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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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Related in: MedlinePlus

Chondrogenic differentiation of rabbit BMSCs isolated by four methods.Collagen type II immunostaining showed that the majority of BMSCs were differentiated into a chondrogenic lineage for 21 days (E–H), and were simultaneously cultured in control medium (A–D). Scale bar  = 50 µm, applies to A–D, and scale bar  = 100 µm, applies to E–H.
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pone-0088794-g005: Chondrogenic differentiation of rabbit BMSCs isolated by four methods.Collagen type II immunostaining showed that the majority of BMSCs were differentiated into a chondrogenic lineage for 21 days (E–H), and were simultaneously cultured in control medium (A–D). Scale bar  = 50 µm, applies to A–D, and scale bar  = 100 µm, applies to E–H.

Mentions: Collagen type II immunostaining confirmed that BMSCs isolated by four unique methods were equally suited for synthesis of high levels of type II collagen in chondrogenic differentiation cultures. As shown in Figure 5, no differences in differentiating potential were observed among the four uniquely isolated groups of BMSCs.


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)

Chondrogenic differentiation of rabbit BMSCs isolated by four methods.Collagen type II immunostaining showed that the majority of BMSCs were differentiated into a chondrogenic lineage for 21 days (E–H), and were simultaneously cultured in control medium (A–D). Scale bar  = 50 µm, applies to A–D, and scale bar  = 100 µm, applies to E–H.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0088794-g005: Chondrogenic differentiation of rabbit BMSCs isolated by four methods.Collagen type II immunostaining showed that the majority of BMSCs were differentiated into a chondrogenic lineage for 21 days (E–H), and were simultaneously cultured in control medium (A–D). Scale bar  = 50 µm, applies to A–D, and scale bar  = 100 µm, applies to E–H.
Mentions: Collagen type II immunostaining confirmed that BMSCs isolated by four unique methods were equally suited for synthesis of high levels of type II collagen in chondrogenic differentiation cultures. As shown in Figure 5, no differences in differentiating potential were observed among the four uniquely isolated groups of BMSCs.

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