Serum Proteomic Analysis Of Gastrointestinal Surgery To Treat Type 2 Diabetes Mellitus

1. BackgroundDiabetes mellitus, gradually, has been one of the most serious society-facing problems. And traditional treatment often can't cure diabetes mellitus effectively. With the development of the obesity surgical treatment, more and more evidence suggest that gastrointestinal surgery not only can cause weigh loss, but can relieve, even cure type 2 diabetes mellitus. And most of these operations could be done with laparoscopy. However, it's still unclear about the detailed mechanisms of the surgical treatment to diabetes. The proteomic approach has offered many opportunities for identifying new therapeutic targets, as well as understanding the mechanisms of the treatment.2. Objectivesâ‘ To get the 2-D gels with a high reproducibility of human sera.â‘¡Using the two-dimensional gel electrophoresis (2-DE) to compare the proteomes expressed in the patients'sera before surgery, one week after surgery and one month after operation. The technology of mass spectrometry (MS) and the methods of bioinformatics were combined to identify the differentially expressed proteins in the sera.â‘¢Two differentially expressed proteins (Rho GDP-dissociation inhibitor 1 and Prohibitin) were further validated to find possible mechanisms of the treatment and to gain new insight into the pathological mechanisms of diabetes formation and development.3. Methodsâ‘ 3 volunteers'sera without diabetes or type 2 diabetes mellitus were collected. Each serum sample was prepared with different method and separated by 2-DE in different condition to find the best method to obtain the 2-D gels with a high reproducibility of human sera.â‘¡12 diabetes patients'sera before surgery, one week after surgery and one month after surgery were collected. Each protein sample was prepared and separated by 2-DE. Protein spots of the gels of before surgery group were compared with that of one week after surgery group and one month after surgery group by 2-DE analysis software, respectively. The common differential protein spots after surgery were selected and analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Peptide mass fingerprints (PMFs) obtained by the MALDI-TOF-MS analysis were used to search SWISS-PROT database by using Mascot software. â‘¢Another 9 diabetes patients'sera before surgery, one week after surgery and one month after surgery were collected.The overexpression of Rho GDP-dissociation inhibitor 1 and Prohibitin was further validated by Western blot analysis.4. Resultsâ‘ 2-D gels of human sera with a high reproducibility were obtained.â‘¡By comparison, there were twenty differentially expressed proteins in the sera after surgery with eight proteins successfully identified by MS. Five proteins were overexpressed after surgery, including Rho GDP-dissociation inhibitor 1, Prohibitin, Alpha-1-antitrypsin precursor, Serotransferrin precursor and Fibrinogen gamma chain precursor. Three proteins were underexpressed after surgery, including MAP3K12-binding inhibitory protein 1,Coronin-1A and Isovaleryl-CoA dehydrogenase.â‘¢Western blot analysis showed the 23 kDa bands of Rho GDP-dissociation inhibitor 1 and the 30 kDa bands of Prohibitin. Sera samples after surgery had higher expression of Rho GDP-dissociation inhibitor 1 and Prohibitin than the samples before surgery.5. ConclusionsThe combination of 2-DE and MS strategy is an ideal platform and powerful approach for differential proteomic analysis of gastrointestinal surgery to treat type 2 diabetes mellitus. Our results of comparative serum proteomics analysis might provide clues for further elucidating the role of these proteins, especially Rho GDP-dissociation inhibitor 1 and Prohibitin, in pathological mechanisms of diabetes formation and development, as well as understanding the mechanisms of the treatment.