Ultrasound Induces Similar Temporal Endothelial Expression Patterns of eNOS and KLF2 as Normal Flow

Jaideep Sahni; Ian S McCue; Adam R Johnson; Morgan A Schake; Luz D Sotelo; Joseph A Turner; Ryan M Pedrigi

doi:10.1016/j.ultrasmedbio.2024.08.017

Ultrasound Induces Similar Temporal Endothelial Expression Patterns of eNOS and KLF2 as Normal Flow

Ultrasound Med Biol. 2024 Sep 20:S0301-5629(24)00328-4. doi: 10.1016/j.ultrasmedbio.2024.08.017. Online ahead of print.

Authors

Jaideep Sahni¹, Ian S McCue¹, Adam R Johnson¹, Morgan A Schake¹, Luz D Sotelo¹, Joseph A Turner¹, Ryan M Pedrigi²

Affiliations

¹ Department of Mechanical and Materials Engineering, University of Nebraska-Lincoln, Lincoln, NE, USA.
² Department of Mechanical and Materials Engineering, University of Nebraska-Lincoln, Lincoln, NE, USA; Nebraska Center for Integrated Biomolecular Communication, University of Nebraska-Lincoln, Lincoln, NE, USA; Department of Cellular and Integrative Physiology, University of Nebraska Medical Center, Omaha, NE, USA. Electronic address: rpedrigi@unl.edu.

PMID: 39306482
DOI: 10.1016/j.ultrasmedbio.2024.08.017

Abstract

Objective: To determine the sensitivity of vascular endothelial cells to long durations of low-intensity pulsed ultrasound (LIPUS) compared to normal flow and identify the duration that maximizes expression of two mechanosensitive genes related to healthy endothelial function, endothelial nitric oxide synthase (eNOS) and Krüppel-like factor 2 (KLF2).

Methods: Custom ultrasound exposure tanks were developed and the acoustic field was characterized. Human umbilical vein endothelial cells were seeded into culture plates and exposed to LIPUS at a frequency of 1 MHz and acoustic pressure of 217 kPa for 20 min, 1 h, 6 h, 9 h, or 24 h. As a comparator, other cells were exposed to normal flow. RT-qPCR was used to assess mRNA expression of eNOS and KLF2.

Results: Maximum eNOS and KLF2 expression occurred at 6 h and was localized to the beam path. Both genes exhibited qualitatively similar patterns of expression under LIPUS compared to normal flow. LIPUS induced a more rapid beneficial response compared to normal flow, but flow induced higher expression of both genes. eNOS expression after 6 h of LIPUS was dependent on RNA yield and culture duration prior to experiments.

Conclusion: Endothelial cells exposed to longer durations of LIPUS than typically employed exhibited greater expression of beneficial genes. The temporal gene expression patterns resulting from LIPUS and normal flow suggest activation of similar signaling pathways. However, LIPUS also caused increased RNA yield that may be linked to proliferation, which would suggest more of a wound healing than atheroprotective phenotype.

Keywords: Acoustic pressure; Atherosclerosis; Biomechanics; Endothelial cells; Low-intensity pulsed ultrasound; Shear stress; Wound healing.