Study On Molecular Mechanisms Of Anti-gastric Cancer Effects Of Rhizoma Amorphophalli Based On Mitochondrial Proteomics

Objective To analysis the influence of Rhizoma Amorphophalli extracts on mitochondrial protein expression of Cell SGC-7901,and identified the differentially expressed proteins by mass spectrometry,then built a network of differentially expressed proteins with software,provided the basis for the study on molecular mechanisms of anti-gastric cancer effects.Methods?The optimum concentration and duration of Rhizoma Amorphophalli extracts on proliferation of Cell SGC-7901 was determined by MTT method;?the influence of Rhizoma Amorphophalli extracts on apoptosis of Cell SGC-7901 was investigated with Flow Cytometry;?2-DE gels of Rhizoma Amorphophalli extracts on Cell SGC-7901 before and after were obtained by two-dimensional gel electrophoresis,then MALDI-TOF-MS peptide mass fingerprinting was used,and differentially expressed proteins were validated with Western-blot method;?interactions of differences protein were analyzed by bioinformatics IPA network.Results ?There were differences among the inhibitory effect on the proliferation of vitro cultured Cell SGC-7901 which was treated by Rhizoma Amorphophalli extracts with differential concentration for 24h.Proliferation of Cell SGC-7901 was not affected when Rhizoma Amorphophalli extracts was lower than of 150?g/ml,while the inhibitory effect was significant when Rhizoma Amorphophalli extracts was higher than of 150?g/ml(P<0.01)especially after 24h(P<0.01).?Rate of Early Apoptotic was improved by 200?g/ml Rhizoma Amorphophalli extracts.?2-DE gels with clear background,high resolution and good reproducibility were obtained by two-dimensional gel electrophoresis.682/643 protein spots were obtained with Image Master 2D Platinum 6.0 software,and 20 differentially expressed proteins which had good repeatability and differences between two groups was greater than or equal to 2 times were identified.?Fingerprints analysis showed that expression content of Zinc finger BED domain-containing protein 5 and Zinc finger protein 562?RNA-binding protein 45?NADH dehydrogenase[ubiquinone]flavoprotein 2?40S ribosomal protein SA?Thyrotropin-releasing hormone-degrading ectoenzyme?Histone acetyltransferase KAT2A?Lysine-specific demethylase 4D?Fumarate hydratase?Peptidyl-prolyl cis-trans isomerase-like 3 was increased while expression content of Enolase 1?3-hydroxyacyl-CoA dehydrogenase type-2?Cyclin B3?ADP-ribosylation factor-like protein 15?GrpE protein homolog 1?CUL7?Ras-related protein Rab-39A?Leucine-rich repeat-containing protein 71?L-lactate dehydrogenase B chain was declined when Cell SGC-7901 was treated by 200?g/ml Rhizoma Amorphophalli extracts for 24h.? Protein interactions network analysis results showed that the above differential proteins could form a protein interaction network,and its main nodal involved of AKT1,ANKH,ARL15,BLVRB,C6orf106,CDKN1A,CNPY2,CUL7,CUL9,ELAVL1,ENY2,FH,FUNDC2,GRPEL1,HSD17B10,KAT2A,KDM4D,KRT2,LDHB,LUZP1,NDUFV2,OBSL1,PPIL3,RAB39A,RBM45,RPS21,RPSA,SLU7,TRHDE,TRMT10C,UBAP2L,UBC,WDHD1,ZBED5,ZNF562,while ubiquitin-conjugating enzymeE2(UBC)located in the center of the network.ConclusionFirstly,Rhizoma Amorphophalli could significantly inhibit the proliferation of cell SGC-7901,and promoted the early apoptosis as well.Secondly,expression content of mitochondrial proteomics were changed when Cell SGC-7901 was treated with Rhizoma Amorphophalli,which inhibited cell proliferation and promoted apoptosis.Thirdly,the possible mechanisms of inhibition of gastric cancer cell proliferation and promoting apoptosis of gastric cancer was as follows:1,tumor cell apoptosis was promoted by regulating protein expression related in the mitochondrial pathway of apoptosis;2,cell maintained homeostasis by regulating protein expression involved in the regulation of gene transcription3,cell cycle progression was inhibited and apoptosis was promoted by regulating protein expression related with the cell cycle regulatory;4,apoptosis of tumor cells was promoted by inducing intracellular oxidative stress response;5,tumor cell proliferation was inhibited by regulating intracellular energy metabolism.Fourthly,apoptosis of tumor cells may be promoted to induce antitumor immunity by Rhizoma Amorphophalli through Ubiquitin conjugating enzyme-mediated mitochondrial apoptotic pathways.