Thrombin stimulates albumin transcytosis in lung microvascular endothelial cells via activation of acid sphingomyelinase

Rationale: Transcellular albumin transport occurs via caveolae which are abundant in lung microvascular endothelial cells. Stimulation of albumin transcytosis by pro-inflammatory mediators may contribute to alveolar protein leak in lung injury, yet the regulation of albumin transport and its underlying molecular mechanisms are so far incompletely understood. Objective: Here, we tested the hypothesis that thrombin may stimulate transcellular albumin transport across lung microvascular endothelial cells in an acid-sphingomyelinase dependent manner. Methods and Results: Thrombin increased the transport of fluorescently labelled albumin across confluent human lung microvascular endothelial cell (HMVEC-L) monolayers to an extent that markedly exceeds the rate of passive diffusion. Thrombin activated acid sphingo-myelinase (ASM) and increased ceramide production in HMVEC-L, but not in bovine pulmonary artery cells which showed little albumin transport in response to thrombin. Thrombin increased total caveolin-1 (cav-1) content in both whole cell lysates and lipid rafts from HMVEC-L and this effect was blocked by inhibition of ASM or de novo protein biosynthesis. Thrombin-induced uptake of albumin into lung microvascular endothelial cells was confirmed in isolated-perfused lungs by real-time fluorescence imaging and electron microscopy of gold-labeled albumin. Inhibition of ASM attenuated thrombin-induced albumin transport both in confluent HMVEC-L and in intact lungs, while HMVEC-L treatment with exogenous ASM increased albumin transport and enriched lipid rafts in cav-1. Conclusions: Our findings indicate that thrombin stimulates transcellular albumin transport in an acid sphingomyelinase-dependent manner by inducing de novo synthesis of cav-1 and its recruitment to membrane lipid rafts.

from #Physiology via Alexandros G.Sfakianakis on Inoreader