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Growth factor and ultrasound-assisted bioreactor synergism for human mesenchymal stem cell chondrogenesis.

Guha Thakurta S, Budhiraja G, Subramanian A - J Tissue Eng (2015)

Bottom Line: Expression of miR-145 was used as a metric to qualitatively assess the efficacy of human mesenchymal stem cell conversion.The combination of growth factor and ultrasound stimulation (group 3) resulted in enhanced COL2A1, SOX-9, and ACAN protein expression when compared to growth factor alone (group 2).No COL10A1 protein expression was noted.

View Article: PubMed Central - PubMed

Affiliation: Chemical & Biomolecular Engineering, University of Nebraska-Lincoln, Lincoln, NE, USA.

ABSTRACT
Ultrasound at 5.0 MHz was noted to be chondro-inductive, with improved SOX-9 gene and COL2A1 protein expression in constructs that allowed for cell-to-cell contact. To achieve tissue-engineered cartilage using macroporous scaffolds, it is hypothesized that a combination of ultrasound at 5.0 MHz and transforming growth factor-β3 induces human mesenchymal stem cell differentiation to chondrocytes. Expression of miR-145 was used as a metric to qualitatively assess the efficacy of human mesenchymal stem cell conversion. Our results suggest that in group 1 (no transforming growth factor-β3, no ultrasound), as anticipated, human mesenchymal stem cells were not efficiently differentiated into chondrocytes, judging by the lack of decrease in the level of miR-145 expression. Human mesenchymal stem cells differentiated into chondrocytes in group 2 (transforming growth factor-β3, no ultrasound) and group 3 (transforming growth factor-β3, ultrasound) with group 3 having a 2-fold lower miR-145 when compared to group 2 at day 7, indicating a higher conversion to chondrocytes. Transforming growth factor-β3-induced chondrogenesis with and without ultrasound stimulation for 14 days in the ultrasound-assisted bioreactor was compared and followed by additional culture in the absence of growth factors. The combination of growth factor and ultrasound stimulation (group 3) resulted in enhanced COL2A1, SOX-9, and ACAN protein expression when compared to growth factor alone (group 2). No COL10A1 protein expression was noted. Enhanced cell proliferation and glycosaminoglycan deposition was noted with the combination of growth factor and ultrasound stimulation. These results suggest that ultrasound at 5.0 MHz could be used to induce chondrogenic differentiation of mesenchymal stem cells for cartilage tissue engineering.

No MeSH data available.


Related in: MedlinePlus

Total glycosaminoglycan (GAG) and collagen (COL) assay: hMSC-seeded scaffolds were cultured for 21 days under mentioned conditions. Total GAG and COL (extrapolated from hydroxyproline content) content per scaffold were estimated at 1, 7, 14, and 21 days. Relative content of (a) GAG to DNA and (b) COL to DNA of group 1, 2, and 3 constructs were plotted. Average with standard deviations are represented (n = 5–8 constructs) (*p < 0.05; **p < 0.01).
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fig5-2041731414566529: Total glycosaminoglycan (GAG) and collagen (COL) assay: hMSC-seeded scaffolds were cultured for 21 days under mentioned conditions. Total GAG and COL (extrapolated from hydroxyproline content) content per scaffold were estimated at 1, 7, 14, and 21 days. Relative content of (a) GAG to DNA and (b) COL to DNA of group 1, 2, and 3 constructs were plotted. Average with standard deviations are represented (n = 5–8 constructs) (*p < 0.05; **p < 0.01).

Mentions: Chondrogenic differentiation of hMSCs seeded on BM scaffolds, cultured for 21 days, was assessed by means of total GAG and collagen content (Figure 5(a) and (b)). Significantly high GAG/DNA (p < 0.01) was observed in group 3 constructs on both day 14 and 21 samples with respect to groups 2 and 3. Collagen/DNA was significantly high for group 3 on day 21 compared to groups 1 and 2.


Growth factor and ultrasound-assisted bioreactor synergism for human mesenchymal stem cell chondrogenesis.

Guha Thakurta S, Budhiraja G, Subramanian A - J Tissue Eng (2015)

Total glycosaminoglycan (GAG) and collagen (COL) assay: hMSC-seeded scaffolds were cultured for 21 days under mentioned conditions. Total GAG and COL (extrapolated from hydroxyproline content) content per scaffold were estimated at 1, 7, 14, and 21 days. Relative content of (a) GAG to DNA and (b) COL to DNA of group 1, 2, and 3 constructs were plotted. Average with standard deviations are represented (n = 5–8 constructs) (*p < 0.05; **p < 0.01).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License 1 - License 2 - License 3
Show All Figures
getmorefigures.php?uid=PMC4300305&req=5

fig5-2041731414566529: Total glycosaminoglycan (GAG) and collagen (COL) assay: hMSC-seeded scaffolds were cultured for 21 days under mentioned conditions. Total GAG and COL (extrapolated from hydroxyproline content) content per scaffold were estimated at 1, 7, 14, and 21 days. Relative content of (a) GAG to DNA and (b) COL to DNA of group 1, 2, and 3 constructs were plotted. Average with standard deviations are represented (n = 5–8 constructs) (*p < 0.05; **p < 0.01).
Mentions: Chondrogenic differentiation of hMSCs seeded on BM scaffolds, cultured for 21 days, was assessed by means of total GAG and collagen content (Figure 5(a) and (b)). Significantly high GAG/DNA (p < 0.01) was observed in group 3 constructs on both day 14 and 21 samples with respect to groups 2 and 3. Collagen/DNA was significantly high for group 3 on day 21 compared to groups 1 and 2.

Bottom Line: Expression of miR-145 was used as a metric to qualitatively assess the efficacy of human mesenchymal stem cell conversion.The combination of growth factor and ultrasound stimulation (group 3) resulted in enhanced COL2A1, SOX-9, and ACAN protein expression when compared to growth factor alone (group 2).No COL10A1 protein expression was noted.

View Article: PubMed Central - PubMed

Affiliation: Chemical & Biomolecular Engineering, University of Nebraska-Lincoln, Lincoln, NE, USA.

ABSTRACT
Ultrasound at 5.0 MHz was noted to be chondro-inductive, with improved SOX-9 gene and COL2A1 protein expression in constructs that allowed for cell-to-cell contact. To achieve tissue-engineered cartilage using macroporous scaffolds, it is hypothesized that a combination of ultrasound at 5.0 MHz and transforming growth factor-β3 induces human mesenchymal stem cell differentiation to chondrocytes. Expression of miR-145 was used as a metric to qualitatively assess the efficacy of human mesenchymal stem cell conversion. Our results suggest that in group 1 (no transforming growth factor-β3, no ultrasound), as anticipated, human mesenchymal stem cells were not efficiently differentiated into chondrocytes, judging by the lack of decrease in the level of miR-145 expression. Human mesenchymal stem cells differentiated into chondrocytes in group 2 (transforming growth factor-β3, no ultrasound) and group 3 (transforming growth factor-β3, ultrasound) with group 3 having a 2-fold lower miR-145 when compared to group 2 at day 7, indicating a higher conversion to chondrocytes. Transforming growth factor-β3-induced chondrogenesis with and without ultrasound stimulation for 14 days in the ultrasound-assisted bioreactor was compared and followed by additional culture in the absence of growth factors. The combination of growth factor and ultrasound stimulation (group 3) resulted in enhanced COL2A1, SOX-9, and ACAN protein expression when compared to growth factor alone (group 2). No COL10A1 protein expression was noted. Enhanced cell proliferation and glycosaminoglycan deposition was noted with the combination of growth factor and ultrasound stimulation. These results suggest that ultrasound at 5.0 MHz could be used to induce chondrogenic differentiation of mesenchymal stem cells for cartilage tissue engineering.

No MeSH data available.


Related in: MedlinePlus