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Comparisons of rabbit bone marrow mesenchymal stem cell isolation and culture methods in vitro.

Zhang W, Zhang F, Shi H, Tan R, Han S, Ye G, Pan S, Sun F, Liu X - PLoS ONE (2014)

Bottom Line: 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 4(th) 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 4(th) generation of cells has the strongest proliferation capacity.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiothoracic Surgery, College of Clinical Medicine, Yangzhou University, Yangzhou, Jiangsu Province, China ; The Research Center for Translational Medicine, Yangzhou University, Yangzhou, Jiangsu Province, China.

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 4(th) 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 4(th) generation of cells has the strongest proliferation capacity.

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Comparison of four primary BMSC growth curves.All growth curves form four ell groups had an “S” form and no isolation method was shown to change cell growth.
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pone-0088794-g006: Comparison of four primary BMSC growth curves.All growth curves form four ell groups had an “S” form and no isolation method was shown to change cell growth.

Mentions: Analysis of average absorbances of cells isolated from four methods. All four groups of primary cells began to grow adherently at about 24 h, and the log growth phase of cells was stable around the 4th-9th day. Growth curves of all four cell groups were “S”-type curves. Primary cells reached 80% confluence at day 10 and at day 13 cells were ∼100% confluent (lag phase). Statistically significant differences among the four primary BMSC growth curves were observed (Figure 6). Data show that untreated whole BM blood isolates were superior to the other 3 groups at the 4th, 7th, 10th, and 13th day (p<0.05) (Figure 7).


Comparisons of rabbit bone marrow mesenchymal stem cell isolation and culture methods in vitro.

Zhang W, Zhang F, Shi H, Tan R, Han S, Ye G, Pan S, Sun F, Liu X - PLoS ONE (2014)

Comparison of four primary BMSC growth curves.All growth curves form four ell groups had an “S” form and no isolation method was shown to change cell growth.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3928292&req=5

pone-0088794-g006: Comparison of four primary BMSC growth curves.All growth curves form four ell groups had an “S” form and no isolation method was shown to change cell growth.
Mentions: Analysis of average absorbances of cells isolated from four methods. All four groups of primary cells began to grow adherently at about 24 h, and the log growth phase of cells was stable around the 4th-9th day. Growth curves of all four cell groups were “S”-type curves. Primary cells reached 80% confluence at day 10 and at day 13 cells were ∼100% confluent (lag phase). Statistically significant differences among the four primary BMSC growth curves were observed (Figure 6). Data show that untreated whole BM blood isolates were superior to the other 3 groups at the 4th, 7th, 10th, and 13th day (p<0.05) (Figure 7).

Bottom Line: 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 4(th) 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 4(th) generation of cells has the strongest proliferation capacity.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiothoracic Surgery, College of Clinical Medicine, Yangzhou University, Yangzhou, Jiangsu Province, China ; The Research Center for Translational Medicine, Yangzhou University, Yangzhou, Jiangsu Province, China.

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 4(th) 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 4(th) generation of cells has the strongest proliferation capacity.

Show MeSH
Related in: MedlinePlus