Comparative Research Of Mesenchymal Stem Cells From Rat Adipose Tissue And Bone Marrow

Mesenchymal stem cells (MSC) represent an archetype of multipotent somatic stem cells that hold promise for application in regenerative medicine. Although MSC were originally isolated from bone marrow, similar populations have been reported in other tissues including adipose tissue. Given the appropriate microenvironment, MSC could differentiate into cells of mesodermal derivation or even cells of nonmesodermal derivation. Many reports suggested that characteristics and multi-potential in MSC isolated from adipose tissues are similar to those in bone marrow. In this study, we compared MSC derived from rat bone marrow and adipose tissue regarding isolation, expansion,idenfication and proteome analysis.Isolation and culture of rat ADSC and BMSC ADSC and BMSC were obtained from the same individual. To isolate rat ADSC, adipose tissues behind rat kidney were digested with 0.25% trypsin and 0.075% collagenaseâ… . At the same time, BMSC from rat thigh-bone were separated by centrifugation in a percoll gradient. Both MSC obtained from rat adipose tissue and bone marrow exhibited a fibroblast-like morphology and expanded easily in vitro. However, ADSC showed a higher proliferation rate and longer survival in culture than BMSC.Idenfication of rat ADSC and BMSC Analysis by flow cytometry of cell surface markers of BMSC and ADSC after there isolation. Both types of MSC were positive for the mesenchymal markers CD29 and CD90.1, but did not express the marker CD49d. Data for the mean fluorescence intensity for the marker CD106 shows difference between both MSC with a mean fluorescence greater for this marker in ADSC than BDSC. To investigate their differentiation potential, ADSC and BMSC were directed toward the osteogenic,adipogenic lineages at the 3 passages. Osteogenic differentiation was confirmed by the detection of an osteogenic phenotype consisting of an increased expression of AP and indicated by the NAP stain. Adipogenic differentiation was demonstrated by the accumulation of neutral lipid vacuoles indicated by the Oil Red O stain. Reverse transcriptase polymerase chain reactions (RT-PCR) determined the expression of a panel of adipocyte and osteoblast associated genes by ADSC and BMSC after 10 days in the appropriate culture condition. The results indicated that under the defined inducing conditions, MSC, isolated from bone marrow and adipose tissue, could differentiate toward the osteogenic and adipogenic lineages in vitro. ADSC showed a higher ability of differentiation than BMSC.Proteome analysis of rat ADSC and BMSC Using the 2-DE technology, 812 spots were mapped in ADSC and 749 spots were mapped in BMSC. By analysis the 2D maps of ADSC and BMSC, 596 spots were matched. 37 protein spots were found to be significant differentially expressed in both MSC and were picked out. To identify these proteins, gel spots were excised and subjected to in-gel trypsin digestion followed by MALDI-TOF MS analysis. The peptide masses obtained were searched in Mascot. Out of the 37 interested protein spots, 18 yielded good Mascot scores(>60) and were considered significant(P<0.05). The identified proteins were either related to cytoskeletal stabilization or stress response or were enzymes of cell metabolism.In these protein spots, structural proteins include Reticulocalbin 3,Ribosomal protein S12,Alpha B-crystallin,Ethylmalonic encephalopathy 1,Tripartite motif protein 9; stress proteins include Rap guanine nucleotideexchange factor (GEF) 4,Reversion induced LIM gene,Chain A, Crystal Structure Of A Mammalian 2-Cys Peroxiredoxin,Heat shock protein 60,Peroxiredoxin 1, Metabolic pathway-associated proteins include Adenylate kinase 1,Peptidylprolyl isomerase A,ATP-dependent RNA helicase,Enoyl Coenzyme A hydratase,ATP synthase. Further analysis suggested ADSC were in more stable and less differentiated conditions with a higher proliferation rate compared to BMSC. The results of comparative proteomic matched what we observed in the culture of ADSC and BMSC.In the present study, we isolated and cultured rat ADSC and BMSC, and compared the their characteristics. Then the proteome analysis revealed some proteins with different expression level in ADSC and BMSC might explain the higher proliferation potential of ADSC . Taking into account all the advantages and disadvantages of both MSC discussed above, further investigations should be continued for their future application.