In Brief: Findings From 5 Publications Harnessing High-Res TRPS Measurements
A brief roundup of findings from 5 studies published in Q2 of 2023 which featured Tunable Resistive Pulse Sensing measurements.
1. EVs derived from hypoxic tumour cells affect neuroblastoma metastasis
While hypoxic signalling is known to influence tumour metastases, including in neuroblastoma, the exact paracrine activity behind it – and the role of extracellular vesicles (EVs) – is not fully understood. Fusco et al. (2023) therefore turned to EVs derived from both normoxic and hypoxic supernatants from neuroblastoma cell cultures. They asked: how does EV-miRNA expression compare across the two groups? Do EVs promote metastatic activity in vitro, and in a zebrafish model? With Tunable Resistive Pulse Sensing (TRPS) measurements of size and concentration part of the EV characterisation toolkit, Fusco et al. came to the following conclusion: EVs from hypoxic neuroblastoma cell cultures (hypoxic EVs) and their miR-210-cargo enrichment appear to play a role in the cellular and microenvironmental changes related to the spread of neuroblastoma. This was based partly on the observation that hypoxic EVs enabled cells to move more quickly in vitro than cells exposed to normoxic EVs, suggesting an increased metastatic invasion capability, findings which were supported by experiments conducted in vivo.1
2. qEV Isolation and TRPS inspected as standardised, reproducible methods for isolating and measuring EVs
In the interest of verifying standardised protocols for the isolation and quantification of EVs from plasma for downstream clinical applications, Diehl et al. (2023) turned to qEV Isolation and TRPS. Both technologies were applied in an analysis of plasma EVs from healthy controls, and from patients with a genetic disorder (Marfan syndrome, with related thoracic aortic aneurysms). They reported the concentration, and the mean and mode diameter of EV-sized particles, alongside an inspection of Western blot markers from EVs and whole cell lysates. TRPS measurements were consistent across replicates and between the two operators, with no differences in EV concentration detected between technicians (median EV concentration detected by Technician 1: 0.81 x 1011 particles/mL, interquartile range 0.59–1.19 x 1011 particles/mL; median EV concentration detected by Technician 2: 0.60 x 1011 particles/mL, interquartile range 0.45–1.10 x 1011 particles/mL; p = 0.606).2
3. Interference with nuclear exporter XPO1 affects EV-RNA cargo profile
It’s widely known that EV heterogeneity depends on many factors including EV-size (a reflection of EV subtype), cell type of origin, and other physiological factors. But what else? Another potential variable lies in the activity of a receptor called exportin-1 (XPO1). As a highly conserved nuclear export receptor, XPO1 binds to proteins containing nuclear export signal sequences, and helps facilitate the export of both proteins and RNAs. With TRPS part of the EV analysis line up, Pálóczi et al. (2023) isolated EVs from conditioned media of different cell lines, either with or without exposure to an XPO1 inhibitor (leptomycin B). Exposing cells to treatments which interfere with XPO1 activity did result in altered RNA profiles, bringing XPO1-mediated RNA transport into the spotlight.3
4. The battle of nanoplastic characterisation continues
The accurate characterisation of nanoplastics, which result from the degradation of bulk plastic litter and intentional use in consumer products, remains a challenge. Using a set of representative test particles of different chemical natures, Huber et al. (2023) describe measurements using a wide range of different analytical methods. Although this is outside the usual application of TRPS, and practical limitations make TRPS an unlikely match for mainstream use (not unique to TRPS: complex, environmental samples are difficult to measure) the high-resolution nature of TRPS translated well to this particular study. Both the Exoid and qNano were used to measure particle size and concentration, highlighting the suitability of TRPS for when high size resolution is needed.4
5. How EVs from skeletal muscle affect macrophage responses
In a study of skeletal muscle-derived EVs, and their impact on macrophage inflammatory responses, TRPS was used to assess EV size distribution and concentration. Specifically, Yamaguchi et al. (2023) showed that myotube-derived EVs exert an anti-inflammatory effect, with RNA-seq results suggesting the upregulation of the IRG1-itaconate pathway, among others.5
- Fusco, P. et al. miR-210-3p enriched extracellular vesicles from hypoxic neuroblastoma cells stimulate migration and invasion of target cells. Cell & Bioscience 13, (2023).
- J. Nathaniel Diehl et al. A standardized method for plasma extracellular vesicle isolation and size distribution analysis. PLOS ONE 18, e0284875–e0284875 (2023).
- Krisztina Pálóczi, Buzás, E. I. & András Falus. Differential impact of exportin-1-mediated nuclear export of RNAs on the RNA content of extracellular vesicle subpopulations. Biologia Futura (2023).
- Huber, M. J. et al. Physicochemical characterization and quantification of nanoplastics: applicability, limitations and complementarity of batch and fractionation methods. Analytical and Bioanalytical Chemistry 415, 3007–3031 (2023).
- Yamaguchi, A. et al. Skeletal myotube-derived extracellular vesicles enhance itaconate production and attenuate inflammatory responses of macrophages. Frontiers in Immunology 14, (2023).