Comparisons of Rabbit Bone Marrow Mesenchymal Stem Cell Isolation and Culture Methods In Vitro

Abstract

Bone marrow mesenchymal stem cells (BMSCs) have great potential in tissue engineering and clinical therapy, and various methods for isolation and cultivation of BMSCs have been reported. However, the best techniques are still uncertain. Therefore, we sought the most suitable among the four most common methods for BMSC separation from rabbits. BMSCs were obtained from untreated whole bone marrow (BM) adherent cultures, 3 volumes of red blood cells (RBC) lysed with ammonium chloride, 6 volumes of RBC lysed with ammonium chloride, and Ficoll density gradient centrifugation. Then, isolated BMSCs were evaluated with respect to primary cell yield, number of CFU-F colonies, proliferative capacity, cell phenotype, and chondrogenic differentiation potential. Our data show that BMSCs were successfully isolated by all four methods, and each method was similar with regard to cell morphology, phenotype, and differentiation potential. However, BMSCs from untreated whole BM adherent cultures had greater primary cell yields, larger colonies, and the shortest primary culture time (P<0.05). Moreover, the 4th generation of cultured cells had the strongest proliferative activity, the fastest growth rate and the most numerous cells compared with other cell passage generations (P<0.05). In conclusion, untreated whole BM adherent cultures are best for rabbit BMSC isolation and the 4th generation of cells has the strongest proliferation capacity.

The potential of 3-dimensional construct engineered from poly(lactic-co-glycolic acid)/fibrin hybrid scaffold seeded with bone marrow mesenchymal stem cells for in vitro cartilage tissue engineering

“…This indicates that the BMSCs were capable of producing major chondrogenic matrices (Leyh et al, 2014;Mauck et al, 2006;Miao et al, 2009;Zhang et al, 2014). Initially, we anticipated that collagen type I would gradually decline towards week 3 as demonstrated by the collagen type I expression on RT-PCR.…”

Comparison of Bone Marrow-derived Mononuclear Cells vs. Mesenchymal Stem Cells in Protecting Random-pattern Skin Flap Survival in Rats

“…Although BMSCs area relatively pure population that might be good for reducing cell-related side-effects, other pro-angiogenic components, such as endothelial progenitor cells and hematopoietic cells [27], are lost during expansion (Figure 3). This might be one reason why no further improvement were observed following BMSC treatment compared with BM-MNC treatment.…”

“…The current standard protocol for BMSC culture is widely used but not ideal [27,29]. Cells are reported to lose their multipotent differentiation potential in later passages [27].…”

Cellular behavior as a dynamic field for exploring bone bioengineering: A closer look at cell–biomaterial interface

“…Although much is known of the potential use of mesenchymal stem cells in regenerative medicine, this promise still depends on more elaborate and assertive studies regarding the identification of the mechanisms and molecules, which control and mediate the differentiation of a specific lineage, the establishment of these cells in a certain tissue of interest and the signaling cascades, which control the survival and proliferation of the cells [62]. For the use of stem cells not to result in further problems for the patient, these questions are eminent and require answers.…”