| Objects: Analysis the difference of serum proteome between the normal TAⅡmiceand TAⅡspontaneous breast cancer mice. Combining the technology ofMALDI-TOF-TOFMS and the methods of bioinformatics, we can find and confirmthe special serum differential protein in spontaneous breast cancer, investigate themolecular mechanism of carcinogenesis in TAⅡspontaneous breast cancer.Methods:1 59 TAⅡspontaneous breast cancer mice were collected from 2005 March to 2006October, female.All animals were sacrificed by cervical dislocation, 75% alcoholsterilized skin, dissected tumor and fixed in formalin,embedded in paraffin,stainedin HE.2 We got the peripheral blood from 5 TAⅡspontaneous breast cancer mice and 5normal TAⅡmice. After centrifugating at 5000rps/min in 10 minutes, we got theupper serum to detect in 2-DE. Immobiline pH gradients isoelectric focusing wasused as 1D PH 3-10, and vertical SDS-PAGE as 2D. Coomassie brilliant blue staining,PDQuest softvcare combined with man-made contrast were used to compare the sera ofthe TAⅡspontaneous breast cancer mice and normal TAⅡmice, the protein spotswhich were more than three times difference in two gels were considered asdiffenential protein, and the ID of the differential protein spots were confirmed.3 The differential protein spots which were confirmed by 2-DE, were analyzed byMALDI-TOF-TOFMS. Peptide mass fingerprint though analyzed by m/z software,and interpreted with Mascot (Matrix Science) against the NCBInr database and thenwe analyzed the function of every differential protein and explain the relationship between the proteins and breast cancer.Results:1 In 59 TAⅡspontaneous breast cancer mice, tumor occurred time was 330±95days, parity was 2.7±1.8, predilection sites of tumor in order were left lowerquadrant,left prothorax, right prothorax, right lower quadrant and the nape.Thetumors of 12 mices were multiple,others were single. Tumors were confirmed to beadenocarcinoma by pathology.2 Results of 2-DE showed that 2-DE serum maps of TAⅡspontaneous breast cancermice and normal TAⅡmice were clear and had high resolution, we successfully goteleven differential protein spots, These eleven protein spots high expressed in TAⅡspontaneous breast cancer mice, while down regulated or unexpressed in normal TAⅡmice.3 The differential protein spots which were confirmed by 2-DE were analyzed byMALDI-TOF-TOFMS, and get eleven peptide mass fingerprint (PMF). Searching inMascot protein database, we confirmed five proteins, such as pregnancy zone protein,Paraoxonase, Modification methylase HemK, Haptoglobin, Serum asyloid Pcomponmnet. All of these proteins were increasedly expressed in TAⅡpontaneousbreast cancer mice, but decreasedly expressed or unexpressed in normal TAⅡmice.Conclusions:1 Through serial passage for a long time,Characteristic of spontaneous breast cancerin TAⅡmice was different from previous reports, the heredity genetype andphenotype were changed,and carcinogenesis of spontaneous breast cancer was morestabilized.2 We identified 5 serum differential proteins between normal TAⅡmice and TAⅡspontaneous breast cancer mice by 2-DE combined with MALDI-TOF-TOFMS.Literature reported that there were relationship between these differential proteins and carcinogenesis of breast cancer. In TAⅡspontaneous breast cancer mices,highlyexpressed pregnancy zone protein may be due to the low immune function inspontaneous tumor mice,it may contribute cancer cells escaped from immune system.High level of serum paraoxonase was likely to play a role in the metabolism ofpotential breast carcinogens,and then incease risk in cancer. Modification methylaseHemK was up-expressed in TAⅡspontaneous breast cancer mites which indicatedthere are relationship between abnormal protein methylate and spontaneous breastcancer. Haptoglobin and Serum asyloid P componmnet were up-expressed inTAⅡspontaneous breast cancer mites which may be due to an acute-phase responseinduced by the tumor that contribute to synthesize these acute phase proteins.
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