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Isolation and characterization of endothelial progenitor cells from Rhesus monkeys.

Sun W, Zheng L, Han P, Kang YJ - Regen Med Res (2014)

Bottom Line: The results showed that nonselective mononuclear EPCs were a better choice for high yield of the target cells.In addition, surface coating of the culture dishes with human fibronectin significantly improved the proliferation and ontogeny of the isolated EPCs.This procedure would help using these valuable cells for regenerative medicine research.

View Article: PubMed Central - PubMed

Affiliation: Regenerative Medicine Research Center, West China Hospital, Sichuan University, Chengdu, Sichuan, 610041 China.

ABSTRACT

Background: Endothelial progenitor cells (EPCs) are increasingly becoming a major focus of regenerative medicine research and practice. The present study was undertaken to establish an appropriate procedure for isolation and characterization of EPCs from Rhesus monkeys for regenerative medicine research.

Result: Selective CD34+ and nonselective mononuclear EPCs were isolated from bone marrow and cultured under varying conditions. The results showed that nonselective mononuclear EPCs were a better choice for high yield of the target cells. The cells grew in M 200 better than in EGM-2, and supplementation with fetal bovine serum promoted cell proliferation; but serum level at 7.5% was better than at 10%. In addition, surface coating of the culture dishes with human fibronectin significantly improved the proliferation and ontogeny of the isolated EPCs. Immunocytochemistry including detection of markers CD34, CD133 and CD31 and double-staining for Ac-LDL and lectin verified the purity of the cultured mononuclear EPCs.

Conclusion: By a thorough analysis, we established a practical procedure for isolation and propagation of EPCs from Rhesus monkeys. This procedure would help using these valuable cells for regenerative medicine research.

No MeSH data available.


Related in: MedlinePlus

The morphology of EPCs isolated from Rhesus monkeys. A. Selected CD34+ mononuclear cells and unselected mononuclear cells were cultured in M 200, observed on day 6 (6D) or day 9 (9D) after culturing and day 4 after the first passage (P1). Both selected CD34+ and unselected mononuclear cells were cerioid shape on day 6, formed cobblestone-appearance colonies on day 9, and become more spindle shape with the loss of stereognosis after first passage. Bar =200 μm. B. Unselected mononuclear cells were cultured in either M 200 or EGM-2 media, observed on day 5 (5D), day 8 (8D) after culturing and day 1 after the first passage (P1). Cells cultured in M 200 proliferated faster than in EGM-2, but cells in EGM-2 had better stereognosis. Bar =100 μm.
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Fig1: The morphology of EPCs isolated from Rhesus monkeys. A. Selected CD34+ mononuclear cells and unselected mononuclear cells were cultured in M 200, observed on day 6 (6D) or day 9 (9D) after culturing and day 4 after the first passage (P1). Both selected CD34+ and unselected mononuclear cells were cerioid shape on day 6, formed cobblestone-appearance colonies on day 9, and become more spindle shape with the loss of stereognosis after first passage. Bar =200 μm. B. Unselected mononuclear cells were cultured in either M 200 or EGM-2 media, observed on day 5 (5D), day 8 (8D) after culturing and day 1 after the first passage (P1). Cells cultured in M 200 proliferated faster than in EGM-2, but cells in EGM-2 had better stereognosis. Bar =100 μm.

Mentions: Mononuclear EPCs isolated from bone marrow by gradient centrifugation were either directly cultured or subjected to CD34-affinity column for further purification. The data presented in Figure 1A show the difference in the morphology of the EPCs in cultures. Adherent CD34+ mononuclear cells grew into cerioid colonies after 5-7 days; cobblestone-appearance colonies were observed after 9-11 days; and spindle shaped colonies existed all the time. On the 4th day after the first passage, which was conducted approximately 2 weeks after the palting, cells grew more uniformly with the increased spindle shaped cells and losing of stereognosis.Figure 1


Isolation and characterization of endothelial progenitor cells from Rhesus monkeys.

Sun W, Zheng L, Han P, Kang YJ - Regen Med Res (2014)

The morphology of EPCs isolated from Rhesus monkeys. A. Selected CD34+ mononuclear cells and unselected mononuclear cells were cultured in M 200, observed on day 6 (6D) or day 9 (9D) after culturing and day 4 after the first passage (P1). Both selected CD34+ and unselected mononuclear cells were cerioid shape on day 6, formed cobblestone-appearance colonies on day 9, and become more spindle shape with the loss of stereognosis after first passage. Bar =200 μm. B. Unselected mononuclear cells were cultured in either M 200 or EGM-2 media, observed on day 5 (5D), day 8 (8D) after culturing and day 1 after the first passage (P1). Cells cultured in M 200 proliferated faster than in EGM-2, but cells in EGM-2 had better stereognosis. Bar =100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4389970&req=5

Fig1: The morphology of EPCs isolated from Rhesus monkeys. A. Selected CD34+ mononuclear cells and unselected mononuclear cells were cultured in M 200, observed on day 6 (6D) or day 9 (9D) after culturing and day 4 after the first passage (P1). Both selected CD34+ and unselected mononuclear cells were cerioid shape on day 6, formed cobblestone-appearance colonies on day 9, and become more spindle shape with the loss of stereognosis after first passage. Bar =200 μm. B. Unselected mononuclear cells were cultured in either M 200 or EGM-2 media, observed on day 5 (5D), day 8 (8D) after culturing and day 1 after the first passage (P1). Cells cultured in M 200 proliferated faster than in EGM-2, but cells in EGM-2 had better stereognosis. Bar =100 μm.
Mentions: Mononuclear EPCs isolated from bone marrow by gradient centrifugation were either directly cultured or subjected to CD34-affinity column for further purification. The data presented in Figure 1A show the difference in the morphology of the EPCs in cultures. Adherent CD34+ mononuclear cells grew into cerioid colonies after 5-7 days; cobblestone-appearance colonies were observed after 9-11 days; and spindle shaped colonies existed all the time. On the 4th day after the first passage, which was conducted approximately 2 weeks after the palting, cells grew more uniformly with the increased spindle shaped cells and losing of stereognosis.Figure 1

Bottom Line: The results showed that nonselective mononuclear EPCs were a better choice for high yield of the target cells.In addition, surface coating of the culture dishes with human fibronectin significantly improved the proliferation and ontogeny of the isolated EPCs.This procedure would help using these valuable cells for regenerative medicine research.

View Article: PubMed Central - PubMed

Affiliation: Regenerative Medicine Research Center, West China Hospital, Sichuan University, Chengdu, Sichuan, 610041 China.

ABSTRACT

Background: Endothelial progenitor cells (EPCs) are increasingly becoming a major focus of regenerative medicine research and practice. The present study was undertaken to establish an appropriate procedure for isolation and characterization of EPCs from Rhesus monkeys for regenerative medicine research.

Result: Selective CD34+ and nonselective mononuclear EPCs were isolated from bone marrow and cultured under varying conditions. The results showed that nonselective mononuclear EPCs were a better choice for high yield of the target cells. The cells grew in M 200 better than in EGM-2, and supplementation with fetal bovine serum promoted cell proliferation; but serum level at 7.5% was better than at 10%. In addition, surface coating of the culture dishes with human fibronectin significantly improved the proliferation and ontogeny of the isolated EPCs. Immunocytochemistry including detection of markers CD34, CD133 and CD31 and double-staining for Ac-LDL and lectin verified the purity of the cultured mononuclear EPCs.

Conclusion: By a thorough analysis, we established a practical procedure for isolation and propagation of EPCs from Rhesus monkeys. This procedure would help using these valuable cells for regenerative medicine research.

No MeSH data available.


Related in: MedlinePlus