Inhibition Of Apoptosis By Two Insect Hemolymph And Analysis Of Three Caspases In Sl-1 Cells

Apoptosis is one of the most popular field in lifo science,the mechanisms of apoptosis in mammalian cells has been studied quite clearly,but the insects apoptosis still have many issues to be considered.This articial will be discussed two topics related to apoptosis.1 Two insect hemolymph's inhibition effect on apoptosis in Spodoptera litura cellsSeveral studies have shown that some hemolymph inhibit not only virus-induced apoptosis but also apoptosis induced by various chemicals such as actinomycin D, camptothecin,staurosporine and sodium butyrate.Moreover,they also inhibit human cell apoptosis.In this study we investigate the inhibition of actinomycin-D-induced and virus-induced apoptosis in S1-1 cells with two insect hemolymph.The main results include:DAPI stain,the rate of apoptotic cells,Caspase-3 activity and release of cytchrome c.S1-1 cells were cultivated in a Grace's medium supplemented with hemolymph of Helicoverpa armigera and Spodoptera exigua at various concentrations(1%,3%,5 %)and 5%fetal bovine serum.Hemolymph can obviously inhibit apoptosis induced by actinomycin D.The result indicated that higher hemolymph concentration showed more positive effect in inhibition of apoptosis,and Spodoptera exigua hemolymph did more effectively than Helicoverpa armigera hemolymph.Preparation of S1-lcells lysis ingredients induced by actinomycin-D for 24 h,using AC-DEVD-AFC to digestion,the result of fluorospectrophotometry shows that both of two insect hemolymph can inhibit the activity of Caspase-3,and the inhibition of Spodoptera exigua expressed more obvious than Helicoverpa armigera.The results by western blotting showed that:there is the release of Cyt c in ActD-S1-1-induced poptosis.Insect hemolymph can also inhibit the release of Cyt c significantly.The inhibition of Spodoptera exigua expressed more obvious than Helicoverpa armigera.At the same time,both of two insect hemolymph can inhibit the activity of Caspase-3 induced by AfMNPV as well.Based on the results of the study,we prove that insect hemolymph inhibit apoptosis extensivly.2 Caspases activity and their relationship induced by AfMNPV in Spodoptera litura cellsIn this study,we investigate the activity of Caspase-3,8,9 and Caspases cascades,and the existence of Caspase-8 and Caspase-9-mediated signaling pathway.S1-1 cells were induced by AfMNPV supplemented with caspse-8 specific fluorescent substrate(Z-IETD-FMK)and caspse-9 specific fluorescent substrate(Z-LEHD-FMK)at various concentrations.The activation of capases in AfMNPV-induced cells was determined in caspses specific fluorescent substrate.The results showed that Caspase-3 activity was measured and not gradually decreased with increased of Caspase-8 inhibitor extension,but decreased with increased of Caspase-9 inhibitor extension.However, activity of Caspase-8 and Caspase-9 had not been tested.Meanwhile,we detected activity of Caspase-8,but not activity of Caspase-9 in positive control.These data indicate that Caspase-8-mediated apoptosis signaling pathway may not exist in S1-1 cells apoptosis. As to whether there is Caspase-9-mediated apoptosis signaling pathway in S1-1 cells apoptosis,further research will be needed.