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

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—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.


Short (1 day)- and long (3 day)-term cell viability assay results of hMSCs incubated with lipid-coated microbubbles (Data are mean ± StdEM, n = 9).
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f3: Short (1 day)- and long (3 day)-term cell viability assay results of hMSCs incubated with lipid-coated microbubbles (Data are mean ± StdEM, n = 9).

Mentions: The cytotoxicity of the lipid-coated MBs synthesized in house was investigated by incubating cells with varying concentrations (0.5, 1, 2, 4, 5 and 10% v/v) of MB suspension for over 72 hours. The viability of the cells incubated with the MB suspension compared to those without MB (0% MB) was determined using the MTS assay and presented in Fig. 3. The results show an increased cell proliferation both after 24 and 72 hour time periods. Due to gas diffusion from the surrounding non-degassed PBS (phosphate buffer saline) during centrifugation, MBs were partially filled with air545556. Observed increased cell proliferation in the presence of MBs might be ascribed to increased oxygen available to the cells. Overall, the results indicated that lipid-coated microbubbles do not cause short-term or long-term cytotoxicity to the cells at the concentrations studied here.


Lipid Coated Microbubbles and Low Intensity Pulsed Ultrasound Enhance Chondrogenesis of Human Mesenchymal Stem Cells in 3D Printed Scaffolds
Short (1 day)- and long (3 day)-term cell viability assay results of hMSCs incubated with lipid-coated microbubbles (Data are mean ± StdEM, n = 9).
© Copyright Policy - open-access
Related In: Results  -  Collection

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

f3: Short (1 day)- and long (3 day)-term cell viability assay results of hMSCs incubated with lipid-coated microbubbles (Data are mean ± StdEM, n = 9).
Mentions: The cytotoxicity of the lipid-coated MBs synthesized in house was investigated by incubating cells with varying concentrations (0.5, 1, 2, 4, 5 and 10% v/v) of MB suspension for over 72 hours. The viability of the cells incubated with the MB suspension compared to those without MB (0% MB) was determined using the MTS assay and presented in Fig. 3. The results show an increased cell proliferation both after 24 and 72 hour time periods. Due to gas diffusion from the surrounding non-degassed PBS (phosphate buffer saline) during centrifugation, MBs were partially filled with air545556. Observed increased cell proliferation in the presence of MBs might be ascribed to increased oxygen available to the cells. Overall, the results indicated that lipid-coated microbubbles do not cause short-term or long-term cytotoxicity to the cells at the concentrations studied here.

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—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.