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Fabricating and Labeling Microbubbles with Fluorescent and Radioactive Tracers
Published on: January 24, 2025
Interaction with leukocytes: phospholipid-stabilized versus albumin-shell microbubbles
Hiroto Takeuchi1, Koji Ohmori, Isao Kondo
1Second Department of Internal Medicine, Kagawa University School of Medicine, 1750-1, Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0793, Japan.
Purpose:
To confirm that BR14 microbubbles (MBs) can be phagocytosed by activated leukocytes, to determine their stability after phagocytosis, and to evaluate how such characteristics influence the fate of neutrophils containing MBs after insonation.
Materials And Methods:
BR14 and human albumin MBs (2 x 10(7)/mL) were incubated with activated human neutrophils (2 x 10(6)/mL) to allow phagocytosis. Deflation rate of the phagocytosed MBs after pulsed insonation (one burst per second for 5 seconds) at 1.8 MHz with peak negative pressure of -540 kPa or -1,340 kPa, lactate dehydrogenase (LDH) leakage, and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling stain-positive cell count after insonation were compared between the two agents.
Results:
At -540 kPa, phagocytosed MBs remained nearly unchanged for both agents after insonation. At -1,340 kPa, although human albumin MBs were disrupted on the first or second burst, BR14 MBs remained undisrupted. After -540-kPa insonation, a similar number of apoptotic cells appeared in neutrophils containing human albumin and BR14 MBs. At -540 kPa, LDH leakage was limited in human albumin MBs and BR14 MBs. At -1,340 kPa, LDH leakage was significantly increased in human albumin MBs and BR14 MBs (P <.01, both vs -540 kPa). Apoptotic cells were significantly decreased in human albumin MBs and BR14 MBs (P <.01, both vs -540 kPa). LDH leakage was lower and apoptotic cell count was greater in BR14 MB-containing neutrophils than in human albumin MB-containing neutrophils (both P <.01).
Conclusion:
Compared with human albumin MBs, BR14 MBs were more stable after phagocytosis with insonation. This stability is associated with less disruption and greater induction of apoptosis in leukocytes after relatively high-pressure insonation in the range for diagnostic use.
Insights
BR14 microbubbles (MBs) are more stable than albumin MBs after phagocytosis and insonation. This enhanced stability in leukocytes leads to less disruption and more apoptosis, particularly at diagnostic pressure ranges.
Area of Science:
- Biomedical Engineering
- Cell Biology
- Pharmacology
Background:
- Microbubbles (MBs) are utilized in various biomedical applications, including drug delivery and diagnostics.
- Understanding the behavior of MBs within cellular environments, particularly after phagocytosis by immune cells like neutrophils, is crucial for optimizing their therapeutic potential.
- Leukocyte phagocytosis of MBs can influence their stability and subsequent response to external stimuli such as ultrasound insonation.
Purpose of the Study:
- To investigate the phagocytosis of BR14 microbubbles by activated human neutrophils.
- To assess the stability of BR14 microbubbles within neutrophils after ultrasound insonation.
- To evaluate the impact of BR14 microbubble stability on neutrophil fate, including cell death and membrane integrity, post-insonation.
Main Methods:
- Human neutrophils were incubated with BR14 and human albumin microbubbles to facilitate phagocytosis.
- Phagocytosed microbubbles were subjected to pulsed ultrasound insonation at varying peak negative pressures (-540 kPa and -1,340 kPa).
- Microbubble deflation rates, lactate dehydrogenase (LDH) leakage (indicating cell membrane damage), and apoptosis (via TUNL staining) were measured.
Main Results:
- BR14 microbubbles demonstrated superior stability compared to human albumin microbubbles under high-pressure insonation (-1,340 kPa) after phagocytosis.
- While both microbubble types showed limited LDH leakage at -540 kPa, significant leakage occurred at -1,340 kPa for both.
- Neutrophils containing BR14 microbubbles exhibited lower LDH leakage and a higher rate of apoptosis compared to those with human albumin microbubbles at -1,340 kPa.
Conclusions:
- BR14 microbubbles exhibit enhanced stability post-phagocytosis and insonation compared to human albumin microbubbles.
- This increased stability is linked to reduced cellular disruption and a greater induction of apoptosis in leukocytes following insonation.
- The findings suggest BR14 microbubbles may offer advantages for diagnostic ultrasound applications involving leukocyte interactions.
