Serum Comparative Proteomic Analysis Of Pediatric B- And T-cell Acute Lymphoblastic Leukemia

ObjectiveUsing Isobaric tags for relative and absolute quantification(i TRAQ)quantitative proteomic technique combined with two-dimensional liquid chromatography tandem mass spectrometry(2DLC-MS/MS)and IPA method we screen the differences in serum of pediatric B-and T-cell Acute Lymphoblastic Leukemia(ALL)to investigate classificatory diagnosis of ALL,the therapeutic target and pathogenesis of T-cell Acute Lymphoblastic Leukemia(T-ALL).Methods1.Serum were collected assigned into acute B-cell acute lymphoblastic leukemia(B-ALL)group and T-ALL group with 20 cases for each group.2.Equal volumes of serum from 2 groups were pooled,respectively.Samples from the 2 groups were depleted of 14 high abundance proteins labeled with i TRAQ tags and then analyzed by 2DLC-MS/MS.We take reverse database to estimate its false positive rate.The i TRAQ experiment is replicated by 3 times.3.According to identificating proteins i TRAQ relative quantitative values to ensure differential proteins between T-ALL and B-ALL groups.At last,the differences in serum proteins were further analyzed by IPA database and get partial signaling pathways and networks.4.The proteomic results were validated by enzyme-linked immunosorbent assay.Results1.The final result shows 468 proteins,which could be used for quantification.With 1.2 and 0.8 as the up-regulation and down-regulation cutoff point for differential proteins in i TRAQ experiment.2.There are 38 differentially expressed proteins in T-ALL group as oppose to B-ALL group,among them,31 proteins are up-regulation and 7 down-regulation.These differentially expressed proteins are related to the function of Cardiovascular Disease ? Lipid Metabolism ? Molecular Transport ? Small Molecular Biochemistry?Protein Synthesis?Cellular Movement?Hematological System Development and Function?Immune Cell Trafficking and Inflammatory Response.3.There are 8 key changing pathways,include LXR/RXR Activation ?Atherosclerosis Signaling?IL-12 Signaling and Production in Macrophages?Production of Nitric Oxide and Reactive Oxygen Species in Macrophages ?Clathrin-mediated Endocytosis Signaling?Acute Phase Response Signaling?FXR/RXR Activation and Complement System.4.There are 3 protein-protein interaction networks,include Lipid Metabolism/Molecular Transport/Small Molecular Biochemistry ? Cellular Movement/Hematological System Development and Function/Immune Cell Trafficking and Cell Death and Survival/Hematological System Development and Function/Cancer.5.ELISA results indicated that s L-selectin was significantly up-regulated in serum of childhood T-ALL compared to B-ALL group.Conclusion1.The iTRAQ-2DLC-MS/MS technology,with the characteristics of easy operation,good repeatability,fast and high throughput of protein changes,has been widely used to tag peptides for multiplexed protein quantification and can be applied to look for different protein between T-ALL and B-ALL.2.IPA analysis can further looking for different proteins related with T-ALL and the interaction networks of signaling pathways.It may be helpful to therapeutic targets for children with T-ALL,and it may provide experimental basis for molecular mechanisms of childhood T-ALL.3.The expression of sL-selectin related to the pathogenesis of childhood T-ALL.L-selectin may be serve as a new potential therapeutic target.