Kumari, T. | Search bioRxiv

bioRxiv2024

LRRC8 complexes are adenosine nucleotide release channels regulating platelet activation and arterial thrombosis

Platelet shape and volume changes are early mechanical events contributing to platelet activation and thrombosis. Here, we identify single-nucleotide polymorphisms in Leucine-Rich Repeat Containing 8 (LRRC8) protein subunits that form the Volume-Regulated Anion Channel (VRAC) which are independently associated with altered mean platelet volume. LRRC8A is required for functional VRAC in megakaryocytes (MKs) and regulates platelet volume, adhesion, and agonist-stimulated activation, aggregation, ATP secretion and calcium mobilization. MK-specific LRRC8A cKO mice have reduced arteriolar thrombus formation and prolonged arterial thrombosis without affecting bleeding times. Mechanistically, platelet LRRC8A mediates swell-induced ATP/ADP release to amplify agonist-stimulated calcium and PI3K-AKT signaling via P2X1, P2Y1 and P2Y12 receptors. Small-molecule LRRC8 channel inhibitors recapitulate defects observed in LRRC8A-null platelets in vitro and in vivo. These studies identify the mechanoresponsive LRRC8 channel complex as an ATP/ADP release channel in platelets which regulates platelet function and thrombosis, providing a proof-of-concept for a novel anti-thrombotic drug target.

cell biology↗

bioRxiv2022

Dapagliflozin improves endothelial integrity and hemodynamics in endotoxin treated mice through an apolipoprotein M dependent pathway

RationaleSodium-glucose co-transporter inhibitors (SGLT2i) are under active clinical investigation in patients with acute inflammatory conditions, based on their clinical cardio-and nephroprotective effects, and a pre-clinical study that demonstrated SGLT2i improve renal outcomes and survival in a lipopolysaccharide (LPS) model. However, a unified mechanism that explains how SGLT2i could prevent hemodynamic consequences of inflammatory conditions has not been described. Apolipoprotein M (ApoM) is inversely associated with mortality in inflammatory conditions and improves cardiac function in endotoxin-treated mice via sphingosine-1-phosphate (S1P) signaling. ObjectiveTest the hypothesis that pre-treatment with SGLT2i dapagliflozin (Dapa) improves hemodynamics in endotoxin-treated mice via the ApoM/S1P pathway. Methods and ResultsMice with diet-induced obesity were gavaged with vehicle or Dapa for 4 days prior to LPS (10 mg/kg, IP). We found that mice receiving Dapa restored circulating ApoM levels, likely by increasing expression of the multi-ligand protein receptor megalin in the proximal tubules. Dapa attenuated LPS-induced reductions in cardiac dysfunction including reductions in ejection fraction, cardiac index, and coronary sinus area as well as vascular permeability as ascertained by intravital microscopy. Using both ApoM transgenic and knockout mice and S1P receptor inhibitors, we show that the ApoM/S1P pathway is important for the beneficial effects of Dapa in the LPS model. ConclusionsIn the setting of acute inflammation, our data suggest that SGLT2i maintains levels of megalin, leading to preservation of ApoM, which in turn promotes endothelial barrier integrity and improves hemodynamics. Our studies suggest a novel mechanism by which SGLT2i can preserve intravascular volume in the acute inflammatory setting.

physiology↗