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Lipid Coated Microbubbles and Low Intensity Pulsed Ultrasound Enhance Chondrogenesis of Human Mesenchymal Stem Cells in 3D Printed Scaffolds

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ABSTRACT

Lipid-coated microbubbles are used to enhance ultrasound imaging and drug delivery. Here we apply these microbubbles along with low intensity pulsed ultrasound (LIPUS) for the first time to enhance proliferation and chondrogenic differentiation of human mesenchymal stem cells (hMSCs) in a 3D printed poly-(ethylene glycol)-diacrylate (PEG-DA) hydrogel scaffold. The hMSC proliferation increased up to 40% after 5 days of culture in the presence of 0.5% (v/v) microbubbles and LIPUS in contrast to 18% with LIPUS alone. We systematically varied the acoustic excitation parameters—excitation intensity, frequency and duty cycle—to find 30 mW/cm2, 1.5 MHz and 20% duty cycle to be optimal for hMSC proliferation. A 3-week chondrogenic differentiation results demonstrated that combining LIPUS with microbubbles enhanced glycosaminoglycan (GAG) production by 17% (5% with LIPUS alone), and type II collagen production by 78% (44% by LIPUS alone). Therefore, integrating LIPUS and microbubbles appears to be a promising strategy for enhanced hMSC growth and chondrogenic differentiation, which are critical components for cartilage regeneration. The results offer possibilities of novel applications of microbubbles, already clinically approved for contrast enhanced ultrasound imaging, in tissue engineering.

No MeSH data available.


Three-week hMSC (a) GAG production and (b) type II collagen. (Data are mean ± StdEM, n = 5). Values significantly different from the control group are indicated by *for p < 0.05 and **for p < 0.01.
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f7: Three-week hMSC (a) GAG production and (b) type II collagen. (Data are mean ± StdEM, n = 5). Values significantly different from the control group are indicated by *for p < 0.05 and **for p < 0.01.

Mentions: In our previous investigation of hMSC growth and chondrogenic differentiation without MBs, scaffolds with square pore geometry performed better than those with hexagonal pore geometry47. Therefore, scaffolds with square pore geometry were chosen here for hMSC chondrogenic differentiation evaluations. We divided the samples into three groups: control group (no LIPUS, no MB), LIPUS only and LIPUS + MB group. Seeded 3D-printed PEGDA scaffolds were evaluated for GAG and type II collagen after three weeks of culture. Glycosaminoglycan (GAG) and collagen type II, which are two key components of a cartilage matrix, were measured using standard assay kits following manufacturer’s instructions. Figure 7(a) shows that all LIPUS treated samples exhibited an increase in GAG production; however, the increase is significantly higher upon incorporation of MBs when compared to the controls. Samples that underwent MB assisted ultrasound excitation exhibited 17% increase in GAG production after 3 weeks. However, samples treated with LIPUS only, had a 5% increase in GAG production after 3 weeks compared to the controls.


Lipid Coated Microbubbles and Low Intensity Pulsed Ultrasound Enhance Chondrogenesis of Human Mesenchymal Stem Cells in 3D Printed Scaffolds
Three-week hMSC (a) GAG production and (b) type II collagen. (Data are mean ± StdEM, n = 5). Values significantly different from the control group are indicated by *for p < 0.05 and **for p < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5121887&req=5

f7: Three-week hMSC (a) GAG production and (b) type II collagen. (Data are mean ± StdEM, n = 5). Values significantly different from the control group are indicated by *for p < 0.05 and **for p < 0.01.
Mentions: In our previous investigation of hMSC growth and chondrogenic differentiation without MBs, scaffolds with square pore geometry performed better than those with hexagonal pore geometry47. Therefore, scaffolds with square pore geometry were chosen here for hMSC chondrogenic differentiation evaluations. We divided the samples into three groups: control group (no LIPUS, no MB), LIPUS only and LIPUS + MB group. Seeded 3D-printed PEGDA scaffolds were evaluated for GAG and type II collagen after three weeks of culture. Glycosaminoglycan (GAG) and collagen type II, which are two key components of a cartilage matrix, were measured using standard assay kits following manufacturer’s instructions. Figure 7(a) shows that all LIPUS treated samples exhibited an increase in GAG production; however, the increase is significantly higher upon incorporation of MBs when compared to the controls. Samples that underwent MB assisted ultrasound excitation exhibited 17% increase in GAG production after 3 weeks. However, samples treated with LIPUS only, had a 5% increase in GAG production after 3 weeks compared to the controls.

View Article: PubMed Central - PubMed

ABSTRACT

Lipid-coated microbubbles are used to enhance ultrasound imaging and drug delivery. Here we apply these microbubbles along with low intensity pulsed ultrasound (LIPUS) for the first time to enhance proliferation and chondrogenic differentiation of human mesenchymal stem cells (hMSCs) in a 3D printed poly-(ethylene glycol)-diacrylate (PEG-DA) hydrogel scaffold. The hMSC proliferation increased up to 40% after 5 days of culture in the presence of 0.5% (v/v) microbubbles and LIPUS in contrast to 18% with LIPUS alone. We systematically varied the acoustic excitation parameters&mdash;excitation intensity, frequency and duty cycle&mdash;to find 30&thinsp;mW/cm2, 1.5&thinsp;MHz and 20% duty cycle to be optimal for hMSC proliferation. A 3-week chondrogenic differentiation results demonstrated that combining LIPUS with microbubbles enhanced glycosaminoglycan (GAG) production by 17% (5% with LIPUS alone), and type II collagen production by 78% (44% by LIPUS alone). Therefore, integrating LIPUS and microbubbles appears to be a promising strategy for enhanced hMSC growth and chondrogenic differentiation, which are critical components for cartilage regeneration. The results offer possibilities of novel applications of microbubbles, already clinically approved for contrast enhanced ultrasound imaging, in tissue engineering.

No MeSH data available.