SELDI-TOF-MS Technology In Researching The Early Proteomics In Nude Mouse Models Of Colorectal Cancer

Objective To build the nude mouse models of HCT-116colorectal cancer in inoculating BALB/C nude mice subcutaneously with colon cancer cell suspension, and observe the changes between the size of the transplanted tumors and the times, and analyze the changes of trace protein profiling in serum of nude mouse models of colorectal cancer after inoculated in different times, search for the tumor-associated differentially expressed proteins, and explore the early diagnostic marker of colorectal cancer.Methods To Recover and cultivate HCT-116colon cancer cell, collect cells in logarithmic growth phase of the3rd or4th generation, adjust cell density into1×107/ml and inoculating in10of3—4weeks’borned nude mice subcutaneously, the dosage was0.1ml each, measure and record the size of the tumor every15days, the control group was also inoculated by RPMI1640culture medium(0.1ml each). The experimental group and the control group were collected the blood sample separately in the20th、40th and60th days after inoculated, using tails-cutting method at the first2times, getting0.1ml blood sample each, use eyeball-taking-off method at the last time, get0.5ml blood sample each, put the nude mice to death and separating the tumor-body, measure the tumors’size, then taking them to the Department of Pathology to have case slice inspection at once. Have the blood samples centrifuged immediately3000rpm in4℃for5min, pack the serum30μL each into3copies and mark on them. Analyze the changes of trace protein profiling in nude mouse models of colorectal cancer through the application of Surface enhanced laser desorption ionization time of flight mass spectrometry(SELDI—TOF-MS), collect data automatically and analyze the expressions of the differentially expressed proteins between experimental group and control group by using Ciphergen Proteinehip3.0and Biomarker Wizard3.1softwares. Screening the colorectal cancer--related trace differentially expressed protein, which would be compared with the data in Protein database.Results The nude mouse models of colorectal cancer were succeed after15days, the tumor formation rate was100%.We could filter out5differentially expressed proteins that had regular changes which were related to the tumor(P<0.01),their molecular weight were2357Da、2424Da、2637Da、8183Da、10133Da. Among them, the2637Da、8182Da、10133Da3kinds of trace proteins turned out to be enhanced expression gradually after nude mouse had been vaccinated, the2357Da、2424Da2kinds decreased expression gradually, and identified as Reactive oxygen species modulator1Leukocyte-specific transcript1protein、Brevinin-2R、Prosalusin、Calcitonin preliminary by retrievaling through the Swiss-Prot protein database. We also could filter out a differentially expressed proteins in the culture medium of the tumor cells, its’molecular weight was4160Da, and identified as WW domain-containing oxidoreductase or Peptide YY. Conclusion1. By building a nude mouse models of HCT-116colorectal cancer in inoculating nude mice subcutaneously with colon cancer cell suspension can be succeed, and has advantages such as easy to operate, a high success rate, the model is stable, which can be great helpful in the study of colorectal cancer.2.It’s effective that researching the early proteomics in nude mouse models of colorectal cancer by using SELDI-TOF-MS Technology, we can find the early serum protein markers of colorectal cancer.3.The new substances we found in cultivating the tumor cells In vitro proved that this isn’t a Isolated process that how the cancer happened but close contacted with outside environment.