Limits...
Combined influence of basal media and fibroblast growth factor on the expansion and differentiation capabilities of adipose-derived stem cells.

Ahearne M, Lysaght J, Lynch AP - Cell Regen (Lond) (2014)

Bottom Line: FGF has a negative effective on passage 4 ASC adipogenesis and high glucose media plus FGF-enhanced osteogenic capacity of passage 4 ASCs.High glucose media plus FGF appeared to be the most beneficial for priming ASCs to induce a keratocyte phenotype.These findings demonstrate the reciprocal effect FGF and basal media have on ASCs.

View Article: PubMed Central - PubMed

Affiliation: Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland ; Department of Mechanical and Manufacturing Engineering, School of Engineering, Trinity College Dublin, Dublin, Ireland.

ABSTRACT

Background: Interest in adipose-derived stem cells (ASCs) has increased in recent years due to their multi-linage differentiation capabilities. While much work has been done to optimize the differentiation media, few studies have focused on examining the influence of different expansion media on cell behavior. In this study, three basal media (low glucose Dulbecco's modified Eagle's medium (DMEM), high glucose DMEM and DMEM-F12) supplemented with or without fibroblast growth factor 2 (FGF) were examined to assess their suitability for expanding ASCs.

Findings: Flow cytometry, colony-forming unit assays (CFU-Fs) and differentiation assays were utilized to study cell behavior. High glucose media CFU-Fs produced fewest colonies while the addition of FGF increased colony size. By passage 2, the majority of cells were positive for CD44, 45, 73, 90 and 105 and negative for CD14, 31 and 45, indicating a mesenchymal phenotype. A sub-population of CD34 positive cells was present among passage 2 cells; however, by passage 4 the cells were negative for CD34. FGF has a negative effective on passage 4 ASC adipogenesis and high glucose media plus FGF-enhanced osteogenic capacity of passage 4 ASCs. FGF supplemented basal media were most suitable for chondrogenesis. High glucose media plus FGF appeared to be the most beneficial for priming ASCs to induce a keratocyte phenotype.

Conclusions: These findings demonstrate the reciprocal effect FGF and basal media have on ASCs. This research has implications for those interested regenerating bone, cartilage, cornea or adipose tissues.

No MeSH data available.


Related in: MedlinePlus

ALDH3A1, αSMA, collagen I and collagen III gene expressions of P2 ASCs cultured from two different donors relative to LG after culture for 14 days in keratocyte differentiation media.n = 3, *significant difference relative to LG, p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4325938&req=5

Fig4: ALDH3A1, αSMA, collagen I and collagen III gene expressions of P2 ASCs cultured from two different donors relative to LG after culture for 14 days in keratocyte differentiation media.n = 3, *significant difference relative to LG, p < 0.05.

Mentions: Following culture in a keratocyte differentiation media for 14 days, ASCs cultured in HG + FGF expressed the highest levels of the keratocyte marker ALDH3A1 lowest levels of the fibrotic markers alpha smooth muscle actin (αSMA/ACTA2), collagen I (COL1A1) and collagen III (COL3A1) (Figure 4). Keratocytes are quiescent cells native to the corneal stroma that are believed to be derived from the neural crest [18] and are characterized by the presence of ALDH3A1 and the absence of αSMA/ACTA2. Upon injury to the cornea, keratocytes become activated and may exhibit fibroblastic or myofibroblastic characteristics. These are normally associated with the loss of corneal transparency and the formation of fibrotic scar tissue. Fibroblastic or myofibroblastic activity result in a downregulation of ALDH3A1 and upregulation of COL1A1, COL3A1 and αSMA/ACTA2 [19, 20]. Gene expression analysis in this study suggests that ASCs expanded in HG + FGF had the most similar phenotype to native quiescent corneal keratocytes.Figure 4


Combined influence of basal media and fibroblast growth factor on the expansion and differentiation capabilities of adipose-derived stem cells.

Ahearne M, Lysaght J, Lynch AP - Cell Regen (Lond) (2014)

ALDH3A1, αSMA, collagen I and collagen III gene expressions of P2 ASCs cultured from two different donors relative to LG after culture for 14 days in keratocyte differentiation media.n = 3, *significant difference relative to LG, p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4325938&req=5

Fig4: ALDH3A1, αSMA, collagen I and collagen III gene expressions of P2 ASCs cultured from two different donors relative to LG after culture for 14 days in keratocyte differentiation media.n = 3, *significant difference relative to LG, p < 0.05.
Mentions: Following culture in a keratocyte differentiation media for 14 days, ASCs cultured in HG + FGF expressed the highest levels of the keratocyte marker ALDH3A1 lowest levels of the fibrotic markers alpha smooth muscle actin (αSMA/ACTA2), collagen I (COL1A1) and collagen III (COL3A1) (Figure 4). Keratocytes are quiescent cells native to the corneal stroma that are believed to be derived from the neural crest [18] and are characterized by the presence of ALDH3A1 and the absence of αSMA/ACTA2. Upon injury to the cornea, keratocytes become activated and may exhibit fibroblastic or myofibroblastic characteristics. These are normally associated with the loss of corneal transparency and the formation of fibrotic scar tissue. Fibroblastic or myofibroblastic activity result in a downregulation of ALDH3A1 and upregulation of COL1A1, COL3A1 and αSMA/ACTA2 [19, 20]. Gene expression analysis in this study suggests that ASCs expanded in HG + FGF had the most similar phenotype to native quiescent corneal keratocytes.Figure 4

Bottom Line: FGF has a negative effective on passage 4 ASC adipogenesis and high glucose media plus FGF-enhanced osteogenic capacity of passage 4 ASCs.High glucose media plus FGF appeared to be the most beneficial for priming ASCs to induce a keratocyte phenotype.These findings demonstrate the reciprocal effect FGF and basal media have on ASCs.

View Article: PubMed Central - PubMed

Affiliation: Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland ; Department of Mechanical and Manufacturing Engineering, School of Engineering, Trinity College Dublin, Dublin, Ireland.

ABSTRACT

Background: Interest in adipose-derived stem cells (ASCs) has increased in recent years due to their multi-linage differentiation capabilities. While much work has been done to optimize the differentiation media, few studies have focused on examining the influence of different expansion media on cell behavior. In this study, three basal media (low glucose Dulbecco's modified Eagle's medium (DMEM), high glucose DMEM and DMEM-F12) supplemented with or without fibroblast growth factor 2 (FGF) were examined to assess their suitability for expanding ASCs.

Findings: Flow cytometry, colony-forming unit assays (CFU-Fs) and differentiation assays were utilized to study cell behavior. High glucose media CFU-Fs produced fewest colonies while the addition of FGF increased colony size. By passage 2, the majority of cells were positive for CD44, 45, 73, 90 and 105 and negative for CD14, 31 and 45, indicating a mesenchymal phenotype. A sub-population of CD34 positive cells was present among passage 2 cells; however, by passage 4 the cells were negative for CD34. FGF has a negative effective on passage 4 ASC adipogenesis and high glucose media plus FGF-enhanced osteogenic capacity of passage 4 ASCs. FGF supplemented basal media were most suitable for chondrogenesis. High glucose media plus FGF appeared to be the most beneficial for priming ASCs to induce a keratocyte phenotype.

Conclusions: These findings demonstrate the reciprocal effect FGF and basal media have on ASCs. This research has implications for those interested regenerating bone, cartilage, cornea or adipose tissues.

No MeSH data available.


Related in: MedlinePlus