisandrin B groups were significantly increased （P<0.01）. Conclusion Schisandrin B has an inhibitory effect on proliferation of the pancreatic cancer Pan02 cells， and its mechanism may be related to the activation of the cysteine aspartic acid protease-3 （Caspase-3） pathway to induce the apoptosis and activating p27 protein to induce the arrest of cell cycle at S phase.

Key words: Schisandrin B, Pancreas neoplasm, Pan02 cell, Apoptosis, Cell cycle