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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.

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Related in: MedlinePlus

Effects of 3-min LIPUS (30 mW/cm2; 1.5 MHz; 20% duty cycle; 200 μs pulse length) at different concentrations of MB suspension on hMSC proliferation after 24 hrs (Data are mean ± StdEM, n = 9). Values significantly different from the control group are indicated by *for p < 0.05.
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f4: Effects of 3-min LIPUS (30 mW/cm2; 1.5 MHz; 20% duty cycle; 200 μs pulse length) at different concentrations of MB suspension on hMSC proliferation after 24 hrs (Data are mean ± StdEM, n = 9). Values significantly different from the control group are indicated by *for p < 0.05.

Mentions: In order to determine the optimal concentration of MBs for ultrasound excitation studies, varying concentrations of MB suspension was added to the cell media. Following the addition of MBs, LIPUS (30 mW/cm2; 1.5 MHz; 200 μs pulse length; duty cycle 20% i.e. PRF 1 kHz) was applied for three minutes. hMSC proliferation, 24 h after LIPUS stimulation, was examined and the results are presented in Fig. 4. It shows that LIPUS alone increased cell proliferation but not significantly. However, LIPUS-treated cells in the presence of 0.5% (v/v) MB suspension resulted in the highest proliferation rate. Ultrasound in the presence of higher concentrations of MBs decreased hMSC proliferation.


Lipid Coated Microbubbles and Low Intensity Pulsed Ultrasound Enhance Chondrogenesis of Human Mesenchymal Stem Cells in 3D Printed Scaffolds
Effects of 3-min LIPUS (30 mW/cm2; 1.5 MHz; 20% duty cycle; 200 μs pulse length) at different concentrations of MB suspension on hMSC proliferation after 24 hrs (Data are mean ± StdEM, n = 9). Values significantly different from the control group are indicated by *for p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

f4: Effects of 3-min LIPUS (30 mW/cm2; 1.5 MHz; 20% duty cycle; 200 μs pulse length) at different concentrations of MB suspension on hMSC proliferation after 24 hrs (Data are mean ± StdEM, n = 9). Values significantly different from the control group are indicated by *for p < 0.05.
Mentions: In order to determine the optimal concentration of MBs for ultrasound excitation studies, varying concentrations of MB suspension was added to the cell media. Following the addition of MBs, LIPUS (30 mW/cm2; 1.5 MHz; 200 μs pulse length; duty cycle 20% i.e. PRF 1 kHz) was applied for three minutes. hMSC proliferation, 24 h after LIPUS stimulation, was examined and the results are presented in Fig. 4. It shows that LIPUS alone increased cell proliferation but not significantly. However, LIPUS-treated cells in the presence of 0.5% (v/v) MB suspension resulted in the highest proliferation rate. Ultrasound in the presence of higher concentrations of MBs decreased hMSC proliferation.

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.


Related in: MedlinePlus