| Cordycepin, a main active composition separated from Cordyceps fungus, has been reported to exert multiple kinds of physiological activity in recent years, such as anti-tumor, anti-aging, anti-virus, enhancing immunity and so on. At present, cordycepin is obtained mainly through the extraction from Cordyceps fruiting body and purified by ion exchange resin or macroporous resin. While, the recovery of cordycepin of these two methods is extremely low, despite of the satisfactory purification effect, which makes it not conducive to industrial production. Cordycepin has been reported to exert anti-tumor activity in a broad spectrum of cancer types. However, there is very few report about the function of cordycepin on human non-small cell lung cancer cells.In our research, Cordyceps Kyushuensis rice culture medium was used as raw material to study the extraction and purification process of cordycepin. First, we explored the impact of different ethanol concentration on cordycepin extraction, and the results indicated that at the concentration of 80% we could receive satisfied quantity of extraction. Then, the cordycepin was purified by AB-8 and acidic alumina, respectively and we found the purity of these two approaches was in close proximity to 35%. While, the yield of cordycepin after purified by acidic alumina could reach up to 83.67% and that of AB-8 was merely 36.78%, so acidic alumina was selected for the purification of cordycepin afterwards. In the extraction and purification process certification of cordycepin,0.89g cordycepin with the purity of 98.20% could be extracted from 1000g raw material, and the total recovery was confirmed to be 39.83%.Furthermore, the anti-tumor activity of cordycepin was deeply investigated. SRB method was used to detect the inhibition effect of cordycepin on A549, H1792, H1299, H460 and H157 after treated for 24h and 48h, respectively. The results showed that H1299, H460 and H157 were more sensitive to cordycepin and the inhibition effect increased with the extension of treated time. Then, flow cytometry was used to examine the apoptosis of H460 and H1299, and the results showed the apoptotic rate were 29.39% and 36.55%, respectively. Western blot essay demonstrated that the cleavage of Caspase8, Caspase9, Caspase3 and PARP increased both in a dose and time dependent manner which showed that cordycepin induced Caspase-dependent apoptosis in human NSCLC cells. Further study revealed pro-apoptosis protein Noxa and Bax could be up-regulated by cordycepin. Meanwhile, Bcl-2 and Mcl-1, the negative modulator of intrinsic apoptosis, were markedly declined after exposure to cordycepin. Therefore, we speculated cordycepin promoted intrinsic apoptosis in human NSCLC cells. What’s more, we found cordycepin triggered extrinsic apoptosis combined with down-regulation of c-FLIPL which suppressed the activity of Caspase8. And ectopic expression of c-FLIPL dramatically prevented cordycepin-caused apoptosis.In addition, mTOR could participate in the regulation of cell proliferation and the phosphorylation level of mTOR downstream protein S6K1 and 4E-BP1 decreased observably after treated by cordycepin which showed cordycepin could inhibit human NSCLC cell proliferation through the suppression of mTOR signaling pathway. Up-regulation of LC3II and down-regulation of p62 demonstrated that cordycepin induced autophagy in human NSCLC cells. What’s more, mTOR negatively regulated autophagy, so we speculated that cordycepin induced autophagy through suppressing mTOR signaling pathway in lung cancer cells. When autophagy was blocked by Atg5 siRNA or LY294002, the levels of apoptosis caused by cordycepin were obviously attenuated. Therefore, we conclude that cordycepin induces pro-apoptotic autophagy in human NSCLC cells. Taking together, our findings provide a novel prospect on the anti-tumor property of cordycepin, which may further prompt cordycepin to serve as a promising therapeutic approach in NSCLC treatment. |
PDF Full Text Request