Differences Analysis Of CD34～+ Cells Proteome In Bone Marrow And Mobilized Peripheral Blood

Objective To clarify the mechanism of clinical differences after hematopoietic stem cells transplantation with hematopoietic stem cells from different sources by comparing the differential expression of proteins in bone marrow and mobilized peripheral blood CD34+ cells.Methods 1. Mononuclear cells(MNCs) in BM and MPB were obtained from the malignant hematopathy patients who were in complete remission phase and just before transplantation, MiniMACS immunomagnetic beads technique was used to separate and purify CD34+ cells from MNCs. 2. The whole cell proteins in CD34+cells were extracted by freeze-thaw lysis and aceton deposit, and applied for two-dimensional electrophoresis(2-DE). Gel images were trans-scanned and analyzed with MelanieS image analysis software to find the differential protein spots . 3. Several obviously comparable protein spots were diged out, and digested in gel, then the extracted peptide segments were analyzed with MALDI-TOF mass spectrometry. 4. Peptide mass fingerprints were researched by using data base to define the proteins.Results The average purity of CD34+ cells was (92.33±2.65 ) % in outputs ,and the average cell number of yields was (1.12±0.42)×106. The 2-DE profiles of proteome were satisfying by optimizing the 2-DE techniques, and different expression of proteins was existed between different CD34+ cells and mainly in low molecular weight zone. Several differential protein spots were defined by MS analysis and data base searching, they were: Chain 2: Complement Clr light chain, Ras-related protein R-Ras2, Chain 1: microfibrillar-associated protein, Chain 1: GRPE protein homolog 1, High mobility group protein 4-like and Tob2 protein.Conclusion There are differences of proteins expression between BMand MPB CD34+ cell. After evaluating the proteins, we get the conclusion that the differences between proteins can directly explain the clinical differences in GVHD, relapse, hemolysis etc after BM and MPB hematopoietic stem cells transplantation. It is a essential supplement to similar nucleic acid level investigation. Especially, the same patient's BM and MPB were used in this investigation, so the results are more significant.