Background The cytokine-like activity of the high mobility group box 1 (HMGB1) has recently been implicated in inflammatory cardiovascular diseases, with roles in cardiac hypertrophy reported . Emerging evidence suggests interplay between HMGB1 and the coagulation enzyme thrombin however the molecular mechanism is unclear . This project aims are to unravel the relationship between thrombin-mediated HMGB1 release from platelet-like particles (PLPs) and extracellular vesicles (EVs) with the view to investigate these events during cardiovascular inflammation. Methods A human megakaryoblastic cell line, MEG-01, was used for in vitro studies to assess PLPs and EVs release. To study pathological HMGB1 release, an in vivo chronic hypertension model was used. Wistar male, weight matched rats were randomly assigned into 3 groups, control (n=3), sham (n=4) and angiotensin II (Ang II, 400ng/kg/min, n=6) and monitored pre-post-surgery for 28 days. Following a lethal overdose with Dolethal (1mg/ml/kg), blood and tissues were taken. Dynamic light scattering (DLS) was used to determine the sizes of PLPs and EVs released with confocal microscopy and Western blotting used to determine HMGB1 localisation and expression in MEG-01 and cardiac tissue samples. Key findings Confocal and Western blotting confirmed HMGB1 expression in MEG-01. Thrombin treatment (1U/ml) increased cytosolic HMGB1 expression in MEG-01 cells 15-minute post-treatment (1.222 ± 0.080-fold compared to unstimulated p= 0.0035, n=5). Thrombin-dependent HMGB1 secretion was evident in the PLP and EVs. DLS confirmed that the MEG-01-derived EVs and rat serum-derived EVs were polydisperse (0.488 ± 0.023 and 0.546 ± 0.046, n=5) with a mean diameter 222.9nm and 408.3nm respectively (n=5). Confocal analysis revealed increased HMGB1 cardiac expression and localisation in the Ang II groups, compared to control and saline groups. Discussion and future work Exposure to thrombin led to a time-dependent secretion of HMGB1 from MEG-01 cells and EVs derived from these cells. These events may be a driver for cardiac inflammation. Pilot data from follow up investigation of serum from Ang II rats has revealed stimuli-dependent changes in EV cargo, particularly HMGB1 expression. The role of thrombin in these events is currently the focus of investigation.
|Number of pages||1|
|Journal||British Journal of Pharmacology|
|Early online date||27 Jun 2023|
|Publication status||Published - 31 Jul 2023|
|Event||19th World Congress of Basic & Clinical Pharmacology 2023 - SEC, Glasgow|
Duration: 2 Jul 2023 → 7 Jul 2023
- inflammatory cardiovascular diseases