Connolly, T. | bioRxiv Science

bioRxiv2025

Barium titanate piezoelectric nanoparticles induce M1 polarization in mouse macrophages via ultrasound in vitro

Macrophages are critical for the maintenance of immune system homeostasis. They differentiate into distinct functional populations, from pro-to anti-inflammatory phenotype, exhibiting remarkable biological plasticity and responding to both chemical and physical cues to achieve these phenotypes. Controlling macrophage cell phenotypes in vivo, with temporal and spatial control, could have significant impact on a wide range of human diseases and ailments associated with inflammation, which range from rheumatoid arthritis and Alzheimers to cancer tumorigenesis. Piezoelectrics, materials in which pressure causes a voltage and vice versa, represent a potential platform for non-invasive and remote modulation of cells and tissues and, in particular, control of immune cell activation. Here, it is demonstrated that RAW264.7 mouse macrophage cells that have taken up piezoelectric nanoparticles (pzNPs) specifically adopt an M1 cellular phenotype and requisite calcium ion influx upon ultrasound stimulation. One can further identify which cells have taken up pzNPs and which cells adopt an M1 polarization in mixed populations of pzNP-loaded and -unloaded cells. The overall goal is to leverage this novel cellular assay to help improve understanding of how biological cells respond to bioelectric stimulation.

biophysics↗

bioRxiv2022

Real-time dynamic single-molecule protein sequencing on an integrated semiconductor device

Proteins are the main structural and functional components of cells, and their dynamic regulation and post-translational modifications (PTMs) underlie cellular phenotypes. Next-generation DNA sequencing technologies have revolutionized our understanding of heredity and gene regulation, but the complex and dynamic states of cells are not fully captured by the genome and transcriptome. Sensitive measurements of the proteome are needed to fully understand biological processes and changes to the proteome that occur in disease states. Studies of the proteome would benefit greatly from methods to directly sequence and digitally quantify proteins and detect PTMs with single-molecule sensitivity and precision. However current methods for studying the proteome lag behind DNA sequencing in throughput, sensitivity, and accessibility due to the complexity and dynamic range of the proteome, the chemical properties of proteins, and the inability to amplify proteins. Here, we demonstrate single-molecule protein sequencing on a compact benchtop instrument using a dynamic sequencing by stepwise degradation approach in which single surface-immobilized peptide molecules are probed in real-time by a mixture of dye-labeled N-terminal amino acid recognizers and simultaneously cleaved by aminopeptidases. By measuring fluorescence intensity, lifetime, and binding kinetics of recognizers on an integrated semiconductor chip we are able to annotate amino acids and identify the peptide sequence. We describe the expansion of the number of recognizable amino acids and demonstrate the kinetic principles that allow individual recognizers to identify multiple amino acids in a highly information-rich manner that is sensitive to adjacent residues. Furthermore, we demonstrate that our method is compatible with both synthetic and natural peptides, and capable of detecting single amino acid changes and PTMs. We anticipate that with further development our protein sequencing method will offer a sensitive, scalable, and accessible platform for studies of the proteome.

biophysics↗